Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareVESICOURETERAL REFLUX - PRESENTATION AND MANAGEMENT AMONG CHILDREN AT A TERTIARY CARE HOSPITAL IN SOUTHWESTERN REGION, SAUDI ARABIA
English0106Abdullah A. MuzallefEnglish Ali M. AlsuheelEnglish Ayed A. ShatiEnglish Saleh M. Al-QahtaniEnglish Zainah A. SabrEnglish Safa M. Al-HaiderEnglish Ahmed H. AlmathamiEnglishObjective: Studying the medically and surgically outcome of Vesicoureteral reflux among pediatric patients in Aseer region. Methodology: This is a retrospective study. We reviewed and analyzed all files for VUR patients who attended the Aseer Central Hospital (ACH) during the period from January 2007 till December 2011. A total of 48 VUR patients were registered. Results: More than half of children with VUR were females (56.3%). The age of these cases ranged from one month to 10 years. Most of these cases were diagnosed at the age of 1-5 years (52.1%), with a mean +SD of 3.93+2.87 years. Conclusions: VUR affects girls more than boys in Aseer Region, starting as early as the first days of life, indicating its congenital etiology. The main presentation for these children is UTI.
EnglishVesicoureteral reflux, Children, Endoscopic deflux injection, Aseer, KSAINTRODUCTION
The Vesicoureteral reflux (VUR) is the retrograde flow of urine from the bladder into the ureter and possibly the kidney according to the grade. Primary VUR is congenital and is not associated with any underling neuromuscular or obstructive phenomena,1 and is usually detected during radiological evaluation of children with urinary tract infection (UTI).2 While in secondary VUR the valvular mechanism is intact and healthy to start with but becomes overwhelmed by raised vesicular pressures associated with obstruction, which distorts the ureterovesical junction. The obstructions may be anatomical or functional. The cause of VUR is a developmental anomaly resulting in an inadequate length of the intravesical submucosal ureter.2 A substantial number of children also have dysfunctional voiding, which may initiate or perpetuate VUR.3 VUR is believed to be present in 1% or less of normal children. Most cases of VUR are diagnosed after occurrence of a UTI. Pediatric clinical practice guidelines recommend screening children for VUR after a UTI.4 In the first seven years, 1.7% of boys and 7.8% of girls have a UTI. Most occur in the first year with a male predominance in the first six months. In uncircumcised boys there is a 10-fold increased incidence.5 Children who have a UTI have a high incidence of VUR, which indicates that VUR predisposes patients to UTI.6 In addition, children who have a UTI and VUR are more likely to have evidence of renal involvement than children without VUR who have a UTI. The principal complications of renal scarring are chronic renal failure and hypertension.7 So, the proper management of VUR should be based on preventing these complications.4 The insult caused by VUR can either be due to the intrusion of infected urine into the renal substance, and this is the most commonly accepted danger related to VUR, or can be due to the abnormal urine pressure exerted by reflux on the papillae, or to abnormal biochemical or immunological reactions caused by the presence of bladder urine in the renal parenchyma. Not one but four factors may play a role in the pathophysiology of VUR: the virulence of bacteria and their reservoirs, incompetent vesico-ureteric junction, renal parenchyma and bladder and bowels dysfunctions.8
Scar formation in children with VUR after UTIs is an important cause of secondary hypertension and can cause chronic renal failure. Risk factors for renal scar formation include higher grade VUR and delay in UTI diagnosis and treatment. The use of prophylactic antibiotics to prevent UTIs also may decrease the risk of scar formation.7 This study aims to identify the pattern of VUR, outcome after deflux injection and antibiotic prophylaxis and complications of VUR among the children population of Aseer Region, Kingdom of Saudi Arabia (KSA) as single center study. MATERIAL AND METHODS This is a retrospective study. We reviewed and analyzed all files for VUR patients who attended the Aseer Central Hospital (ACH) during the period from January 2007 till December 2011. A total of 48 VUR patients were registered. Voiding cystourethrography (VCUG) was used to confirm the diagnosis of VUR. A dimercaptosuccinic acid (DMSA) renal scan is used to evaluate for any renal scar. Until the reflux resolves or the reflux is surgically treated, the patient should undergo monitoring with cystography (VCUG) every 12-24 months.9 Aseer Central Hospital (ACH) is the tertiary care hospital that serves all referred patients in Aseer Area. VUR was graded according to Dähnert as follows:10 • Grade I: Urine backs up into the ureter only, and the renal pelvis appears healthy, with sharp calyces. • Grade II: Urine backs up into the ureter, renal pelvis, and calyces. The renal pelvis appears healthy and has sharp calyces. • Grade III: Urine backs up into the ureter and collecting system. The ureter and pelvis appear mildly dilated, and the calyces are mildly blunted. • Grade IV: Urine backs up into the ureter and collecting system. The ureter and pelvis appear moderately dilated, and the calyces are moderately blunted. • Grade V: Urine backs up into the ureter and collecting system. The pelvis is severely dilated, the ureter appears tortuous, and the calyces are severely blunted, see (Figure -1).
RESULTS
More than half of children with VUR were females (56.3%). The age of these cases ranged from one month to 10 years. Most of these cases were diagnosed at the age of 1-5 years (52.1%), with a mean +SD of 3.93+2.87 years, as shown in Table (1). Presentation was mainly symptoms of UTI (79.2%) and the rest was accidentally discovered to have VUR who were diagnosed to have hydronephrosis antenatally. The left side was more affected (39.6%) than the right side (22.9%). However, 37.5% of cases had bilateral VUR. Half of cases had grade III VUR (50%), while 18.8% had grade IV and 12.5% had grade V. Ultrasonography showed abnormal findings in 45.8% as a first tool for screening which was showing hydronephrosis. About one third of cases were managed medically (37.5%) by using antibiotics prophylaxis alone (amoxicillin was used for infant less than 6 weeks and co-trimoxazole for whom older than 6 weeks), while 62.5% were managed medically by using antibiotics prophylaxis and surgically by injecting bulking agent (deflux) underneath the intravesical portion of the ureter in a submucosal location, either once (50%) or more than once (12.5%), as shown in Table(3). Patients who required deflux injection more than once were having neurogenic bladder, three of them was due to mylomeningocele and others were idiopathic. All patients were continued on antibiotics and outpatient follow-up. VCUG and renal scan were done during follow up after 6-12 months from the procedure date. VCUG showed residual reflux in 37.5% of cases. DMSA revealed renal scarring in 50% of cases. Chronic kidney disease was present in 25% (12) in form of proteinuria and rising of urea and creatinine, and 8.3% (4) were hypertensive, as shown in figure (1). Comparing the male patients with female patients revealed a higher prevalence of renal scarring among females than males (59.3% vs. 38.1%, respectively). Incidence of recurrent UTI was higher among males (66.7%) than females (48.1%) after Deflux. Voiding cystourethrogram (VCUG) showed a significantly higher prevalence of reflux among females than males (51.9% vs. 19%, respectively, p=0.020), as shown in Table (4).
DISCUSSION
VUR is a congenital abnormality, which affects approximately 1% of infants and children. It may predispose a child with UTI to the development of pyelonephritis, which may lead to renal scarring and hypertension.11 It is the most common hereditary disorder of the genitourinary tract and some transmitted in an autosomal dominant fashion.12 This study showed an early age at diagnosis of VUR (mean age is 3.93 years) and also the more incidence among girls (56.3%) than boys (43.7%). These findings in Aseer, KSA are similar to those reported in other countries. Sharbaf et al. (2007), in Iran, reported a mean age at VUR diagnosis of 4.1 years, with a range from 54 days to 16 years and the male-to-female ratio was 0.21 (girls were 262 while boys were 57). Leroy et al. in England, studied 117 VUR children (age range, 0.0–13.9 years) of whom, 46 (39.3%) were boys and 71 (60.7%) were girls.13 Greenbaum and Mesrobian noted that girls are more commonly diagnosed with VUR because they are more likely to have a UTI.5 Wald explained the early presentation of VUR cases by its congenital nature. He added that these cases can be diagnosed in utero by ultrasonography.14 Experimental studies suggest that abnormal insertion of the ureteral bud could induce abnormal differentiation of metanephros (“renal dysplasia”).8 Smellie et al. added that VUR has a natural tendency to resolve as the intravesical part of the ureter lengthens with growth. By age 10 about 75% of VUR has resolved.15 This study showed that the main presentation for VUR cases was that of UTI (79.2%). This finding is in accordance with that reported by several authors. VUR and UTI are often associated. 16 Eighty eight percent of urine culture performed in children with VUR have sterile urine at the time of diagnosis which does not confirm that UTI causes VUR, and 60% of febrile UTI have no demonstrable VUR. Mild VUR does not increase the incidence of UTI, pyelonephritis, or renal scarring after acute pyelonephritis.17 Williams et al. stated that VUR is diagnosed in 20-30% of children with a first UTI.18 Godley noted that VUR and UTI seem to be independent pathological factors that may potentiate each other.16 The main infection pathway is the ascent from the lower to the upper urinary tract although rarely demonstrated. Other pathways have been suggested such as hematogenous or lymphatic, but none have been validated yet.8 Results of this study indicated that VUR affected the left side more than the right side, while, 37.5% of VUR cases were bilateral. This is in accordance with that reported by Jang et al., who found that 32% of VUR cases occurred on the left side, 27.9% were on the right side while 41.1% were bilaterally affected.19 This study revealed that 4.2% of VUR cases had grade I, 14.6% had grade II, 50% had grade III, while 18.8% had grade IV and 12.5% had grade V. In Korea, the series of VUR cases of Jang et al. were classified as VUR grade I in 9.2%, grade II in 36.4%, grade III in 27.7, grade IV in 16.5, and grade V in 10.1%. Differences in VUR grades among different studies may be due to variable study selection criteria and timing of cases presentation (i.e., early or late).19 It is to be noted that grades of VUR may constitute an important prognostic variable. Greenbaum and Mesrobian reported that rate of VUR resolution varies.4 Grades I and II reflux eventually cease in more than 80% of affected ureters, with a resolution rate of 10% to 25% per year. Grade III reflux resolves in more than 50% of cases, and grade IV VUR resolves in approximately 30% of cases. Grade V reflux is unlikely to resolve spontaneously. In the present study, imaging diagnostic modalities were applied. Ultrasonography showed abnormal findings in 45.8%which was done as first tool for workup of a patient with UTI . DMSA revealed renal scarring in 50% of cases. Several recent studies discussed the role and benefits of radiography in VUR. Demède and Mouriquand stated that when the renal sonography is normal, the cystography should be reserved for those cases where an abnormal DMSA scan is found or if surgery is contemplated.8 When the renal sonography is abnormal, a cystography should be performed in all infants irrespective of the findings of the DMSA scan. Leroy et al. added that renal ultrasonographic findings are believed to be good predictors of VUR. 13 Craig et al. noted that the American Academy of Pediatrics recommends ultrasonography and either voiding cystourethrography or radionuclide cystography.20 Given the good prognosis for children with VUR and the absence of good evidence for improved outcomes, the invasive, unpleasant nature of a VCUG outweighs the possible benefit of prophylactic treatment.18 Farhat et al. noted that the widespread use of routinely performing prenatal ultrasonography has led to more frequent detection of antenatal hydronephrosis. When screened, approximately 10% to 20% of these children proved to have VUR.21 Howard et al. stated that the correlation between renal scarring and VUR has been shown to vary between 23% and 75% and is higher in patients with a high grade of VUR.22 So, the high prevalence of renal scarring among VUR patients in the present study may be explained by several reasons, i.e., delayed presentation, late diagnosis and management as well as a high VUR grade among cases. Results of this study indicated that Chronic kidney disease was present in 25% (12) of VUR cases in form of proteinuria, raising of urea and creatinine, while 8.3% (4) were hypertensive as showed in figure (1) . McLaren et al. noted that VUR may result in hypertension and end-stage renal disease.23 Rodriguez et al. explained that VUR may predispose a child with a bladder infection to the development of pyelonephritis, which may lead to renal scarring and hypertension.11 Similarly, Lim emphasized that the current standards of care in the United States recommend that infants and young children with first-time UTI undergo imaging tests to evaluate for VUR.24 The severity of VUR is thought to correlate to the risk of developing permanent renal scarring that may lead to serious sequelae later in life, such as hypertension, proteinuria or end-stage renal disease. Interventions, such as follow-up imaging, antibiotic prophylaxis and surgical correction are thought to reduce the incidence of these complications. Sharbaf et al. added that complication such as hypertension and renal failure are common complication of VUR, therefore arterial blood pressure and renal function need to be continuously monitored in these patients.2 About one third of cases in the present study were managed medically, while two thirds were managed medically and surgically by anti-reflux surgery using injection of bulking agent (deflux ), either once or more than once. It was noticed that the main reason for deflux injection failure was due to neurogenic bladder. Callewaert stated that surgical treatment of VUR as well as medical treatment by prophylactic antibiotics prevent infections and renal scarring.25 Gill et al. noted that surgical techniques for VUR apply the basic principle of creating an anti-reflux mechanism by increasing the portion of the distal ureter lying in a submucosal tunnel between the detrusor muscle and the bladder mucosa.26 Sharbaf et al. added that anti-reflux surgery offers no short-term advantages other than abolishing the reflux. It also does not result in improved renal function or renal growth, and does not affect the rate of new scar formation or the incidence of hypertension.2 Comparing the male patients with female patients revealed a higher prevalence of renal scarring among girls than boys, higher incidence of recurrent UTI among boys, higher prevalence of kidney abnormalities among girls and a significantly higher prevalence of residual reflux among girls. Variable and contradicting results were reported by different authors. Sharbaf et al. stated that VUR is more common, yet less severe, in girls than boys.2 They also reported that incidence of renal scaring is significantly higher among boys than girls. Alova and Lottmann reported that renal parenchymal lesions were higher among boys than girls.27 Greenbaum and Mesrobian noted that girls with VUR are more likely to have UTI than boys,4 while Lim reported that incidence of UTI is higher among boys than girls.25 This variability in reported results may be due to differences among studies in sampling and methodology. In conclusion, VUR affects more girls than boys in Aseer Region, starting as early as the first days of life, indicating its congenital etiology. The main presentation for these children is UTI. Most cases have high grades of VUR (i.e., grades III and higher), and prevalence of renal scarring is quite high, indicating a late presentation. Imaging is important in assessment of VUR cases. Girls experience significantly higher residual reflux than boys. It is recommended to routinely screen by ultrasound all children with UTI for VUR. The awareness of general practitioners regarding VUR should be raised and they should be advised to refer immediately any child with UTI to a pediatrician. Abbreviations VUR: Vesicoureteral reflux UTI: Urinary tract infection VCUG: Voiding cystourethrography DMSA: 99 m Technetium-dimercaptosuccinic acid
ACKNOWLEDGEMENT
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors/editors/publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed.
Source of Funding: None
Conflict of Interest: None
Englishhttp://ijcrr.com/abstract.php?article_id=679http://ijcrr.com/article_html.php?did=6791. Rushton HG. Vesicouretral reflux and scaring. In: Avner ED, Harman WE, Niaudet P. Pediatric Nephrology. 5th edn, Philadelphia Lippincott Williams and Wilkins, 2004; 1027- 1048.
2. Sharbaf FG, Fallahzadeh MH, Modarresi AR, Esmaeili M. Primary Vesicoureteral Reflux in Iranian Children. Indian Pediatrics 2007; 44:128-130.
3. Herndon CDA, DeCambre M, McKenna PH. Changing concepts concerning the management of vesicoureteral reflux. J Urol 2001;166:1439–43.
4. Greenbaum LA, Mesrobian HO. Vesicoureteral Reflux. Pediatr Clin N Am 53 (2006) 413– 427.
5. Chand DH, Rhoades T, Poe SA, et al. Incidence and severity of vesicoureteral reflux in children related to age, gender, race and diagnosis. J Urol 2003; 170:1548–50.
6. Cleper R, Krause I, Eisenstein B, et al. Prevalence of vesicoureteral reflux in neonatal urinary tract infection. Clin Pediatr (Phila) 2004;43(7):619– 25.
7. Ardissino G, Dacco V, Testa S, et al. Epidemiology of chronic renal failure in children: data from the ItalKid project. Pediatrics 2003;111(4 Pt 1):e382-7.
8. Demède D, Mouriquand P.Vesicoureteral Reflux: Why we Can’t Agree on its Management! An Evidence Based Approach. Arch Esp Urol, 2008; 61 (2):160-6.
9. Estrada JR CR. Vesicoureteral Reflux. http://emedicine. medscape.com/article/439403-overview. date retrieved: 13 November 2013
10. Dähnert W. Radiology Review Manual. Lippincott Williams and Wilkins, 2011. 1
1. Rodriguez E Jr, Weiss DA, Copp HL. Adherence to Antibiotic Prophylaxis in Children with Vesicoureteral Reflux. Advances in Urology 2011; vol. 2011, Article ID 134127, 6 pages, 2011.
12. Kaefer M, Curran M, Treves ST. Sibling vesicoureteral reflux in multiple gestation births. Pediatrics 2000; 105: 800- 804.
13. Leroy S, Vantalon S, Larakeb A, Ducou-Le-Pointe H, Bensman A. Vesicoureteral Refl ux in Children with Urinary Tract Infection: Comparison of Diagnostic Accuracy of Renal US Criteria. Radiology 2010; 255(3): 890-98.
14. Wald ER. Vesicoureteral Reflux: The Role of Antibiotic Prophylaxis. Pediatrics 2006;117;919-22.
15. Smellie JM, Jodal U, Lax H, Writing Committee for the International Reflux Study in Children (European Branch), et al. Outcome at 10 years of severe vesicoureteric reflux managed medically: report of the international reflux study in children. J Pediatr 2001; 139:656–63.
16. Godley ML. Vesicoureteral reflux: Pathophysiology and experimental studies. Gearhart JP, Rink RC, Mouriquand PDE, eds. Pediatric Urology, Philadelphia: W.B. Saunders Company, pág. 359-381, 2001.
17. Garin EH, Olavarria F, Nieto VG, et al. Clinical significance of primary vesicoureteral reflux and urinary antibiotic prophylaxis after acute pyelonephritis: A multicenter, randomized, controlled study. Pediatrics, 117: 626, 2006.
18. Williams G, Fletcher JT, Alexander SI, Craig JC. Vesicoureteral reflux. J Am Soc Nephrol 2008; 19 (5): 847 – 862.
19. Jang HC, Lee KH, Park JS. Primary Vesico-Ureteral Reflux: Comparison of Factors between Infants and Children. Korean J Urol 2011;52:206-209
20. Craig JC, Irwig LM, Knight JF, Roy LP: Does treatment of vesicoureteric reflux in childhood prevent end-stage renal disease attributable to reflux nephropathy? Pediatrics 105: 1236–1241, 2000
21. Farhat W, McLorie G, Geary D, et al. The natural history of neonatal vesicoureteral reflux associated with antenatal hydronephrosis. J Urol 2000;164(3 Pt 2):1057 –60.
22. Howard RG, Roebuck DJ, Yeung PA, Chan KW, Metreweli C. Vesicoureteric reflux and renal scarring in Chinese children. Br J Radiol. 2001;74:331–334.
23. McLaren CJ, Simpson ET. Vesico-ureteric reflux in the young infant with follow-up direct radionuclide cystograms: the medical and surgical outcome at 5 years old. BJU Int. 2002;90:721–724.
24. Lim R. Vesicoureteral Reflux and Urinary Tract Infection: Evolving Practices and Current Controversies in Pediatric Imaging. AJR 2009; 192:1197–1208
25 Callewaert PRH. What is new in surgical treatment of vesicoureteric reflux? Eur J Pediatr 2007; 166:763–768.
26. Gill IS, Ponsky LE, Desai M, Kay R, Ross JH (2001) Laparoscopic cross-trigonal Cohen ureteroneocystostomy: novel technique. J Urol 166(5):1811–1814.
27. Alova I, Lottmann HB. Vesico ureteral reflux and elimination disorders. Arch Esp Urol 2008; 61(2): 218-28.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareNANOTECHNOLOGY - A NEW ERA IN MEDICINE AND DIAGNOSTICS
English0710Evarisalin MarbaniangEnglish DonboklangEnglishAn era of new and constantly advancing techniques is revolutionizing our world in the present times. We are in a phase where technology is evolving at its best. In the field of medicine and particularly, in the diagnostic realm ,we have seen the developing molecular diagnostics e.g polymerase chain reaction (PCR), hybridization techniques (flourescent in situ hybridization,etc), array based comparative genomic hybridization, so on and so forth. We are again fortunate to witness the evolution of another technology termed as NANOTECHNOLOGY. It has a wide variety of applications in diagnostics, cancer treatment, drug delivery, and tissue engineering. It is a vast field of modern science where it can be applied in organic chemistry, nuclear reactors, robotics, space applications, telecommunications, satellites, heavy industry and even cosmetics. The list goes on, but a word of caution as we humans are always crossing boundaries and may be using the technology for ignoble causes. So it is up to us to turn nanotechnology into a blessing or a curse.
EnglishNanoparticles, Nanosensors, Magnetic nanoparticlesINTRODUCTION
“Nanotechnology” was first defined by Tokyo Science University, Norio Taniguchi in 19741 . Although the application of nanotechnology to medicine appears to be a relatively recent trend, the basic nanotechnology approaches for medical application dates back to several decades2 . Lipid vesicles which were named as liposomes, were described in 19653 . Nanotechnology can be defined as the science and engineering involved in the design, synthesis, characterization, and application of materials and devices whose smallest functional organization, in at least one dimension, is on the nanometer scale or one billionth of a meter. At these scales, consideration of individual molecules and interacting groups of molecules in relation to the bulk macroscopic properties of the material or device becomes important, as it has a control over the fundamental molecular structure, which allows control over the macroscopic chemical and physical properties4 . These technologies include nanoarrays, protein arrays, nanopore technology, nanoparticles (NPs) as a contrivance in immunoassays and nanosensors, among others. Gold NPs and quantum dots (semiconductors) are the most widely used, but new materials are becoming available as more molecular entities are discovered as amenable to nanoscale design and fabrication. Crystal materials like those of gallium, phosphate, quartz, and ceramic are chosen for their durability and piezoelectric properties of developing and retaining an electric potential (charge) when subjected to mechanical stress. Another area of development is nanobiosensors, in which antibody-based piezoelectric nanobiosensors are well developed5-6. Nanotechnology is rapidly evolving and is taking medicine forward in which cancer treatment, diagnostics and research has been reduced to a nanoscale. These applications include fluorescent biological labels, drug and gene delivery, bio-detection of pathogens, detection of protein, probing of DNA structure, tissue engineering, tumor detection, separation and purification of biological molecules and cells, MRI contrast enhancement and phagokinetic studies7 .
APPLICATIONS
1. Viral diagnostics - The use of nanoparticles as tags or labels allows for the detection of infectious agents in small sample volumes directly in a very sensitive, specific, and rapid format at lower costs than current in-use technologies8 . A microfluidic platform-based detection system is now described. The term refers to precise control and manipulation of fluids contained typically in sub-millimetre scale volume. The assay can detect the avian influenza virus H5N1 in throat swab samples by using magnetic forces to manipulate a free droplet containing superparamagnetic particles (ferric oxide-labelled antibody) to concentrate the viruses9 . A method for the respiratory syncytial virus (RSV) consisting of functionalized NPs conjugated to monoclonal antibodies can be used to rapidly and specifically detect RSV in clinical samples with a great degree of sensitivity10. A new nanoparticle-based biobarcode amplification (BCA) assay has been developed for early and sensitive detection of human immunodeficiency virus (HIV)-1 capsid (p24) antigen11. The hepatitis B virus (HBV), hepatitis C virus (HCV), and HBV/HCV gene chips with gold/silver NP staining amplification method were shown to be useful in detecting these viruses in patients’ samples12. An array-based nano-amplification technique method for the detection of hepatitis E virus (HEV) has been developed by utilizing nano-gold-labelled oligonucleotide probes coupled with silver stain enhancement and the microarray technique. The microarray was shown to detect 100 fM of amplicon with the image development time as short as 2 minutes. A similar technique is also described for hepatitis A virus (HAV) 13-14. Detection of herpes simplex virus (HSV)-1 virus particles is achieved by exposing the virus-containing sample to a sensor surface coated with a specific antibody against HSV-1. The Young interferometer sensor was shown to detect HSV-1 at very low concentrations (850 particles/ mL) and even directly in serum samples5 . A novel signal amplification technology for a human papillomavirus (HPV)-DNA hybridization assay based on fluorescein diacetate (FDA) nanocrystals has been developed. This approach resulted in high selectivity, short incubation times, and high sensitivity. This innovative method allows rapid detection of small amounts of target sequence in a fewer number of PCR cycles15. Diarrhoea-causing viruses today are a major public health concern, and newer agents with potential for large food-borne outbreaks have been described. Norovirus is a leading cause of gastroenteritis in many parts of the world. A matrix-assisted laser desorption ionization and nanospray mass spectrometry was developed and evaluated for norovirus detection using different approaches16. 2. Molecular imaging has emerged as a powerful tool to visualize molecular events of an underlying disease, sometimes prior to its downstream manifestation. The merging of nanotechnology with molecular imaging provides a versatile platform for the novel design of nanoprobes that will have tremendous potential to enhance the sensitivity, specificity and signalling capabilities of various biomarkers in human diseases17. Nanoparticle probes can endow imaging techniques with enhanced signal sensitivity, better spatial resolution and the ability to relay information on biological systems at molecular and cellular levels. Simple magnetic nanoparticles can function as magnetic resonance imaging (MRI) contrast enhancement probes. These magnetic nanoparticles can then serve as a core platform for the addition of other functional moieties including fluorescence tags, radionuclides and other biomolecules, for multimodal imaging, gene delivery and cellular trafficking. An (MRI) with hybrid probes of magnetic nanoparticles and adenovirus can detect target cells and monitor gene delivery and expression of green fluorescent proteins optically18. Nuclear techniques such as positron-emission tomography (PET) potentially provide detection sensitivities of higher magnitude, enabling the use of nanoparticles at lower concentrations than permitted by routine MRI. Furthermore, a combination of the high sensitivity of PET with the anatomical detail provided by computed tomography (CT) in hybrid imaging, has the potential to map signals to atherosclerotic vascular territories19. 3. Drug delivery-Nanotechnology plays an important role in advanced biology and medical research particularly in the development of potential site specific delivery systems with lower drug toxicities and greater efficiencies20. The era of nanotechnology has allowed novel research strategies to flourish in the field of drug delivery. Nanotechnology designed drug delivery systems have been seen to be suitable for treating chronic intracellular infections21. One of the major applications of nanotechnology in relation to medicine is drug delivery. The problems with the new chemical entities such as insolubility, degradation, bioavailability, toxicologic effects, targeted drug delivery, and controlled drug release are solved by nanotechnology. For example, encapsulated drugs can be protected from degradation. Specific nanosized receptors present on the surface of the cell can recognize the drug and elicit appropriate response by delivering and releasing the therapy exactly wherever needed. Because of their small size and large surface area relative to their volume, nanoscale devices can readily interact with biomolecules. Nanoscale devices include: nanoparticles, nanotubes, cantilevers, semiconductor nanocrystals, and liposomes22.
APPROVED PRODUCTS OF NANOTECHNOLOGY:
Due to the stringent food and drug act (FDA) regulations, only a few products based on nanotechnology are available for clinical use. Doxil® (Centocor Ortho Biotech Products L.P, New Jersey, USA.) and Abraxane® (Abraxis Bioscience, Los Angeles, USA.) are among the two available for clinical use23.
CANCER DETECTION AND TARGETING :
Detection and targeting of the cancerous tissues or cells have always been a challenge to the formulator. Cancerous tissues or cells being self becomes very difficult to target the specific cells or organs; as a result, many normal cells are being killed in the process. Many devices based on nanotechnology have come to the rescue of the formulators, wherein, using biomarkers, the anticancer agents can be targeted only to specific cells or organs24-25. One such method to detect cancer is use of Photodynamic Therapy (PDT) using 5-aminolaevulinic acid which is metabolized in body to protoporphyrin IX which is a photosensitizer26. Quantum dots (QD) are very useful in lymph node mapping which is an important technique for cancer mapping during surgery and in vivo cancer imaging using semiconductor QD is also well documented in the literature27-28.
DISCUSSION
Nanotechnology is a part of science where the engineer and the scientist functions together as a unit. The potential of its applications are so diversified that it is not possible to address all of them here. In this review, we are dealing more with the nanomedicine aspects i.e the medical applications of nanotechnology. Due to nanoscale effects and increased surface area, nano-materials have been investigated as promising tools for the advancement of diagnostic biosensors, drug and gene delivery, and biomedical imaging29. The use of nanoparticles as tags or labels allows for the detection of infectious agents in small sample volumes directly in a very sensitive, specific, and rapid format at lower costs than current in-use technologies. This advance in early detection enables accurate and prompt treatment. The quantum dot technology is currently the most widely employed nanotechnology. The technology strengthens and expands the DNA and protein microarray methods and has applications in genomic analysis, proteomics, and molecular diagnostics. Nanosensors are the new contrivance for detection of bioterrorism agents8 . Imaging modalities like MRI etc with low sensitivity, nanoparticles bearing multiple contrast groups provide signal amplification. The same nanoparticles can, in principle, deliver both the contrast medium and the drug, allowing monitoring of the bio-distribution and therapeutic activity simultaneously (referred to as theranostics)30. Such nanofiber-based scaffolds are available in a wide range of pore size distribution, high porosity and high surface area-to-volume ratio. Such a wide range of parameters are favourable for cell attachment, growth and proliferation, and also provide a basis for the future optimization of an electrospun nanofibrous scaffold in a tissue-engineering application. The genesis of nanotechnology can be traced to the promise of revolutionary advances across medicine, communications, genomics and robotics31. Target specific nature of the delivery systems developed applying nanotechnology principles have been able to reduce the amount of drug that needs to be loaded and hence prevent many dose-related adverse reactions. Currently, not many products are available for clinical use, but looking at the amount of research activity happening in this field, the next few years will witness the outburst of nano-medical devices, therapeutic aids, and many products being launched in the market for clinical use22. Nanotechnology is a multifaceted weapon as its applications is evolving and developing in many fields of medicine, both diagnostic and therapeutic especially targeted therapy for the treatment of cancer. In view of its varied uses, the need arises for more research and trials along with regulations so the suffering of mankind may be alleviated in this already disease filled universe.
CONCLUSION
The future of medicine is bright and hopeful with the latest technologies particularly nanotechnology emerging to give us an optimistic view of solutions relating to diseases and their dilemmas. Its utility in organic chemistry, nuclear reactors, robotics, space applications, telecommunications, satellites, heavy industry and even cosmetics has been documented. Nanotechnology is here to stay and its uses and applications are unlimited provided we use it wisely and ethically to benefit mankind.
ACKNOWLEDGEMENT
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors / editors / publishers of all those articles, jour-nals and books from where the literature for this article has been reviewed and discussed. Author(s) contributions: Evarisalin Marbaniang conceived and wrote the review, Donboklang Lynser gathered materials and together contributed equally in writing the paper.
Source of funding: nil
Conflict of Interest: None
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Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareA DETAILED STUDY OF HYPEREOSINOPHILIA OR SEVERE EOSINOPHILIA IN PATIENTS WITH HOOKWORM INFECTION FOUND WHILE DOING UPPER GASTRO INTESTINAL ENDOSCOPY IN A TERTIARY CARE HOSPITAL
English1115Govindarajalu GanesanEnglishObjective: Eosinophilia occurs very commonly in hookworm infection. But so far detailed study was not done to know about the occurrence of hypereosinophilia in various grades of hookworm infection with or without anaemia. Hence a detailed study was done to know about the occurrence of hypereosinophilia in various grades of hookworm infection found in patients while doing upper gastro-intestinal endoscopy in our institute. Methods: A study of 1259 patients who had undergone upper gastro-intestinal endoscopy for a period of 5 years from May 2009 to April 2014 was carried out in our institute. In each of these 1259 patients, the first and second part of duodenum were carefully examined to find out the presence of hookworms. In all the patients found to have hookworms in duodenum, investigations were done to know about the presence or absence of hypereosinophilia and anaemia except in the very few patients who were lost for follow up. The results were found as given below. Results: Of these 1259 patients, as many as 18 patients were found to have hookworms in duodenum while doing upper gastrointestinal endoscopy. Of these 18 patients, 4 patients were lost for follow up and full details about their investigations were not available. The remaining 14 patients were taken into consideration for our study. Of these14 patients, as many as 10 patients were found to have eosinophilia(71%). Severe eosinophilia or hypereosinophilia was found in 4 out of 10 patients with eosinophilia(40%).3 out of 4 patients with hypereosinophilia with hookworm infection in our study had anaemia, but one patient with hypereosinophilia did not have anaemia. Conclusion: Hence, hypereosinophilia is an extremely important indicator of hookworm infection with or without anaemia. Hence upper gastro-intestinal endoscopy should be done to confirm the presence of hookworms in all patients with hypereosinophilia even when there is no anaemia in tropical and subtropical countries.
EnglishHypereosinophilia, Hookworm infection, Upper gastro-intestinal endoscopy.INTRODUCTION
Eosinophilia occurs very commonly in hookworm infection (1to13). But so far detailed study was not done to know about the occurrence of hypereosinophilia in various grades of hookworm infection with or without anaemia. Hence a detailed study was done to know about the occurrence of hypereosinophilia in various grades of hookworm infection found in patients while doing upper gastro-intestinal endoscopy in our institute.
MATERIALS AND METHODS
This study was conducted in the department of general surgery, Aarupadai Veedu Medical College and Hospital, Puducherry. A study of 1259 patients who had undergone upper gastro-intestinal endoscopy in our institute for a period of 5 years from May 2009 to April 2014 was carried out. In each of these 1259 patients,the first and second part of duodenum were carefully examined to find out the presence of single or multiple hookworms. In all the patients found to have hookworms in duodenum,investigations were done to know about the presence of hypereosinophilia and anaemia except in the very few patients who were lost for follow up. Anaemia is defined as haemoglobin < 12g/dl or 12g% in women and haemoglobin < 13g/dl or13g% in men. Mild anaemia is taken as haemoglobin 10to12g/dl or g%, moderate anaemia is taken as haemoglobin 7 to10g/dl or g% and severe anaemia is taken as haemoglobin or = 500 cells/cu.mm (14). Severe eosinophilia or hypereosinophilia is defined as eosinophils>1000 cells/cu.mm (4). The results were found as given below.
RESULTS
1. Of these 1259 patients, as many as 18 patients were found to have hookworms in duodenum while doing upper gastro-intestinal endoscopy.
2. Of these 18 patients, 4 patients were lost for follow up and full details about their investigations were not available. The remaining 14 patients were taken into consideration for our study.
3. Of these14 patients, as many as 10 patients were found to have eosinophilia(71%).
4. Severe eosinophilia or hypereosinophilia was found in 4 out of 10 patients with eosinophilia(40%).
5. Of the 4 patients with hypereosinophilia or severe eosinophilia,
a. One patient did not have anaemia.
b. One patient had mild anaemia.
c. One patient had moderate anaemia.
d. One patient had severe anaemia.
Hence the 4 patients with hypereosinophilia or severe eosinophilia can be divided into 4 groups as follows
a. Hypereosinophilia or severe eosinophilia without anaemia( 1patient) . b. Hypereosinophilia or severe eosinophilia with mild anaemia( 1 patient). c. Hypereosinophilia or severe eosinophilia with moderate anaemia( 1 patient). d. Hypereosinophilia or severe eosinophilia with severe anaemia( 1 patient).
a. Hypereosinophilia or severe eosinophilia without anaemia [1patient]
1. One patient with hypereosinophilia with hookworm infection did not have anaemia(haemoglobin 18g%, absolute eosinophil count - 1000cells/ cu.mm) which is of extremely great significance.
2. This patient with hypereosinophilia had adequate amount of haemoglobin(18g%) indicating that there was only very minimal loss of blood due to very early stage of hookworm infection.
3. Thus hypereosinophilia or severe eosinophilia can be present when there is no anaemia due to very early stage of hookworm infection.
4. Hence hypereosinophilia or severe eosinophilia can be the earliest sign of hookworm infection and can be present in the very early stage of hookworm infection even before the onset of anaemia. b. Hypereosinophilia or severe eosinophilia with mild anaemia [1 patient]. 1. Hypereosinophilia or severe eosinophilia occurring along with mild anaemia was present in one patient in our study(haemoglobin 11.7g%, absolute eosinophil count - 1248cells/cu.mm). 2. Multiple hookworms in duodenum with hypereosinophilia or severe eosinophilia[absolute eosinophil count - 1248cells/cu.mm)] and with mild anaemia( haemoglobin 11.7g %) seen in this patient indicating mild hookworm infection due to mild anaemia despite multiple hookworms is shown in Fig1, 2. c. Hypereosinophilia or severe eosinophilia with moderate anaemia [1 patient]. 1. Hypereosinophilia or severe eosinophilia with moderate anaemia was present in one patient in our study( haemoglobin 8.6g%, absolute eosinophil count - 1260cells/cu.mm) . 2. Moderate anaemia is due to significant loss of blood due to more number of hookworms and is associated with stage of moderate hookworm infection. 3. One study has also shown the occurence of moderate anaemia with hypereosinophilia or severe eosinophilia (15).
d. Hypereosinophilia or severe eosinophilia with severe anaemia [1 patient].
1. Severe anaemia is due to significant loss of blood due to large number of hookworms and very heavy burden of hookworm infection and indicates late stage of hookworm infection or stage of severe hookworm infection.
2. Hypereosinophilia or severe eosinophilia with severe anaemia was present in one patient in our study(haemoglobin 3.2g%, absolute eosinophil count - 1100cells/cu.mm) .
3. Multiple hookworms in duodenum with hypereosinophilia or severe eosinophilia(absolute eosinophil count - 1100cells/cu.mm) and with severe anaemia (haemoglobin 3.2 g%)indicating late stage of hookworm infection or stage of severe hookworm infection is shown in Fig 3, 4.
4. Other studies have also shown the occurence of severe anaemia with hypereosinophilia or severe eosinophilia (2, 9, 13, 22).
5. But stool examination was negative for hookworm ova in this patient despite heavy burden of hookworm infection with severe anaemia. Many studies have also shown negative stool examination for hookworm ova despite heavy burden of hookworm infection with severe anaemia (1, 9, 13, 16 to22).
6. Hypereosinophilia or severe eosinophilia persisted in this patient throughout the period of 6 months despite her improvement of haemoglobin to 9.2g% and negative stool examination for hookworm ova .
7. This is of extremely great significance since persistent hypereosinophilia or severe eosinophilia was the only indicator of hookworm infection due to which upper gastro-intestinal endoscopy was done and hookworm infection was confirmed in this patient.
DISCUSSION
Persistent hypereosinophilia helping in the diagnosis of hookworm infection which was greatly delayed due to persistent negative stool examination for hookworm ova
In this female patient with severe anaemia and hypereosinophilia or severe eosinophilia(hemoglobin 3.2g%, absolute eosinophil count - 1100cells/cu.mm) , upper gastro intestinal endoscopy was done only after 6 months of the initial presentation since repeated stool examination done in these 6 months were persistently negative for hookworm ova in this patient. At her initial admission , despite severe anaemia and hypereosinophilia, upper gastro intestinal endoscopy was not done since the stool examination was negative for hookworm ova. She was treated with two pints of blood transfusion and haematinics and her haemoglobin improved significantly to 9.2g% after 6 months. But stool examination was again negative for hookworm ova even after 6 months. But despite her improved haemoglobin of 9.2g% and persistent negative stool examination for hookworm ova, her hypereosinophilia or severe eosinophilia( absolute eosinophil count >1000 cells/cu.mm) never resolved and persisted throughout this period of 6 months. Due to the persistent severe eosinophilia or hypereosinophilia,upper gastro intestinal endoscopy was finally done and the diagnosis of hookworm infection was finally made after a delay of 6 months which occured due to the persistent negative stool examination for hookworm ova. Hence upper gastro-intestinal endoscopy should always done in all patients with hypereosinophilia to confirm the presence of hookworms in tropical and subtropical countries even when there is no anaemia and even when stool examination is negative for hookworm ova. The diagnosis of hookworm infection was completely missed at her initial admission since upper gastro-intestinal endoscopy was not done due to the negative stool examination for hookworm ova. Hence stool examination should never be considered as a single diagnostic criterion to diagnose hookworm infection.
CONCLUSION
Severe eosinophilia or hypereosinophilia was found in 40% of our patients indicating that significant number of patients with hookworm infection had very high eosinophil count. Hypereosinophilia or severe eosinophilia can be the earliest sign of hookworm infection and can be present in the very early stage of hookworm infection even before the onset of anaemia. Hypereosinophilia or severe eosinophilia can also be present in the late stage of hookworm infection with severe anaemia. In the female patient with severe anaemia due to severe hookworm infection with hypereosinophilia or severe eosinophilia with multiple hookworms in duodenum, stool examination was negative for hookworm ova. But, hypereosinophilia or severe eosinophilia persisted in this patient throughout the period of 6 months despite her improvement of haemoglobin to 9.2g% and negative stool examination for hookworm ova. This is of extremely great significance since persistent hypereosinophilia or severe eosinophilia was the only indicator of hookworm infection due to which upper gastro-intestinal endoscopy was done and hookworm infection was confirmed in this patient. Hence upper gastro-intestinal endoscopy should always done in all patients with hypereosinophilia to confirm the presence of hookworms in tropical and subtropical countries even when there is no anaemia and even when stool examination is negative for hookworm ova.
ACKNOWLEDGEMENT
The author acknowledges the immense help received from the scholars whose articles are cited and included in references of this manuscript. The author is also grateful to authors / editors / publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed. The author is extremely grateful to IJCRR editorial board members and IJCRR team of reviewers who have helped to bring quality to this manuscript.
Englishhttp://ijcrr.com/abstract.php?article_id=681http://ijcrr.com/article_html.php?did=6811. Kato T, Kamoi R, Iida M, Kihara T.Endoscopic diagnosis of hookworm disease of the duodenum J Clin Gastroenterol. 1997 Mar;24(2):100-102
2. Anjum Saeed, Huma Arshad Cheema, Arshad Alvi, Hassan Suleman. Hookworm infestation in children presenting with malena -case seriesPak J Med Res Oct - Dec 2008;47(4) ):98-100
3. Verboeket SO1, van den Berk GE., Arends JE, van Dam AP, Peringa J, Jansen RR. Hookworm with hypereosinophilia: atypical presentation of a typical disease. J Travel Med. 2013 Jul-Aug;20(4):265-7
4. Heukelbach J, Poggensee G, Winter B, Wilcke T, Kerr-Pontes LR, Feldmeier H Leukocytosis and blood eosinophilia in a polyparasitised population in north-eastern Brazil. Trans R Soc Trop Med Hyg. 2006 Jan;100(1):32-40. Epub 2005 Sep 23.
5. Nutman TB, Ottesen EA, Ieng S, Samuels J, Kimball E, Lutkoski M, Zierdt WS, Gam A, Neva FA. Eosinophilia in Southeast Asian refugees: evaluation at a referral center. J Infect Dis. 1987 Feb;155(2):309-
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8. Wang CH, Lee SC, Huang SS, Chang LC. Hookworm infection in a healthy adult that manifested as severe eosinphilia and diarrhea. J Microbiol Immunol Infect. 2011 Dec;44(6):484-7. Epub 2011 May 23.
9. Yan SL, Chu YC. Hookworm infestation of the small intestine Endoscopy 2007; 39: E162±163 .
10. Lee VJ1, Ong A, Lee NG, Lee WT, Fong KL, Lim PL.Hookworm infections in Singaporean soldiers after jungle training in Brunei Darussalam. Trans R Soc Trop Med Hyg. 2007 Dec;101(12):1214-8. Epub 2007 Oct 4.
11. Kucik CJ, Martin GL, Sortor BV. Common intestinal parasites. Am Fam Physician. 2004Mar 1;69(5) ):1161-8 12. Mawhorter SD. Eosinophilia caused by parasites. Pediatr Ann. 1994 Aug;23(8):405, 409-13.
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17. Kalli T1, Karamanolis G, Triantafyllou K Hookworm infection detected by capsule endoscopy in a young man with iron deficiency. Clin Gastroenterol Hepatol. 2011 Apr;9(4):e33
18. Chen JM1, Zhang XM, Wang LJ, Chen Y, Du Q, Cai JT. Overt gastrointestinal bleeding because of hookworm infection. Asian Pac J Trop Med. 2012 Apr;5(4):331-2.
19. Cedrón-Cheng H, Ortiz C (2011) Hookworm Infestation Diagnosed by Capsule Endoscopy. J Gastroint Dig Syst S1:003. doi: 10.4172/2161-069X.S1-003.
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21. Chao CC1, Ray ML. Education and imaging. Gastrointestinal: Hookworm diagnosed by capsule endoscopy. J Gastroenterol Hepatol. 2006 Nov;21(11):1754.
22. Christodoulou, D. K., Sigounas, D. E., Katsanos, K. H., Dimos, G., and Tsianos, E. V. Small bowel parasitosis as cause of obscure gastrointestinal bleeding diagnosed by capsule endoscopy. World journal of gastrointestinal endoscopy, 2010;2(11),: 369.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareISOLATED AND UNILATERAL DELAYED ONSET MUSCLE SORENESS AFFECTING GASTROCNEMIUS -MAGNETIC RESONANCE IMAGING VALIDATION AND TREATMENT
English1618Ganesh Singh DharmshaktuEnglish Irfan KhanEnglishThe Delayed onset muscle soreness (DOMS) is a response to musculoskeletal insult following bouts of extraordinary physiological overload. The condition can take a morbid course if neglected or undertreated. Often considered a primer for musculoskeletal reparative process, the phenomena should be managed with holistic approach. The presented case is an uncommon pattern of DOMS in common settings and its appropriate management. It also highlights the importance of preventive and rehabilitative aspect of the condition.
EnglishDelayed onset muscle soreness, Magnetic resonance imaging, Physical therapyINTRODUCTION
The Delayed onset muscle soreness (DOMS) results from intensive, unaccustomed eccentric muscle activity and is a cause of variable clinical presentation ranging from mild pain to severe morbidity.[1] Initiation of a sports or endurance activity especially after a period of inactivity is common cause. Usually a self limiting event can take a chronic course if not managed early. Usually the DOMS affects multiple muscle groups at the same time and isolated and unilateral involvement after physical endurance is an uncommon occurrence.
CASE REPORT
A 23 year old male presented to us with complaints of left calf pain that was mild and intermittent to begin with but gradually increasing in severity and duration. There was history of involvement in military recruitment camp two days back. There was history of intense physical exertion including drills, running and other physical tests during the recruitment drive. He was having mild but tolerable pain which he dismissed as a transient episode. The pain got intense by third day and the patient found unable to bear weight and ambulate with the affected limb. The pain was severe and affecting the activities of daily living. The patient consulted a local practitioner where he was advised some pain medication including muscle relaxant. He consulted us a day later after he found no relief with a short period of rest, limb elevation and medications. On clinical examination, the patient has spasm over his left upper calf muscle and relative localized swelling at painful area when compared to contra lateral limb. The affected calf and gastrocnemius muscle was tender. The local temperature was normal and there was no history of other ‘red flags’. The pain was localized, increased on use of the limb and without any referral pattern, radiation or diurnal variation. The provisional diagnosis of muscular spasm of calf was made and patient was admitted for supervised physical therapy and appropriate supportive management. Magnetic resonance imaging (MRI) was done by patient out of comprehension to rule out serious underlying disorder.(Fig. 1) The MRI correlated with DOMS affecting gastrocnemius region with myofascial edema in medial and lateral head of gastrocnemius with perifascial fluid.(Fig. 2,3) The patient was managed by stretching and warm massage under supervised care. A schedule of five days of therapy resulted in dramatic improvement in pain scores and morbidity.
RESULT
The pain profile as measured by 10 point visual analogue score (VAS) and 100 point numerical rating scale (NRS) was substantially improved with patient pain free and performing activities of daily living in follow up at 6 and 12 weeks and finally at 3 months.
CONCLUSION
The anticipation of DOMS and its early management can save agony and pain involved in delay of appropriate care. At times, advance imaging modalities like MRI can be instrumental in diagnosis as well as to rule out other sinister disorders.
DISCUSSION
Micro-structural tear within muscle and connective tissues, lactic acidosis, inflammation and spasm has been attributed to clinical symptoms of DOMS.[1] Often peaking 1-3 days after unaccustomed physical exertion, the exact etiology of disease is unclear with only few hypotheses described. Structural damage to contractile tissue, disruption of calcium homeostasis and certain chemical products accumulation in interstitium stimulating free nerve endings is one such proposed mechanism in the previous work. [2] The role of inflammatory mediators in the causation of the disease is also an area of concern. [3] Quadriceps, hamstrings and triceps surae are commonly involved muscle groups. It may be considered sign of regenerative process helping build stronger muscular tissues.[4] There is no universal guidelines to treat DOMS owing to multiple treatment modalities each with inconsistent efficacy.[5] Warm up, stretching and massage has been widely used physical treatment modalities with good outcome to treat DOMS.[6]
ACKNOWLEDGEMENT
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors / editors / publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed.
Conflict of interest – None.
Source of funding – None
Englishhttp://ijcrr.com/abstract.php?article_id=682http://ijcrr.com/article_html.php?did=6821. Cheung K, Hume P, Maxwell L. Delayed onset muscle soreness: treatment strategies and performance factors. Sports Med. 2003;33(2):145-64.
2. Armstrong RB. Mechanisms of exercise-induced delayed onset muscular soreness: a brief review. Med Sci Sports Exerc.1984;16(6):529-38.
3. MacIntyre DL, Reid WD, McKenzie DC. Delayed muscle soreness.The inflammatory response to muscle injury and its clinical implications.Sports Med. 1995;20(1):24-40.
4. Coudreuse JM, Dupont P, Nicol C. Delayed post effort muscle soreness. Ann Readapt Med Phys. 2004;47(6):290-8.
5. Connolly D.A.J. ,Sayers S.P., Mchugh M.P. Treatment and Prevention of Delayed Onset Muscle Soreness. Journal of Strength and Conditioning Research, 2003, 17(1), 197– 208.
6. Rodenburg, J.B., Steenbeck D, Schierect P, Bar PR. Warmup, stretching and massage diminish harmful effects of eccentric exercise. Int. J. Sports Med. 15:414–419. 1994.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareELECTIVE SURGICAL CASE CANCELLATION - AN AUDIT
English1923Nikhil Nanjappa B. A.English Kirti Katherine KabeerEnglish S. Robinson SmileEnglishBackground: Elective surgical case cancellation refers to any surgical case that is booked into the operation theatre list on the day prior to surgery, but is not operated upon as scheduled [1]. The reasons for cancellation of elective surgical cases are many; they are as unique as they are similar. Aims and Objectives: The study aims to assess the magnitude of elective case cancellation at our hospital. This study tries to delineate the reasons for elective case cancellations across all surgical departments. The study aims to suggest necessary steps to alleviate the problem. Methodology: This is a prospective observational hospital based audit done between 23rd February 2012 and 5th May 2012. Observations and Results: The total number of elective cases booked into the operative list during the study period was 725, of which 197 (27.2%) was cancelled on the day of surgery. General surgery had the highest percentage share of total cancellations at 54.8%, followed by obstetrics gynecology (16.2%), orthopedics (13.1%), ENT (9.1%), urology (3.5%), pediatric surgery (2%), CTVS and neurosurgery (0.5% each). There were 9 cases that were cancelled more than once. They commonest reason for case cancellation was list over run (73%), followed by lack of patient fitness (14%). Discussion: Case cancellations on the day of surgery lead to underuse of OT’s, increases waiting period for the patients, frustration and mental stress to the patients and their families, and increases cost and wastage of hospital consumables [1-6]. Cancellation of booked elective surgeries is a common problem across all hospitals in our country and around the world. The incidence of cancellation of elective surgical operations has been reported in literature to range from 20% to 40% [3, 5, 7-10]. Improving the scheduling and admission procedure is of paramount for better use of hospital resources [16].
EnglishSurgical case, CTVS, NeurosurgeryINTRODUCTION
The operation theatre, among the surgical staff is regarded as the heart and soul of the hospital. Most hospital administrators agree that a considerable allocation of resources is made for the proper functioning of the operating room (OR). Also, it is the operation theatre that generates a substantial part of the hospital revenue. Elective surgical case cancellation refers to any surgical case that is booked into the operation theatre list on the day prior to surgery, but is not operated upon as scheduled [1]. The reasons for cancellation of elective surgical cases are many; they are as unique as they are similar. Unexpected OR cancellations are usually divided into avoidable cancellations (e.g., scheduling errors, equipment shortages, list overrun, and cancellation due to inadequate preoperative evaluation) and unavoidable cancellations (e.g., emergency case superseding the elective schedule, unexpected changes in the patient’s medical status, or patient nonappearance)[1]. The most common factor that leads to cancellation is lack of OR time [1]. The cancellation of patients from elective theatre operating lists considerably increases cost, duplicates workload, decreases efficiency and wastes operating room time [2, 3]. The major brunt of case cancellation is borne by the patients and their families. It causes significant emotional trauma to the patients as well as their families [4]. Elective surgery cancellations will directly and indirectly increase the patients’ treatment expenses due to longer hospital stay and in many cases, repetitions of pre-operative preparations and management [5, 6]. Repeat cancellations result in poor patient satisfaction, staff morale, hospitalpatient relationship and training [3, 5, 6]. The incidence of cancellation of elective surgical operations has been reported in literature to range from 20% to 40% [3, 5, 7-10]. With the end objective to reduce the number of case cancellations in our hospital, the first step would be to assess the magnitude of the problem.
AIMS AND OBJECTIVES
1. To assess the magnitude of elective case cancellation at our hospital.
2. To study and evaluate reasons for elective case cancellations across all surgical departments.
3. To suggest necessary steps to alleviate the problem.
METHODOLOGY
This was a prospective observational hospital based audit done between 23rd February 2012 and 5th May 2012. All elective cases cancelled on the day of surgery from the departments of General Surgery, Orthopedics, Obstetrics and Gynecology, ENT, Pediatric Surgery, Cardiovascular and Thoracic Surgery (CTVS), Urology and Neurosurgery during the study period were included. At Mahatma Gandhi Medical College and Research Institute, we have 10 elective theatres that function six days of the week (Monday-Saturday) from 8:30 AM to 2 PM, and one Emergency theatre common to all departments that functions 24hours a day, 7 days of the week. Of the 10 elective theatres, two are allotted to General Surgery, two to Orthopedics, one that is shared between Urology and Pediatrics, one between OMFS and ENT, one between CTVS and Neurosurgery and three for Obstetrics and Gynecology. The anesthetist in the pre-anesthetic check-up room assesses all elective cases on the previous day. Cases that are provisionally cleared for surgery will then be entered in the OT booking list. A copy of this list will be sent to the Anesthetic department and to the respective ward. The concerned anesthetist in charge of a particular OT for the next day, follow up these cases in the ward, the evening prior to the surgery. A list of all cancelled cases on the day of surgery were studied and analyzed by a surgical resident. Case cancellation of individual departments were taken into account and each case was allotted into one of the 13 causes for case cancellations which are patient unfit, operation not necessary, list over run, patient unwilling, attender not available, patient not fasted, procedure done in ward, staff not available, equipment/consumable problem, patient not affordable, blood not available, list interrupted by emergency and strikes caused by the hospital staff for various reasons.
OBSERVATIONS AND RESULTS
Total number of elective surgical cases posted for surgery during the study period was 725 in 61 working days. The mean was 11.9 elective cases per working day across all surgical departments. General surgery had maximum cases during the study period, accounting for 40.9%, followed by Obstetrics and Gynecology 19.8%, Orthopedics 15.1%, ENT 8.9%, Urology 4.9%, CTVS 3.8%, pediatric surgery 3.7% and neurosurgery 2.4%. [Table 1] The total number of cases cancelled was 197 (27.2%). The number of cases cancelled by individual departments is summarized in this section. Department of general surgery had the highest rate of cancellations, i.e. 36.3% of their 297 elective cases posted during the study period, followed by ENT (27.6%), orthopedics (23.6%), obstetrics and gynecology (22.2%), urology (19.4%), pediatric surgery (14.8%), neurosurgery (5.5%) and CTVS (3.5%). [Table 1] General surgery also had the highest percentage share of total cancellations at 54.8%, followed by obstetrics gynecology (16.2%), orthopedics (13.1%), ENT (9.1%), urology (3.5%), pediatric surgery (2%), CTVS and neurosurgery (0.5% each). [Table 1] The most common reason stated for case cancellation was list over run (73%), followed by patient being unfit for surgery (14.2%), strikes (4.5%). The other reasons contributed to a very small share and are listed on table 2. Surgeries for 9 of 197 patients were cancelled on more than one occasion. General surgery had 7 such incidents, followed by obstetrics gynecology and urology with 1 each. [Table 3] The reasons for case cancellation was studied separately for the department of general surgery, as it the highest share of cancellations. The commonest reason was list over run at 78.7%, followed by strikes (7.4%), patient unfit (5.5%), among other causes listed in table 4.
DISCUSSION
Elective surgical operations require a multidisciplinary approach between surgical team, hospital staff and hospital administration. Case cancellations on the day of surgery lead to underuse of OT’s, increases waiting period for the patients, frustration and mental stress to the patients and their families, and increases cost and wastage of hospital consumables [1-6]. Cancellation of booked elective surgeries is a common problem across all hospitals in our country and around the world. The cancellation rates vary significantly in various reports. The incidence of cancellation of elective surgical operations has been reported in literature to range from 20% to 40% [3, 5, 7-10]. In our study we had case cancellation of 27.2%. Though the number seems high, it is within range that is reported from most developing countries. Interestingly, the rates in developed countries are not significantly lower. The reasons for case cancellation are as varied as they are unique to each hospital. The reasons are usually categorized as modifiable and non-modifiable. Increased bed usage by medical specialties is one important factor [9]. Lack of theatre space and facilities have been cited as reasons by many researchers [5, 8, 14]. Cancellations of elective operations due to patient related factors accounted for 6.2% of cases. The most common reason for this was financial constraints [14]. “No show up of patients was the leading cause of cancellations in elective surgery” [1]. However, in our hospital day care surgery doesn’t contribute to significant proportion of elective cases, and patient ‘no show’ wasn’t a problem in our hospital. Other miscellaneous reasons are inadvertent discharge of the patient, non-fasting, anaphylactic shock, patient refused surgery, patients with prolonged coagulation profile [1]. In our study the single most important reason for case cancellation was ‘list over run’. It accounted for nearly three quarters of all case cancelations. Being a teaching hospital, most surgeries take longer to perform, as both surgical and anesthesiology residents are being mentored during the surgeries and this contributes to ‘list over run’. The scenario is common all teaching hospitals across the country and around the world. The most common cause for case cancellations on the day of surgery in all departments studied is due to list over run (73%), General Surgery having the most number of cancellations. Chalya et. al observed that case cancellations due to list over run were often due to surgeons underestimating the operating time required per case and also depended on the performing surgeon (less experienced surgeons/trainees) [14]. ‘List over run’, was followed by ‘patient unfit’. The reasons for this could vary from poor glycemic control, inadequately control of blood pressure, dyselectrolytemia, etc. In our study, the highest cancellation rate was in the general surgery department. 36.8% of all booked elective cases were cancelled. General surgery accounted for 54.8% of all case cancellations at our hospital during the study period. At distant second and third were departments’ of obstetrics and gynecology and orthopedics. The least number of cancellations were in departments of CTVS and neurosurgery. A study from Tanzania also found that most cancellations were in the general surgery section followed by orthopedic surgery and otorhinolaryngology, and the least cancellation was in the ophthalmology and cardiothoracic sections [14]. However, three other studies reported higher cancellation rate among the urology, orthopedics and gynecology than in general surgery [7, 15, 16]. Department of general surgery performs nearly 40% of all elective surgeries in our hospital; and also has the highest number of resident surgeons; hence the result was not very surprising. The commonest reason for case cancellation in the department was also ‘list over run’. General surgery also had the highest proportion of repeat cancellations (7/9), needless to say that repeat cancellations multiply patient anxiety, stress and financial burden. The cancellation of surgery creates untold hardship for patients who plan their working and family lives around the proposed operation date. Most are cancelled at less than 24 hours notice [13]. The cost implications to the community are immense but have not been calculated. Cancellations of elective operations due to lack of theatre space and theatre facilities can be prevented through careful planning and efficient utilization of the already limited hospital resources including the operating room, theatre facilities and valuable manpower. Improving the scheduling and admission procedure is of paramount for better use of hospital resources [16].
ACKNOWLEDGEMENTS
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors / editors / publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed.
Englishhttp://ijcrr.com/abstract.php?article_id=683http://ijcrr.com/article_html.php?did=6831. Dawlatly AA, Turkistani A, Aldohayan A, Zubaidi A, Ahmed A. Reasons of Cancellation of Elective Surgery in a Teaching Hospital. Intern J Anesthesiol. 2008;15:2.
2. Zafar A, Mufti TS, Griffin S, Ahmed S, Cancelled elective general operations in Ayub Teaching hospital. J. Ayub Med Coll Abbollabad 2007; 19 (3): 64-66.
3. Robb WB, O’Sullivan MJ, Brannigan AE, Bouchier-Hayes DJ. Are elective surgical operations cancelled due to increasing medical admissions? Irish J Med Sci. 2004;173(3):129.
4. Miller GG. Waiting for an operation: parent’s perspectives. Can J Surg 2004; 47(3):167-9. and Kolawole I.K., Bolaji B.O. Reasons for cancellation of elective surgery in Ilorin. Nig .J Surg. Res 2002; 4 (1-2): 28-33.
5. Ojo E.O., Ihezue C.H. An Audit of Day Case Cancellations In A Nigerian Tertiary Hospital Based Day Case Unit East and Central African Journal of Surgery 2008; 13 (2); 150- 153.
6. Kolawole I.K., Bolaji B.O. Reasons for cancellation of elective surgery in Ilorin. Nig .J Surg. Res 2002; 4 (1-2): 28-33.
7. Lacqua MJ, Evans JT. Cancelled elective surgery: an evaluation. American Surgeon 1994;60:809-11.
8. Rai M, Pandit JJ. Day of surgery cancellation after nurse led pre-assessment in an elective surgical centre: the first 2 years. Anesthesia 2003; 58:692-9.
9. Dakum N. K., Ramyil V. M, Misauno M. A., Ojo E.O., Ogwuche E. I., Sani A. A. Reasons for cancellations of urologic day care surgery. Nigerian of Surgical Research 2006; 8 (1 – 2): 30- 33.
10. El-Bushra A.D., Mohamed I.M., Awadalla M.A., Mohamed Y.B., Salah E.M. Cancelled elective surgical operations at El Obeid Hospital, Western Sudan. Sudan Med. J. 2008; 44 (1, 2 and 3):56-61
11. Are elective surgical operations cancelled due to increasing medical admissions? Robb WB, O’Sullivan MJ, Brannigan AE, Bouchier-Hayes DJ. Ir J Med Sci. 2004 JulSep;173(3):129-32.
12. Elective surgery--cancellations, ring fencing and efficiency. Aaserud M, Trommald M, Boynton J. Tidsskr Nor Laegeforen. 2001 Sep 10;121(21):2516-9.
13. Impact of emergency admissions on elective surgical workload. Nasr A, Reichardt K, Fitzgerald K, Arumugusamy M, Keeling P, Walsh TN. Ir J Med Sci. 2004 JulSep;173(3):133-5.
14. Chalya PL, Gilyoma JM, Mabula JB, Simbila S, Ngayomela IH, Chandika AB, Mahalu W. Incidence, causes and pattern of cancellation of Elective surgical operations in a University Teaching Hospital in the Lake Zone, Tanzania. African Health Sciences Vol 11 No 3 September2011: 438-43.
15. Jonnalagadda R, Walrond ER, Marinara S, et al. Evaluation of the reasons for cancellations and delays of surgical procedures in a developing country. Int Clin Pract. 2005;59:716–720.
16. Venkartaraman S, Seriam K. Cancelled elective surgery: study in an Indian Corporate Hospital. Indian Surg. 1997;59:372–376.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareNORMATIVE VALUES OF TANDEM AND UNIPEDAL STANCE IN SCHOOL CHILDREN
English2430Sneha D. DhananiEnglish Lata D. ParmarEnglishBackground: There were very few studies done on unipedal stance in school children and there was no study found on tandem stance in school children to the best of our knowledge Aim: To establish normative values of tandem and unipedal stance in school children. Study Design: Descriptive study Methodology: 250 subjects, meeting inclusion-exclusion criteria were included Each participant then completed timed unipedal balance test on right and left foot, and tandem stance test on right foot in front of left foot. The tests were performed barefoot on a floor surface. The timed measurements were completed using a stopwatch. Results: Mean age of participants was 11.76±3.499. Mean range of unipedal stance (rt. and lt.) between 6-17 years (total participants) was 1.14±0.352 to 2.55±0.702 seconds. Mean range of tandem stance between 6-17 years was 1.47±0.520 to 3.26±0.953.The results showed that age had statistically significant correlation with the both static balance tests (p value 0.000 and r value >0.5), but gender did not show any significant correlation for balance performance in both these static balance tests. BMI also did not show any statistically significant correlation for both the static balance tests. (p value 0.000, r value EnglishBalance skills, Dynamic balance, Static balanceINTRODUCTION
Balance is a complex process involving the reception and integration of sensory inputs and the planning and execution of movement to achieve a goal requiring upright posture. It is the ability to control the center of gravity (COG) over the base of support in a given sensory environment.1 Balance control is temporally organized from the head to feet, according to a descending organization. In motor development, the first period is characterized by the development of postural responses along a cephalocaudal gradient. Overall balance ability and postural stability increase and mature by 6 to 10 years of age2 there are many systems within the body that work in concert to keep the centre of mass (COM) within the base of support (BOS) when maintaining static postures and to move the COM in relation to the BOS in a controlled manner when engaged in dynamic tasks.3 Before birth, different brain regions develop at different rates, and sensory organs initially develop independently of the brain regions to which they will later be connected. The vestibular sense or proprioception begins to function before the fetus first shows a righting reflex at 25 weeks and probably plays a part in bringing the fetus into a head-down position before birth. 4, 5 Balance and posture evaluations address whether sensory information from the vestibular system coordinates and integrates with vision and somatosensory information from the muscles and joints. Such tests require the child to balance under various vision conditions (e.g., with eyes open and closed or with a moving visual field) and varied standing surfaces (e.g., on a firm, soft, or moving surface).6 Numerous techniques have been described to measure standing balance, with varying levels of challenge in different populations. The Timed unipedal stance test (also referred to as timed single limb stance, unipedal balance test, one leg stance test, and one-leg standing balance) is a simple test for measuring static aspects of balance that can be used in a variety of settings and requires minimal equipment or training.7 Standing balance is the necessary precursor to all upright loco motor skills.7 Static balance is the ability to maintain a posture in a resting position, while dynamic balance is the ability to maintain postural control during the performance of functional tasks.8, 9 The task of standing on one leg requires initially voluntary shift of the COM to the leg which will be stood on, followed by maintenance of postural orientation in space, by controlling weight, maintaining the vertical alignment of the rest of the body and equilibrium. One leg standing assesses postural steadiness in a static position by a quantitative method, measurement being the number of seconds that an individual can maintain a one leg stance. The underlying assumption being that the better one’s postural steadiness, the longer such a position can be held. Tandem stance is the ability to stand in a heel-to-toe position, reflects the degree of postural steadiness when the BOS in the medial/lateral direction is narrow, also known as the sharpened Romberg test. Tandem stance was developed on the basis of classic Romberg’s sign and originally utilized as a bedside indicator of abnormality of the proprioceptive system.8, 10, 11 There is an information deficiency of norms regarding balance skills, especially in our country and taking into consideration that such information will contribute to both the planning of developmentally adequate movement programs and the study of developmental specificities during such critical years as the school age, the aim of the present research was to establish normative value of tandem and unipedal stance. AIM To establish normative value of Tandem stance and Unipedal stance in school children, age group between 6 to 17 years.
OBJECTIVE
• To establish the minimum duration of Tandem stance.
• To establish the minimum duration of Unipedal Stance.
MATERIALS NAD METHODS
Ethical clearance was received from Sumandeep Vidyapeeth Institutional Ethical Committee (SVIEC). Study Design: Descriptive Study Study Population: school children
Inclusion criteria:
All School going children – age group between 6-17 years were eligible for inclusion.
Exclusion criteria:
Children with any neurological, musculoskeletal, and cardiovascular deficit that affect balance.
• Any history of balance impairment.
Outcome measures
• Duration in seconds in Tandem stance test.
• Duration in seconds in Unipedal stance test. Methodology: Following ethical permission from Sumandeep Vidyapeeth Ethical Committee (SVIEC), various schools were approached for official permission from the Head / Principal of the School. The Principal / teachers and students were explained about the project and requested for informed consent. Verbal assent was ensured from small children apart from written permission from Principal / teacher.
Methodology:
Following ethical permission from Sumandeep Vidyapeeth Ethical Committee (SVIEC), various schools were approached for official permission from the Head / Principal of the School. The Principal / teachers and students were explained about the project and requested for informed consent. Verbal assent was ensured from small children apart from written permission from Principal / teacher.
Each participant was enquired of baseline questions and checked for inclusion criteria. First of all, the anthropometric measurements of the subjects were taken. Prior to balance testing, participants were familiarized with the balance test and provided practice sessions on the testing procedures to decrease the chance of a learning effect occurring during testing. The tests were conducted at the same times of the day (between 10.00 am to 5.00 pm) when the body was in rest, and measures were taken to prevent distraction due to environmental factors (anyone talking, any noise). The procedure lasted 10-12 minutes for each child. The tests of whole group were completed within 15-20 days. Each participant then completed timed unipedal balance test i.e. standing on right and left foot alternatively, and tandem stance test on right foot in front of left foot. The tests were performed barefoot on a floor surface. For two balance tests, each subject completed 3 trials on each leg. A 15-second rest was given between trials set to avoid fatigue. For all trials, the participants placed their hands across the chest and time started upon elevation of the opposite foot from the floor. Participants focused on a target placed at eye level, in front of them. A stopwatch was used to time the test. This study was carried out in School’s class room.Tests were done in separate room where source of light was good and also in secured place so that risk of fall during tests was prevented. For unipedal stance, participants were asked to stand barefoot on the limb of their choice, with the other limb raised so that the raised foot is near but not touching the leg / ankle of their stance limb. Prior to raising the limb, the subject was instructed to cross his arms over the chest as shown in figure 1and2. The stopwatch was used to measure the amount of time the subject was able to stand on one limb. Time commenced when the subject raised the foot off the floor and time ended strictly when the subject either: (1) used his arms (i.e., uncrossed arms), (2) used the raised foot (moved it toward or away from the standing limb or touched the floor), (3) move the weight-bearing foot to maintain his balance (i.e., rotated foot on the ground). The procedure was repeated 3 times the mean of the 3 trials were recorded.7 For tandem stance, participants were made to stand with feet in a heel-to-toe position on straight line drawn with chalk stick on the floor, and arms across the chest, with eyes open as shown in the figure 3 and 4. Time commenced when the subject placed the rt. foot in front of left foot on the straight line and time ended strictly when the subject either: (1) use his arms (i.e., uncrossed arms), (2) displace any foot,(3) movement of the foot from original position/ stepping. The procedure was repeated 3 times and the mean of the 3 trials was taken.
RESULTS
There were total of 250 school children included, of them 123 were males and 127 were females (fig.5). Figure 6 shows the percentage of distribution in the three groups age wise, 38% were in 6-10 years, 43% in age group 11- 15 years and 19% >15 years of age. Only 24 of 250 were with left hand dominance. Mean difference of Height, weight and BMI is shown in figure 7 Figure 8 shows Mean values of Tandem and Unipedal stance between males and females. Unipedal stance is studied of both the lower limbs. 1) Tandem Stance(Rt. foot in front of Lt. foot): • 6-10 yrs: 1.37 - 1.58 sec. • 11-15 yrs: 2.28 - 2.60 sec. • ≥15 yrs: 2.98 - 3.53 sec. 2) Unipedal Stance on maximum range Weight bearing (both) feet: • 6-10 yrs: Min. =1.21-1.30 sec. • 11-15 yrs: Min. = 1.92- 2.09 sec. • ≥15 yrs: Min. = 2.72 - 2.76 sec. Table 1 shows correlation between Age and Tandem and unipedal stance. Highly significant relationship is seen with r value > 0.5 and p- Value 0.000 Similarly Table 2 and 3 show significantly high correlations between height and the two balance tests. Table 4 shows that unipedal stance between right and eft lower limbs were highly correlated with r=.817**
DISCUSSION
The study aimed to establish normative values of tandem and unipedal stance in normal school children. There were very few studies done on unipedal stance in school children and there was no study found on tandem stance in school children to the best of our knowledge, these two tests of balance were therefore chosen for the study. In the present study, the total numbers of participants were 250, of which 123 were males and 127 were female. The tandem stance and unipedal stance was studied in school children of mean age 11.76±3.499. It was found that, age had statistically significant correlation with the static balance tests (p value 0.000 and r value >0.5) but gender did not show any significant correlation for balance performance in both these static balance tests. Similarly, other studies7, 8, 9, 13, have also suggested that age have greater significant correlation with balance than gender. However, significant differences between rural and urban school boys’ coordinative and balancing abilities have been reported by Raghupathi K etal.9 Barbara A. Springeret et al, 7 studied 549 participants, in which 258 were female, and 291 male (aged 18- 80+). They have also taken mean and best of 3 Unipedal Stance Test times (similar to the present study) for males and females of 6 age groups and found that there is no interaction between gender and test condition, and also no significant difference was found for gender. But there were significant main effects for age category and they have also concluded that difference in Unipedal Stance Test times is not gender specific but is related to age. 7 The authors7 have interpreted that elderly subjects had difficulty maintaining balance in the static phase due to difficulty adjusting postural control during the initial dynamic phase of one-leg stance. Another possibility for the decrease in stance times for elderly subjects is a decrease in lower extremity muscular strength and endurance. 7 Fotini Venetsanou et al,8 have taken balance subtest of the Bruininks-Oseretsky Test of Motor Proficiency and found that age have great significant correlation with both the subtest than gender. They have also concluded that age seems to be a significant factor for preschoolers’ performance in terms of balance skills, while gender does not and interpreted that it can be by the rapid progress caused by the biological processes of development during the period between four and eight years of age but for gender differences, at the preschool age, the biological characteristics of boys and girls are similar rather than diverse and differences between genders can be found- may be, in “modern” societies, qualitative differences in encouragement, support, and opportunities regarding participation in play-game type of activities can be identified and may have greater influence on balance, and also balance is a multidimensional structure and as such cannot be affected by only a couple of factors. 8 V. Hatzitaki et al, 12concluded that balancing under static conditions was strongly associated with the ability to perceive and process visual information, which is important for feedback-based control of balance. Adults, 11- to 13-year-old children have the ability to select varying balance strategies (feedback, feed forward, or both), depending on the constraints of a particular task.12 Oya Erkut At?lgan et al, 14 studied 60 children, aged 9-11 years. They have used static balance tests: Bipedal static balance with Eyes open (EO) and eyes closed (EC) and Unipedal static balance with respectively on right and left foot, eyes open for 30 seconds. They found that boys had good bilateral and unilateral balances than the girls but there was no statistical significant differences found for Romberg test and unilateral (right leg) static balance of the subjects between genders and concluded that unilateral static balance (left leg) parameters of the boys were statistically significant while there wasn’t any significant difference of unilateral static balance (right leg) of the boys and the girls and interpretated that boys have a more active life than the girls in Turkey, 14 they play games on the streets more. The authors of this study therefore state that static balance (EO-EC- Left foot) of the boys was better than the girls because they did more physical activities. Soccer is also a very popular sport branch in their country and boys play soccer on the streets in early ages with their friends.14 In present study however, right and left were highly correlated on unipedal stance (r=.817) In present study height was found to be significantly correlating with the balance performance for both the tests. (p value 0.000 and r value >0.5). This has been interpreted as due to the rapid progress caused by the biological processes of development during the period between four and eight years of age as explained by Fotini Venetsanou etal.8 However, body mass index (BMI) did not show any statistically significant correlation with balance performance for both the static balance tests (r value Englishhttp://ijcrr.com/abstract.php?article_id=684http://ijcrr.com/article_html.php?did=6841. Darcy A. Umphred, NEUROLOGICAL REHABILITATION, Fifth Edition, ch.23, pg no.740.
2. Chemin Joseph Aiguier et al, Development of Locomotor Balance Control in Healthy Children, Vol. 22, No. 4,1998,pp. 527–532.
3. Westcott SL et al, Evaluation of postural stability in children: current theories and assessment tools. Phys Ther. 1997; 77:629-645.
4. Parncutt, R. et al (2006), prenatal development, The child as musician Oxford, England: Oxford University Press, pp1-31.
5. Susan J. Herdman, Vestibular Rehabilitation, Third Edition, Ch.1, pg no.2, 3.
6. Gaye W. Cronin et al, Pediatric Vestibular Disorders Recognition, Evaluation and Treatment, Vestibular Disorders Association.
7. Barbara A. Springer et al, Normative Values for the Unipedal Stance Test with Eyes Open and Closed,Journal of Geriatric Physical Therapy Vol. 30;1:07.
8. Fotini Venetsanou et al , THE EFFECTS OF AGE AND GENDER ON BALANCE SKILLS IN PRESCHOOL CHILDREN, Physical Education and Sport Vol. 9, No 1, 2011, pp. 81 – 90.
9. Raghupathi K et al, Comparative Analysis of Coordinative and Balancing Abilities Among 10-15 Years of Rural and Urban School Boys, Volume: 2, Issue: 5, May 2013, ISSN No 2277-8160.
10. Maria Nida c. et al, Development of Lower Extremity Kinetics for Balance Control in Infants and Young Children, Journal of Motor Behavior,20011,Vol.33, No.2,180-192.
11. Carolyn A Emer et al, Development of a Clinical Static and Dynamic Standing Balance Measurement Tool Appropriate for Use in Adolescents, Physical Therapy, Volume 85, Number 6, June 2005
12. V. Hatzitak et al, Perceptual-Motor Contributions to Static and Dynamic Balance Control in Children, Journal of Motor Behavior, 2002,Vol. 34, No. 2, 161–170.
13. Stanley N. Graven et al , Auditory Development in the Fetus and Infant, VOLUME 8, NUMBER 4, www.nainr.com.
14. Oya Erkut At?lgan et al, The effects of postural control to gender differences in children, Volume: 9 Issue: 2 Year: 20.
15. Eva D’Hondt et al, Relationship Between Motor Skill and Body Mass Index in 5- to 10-Year-Old Children, Physical Activity Quarterly, 2009, 26, 21-37.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareASSESSMENT OF ARTERIAL STIFFNESS AMONG MIDDLE AGED OFFSPRING OF DIABETIC PARENTS
English3134Niruba R.English Subha K. C.English Vijayalakshmi I.EnglishBackground: Genetic susceptibility appears to play an important role in the occurrence of Type 2 Diabetes with high prevalence among first degree relatives .Subjects with type 2 diabetes have 2-3 fold increased risk of cardiovascular disease compared with normal population. Aim: To compare the arterial stiffness between normotensive normoglycemic middle aged offspring of diabetic parents (Cases) with normotensive normoglycemic middle aged offspring of Non- diabetic parents (Controls). All participants had a sedentary life style.BMI, Blood pressure and glycemic status was matched between cases and controls. Alcoholics, smokers, subjects with peripheral vascular disease and other illness were excluded. Arterial stiffness was assessed from Reflective Index (RI) and Stiffness Index (SI). Reflective index and Stiffness index was measured using IR1 model digital finger tip photo pulse plethysmograph. Results: Controls RI- MEAN – 47.9 ±2.8%,SI –MEAN 6 ±.7 m/s and Cases -RI- MEAN -55.1± 3.6%,SI MEAN 7±.9 m/s. Paired ‘t ‘test was applied and it showed statistical significance with p value < .001.We conclude that vascular endothelial dysfunction and
arterial stiffness is present from an early stage in subjects with family history of diabetes. Life style modifications are suggested to improve the vessel wall compliance and prevent further progression.
EnglishStiffness index, Reflective index, Diabetes mellitusINTRODUCTION
Diabetes has become a worldwide epidemic. In the present scenario early detection and measures to prevent the morbidity and mortality due to diabetes is absolutely necessary .Cardiovascular diseases are the most common cause of disability and death among subjects with non– insulin-dependent diabetes mellitus (1). The atherosclerotic process begins during the prediabetic phase characterized by impaired glucose tolerance, hyperinsulinemia, and insulin resistance. Studies have suggested that glucose and insulin can substantially alter the structure and function of the arterial wall and affect the development of atherosclerosis (2).The adverse association of cardiovascular risk factors in both children and adults with parental history of diabetes is well recognized (3,4). Increased risk of atherosclerosis is found in prediabetic individuals (5), and in populations at high risk of Coronary Heart Disease, almost half of middle-aged men and women with NIDDM have symptomatic Coronary heart disease at the moment their diabetes is diagnosed (6). Recording of peripheral pulse wave and digital volume pulse is convenient method to measure stiffness index which can assess large artery stiffness. Reflection index can assess medium and small artery stiffness (7).
RATIONALE OF THE STUDY
There is paucity of data in our country on the vessel wall compliance among the middle aged individuals who are genetically prone to develop diabetes. And also there is lack of simple cost effective screening procedures to assess vascular function and reduce the cardiovascular morbidity.
AIM
To assess the vessel wall compliance among the middle aged off spring of diabetic parents.
MATERIALS AND METHOD
All participants gave a written informed consent to participate in this study. Institutional ethical committee clearance was obtained. Information details about socio demographic characteristics, disease history, alcohol consumption, cigarette smoking, drug intake and occupational history were obtained by a structured questionnaire. All participants had a sedentary lifestyle. Inclusion criteria: Sex = Males, Age= 35- 45 yrs, Blood pressure = < 140/90 mmHg, Fasting blood sugar = 90-100 mg/dl. Exclusion Criteria: Alcoholics, smokers, subjects with peripheral vascular disease and other illness were excluded from the study. Cases – 50 healthy volunteers normoglycemic , normotensives males with paternal or maternal history of diabetes more than 10 years. Controls- 50 healthy volunteers normoglycemic normotensive males with no paternal or maternal history of diabetes. Anthropometric measurements were taken. Height was measured using stadiometer and weight was measured using precalibrated weighing machine.. Quetlet’s index was used to calculate Body mass index (weight/height in m2 ).Blood pressure was measured using a standard mercury sphygmomanometer. Fasting blood glucose was measured to rule out diabetic mellitus. Recording of digital volume pulse (DVP): Digital volume pulse was recorded by in house built instrument IR1 model digital finger photoplethysmography (8) ,using infra- red light with wave length 0f 940 nm; placed on the right index finger of the subjects. The signal from the instrument was digitalized by digital converter with a frequency of 100 Hz which was connected to the computer. DVP was analyzed by software virtual oscilloscope. Digital volume pulse contains 2 peaks: Fig-1 1. Systolic peak. 2. Diastolic peak. Early systolic peak is formed by pulse wave transmitted from the left ventricle to the finger directly. Second peak or diastolic peak arises from pulse wave transmitted along the aorta to the small arteries in the lower body, from where they are again reflected along the aorta as a reflected wave .This path length is proportional to the subjects height (h) (fig-1). Pulse transit time (PTT or ?T) is the time interval between systolic peak and diastolic peak. It was measured by software image tool (which was provided by Stanford university). Magnitude of systolic and diastolic peak were also measured Stiffness index is based on the subjects height (9).Stiffness index and Reflective index were calculated by the following formulas.
RESULT
Statistical analysis was done Using SPSS Software 16.0 and Paired t- Test applied. Table-1 shows the descriptive statistics of BMI, Blood sugar, Systolic blood pressure, Diastolic blood pressure. Table 2- shows arterial stiffness indices of both control and study group. Paired ‘t’ test applied between stiffness index and reflective index of controls with study group and it showed a statistically significance with p value < 0.001.
DISCUSSION
In our present study Reflective index and Stiffness index of normoglycemic healthy offspring of type 2 diabetic parents, was found to be greater than the control group with no family history of diabetes. Stiffness index and Reflective index are reliable indicator of Arterial stiffness (10,11,12). Arterial stiffness, an early feature of diabetic vasculopathy, has been recognised as a powerful and independent predictor for Cardio vascular disease (13,14,15) and considered as a “transitional medicine biomarker for CV risk factor”(16). Thus, subjects with a pronounced familial background has high probability of developing diabetes (17) show arterial stiffening already at a stage in which blood glucose is normal. Giannattasio et al reported that arterial stiffening can increase the risk of cardiac and vascular morbid or fatal events presumably because a reduction in the ability of the arteries to distend increases blood pressure and is associated with an increase in arterial impedance and, thus, in the after load to the heart, and endothelial damage because a reduced distensibility enhances the traumatic effect of the intravascular pressure on the vessel wall (18). Similar study by Cristina et al and Rahman et al says subjects with predisposition to diabetes show carotid artery stiffening and alterations in arterial mechanical properties even in the absence of blood pressure alterations, as well as substantial alterations of glucose metabolism, body mass index, and changes in carotid wall thickness and further added that normoglycemic offspring of newly diagnosed parents have increased arterial stiffness, indicating early manifestations of preclinical vasculopathy, which may predate the onset of type 2 diabetes (19,20) .The mechanisms responsible for the early arterial stiffening of normoglycemic offspring of diabetic parents are not clarified by our study as the glucose values were normal and similar between the offspring of diabetic parents and control subjects, the arterial stiffening probably could not be due to any adverse effect of glucose on the functional characteristics of the vessel wall. Laurent et al says Genetic influences operating through other mechanisms may be responsible in the majority and further added that diabetes is accompanied by an increased wall thickness even in the absence of microvascular and macrovascular complications (21). Thus in this context early detection and screening of predisposed group can be beneficial.
CONCLUSION
Arterial stiffness assessed by reflective index and stiffness index are increased in the offspring of diabetic parents. Implication: Our study was designed to prevent the morbidity and mortality in the offspring of diabetic parents due to their genetic influence. Noninvasive, cost effective, portable and rapid method used in our study can be used for screening measures, based on which life style modifications can be suggested to delay and improve the vascular function.
ACKNOWLEDGEMENT
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors / editors / publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed. Authors sincerely thank Dr K N Maruthy, Professor, Narayana medical college, Nellore for his extensive guidance. Funding for the project was shared by the authors and there is no conflict of interest.
Englishhttp://ijcrr.com/abstract.php?article_id=685http://ijcrr.com/article_html.php?did=6851. Barrett-Connor E, Orchard T. Diabetes and heart disease. In: Harris M, ed. Diabetes in America. Washington, DC: NIH; 1984; XVI :1-41.
2. Veikko Salomaa, MWard Riley,Jeremy D et al Non–InsulinDependent Diabetes Mellitus and Fasting Glucose and Insulin Concentrations Are Associated With Arterial Stiffness Indexes,Circulation.1995; 91: 1432-1443.
3. Blonde CV, Webber LS, Foster TA et al Parental history and cardiovascular disease risk factor variables in children. Prev Med.1981;10:25-37.
4. Burke GL,Savage PJ,Sprafka JM et al. Relation of risk factor levels in young adultwood to parental history of disease: the cardia study; Circulation 1991;84:1176-1187.
5. Haffner SM, Stern MP, Hazuda HP et al Cardiovascular risk factors in confirmed prediabetic individuals: does the clock for coronary heart disease start ticking before the onset of clinical diabetes? JAMA.1990; 263;2893-2898.
6. Uusitupa M, Siitonen O, Aro A et al , Prevalence of coronary heart disease, left ventricular failure and hypertension in middle-aged, newly diagnosed type 2 (non-insulindependent) diabetic subjects. Diabetologia.1985;28:22-27
7. Millasseau SC,Kelly RP,Ritter JM et al .Determination of age related increases in large artery stiffness by digital pulse contour analysis. Clin Sci (lond) 2002;103:371-7.
8. G Sivagami,Milind Bhutkar,A Comparitive study of arterial stiffness indices between normotensive and hypertensive subjects-NJBMS,Volume 4 ;issues-3:pg-177-179.
9. Chen CH, Nevo E ,Fetics B et al .Estimation of central aortic pressure waveform by mathematical transformation of radial tonometry pressure.Validation of generalized transfer function. Circulation 1997;95:1827-36.
10. Cohn JN, Finkelstein S, McVeigh G, et al. Noninvasive pulse wave analysis for the early detection of vascular disease. Hypertension1995; 26:503–8.
11. McVeigh GE, Bratteli CW, Morgan DJ et al Age-related abnormalities in arterial compliance identified by pressure pulse contour analysis: aging and arterial compliance. Hypertension1999; 33:1392–8.
12. Chowienczyk PJ, Kelly RP, MacCallum Het al Photoplethysmographic assessment of pulse wave reflection. J Am Coll Cardiol 1999; 34:2007–14.
13. Woodman RJ, Watts GF. Measurement and application of arterial stiffnessin clinical research: focus on new methodologies and diabetes mellitus. Med Sci Monit 2003;9:RA101- 10.
14. M eaume S, Rudnichi A, Lynch A et al. Aortic pulse wave velocity as amarker of cardiovascular disease in subjects over 70 years old. J Hypertens 2001;19:871-77.
15. Hopkins KD, Lehmann ED, Gosling RG. Aortic compliance measurements:a non-invasive indicator of atherosclerosis? Lancet 1994; 343:1447.
16. Wang X, Keith JC, Struthers AD et al. Assessment of arterial stiffness, a translational medicine biomarker system for evaluation of vascular risk. Cardiovasc Ther 2008;26:214- 23.
17. Polonsky KS, Sturis J, Belle GI. Non-insulin-dependent diabetes mellitus- a genetically programmed failure of the beta cell to compensate for insulin resistance. N Engl J Med. 1996; 334: 777–783.
18. Giannattasio C, Failla M, Piperno A et al Early impairment of large artery structure and function in type I diabetes mellitus. Diabetologia. 1999; 42: 987–994.
19. Cristina Giannattasio, Monica Failla, Anna Capra et al Increased Arterial Stiffness in Normoglycemic Normotensive Offspring of Type 2 Diabetic Parents Hypertension. 2008; 51: 182-187
20. Sayeeda Rahman, Aziz Al-Safi Ismail,Shaiful Bahari Ismail et al Increased arterial stiffness in normoglycaemic offspring of newly diagnosed, never treated type 2 diabetic and impaired glucose tolerance parents Br J Diabetes Vasc Dis 2009;9: 65–68.
21. Laurent S, Cockcroft J, Van Bortel L, Boutouyrie P, Giannattasio C, Hayoz D, Pannier B, Vlachopoulos C, Wilkinson I, Struijker-Boudier H; European Network for Noninvasive Investigation of Large Arteries. Methodological issues and clinical applications. Eur Heart J. 2006; 27: 2588–2605.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareMATHEMATICAL RELATIONSHIP AMONG AGE, BMI AND VISUAL REACTION TIME IN NORTHERN INDIAN CHILDRENS
English3537Hom Nath DhunganaEnglish Seema SinghEnglish Pooja TripathiEnglish Jay prakashEnglishReaction time has been defined as the time interval between the application of a stimulus and the response by the subject. Various factors affect reaction time like handedness, age and gender of the person, BMI, type of receptor system involved. It is well known that there exist relationship among Visual Reaction Time, age and BMI. The degree of correlation among these variables is found to be higher. In this paper we develop a multivariate regression line in the form of z = ax +by +c, where x, y are two independent variable (x=age, y=BMI) and z is the measure of visual reaction time.
EnglishVisual reaction time, Correlation, Regression lineINTRODUCTION
Visual reaction time (VRT) is the time that elapses from the initiation of a stimulus until a response is achieved. Reaction is a purposeful voluntary response to external stimulus. There is certain time period between application of stimulus and appropriate motor response. Reaction time can be divided into three parts. The first part is perception time, the time for the application and perception of stimulus. Second part is decision time, which signifies time for giving a suitable response to the stimulus. The third part is motor time, which is the time for the compliance to the order received1 . Reaction is a purposeful voluntary response to external stimulus. Reaction time is defined as interval of time between presentation of stimulus and appearance of appropriate voluntary response in a subject. Our emotions, attention, memories these and all added reaction we make are responses to stimuli which play upon us2 . Thus it indicates the time taken by an individual to react to external stimulus3 . One measure of information processing is reaction time and is used to judge the ability of the person to concentrate and coordinate. Many studies revealed that reaction time task is a good indicator of sensor motor performance of an individual, as the young individuals performed better in the reaction time tasks than elderly individuals who have the tendency to be more careful and monitor their responses more thoroughly. It is one of the important methods to study a person’s central information processing speed and fast coordinated peripheral movement response. Many research works have been published on reaction time and it is reported that there are many factors on which the reaction time depends. Some factors are Age, BMI, Sex, Fatigue condition etc. But there are very few research works available on the development of mathematical models based on reaction time visual reaction time with respect to age, BMI, sex etc. The application of these models may be wide not only to predict or estimate the visual reaction time but also to know the contribution of these factors on reaction time.
MATERIAL AND METHODS
The study subjects were taken randomly from Techno Academic School and integral Institute of Medical Sciences and Research, Lucknow with a permission of Director/Principal .The inclusion criterion was all disease free normal children and young adults aged between 6 – 21 years. Before the collection of data it was insured that subjects had taken no medication while we excluded those children’s who were any addiction such as smoking, tobacco or alcohol, any history of drug abuse and any history of hereditary or chronic illness. The total number of subjects was calculated by the help of statistician which was 104.The very common, efficient and easy machine “IMCORP Ambala Reaction Time Instrument” was used to acquire the simple reaction time data in children. The Reaction Time tests consist of two colored lights (red and green) and two response buttons to which a digital timer is connected. The specifications of reaction timer machine are, 1. Inbuilt chronoscope – 4 digit chronoscope with least count of 1/1000 seconds. 2. It works on – 230 volts AC. Subjects were present randomly with two visual stimuli i.e. red and green light. The operating process was demonstrated priory to reduce errors .Three readings of each stimulus noted after giving three practical trials and the lowest taken as the reaction time. The reaction time measured for both red and green colour. The process of pressing the buttons was explained to children to minimise the error.The study was cleared by the Institutional ethical committee.After the collection of data it was analyzed by using SPSS of version 20 to calculate descriptive statistics and regression constants.
RESULTS
The mean VRT score of green and red light was calculated. The average VRT (AVRT) is dependent variable in linear regression analysis while the age and BMI of children are independent variables. Table 1 shows the descriptive statistics. The average of average visual reaction time (AVRT) is 210.36 with standard deviation 62.53; the average age of children is 13.45 years with standard deviation 4.62 and means BMI is 145.78 with standard deviation 16.46.The mathematical linear relationship among variable is calculated as AVRT= -11.6 ×Age – 0.38×BMI + 423.The coefficient of determination (R2 ) is 0.89.The high value of R2 shows that the high degree of the data are to the fitted regression line is. The ANOVA obtained from regression analysis is found statistically significant (F=400.48, p Englishhttp://ijcrr.com/abstract.php?article_id=686http://ijcrr.com/article_html.php?did=6861. Tripo RS (1965). How fast can you react? Sci Dig, 57:50.
2. Joseph Tiffin, Federic B Knight and Eston Jackson Asher (1946).The Psychology of normal people. 2nd edition , D.C. Health and company, Boston 415-431.
3. Mishra N, Mahajan KK, Maini BK. Comparative study of visual and auditory reaction time of hands and feet in males and females. Indian Journal of Physiol Pharmacol 1985; 29: 213–218
4. Dhungana H. N.et al., “Visual Reaction Time and its Relationship with Sex, Age and BMI in Northern Indian Children’s”. Research and Reviews: Journal of Medical Science and Technology Volume 3, Issue 2.page 1-6, 2014
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20Healthcare16S rRNA BASED T-RFLP ANALYSIS OF METHANOGENS IN BIOGAS PLANT WITH P. HYSTEROPHORUS L. AS SUBSTRATE
English3842Ramya R.English Shree M. P.EnglishBackground: Methanogenic microbiome plays an important role in contributing to global renewable energy resource. Globally, biogas production totally estimates to 1/4th of total consumption of fossil fuel. The bioconversion of organic material by anaerobic digestion process to a valuable energy source has a promising outlook. Though there is lot of research contribution to the biogas technology, the knowledge about the microbiome involved in the bioconversion of plant biomass is limited. Objective: In this study, batch fermenter with P. hysterophorus as mono-substrate and along with co-additives of bovine animal faeces at different ratios is presented in the context of molecular data on the microbial composition. Material and methods: A slightly modified protocol of the kit RKN15 to extract the metagenomic DNA from the microbial community participating in the bioconversion of the mono-substrate P.hysterophorus, P. hysterophorus with cow dung and P. hysterophorus nwith goat dropping was performed. Thus extracted and purified DNA from the microbial consortium was subjected for 16S rRNA analysis by terminal restriction fragment length polymorphism (T-RFLP).
Result: The minimum estimate number of bacterial community present among the samples with high methane yield revealed the presence of 18 and 14 different types of microbes correspondingly.
EnglishBioconversion, Co-additive, Methanogenic, Microbiome, P. hysterophorusINTRODUCTION
The diversity and functional analysis of the microbial community involved in the biogas technology is the principle aspect of upgrading the production efficiency in terms of methane yield. Hence a better understanding regarding the composition and abundance of the microorganisms involved in these systems becomes the need of the hour. Though there are reports of cultivable methanogens from diverse habitats (Garcia, 1990), cultivation of indigenous methanogens were unsuccessful (Williams and Crawford 1985; Goodwin and Zeikus, 1987). For the past few decades, researchers depend on culture independent methods of microbial community analysis using molecular techniques to isolate, identify and compare gene sequences (Atlas et. al., 1992; Madson, 1998; Schneegurt and Kulpa, 1998). Anaerobic digestion, an attractive option for the weed management at ambient environmental conditions offers technical and economical benefits. The potential limitations previously associated with weed biomass batch reactor operation have been largely overcome by optimizing various pretreatments and co-additive options. However, much scope still exists via application of the ambient microbial consortium which will allow in improvising the efficiency in terms of methane yield. In the present study, batch reactor fermenter operations were investigated by monitoring the effect of pretreatments and co-additives on reactor performance over hydraulic retention time of 35 days. DNA extracted from the whole microbial consortium was subjected to PCR amplification and 16S rRNA analysis by T-RFLP was performed.
MATERIALS AND METHODS
Batch Reactor Set-up
Biochemical Methane Potential (BMP) vials of 100ml were used as reactor bottles with P. Hysterophorus as substrate, which was pretreated (Ramya R and Shree M. P., 2014a). Inoculum substrate (I/S) ratio of 0.5%, 1.0%, 2.0% were done in triplicates. The inoculum was collected from a plant-litter based biogas plant, which was filtered with muslin cloth and stored at refrigeration temperature until use. Substrate along with co-additives (C/S ratio), cow dung and goat dropping at ratio of 1:0.25, 1:0.5 and 1:1 were performed in triplicates for a hydraulic retention time of 35 days in batch fermentation process at ambient temperature.
Biogas analysis
The composition of the biogas collected in the head space of the digester was analyzed using a Nano HP-1 Model BG 1000 Gas Chromatograph equipped with a Flame Ionization Detector (FID) and Thermal Conductivity Detector (TCD). Argon (Ar) was used as carrier gas with flow rate of 8ml/min. Oven temperature was set up to 60o C. Methanator temperature was set at 360o C. All the digesters were operated in triplicate. Biogas was sampled by directly inserting the intra venous syringe into the Anaerobic Digestion (AD) vial and volumetric composition of biogas was analyzed by using gas chromatography. Gas chromatograph (Nano HP-1) equipped with FID was used to analyze gas composition (Ramya R and Shree M. P., 2014b).
Extraction of metagenomic DNA from digested slurry
In this study, extraction of DNA of two samples PT4cCDI and PT4cGDIII were performed using the bacterial genomic DNA isolation kit RKN15 with slight modification. 1.5 ml of bacterial culture was taken in 2 ml vial after spinning the BMP vial at 70rpm for 10 mins. The supernatant thus obtained was spun at 10,000 rpm for 2 min. Supernatant was completely discarded. To the pellet 750 µl of 1X suspension buffer was added and subjected for vortexing. 5 µl of RNase was then added and incubated at 65°C for 10 min with intermittent mixing. To this, 1 ml of lysis Buffer was added and mixed by inverting the tube 10-12 times. The tubes were incubated at 65°C for 15 min and spun at 10,000 rpm for 5min. The clear lysate was loaded (unlysed Cells will settle as pellet leaving clear supernatant) onto spin column. The mini-spin column was placed into 2ml collection tube and 600µl of the lysate was loaded on the spin column each time, spun at 10,000 rpm for 1min at room temperature. The content of the collection tube was discarded and the spin column was placed back in the collection tube. 500 µl of 1X wash buffer was added to the column, spun at 10,000 rpm for 1 min at room temperature. The contents were discarded of the collection tube. The spin column was placed back in the same collection tube and the procedure was repeated. The empty column was spun at 10,000 rpm for 3 min at room temperature. 50 µl of warm elution buffer was added to the spin column placed in a fresh 1.5ml vial, at 65°C for 1 min and spun at 10,000 rpm for 1 min at room temperature. The elution was collected in the same vial. Thus obtained DNA concentration was determined by quantitative analysis on 1% (w/v) agarose gel with 0.5xTBE as electrode buffer as described elsewhere (Sun, et al., 2009).
T-RFLP analysis
The ~1.5 Kb bacterial 16s rRNA fragment was amplified from the metagenomic DNA extracts by using high–fidelity Polymerase Chain Reaction (PCR) polymerase as described by Sun, et. al., 2009, with the labelled primers marked with 6-carboxyfluorescein (FAM). The PCR products were subjected to restriction digestion with a 4-base cutter (HpaII). The fluorescent labelled t-RFs (terminal restriction fragments) were size separated on an ABI 3130 automated sequencer (Applied Biosystems) using an internal size standard (LIZ-500). t-RFLP (terminal restriction fragment length polymorphism) electropherograms were analyzed with GeneScan 3.7 software (Applied Biosystems) at chromous biotech Pvt Ltd, Bangalore (Wayan Suardana, 2014). The numbers of peaks obtained in t-RFs represents the minimum number of bacteria present in the sample.
RESULT
The biogas compositions of the samples subjected for AD are presented in Table 1 and 2. It is very evident from the GC result that the PT4 at the I/S ratio of 2%, C/S ratio of 1:1 with CD and C/S ratio of 1:0.25 with GD showed the maximum methane yield of 149.24% and 120.81%. Thus these two samples where further subjected for extraction of metagenomic DNA (Fig 1 and 2) and analysis by T-RFLP (Fig 3 and 4).
DISCUSSION
In our study, the DNA extracted from the BMP vials using slightly modified protocol of RKN15 kit gave us a good quality DNA which was amplified by PCR. T-RFLP is a technique widely used for study of microbial diversity in environmental samples (Salvador Embarcadero-Jimé- nex, et al., 2014). With the metagenomic DNAs obtained in this study, 35-500 T-RFs were obtained from the 16S rRNA amplicons digested by HpaII restriction endonuclease. Fig 3 and 4 indicated that the bacterial communities in the anaerobically digested P.hysterophorus present in the 100ml BMP vials were very diverse and composed of many species.
CONCLUSION
The effect of pretreatment and co-additive on the biomethanation efficiency of P.hysterophorus was studied by performing batch anaerobic digestion process experiment using BMP vials. The experimental results in this study demonstrated that the suitable pretreatment for biomethanation of P.hysterophorus for a HRT time of 35 days is PT4 with the maximum methane yield of 149.24% with CD as co-additive and 120.81% with GD as co-additive. The most important finding of this research is the microbial consortium involved in the biomethanation process. Therefore, the evaluation process with respect to the microbial consortium should be considered carefully. In the present study, extraction of high quality metagenomic DNA from anaerobically digested BMP vials was performed. Thus obtained DNA samples were subjected for PCR amplification and T-RFLP analysis, which showed the presence of diverse bacterial communities. Studies related to the diversity are still under progress.
6-carboxyfluorescein (FAM)-labeled primers gives blue color in Genescan analysis. (Blue colored peaks which corresponds to tRF’s are sized by internal size standard) Orange peaks are internal size standard (LIZ 500, Chromous)
The GeneScan™ 500 LIZ® Size Standard is a fifth dyelabelled size standard for the reproducible sizing of fragment analysis data. This size standard is used for fragments between 35 and 500 bp. The standard contains 16 LIZ® dye-labelled, single-stranded DNA fragments. Since the standard is labeled with the fifth dye, can genotype a greater number of markers in a given lane, compared to the four-dye system.
Size Fragments in the 35-500 Nucleotides Range:
GeneScan™ 500 LIZ® Size Standard is designed for sizing DNA fragments in the 35-500 nucleotides range and provides 16 single-stranded labeled fragment of: 35, 50, 75, 100, 139, 150, 160, 200, 250, 300, 340, 350, 400, 450, 490 and 500 nucleotides. The sizing curve generated from these fragments make the GeneScan™ 500 LIZ® Size Standard ideal for a variety of fragment analysis applications such as Microsatellites, Fragment Length Polymorphisms and Relative Fluorescent Quantitation. Each of the DNA fragments is labelled with the LIZ® fluorophore which results in a single peak when run under denaturing conditions. With the 5th dye LIZ® your marker fragments can be labelled with the dyes FAM™, VIC™, NED™ or PET®.
ACKNOWLEDGEMENT
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors/editors/publishers of all those articls, journals and books from where the literature for this article has been reviewed and discussed.
Source of Funding: No sources of funding
Conflicts of Interest: None
Englishhttp://ijcrr.com/abstract.php?article_id=687http://ijcrr.com/article_html.php?did=6871. Atlas R. M., Sayler G., Burlage R. S., and Bej A. K. Molecular approaches for environmental monitoring of microorganisms. BioTechniques, 1992;12:706-717.
2. Madsen E. L. Epistemology of environmental microbiology. Environ. Sci. Technol. 1998;32:429-439.
3. Garcia, J.L. Taxonomy and ecology of methanogens. FEMS Microbiol. Lett. 1990;87:297-308.
4. Goodwin S. and Zeikus J. G. Ecophysiological adaptations of anaerobic bacteria to low pH: analysis of anaerobic digestion in acidic bogs sediments. Appl. Environ. Microbiol. 1987;53:57-64.
5. Ramya. R, Shree. M. P. Comparative efficiency of pretreatment methods on Parthenium hysterophorus L., as a potential feed stock, “International Journal of Scientific and Research Publications, 2014.Vol. 4, Issue 9, September Edition”. pp 1-3 ISSN 2250-3153.
6. Ramya R, Shree M. P. Impact of C/N ratio of pretreated P. hysterophorus L., on Biomethanation, “Internation Journal of Scientific Research, 2014. Vol:III, Issue:XI, November Edition”, Journal DOI:10.15373/22778179.
7. Salvador Embarcadero-Jiménez, Feng Long Yang, Raquel Freye-Hernàndez, Yanelly Trujillo-Cabrera, Flor N. Rivera Orduña, Hong Li Yuan and En Tao Wang. An improved protocol for extraction of metagenomic DNA from high humus, alkaline and saline soil of Chinampa for T-RFLP fingerprinting analysis. British Microbiology Research Journal. 2014;4(7):821-830.
8. Schneegurt M. A. and Kulpa C. F. Review: The application of molecular techniques in environmental biotechnology for monitoring microbial systems. 1998;27:73-79.
9. Sun Y.M., Zhang N.N., Wang E.T., Yuan H.L., Yang J.S., Chen W.X. Influence of intercropping and intercropping plus rhizobial inoculation on microbial activity and community composition in rhizosphere of alfalfa (Medicago sativa L.) and Siberian wild rye (Elymus sibiricus L.) FEMS Microbiol. Ecol. 2009;70:62-70.
10. Wayan Suardana I. Analysis of Nucleotide Sequences of the 16S rRNA Gene of Novel Escherichia coli Strains Isolated from Feces of Human and Bali Cattle. Jornal of Nucleic acids, 2014. Vol (2014), Article ID 475745.
11. Williams R. T. and Crawford R. L. Methanogenic bacteria including an acid-tolerant strain, from peatlands. Appl. Environ. Microbiol. 1985;50: 1542-1544.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcareBIOCHEMICAL CHANGES HUMAN IMMUNODEFICIENCY VIRUS INFECTED PATIENTS BEFORE AND AFTER ANTIRETROVIRAL THERAPY
English4347Usha kiran P.English Komala P.EnglishObjectives: The aim of present study was to evaluate the levels of liver enzymes and lipid abnormalities in patients receiving antiretroviral therapy compared to controls. Lactic acidosis was one of the known complications of ART which is the major cause of discontinuation of treatment. Methods: Siemens Dimension Expand Plus fully automated random access analyzer was used for analyzing different parameters. Results: Mean plasma levels of ALT and AST were more in HIV infectedpatients than control group. These parameters were further increased in HIV infected patients after ART. Mean plasma levels of cholesterol and HDL are lower than control group in HIV infected patients before treatment. Cholesterol levels increased after HAART whereas HDL level decreased. Mean plasma levels of triglycerides, LDL and lactic acid are high in HIV infected patients than control and they are further increased after ART. Conclusion: Serum liver enzymes are significantly increased after ART and may be used as prognostic marker of lactic acidosis. This study helps in diagnosing lactic acidosis which can be treated successfully so as to render patients to adhere to treatment to decrease mortality.
EnglishHIV, ART, Lactic acid, Liver enzymes, Lipid profileINTRODUCTION
Most antiretroviral agents have been associated with hepatic toxicity. NRTIs can cause hepatic steatosis generally after more than 6 months of therapy1 , probably via mitochondrial toxicity. NRTIs can cause hepatitis in first 2-3 months of therapy, sometimes as part of a hypersensitivity reaction. Protease inhibitors can also cause hepatitis2 by an unknown mechanism, particularly in patients co-infected with hepatitis B or hepatitis C and with raised hepatic aminotransferases concentration and less commonly have been associated with variceal bleeding in patients with cirrhosis3 . Some cases of hepatitis with antiviral seem to represent a side effect of an improved immune response, where immune restoration leads to recognition of hepatitis B4 or C in chronic carriers and results in a clinical episode of hepatitis with sero conversion. What proportion of hepatic reaction to any retroviral is caused by such mechanism is not known. Unconjugated hyperbilirubinemia can occur with indinavir (about 7%) but does not represent liver toxicity5 . Pre – HAART dyslipidemia was caused by an initial decrease in serum levels of total cholesterol, HDL and LDL followed by increase in triacylglycerol during advanced stages of HIV infection.6, 7 The severity of observed dyslipidemia was related inversely to decreased CD4 lymphocyte count particularly when CD4 counts were below 200.8 In contrast, correlation between CD4 lymphocyte count / viral load levels and hyperlipidemia in patients taking PI containing HAART regimen have not been found.9, 10 Several cross-sectional studies have established a relationship between PI use and hypercholesterolemia and hypertriglyceridemia.11,12 Studies which followed patients before and after PI exposure noted higher cholesterol and triglyceride levels after PI initiation.13,14 The average increase of total cholesterol and triglyceride levels was 28% and 29% respectively from baseline pretreatment levels.15 Following a sample of 221 HIV- infected patients for 5 years, Tsiodras and colleagues 2000 found that PI therapy was associated with a 2.8 fold increased rate of hypercholesterolemia and a 6 fold increased rate of hypertriglyceridemia. Lactic acidosis Lactic acidosis results from an increase in blood lactate levels when lactate production exceeds consumption and body buffer systems become overburdened. Lactic acidosis is classified by Cohen and Woods into 2 broad categories Type A lactic acidosis is due to decreased tissue perfusion may be due to over production or underutilization. Type B2 lactic acidosis is due to drugs without evidence of poor tissue perfusion or oxygenation or with occult tissue hypo perfusion. NRTIs have been associated with functional and structural mitochondrial abnormalities leading to lactic acidosis.16 Mild to moderate asymptomatic hyperlactatemia has been frequently reported in patients treated with an estimated prevalence between 15% and 35%. On the contrary, symptomatic, severe hyperlactatemia and lactic acidosis are less common, with an incidence ranging from 1.7 to 25.2 cases per 1000 person- years of antiretroviral treatment, and are associated with a remarkable mortality rate, which varies from 30% to 60% in different studies.17
MATERIALS AND METHODS
The present study was conducted in department of Biochemistry, Rangaraya Medical College, Kakinada, Andhra Pradesh, India. The present study was undertaken to determine biochemical changes in HIV infected patients before and 2 months after ART in venous blood consisting of 100 patients. These values are compared with 50 normal healthy persons. All of these subjects were taken from ART centre, Government General Hospital, Kakinada. IRB/ Ethics Committee decided approval was not required for this study. Lactic acid is estimated by Marbach and Weil method. Principle: L-Lactate + NAD LDH Pyruvate + NADH + H+ Liver enzymes are estimated by Reitmane Frankel method. Cholesterol is estimated by CHOD-PAP method.
STATISTICAL METHODS
The statistical software used was graphpadquickcalcs. Student t test was used. RESULTS Mean plasma levels of ALT and AST were more (51.88 ± 28.52), (53.69 ± 31.25) than control group (30.14 ± 9.25), (23.02 ± 6.46). These parameters were further increased in HIV infected patients after ART (102.11 ± 29.50), (113.47 ± 45.34). Mean plasma levels of cholesterol were lower (158.40 ± 68.57), than the control group (175.14 ± 30.61) in patients before treatment and it is more than the control group after ART (217.10 ± 116.31). Mean plasma levels of triglycerides were more (117.86 ± 47.98) than the control group (92.36 ± 15.90) before ART and it is further increased after ART (165.71 ± 46.92). Contrarily the mean levels of HDL cholesterol were high in control group (34.30 ± 8.50) than in HIV infected patients (29.69 ± 10.97) and it is further decreased in HIV infected patients (20.11 ± 5.48) after ART. Mean levels of LDL cholesterol were high in HIV infected patients (73.68 ± 26.15) than the control (67.78 ± 14.62) and it is further increased in HIV infected patients after ART (111.69 ± 35.59). Mean levels of lactic acid were high in HIV infected patients (1.913 ± 1.066) than the control (1.398 ± 0.445) and it is further increased in HIV infected patients after ART (4.553 ± 0.987).
DISCUSSION
Highly Active Antiretroviral therapy (HAART) has led to decreased morbidity and mortality from HIV infection due to immune reconstitution and viral suppression and increased recognition of both acute and long term toxicities of ART. Hepatitis, lipid abnormalities and lactic acidosis are the common complications seen with HAART. In the present study serum ALT and AST values were significantly elevated in HIV infected patients when compared with controls (p< 0.001). Serum total cholesterol levels, tri-glyceride levels and LDL levels were also significantly elevated when compared with controls (p< 0.001). These results suggest that elevated serum liver enzymes and lipid levels are complications of ART. Similar observations are also made by M.Nunez, R.Lana, J.L.Mendoza, L.Marttin- Carbonero and V. Soriano. 18 The raised serum ALT and AST enzymes are markers of liver injury due to antiretroviral therapy. The total cholesterol and triglycerides which are increased are markers of cardiac risk for the patients receiving antiretroviral therapy in future on long term treatment. The increased cholesterol and triglyceride levels in turn lead to excess formation of pyruvate. This leads to excess production of lactate leading to lactic acidosis. Lactic acid levels are increased significantly (pEnglishhttp://ijcrr.com/abstract.php?article_id=688http://ijcrr.com/article_html.php?did=6881. M. Nunez and V. Soriano, “Hepatotoxicity of antiretrovirals: incidence, mechanisms and Management,” Drug Safety, vol.28, no. 1, pp. 53–66, 2005.
2. Lactic Acidosis and Hepatic Steatosis Associated with Use of Stavudine: Report of Four Cases Kirk D. Miller, MD; Miriam Cameron, MD; Lauren V. Wood et al; Ann Intern Med. 2000;133(3):192-196.
3. M. S. Sulkowski, D. L. Thomas, S. H. Mehta, R. E. Chaisson,and R. D. Moore, “Hepatotoxicity associated with nevirapine or efavirenz-containing antiretroviral therapy: role of hepatitis C and B infections,” Hepatology, vol. 35, no. 1, pp. 182–189,2002.
4. A. R. Lai, K. T. Tashima, and L. E. Taylor, “Antiretroviral medication considerations for individuals coinfected with HIV and hepatitis C virus,” AIDS Patient Care and STDs, vol.20, no. 10, pp. 678–692, 2006.
5. M. Bonacini, “Liver injury during highly active antiretroviral therapy: the effect of hepatitis C coinfection,” Clinical Infectious Diseases, vol. 38, supplement 2, pp. S104–S108, 2004.
6. Stein JH.Dyslipidemia in the era of HIV protease inhibitors. Prog Cardiovasc Dis. 2003 Jan-Feb; 45(4):293-304.
7. Penzak SR, Chuck SK, Stajich GV. Safety and efficacy of HMG CoA Reductase inhibitors for treatment of hyperlipidemia in patients with HIV infection. Pharmacotherapy, 2000; 1066-1071
8. Constans J, Pellegrin JL, Peuchant Eet al (1993) High plasma lipoprotein (a) in HIV –positive patients. Lancet 341; 1099-1100
9. Tsiodras S, Mantzoros C, Hammer S, Samore M. Effects of protease inhibitors on hyperglycemia, hyperlipidemia and lipodystrophy. Arch Intern Med.200; 160:2050-2056
10. Galli, M., Cozzi-Lepri, A., Ridolfo, A. L., Gervasoni, C., Ravasio,L., Corsico, L., et al. (2002). Incidence of adipose tissue alterations in first-line antiretroviral therapy: The LipoICoNaStudy.Archives of Internal Medicine162(22), 2621-2628.
11. Koppel K. Bratt G. Eriksson M. Sandstrom E. Serum lipid levels associated with increased risk for cardiovascular disease is associated with highly active antiretroviral therapy (HAART) in HIV-1 infection.Int J STD AIDS. 2000;11:451– 455. [PubMed]
12. Behrens G, Dejam A, Schmidt H,et al: Impaired glucose tolerance, beta cell function and lipid metabolism in HIV patients under treatment with protease inhibitors. AIDS 1999; 13: F63- F70
13. Sergerer, S and Bogner, JR and Walli, R and Loch, O and Goebel, FD, “Hyperlipidaemia under treatment with protease inhibitors”, Infection, vol. 27, 1999, p.77-81
14. Periard, D and Telenti, A and Sudre, P, “Atherogenic dyslipidemia in HIV-infected individuals treated with protease inhibitors. The Swiss HIV Cohort Study”, Circulation, vol. 100, 1999, p.700-705
15. Friis-Møller N1 , Weber R, Reiss P, et al.Cardiovascular disease risk factors in HIV patients--association with antiretroviral therapy. AIDS. 2003 May 23;17(8):1179-
16. Joly V, Flandre P, Meiffredy V et al. Assessment of lipodystrophy in patients previously exposed to AZT, ddI or ddC, but naive for d4T and protease inhibitors (PI), and randomized between d4T/3TC/ indinavir and AZT/3TC/ indinavir (NOVAVIR Trial). VIII Conference on Retroviruses and Opportunistic Infections. Chicago, IL, February, 2001 [Abstract 539].
17. Brinkman K, Hofstede H.J, Veerkamp M.J, Kolke H.J, Williams J.L and Wesserling P. (1998). Fatal lactic acidosis following HAART containing stavudine (d4T), lamivudine (3TC) and saquinavir. In program and Abstracts of the Twelfth World AIDS Conference, Geneve, 1998.Abstract 60998
18. M. Núñez, R. Lana, J. L. Mendoza, L. Martín-Carbonero, and V. Soriano, “Risk factors for severe hepatic injury after introduction of highly active antiretroviral therapy,” Journal of Acquired Immune Deficiency Syndromes, vol. 27, no. 5, pp. 426–431, 2001.
19. Jansen TC, van Bommel J, Bakker J. Blood lactate monitoring in critically ill patients: a systematic health technology assessment. Crit Care Med. 2009;3(10):2827–2839.
20. Dubé MP, Stein JH, Aberg JA, et al. Guidelines for the Evaluation and Management of Dyslipidemia in Human Immunodeficiency Virus (HIV)–Infected Adults Receiving Antiretroviral Therapy: Recommendations of the HIV Medicine Association of the Infectious Disease Society of America and the Adult AIDS Clinical Trials Group.* Clin Infect Dis. 2003;37:613–627.
21. Nerurkar PV, Pearson L, Frank JE, Yanagihara R, Nerurkar VR. Highly active antiretroviral therapy (HAART)-associated lactic acidosis: in vitro effects of combination of nucleoside analogues and protease inhibitors on mitochondrial function and lactic acid production. Cell Mol Biol. 2003; 49:1205–1211.
22. Miro O, Gomez M, Pedrol E, Cardellach F, Nunes V, Casademont J. Respiratory chain dysfunction associated with multiple mitochondrial deletions in antiretroviral therapyrelated lipodystrophy AIDS:18 August 2000 - Volume 14 - Issue 12 - pp 1855-1857 Research Letters.
23. Shikuma CM, Shiramiju B. Mitochondrial toxicity associated with nucleoside reverse transcriptase inhibitor therapy. Current Inf Dis Reports 2001; 3:553-560.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241624EnglishN2014December20HealthcarePREVALENCE OF METALLO BETA LACTAMASE PRODUCING PSEUDOMONAS AERUGINOSA IN HOSPITALIZED PATIENTS
English4853Vineeta MittalEnglish Fareya HaiderEnglishAims: Study the prevalence of multidrug resistant Ps. aeruginosa in hospitalized patients by antibiotic sensitivity test and detection of MBL producing Ps. aeruginosa by two phenotypic methods.
Methods: Total numbers of 83 samples were taken from patients admitted in wards of different departments. Specimens were inoculated on Mac Conkey agar and sheep blood agar. Identification of isolates was done by colony characteristics and biochemical reactions. These isolates were then subjected to susceptibility testing against various antibiotics by Kirby-Bauer disc diffusion test as per CLSI guidelines. Imipenem resistant isolates were selected for the detection of metallo-beta-lactamases production by two phenotypic methods ie Modified Hodge Test and Imipenem-EDTA combined disc test and confirmed by E test
Results: In this study eleven imipenem resistant isolates were found. Out of which six were found positive for MBL by modified Hodge test, Disc synergy test and MBL E test .These MBL positive isolates were mostly from urine samples.
Conclusion: Clinical microbiology laboratories should use proper phenotypic screening methods in all imipenem resistant isolates. In patients admitted in hospital, there should be careful and conservative use of Carbapenems. Most of MBL producing Ps.aeruginosa were isolated from urine specimen coming from surgery wards, so strict vigilance of catheterized operated patients is needed..
EnglishCarbapenem resistance, Metallo-beta-lactamase, Pseudomonas aeruginosaINTRODUCTION
Carbapenems are used as last choice for treating serious infections caused by multidrug resistant organisms especially Pseudomonas aeruginosa (Ps.aeruginosa) and Acinetobacter spp . But later on there has been a substantial rise in the reporting of carbapenemase, most notably Metallo β-Lactamases1 . Metallo β lactamase (MBL) belongs to Ambler class B of β-lactamase which requires divalent cations of zinc as cofactors for enzyme activity. They can hydrolyse a wide variety of β-lactams, including penicillins, cephalosporins and carbapenemes.2 MBL-producing Ps.aeruginosa isolates have been reported to be important causes of nosocomial infections across the world. These constitute 20-42 per cent of all nosocomial isolates.3. We undertook this study to determine prevalence of MBL producing Ps. aeruginosa in patients admitted to tertiary care centre over a period of one year.
MATERIAL AND METHOD
Setting: The study was carried out in the department of microbiology of a tertiary care centre of Lucknow, India. Clinical samples were taken from patients admitted in wards of different departments of a tertiary care centre. Duration of study: One year, from February 2011 to March 2012 Study design: Epidemiological Study Sample size: Total 83 isolates of Ps.aeruginosa were enrolled in one year in this study Selection of subjects: Clinical details were collected from all patients whose culture was positive for Ps.aeruginosa. We included patients who were admitted in the medical and surgical wards and ICU. Information regarding patient’s age, sex, and etiology of disease was noted. Clinical details and history of antibiotic intake at the time of admission were taken. Informed written consent was taken from each patient. Specimen collection: Specimens were processed according to site of lesion e.g. respiratory secretions, swabs pus/ wound, urine, blood culture or CSF. With Universal safety precautions, samples were collected from hospitalized patients, transported and processed in the laboratory without delay. Specimen processing Isolation: Specimens were inoculated on MacConkey agar and sheep blood agar. Plates were seen for growth after 48 hrs of aerobic incubation at 37o C. Blood culture bottle was incubated at 37ºC for 24 hr then subculture on Mac conkey agar and sheep blood agar media on next day and after 7 days. Isolate was gram stained. Identification: Identification of Ps.aeruginosa, was done by biochemical tests as Catalase, Oxidase, Indole, Methyl red, NO3 reduction, Lactose and Maltose. Antibiotic sensitivity test: Antibiotic sensitivity test was done by disk diffusion methods on Mueller- Hinton - agar (Hi-Media, Mumbai) recommended by CLSI incorporating standard stains of P. aeruginosa (ATCC 27853) with the following antibiotics discs amikacin (30 µg), gentamicin (10 µg), netilmicin (30 µg), tobramicin (10 µg)],cephalosporin’s [cefepime (30 µg), ceftazidime (30 µg), ceftriaxone (30 µg)], floroquinolones [ciprofloxacin (5 µg),],carbapenems [imipenem (10 µg), meropenem (10 µg)], and piperacillin/tazobactum (100 µg /10 µg) and colistin (10 µg ). The zone of inhibition was determined after incubation of 24 hours at 37°C. 2. MIC: MICs of Imipenem was determined by E Test (BioMeriux). [Figure 1] Result was recorded after incubation for 18 hrs at 37o C. Eleven Imipenem resistant isolates showed MIC > 4 µg/ml which is resistant according to CLSI guidelines 2012. 4 3. MBL identification: All Imipenem resistant isolates were screened for carbapenemase by modified Hodge test and for MBL production by double disc synergy test. MBL production was confirmed by E test. A. Modified Hodge test5 Modified Hodge test was carried out on Mueller -Hinton agar. The plate was inoculated using a cotton swab dipped in an overnight culture suspension of E.coli ATCC25922. (Opacity of the tube was adjusted by comparing with a 1:10 dilution of 0.5 McFarland opacity standards). After brief drying, 10 µg imipenem disc was placed at the center of the plate and test strain was streaked from the edge of the disc to the periphery of the plate in four different directions. After overnight incubation the plates was observed for the presence of a clover leaf of shape zone of inhibition. The plates with such zones were interpreted as Modified Hodge test positive. [Figure 2] B. Zone enhancement with EDTA (Double Disc Synergy test)5 Impregnated imipenem and Ceftazidime Discs: Ps.aeruginosa was inoculated on to plates with Muller-Hinton agar as recommended by CLSI. A 0.5 m EDTA.2H2 O mix in 1000 ml of distilled water and adjust it to pH 8.0 by using NaOH. The mixture was sterilized by autoclaving. Two 10 µg imipenem discs and two-30µg ceftazidime disc was placed on the surface of an agar plate and EDTA solution was added to one of them to obtain a desired concentration of 750µg. The inhibition zone of imipenem, ceftazidime, and imipenem EDTA and ceftazidime EDTA disks were compared after 16-18 hrs of incubation at 35 º C.[Figure 3] C. MBL E test (HiMedia) E-test metallo-beta-lactamase strips (Hi Media) consist of a double sided seven dilution range of imipenem IP (4 -256 µg/ml) and IP (1 - 64 µg/ml) overlaid with a constant gradient of EDTA. Individual colonies were picked from overnight agar plates and suspended in 0.85% saline to a turbidity of 0.5% McFarland’s standards. A sterile cotton swab was dipped into the inoculum suspension. The excess moisture was allowed to be absorbed for about 15 min before the MBL E-test strip was applied. Plates were incubated for 16 to 18 hrs at 37°C. The MIC end points were read where the inhibition ellipses intersected the strip. When the ratio of the value obtained for Imipenem (IPM): the value of IPM + EDTA is more than 8 or zone was observed on the side coated with IPM+EDTA and no zone was observed on the opposite side created with IPM, the culture was interpreted as MBL positive. [Figure 4] Ethics: The study was carried out after approval with institutional review board and taking informed consent. Statistical analysis: Statistical analysis was done by Gaussian test of proportion and Fisher’s exact test.
RESULTS
In our study 83 Ps. aeruginosa strains were isolated from various clinical specimens which were identified by biochemical methods. These Ps. aeruginosa isolates showed resistance to even the most recent antibiotics like third generation cephalosporins, antipseudomonal penicillins, aminoglycosides and flouroquinolones. In this study, around 22 isolates (26.5%) were MDR ie. resistant to all antibiotics. Colistin was most effective antibiotic recording 0% resistance. Ceftazidine, antipseudomonal reserve drug was found 33% resistant in this study. Another drug is piperacillintazobactum .We observed 29% resistant against this drug. Among aminoglycosides Netilmycin showed least resistance (27%) as compared to tobramycin (33%), amikacin (34%) and Gentamycin (37%). Quinolone, particularly ciprofloxacin was sensitive in 65% of Ps. aeruginosa. [Figure 5] In this study carbapenem resistance was found in eleven (13%) isolates. All the eleven imipenem resistant isolates were resistant to most of the antimicrobials tested including Amikacin, Gentamicin, Ciprofloxacin, Ceftriaxone, Ceftazidime, Cefipime, Tobramycin, Netilmycin and Pipercillin- tazobactum. MIC of Imipenem by E test was done. Complete correlation with dilution and diffusion method was found. In these eleven imipenem resistant isolates, six were positive for MBL by Modified Hodge test, and Imipenem -EDTA disc synergy tests, Ceftazidime EDTA disc synergy tests and five negative by all tests. These six isolates were found positive by MBL E test (Hi Media) for MBL production. DISCUSSION Ps. aeruginosa is a pathogen associated with numerous nosocomial infections in immuno compromised patients. Ps. aeruginosa strains are intrinsically resistant to various antimicrobial agents. Nowdays, Ps. aeruginosa isolates are resistance to the most recent antibiotics also. . In this study, 22 out of 83 isolates were resistant to all antibiotics except colistin which is major concern emerging from our study. Carbapenems are the drugs of choice for multidrug resistant Ps. aeruginosa and ESBL producing organisms. However, resistance to carbapenems due to reduced uptake of drug leads to imipenem/meropenem resistant isolates.6 In this study carbapenem resistance was found in 13% isolates and all the 11 imipenem resistant isolates were MDR. In this study most of the carbapenem resistance isolates were from surgery wards followed by NICU, medicine ward. [Table1] Ps. aeruginosa producing MBL was first reported from Japan in 1991.7 Navneeth et al 8 reported MBL production in Ps. aeruginosa first time from India in 2002. They reported 12% prevalence of MBL producing Ps. aeruginosa. In other studies of India prevalence ranging from 8 to 14% has been reported.9-11. In this study the overall prevalence of MBL positives Ps. aeruginosa was 7.23%. There are various methods by which MBL production can be identified. In which PCR is most sensitive and accurate method but PCR can identify specific type of MBL only and PCR could not be done in all laboratories due to its cost effectiveness. So different phenotypic methods are useful for MBL detection like modified Hodge test, imipenem-EDTA combined disk test, double-disk synergy test using imipenem or ceftazidime and EDTA and E-test.5, 12-13, In this study we used three different methods for detection of MBL producing Ps. aeruginosa; Modified Hodge test, Double disk synergy test by Imipenem and Ceftazidime with EDTA and MBL- E-test.[Figure 6] A total of 83 strains were isolated in this study over a period of one year. Eleven (13.25%) were resistant to Imipenem and six (7.23%) were found to be MBL producers by Double disk synergy test. In this study all the six isolates gave MBL positive results by both ImipenemEDTA disc synergy test and Ceftazidime EDTA disc synergy test. Bashir etal 14 showed different results by both methods in their study. This study is more or less similar to study conducted by Behera200812 who reported 14.47% imipenem resistances and 10.52% MBL production in Ps. aeruginosa. All the six isolates on which MBL-E-test was performed were MBL positive. Thus 100% positive results were obtained in this study which is comparable to various studies. In other studies, E test had a sensitivity of 92% 15 and 96% 16 in detecting MBL. Behra etal found 100% positivity by E test.12 In this study we screened carbapenem resistant isolates only which might have accounted for very high sensitivity of the test. Double disk synergy test and E-test were found to be equally sensitive in detecting MBL positive isolates. E-test is a simple screening test and can be performed along with routine susceptibility testing. The maximum number of MBL positive isolates were obtained from urine (85%) followed by pus swab (15%). Hirakata et al. (1998)11 reported in their study that the predominant source of isolation for MBL positive Ps. aeruginosa was urinary tract (40.0%) followed by respiratory tract (18.8%) and abscesses, pus and wounds (15%) .In the present study 6.02% MBL positive isolates were recovered from urine (85% of total MBL positive isolates) and the association was statistically significant. [Table 2] Most of these patients were having indwelling urinary catheter. This finding is consistent with another study reported by Hirakata et al. (2003). 16 MBL producing isolates are associated with a higher morbidity and mortality. Moreover given the fact that MBLs will hydrolyze all classes of ß-lactams and that were several years away from the development of a safe therapeutic antibiotic; their continued spread would be a clinical disaster.13 The occurrence of an MBL positive isolate poses not only a therapeutic problem but is also a serious concern for infection control management. As a result of being difficult to detect, such organisms pose significant risks particularly due to their role in unnoticed spread within institutions and their ability to participate in horizontal MBL gene transfer, with other pathogens in the hospital.17 In this study also, all the patients having MBL positive Ps. aeruginosa showed severe morbidity and did not recover well and were shifted to higher tertiary centre.
CONCLUSION
Clinical microbiology laboratories should use proper phenotypic screening methods in all imipenem resistant isolates. In patients admitted in hospital, there should be careful and conservative use of Carbapenems. Most of MBL producing Ps.aeruginosa were isolated from urine specimen coming from surgery wards, so strict vigilance of catheterized operated patients is needed.
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