International Journal of Current Research and Review
ISSN: 2231-2196 (Print)ISSN: 0975-5241 (Online)
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IJCRR - 4(11), June, 2012

Pages: 43-51

Date of Publication: 18-Jun-2012


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DETECTION OF VIRUS STRAIN THAT CAUSED FOOT AND MOUTH DISEASE OF BASRAH MARSHES CATTLE BY USING PCR TECHNIQUE

Author: Khitam Jassim Salih, Majeed Hussein Majeed

Category: Healthcare

Abstract:Although, the disease has been controlled successfully in many parts of the world by regular vaccination
of susceptible animals and slaughtering of infected animals, no country has been considered safe, because
of the highly contagious nature and rapid spread of the infection for the effective control of the disease,
outbreaks should be detected at an early stage and persistent infections should also be recognized to
prevent further transmittance. The purpose of this study was to determine the virus strain that caused
FMD in cattle of Basrah marshes by amplified VP3gene in seven strains of FMDV deposited in the Gen
Bank database. The results revealed that the O strain type was appear in a total of cases (100%) of the
virus strain that caused foot and mouth disease in cattle of Basrah marshes by amplified VP3gene from
seven serotype of FMDV. From the total cases 8% were ASIA1 serotype , 4% SAT1, 2% SAT2, while
the other strains A, C and EUR were 0% . The results showed interaction among the strains in appearance
of FMD, the interaction between O and ASIA1strain was 7%; O and SAT1 was 4%; O and SAT2 was
2%; O, ASIA1 and SAT1 were 4%; O, ASIA1 and SAT2 were 2%; O, ASIA1, SAT1 and SAT2 were
2%. The interaction between ASIA1 and SAT1 was 4%; ASIA1 and SAT2 was 2%; ASIA1 , SAT1 and
SAT2 were 2%.

Keywords: FMD, Bsrah marshes, PCR, Viruse strains of Foot and Mouth Disease

Full Text:

INTRODUCTION Foot-and-mouth disease or hoof-and-mouth disease (Aphtae epizooticae) is an infectious and sometimes fatal viral disease that affects clovenhoofed animals, including domestic and wild bovids. The virus causes a high fever for two or three days, followed by blisters inside the mouth and on the feet that may rupture and cause lameness. Foot-and-mouth disease is a severe plague for animal farming, since it is highly infectious and can be spread by infected animals through aerosols, through contact with contaminated farming equipment, vehicles, clothing or feed, and by domestic and wild predators (CFIA, 20011). The FMD virus is a member of the genus Aphthovirus in the family Picornaviridae. There are seven immunological distinct serotypes O, A, C, SAT1, SAT2, SAT3 and Asia 1and over 60 strains within these serotypes(Knowles et.al., 2003). The virus responsible for the disease is a picorna virus, the prototypic member of the genus Aphthovirus. Infection occurs when the virus particle is taken into a cell of the host. The cell is then forced to manufacture thousands of copies of the virus, and eventually bursts, releasing the new particles in the blood. The virus is highly variable(Martinez-Salas et.al.,2008). FMD occurs throughout much of the world, and whilst some countries have been free of FMD for some time, its wide host range and rapid spread represent cause for international concern. After World War II, the disease was widely distributed throughout the world. In 1996, endemic areas included Asia, Africa, and parts of South America(FMD, 2007). FMD generally involves mortality rates below 5%, but even so it is considered the most important disease of farm animals since it causes huge losses in terms of livestock productivity and trade. Although FMDV rarely causes death in adult animals, the virus can cause severe lesion in the myocardium of young animals, leading to high mortality rates (Sharma and Das, 1984; Domingo et.al., 1990 and Woodbury, 1995) Aim of study: The purpose of this study was to determine the virus strain that caused FMD in cattle of Basrah marshes by amplified VP3gene in seven strains type of FMDV deposited in the GenBank database. MATERIALS AND METHODS Sampling fluid from vesicles and saliva: One hundred cases of cattle(cows and buffalo) infected with FMD were used to collect the fluid from vesicle and saliva using a sterile tubes, needles and syringes. The fluid kept in transport medium (normal saline at pH 7-8.in 4-10 °C) and transport to laboratory within 24 hours . RNA Extraction: RNA samples were extracted using the total RNA Mini kit (tissue) following the manufacturer's instructions. Briefly, 400 μl of the RB buffer that is included in the kit added to the tubes that containing the FMD fluid ,then 4 μl of ß-mercaptoethanol followed by 400 μl of 70% ethanol, then transferred to a Mini RNase column previously inserted into a 2ml collecting tube. RNA was immobilized in the column by centrifugation, sequentially washed, and diluted in 50 μl of RNase free water. Reverse transcription polymerase chain reaction: The following primers were used F5'- ACTGGGTTTTACAAACCTGTGA-3' and R5'- GCGAGTC CTGCCACGGA-3' along with the probe 5'-TCCTTTGCACGCCGTGGGAC-3'in the one-step RT-PCR amplification started with reverse transcription for 1 hr at 60°C, followed by PCR with the following parameters: 55 cycles of 2 sec at 95 °C and 30 sec at 60°C. (Knowles et al ,2005). The amplified PCR products (672 bp) of the expected length were subjected to electrophoresis in a 1% agarose gel and visualized by staining with ethidium bromide under UV light. The other RT-PCR protocol for VP3 gene amplification by used a kit of green master mix, the reaction mix include green master mix 12.5μl, forward primer and reverse primer(modified from)(Gelagay et.al.,2009). (table:1) each 1μl, DNA 5μl, D.W. 5.5μl, then PCR amplification according to the following thermal profile: initial denaturation at 95°C for 5 min; 94°C for 30 sec, annealing at 50°C for 30 sec and extension at 72°C for 2.5 min, for 30 cycles. The final extension step of 72°C for 10 min. The products were 320bp analyzed by 1% agarose gel electrophoresis and visualized under UV light after staining with ethidium bromide 

RESULTS AND DISCUSSION
The primary diagnosis according to the case history and the cardinal signs and symptoms of FMD which are wobble between increase the body temperature ,salivation, anorexia, laminas and appear of vesicles in mouth and foot(between digits space of hooves); the FMD was determine (figure.1-8).

The laboratory diagnosis depended on PCR technique by amplified of the 3AB gene of FMDV which is a member of the genus Aphthovirus in the family Picornaviridae. The whole volume of gene were 672 bp (Fig 9,10) which fixed that the infections were FMD not another diseases. The results revealed that the O strain type was appear in a total of cases (100%) of the virus strain that caused foot and mouth disease in cattle of Basrah marshes by amplified VP3gene from seven strain type of FMDV(figure 11.table 2).From the total cases 8% were Asia1 strain type(figure 12. table 2) 4% SAT1, 2% SAT2, (figure 13,14.table 2)while the other strains A, C and EUR were 0% (figure.15- 17.table 2). The results showed interaction among the strains in appearance of FMD, the interaction between O and ASIA1strain was 7%; O and SAT1 was 4%; O and SAT2 was 2%; O, ASIA1 and SAT1 were 4%; O, ASIA1 and SAT2 were 2%; O, ASIA1, SAT1 and SAT2 were 2%. The interaction between ASIA1 and SAT1 was 4%; ASIA1 and SAT2 was 2%; ASIA1 , SAT1 and SAT2 were 2% (table. 2).

Infection with foot and mouth disease tends to occur locally, the virus is passed on to susceptible animals through direct contact with infected animals or with contaminated pens or vehicles used to transport livestock. The clothes and skin of animal handlers such as farmers, standing water, and uncooked food scraps and feed supplements containing infected animal products can harbor the virus as well. The absent of control measures such as quarantine and destruction of infected livestock, and export bans for meat and other animal products to countries not infected with the disease. Almost the viral disease laboratory diagnosed by use the ELISA assay, but this way of laboratory lack of the high sensitivity, so the PCR is the standard laboratory assays which applied to detect FMDV by detect DNA/RNA. Although the program of the vaccination against FMD is in continue in Iraq but this disease become endemic specially in marshes cattle. The failed in immunization may be due to lack the athletic with the virus strain that endemic in regions or due to activate the strain which is occurred by the vaccine not originally present in the region; Also one of the difficulties in vaccinating against FMD is the huge variation between and even within serotypes. The lose of crossprotection between strain types which meaning that a vaccine for one serotype won't protect against any others and in addition, two strains within a given serotype may have nucleotide sequences that differ by as much as 30% for a given gene; so the FMD vaccines must be highly specific to the strain involved. As we know the vaccination only provides temporary immunity that lasts from months to years and this idea agree with Tamilselvan et.al., (2009) the main constraints in controlling this disease and why it is considered as the most dreaded viral disease are its high contagiousness, wide geographical distribution, broad host range, ability to establish carrier status, antigenic diversity leading to poor cross-immunity, and relatively short duration of immunity. Poor surveillance and diagnostic facilities as well as inadequate control programs are major problems in control of this disease in the country. Although, the disease has been controlled successfully in many parts of the world by regular vaccination of susceptible animals and slaughtering of infected animals, no country has been considered safe, because of the highly contagious nature and rapid spread of the infection for the effective control of the disease, outbreaks should be detected at an early stage and persistent infections should also be recognized to prevent further transmittance. These can be achieved when vaccination is regular and effective and when diagnostic tools available are specific and sensitive and at the same time rapid(Bruner and Gillespie, 1973). Analysis of the viral genome is importance to monitor the field isolates in areas where the disease is endemic The virus particle which sediments at 146S consists of a single stranded positive sense RNA molecule of about 8.5 kb with a molecular weight of 2.6 ´ 106 daltons enclosed in a capsid which is composed of 60 copies each of four structural proteins named VP1, VP2, VP3 and VP4. VP1 is exposed on the surface of the virion and has immunogenic property. (Suryanarayana et.al., 1999). The results of the current study secure the O strain is the main strain that causes FMD in Iraq marshes cattle depend on genetic diagnosis of VP3 gene. Also the results showed that the other strain such as ASIA1 ,SAT1 and SAT2 respectively are combined with strain O in FMD accident. While the genetic diagnosis that used in the current study discovered the strains A,C and EUR not have any role in FMD occurrence. According to Global Animal Health–International Disease Monitoring Preliminary Outbreak Assessment(2009) the middle east specially Iran was endemic in FMD strain type O and Iraq not endemic with this disease, so we think the main causes that make Basrah marshes cattle become endemic in FMD from 2009-2011 is as a result of animal contraband between Iran and Iraq , random greasing with neighbouring countries, contaminated of marshes water that are link and sharing between Iran and Iraq and used the vaccine that prepare in Iran which contain different strain.

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

References:

1. Bruner DW and Gillespie JH (1973). The family Picornaviridae. in Hagan’s Infectious Disease of Domestic Animals, pp. 1207– 1028, 6th edition,.

2. Domingo E, Mateu MG, Martínez MA, Dopazo J, Moya A and . Sobrino F (1990). Genetic variability and antigenic diversity of foot-and-mouth disease virus, in Applied Virology Research, vol. 2, pp. 233–266.

3. Gelagay A, Mana M , Esayas G, Berhe GE, Tesfaye R, Mesfin S, Nigel PF, Jemma W, Geoffrey HH, and Nick J (2009). Genetic Characterization of Foot-and-Mouth Disease Viruses, Ethiopia, 1981–2007. Emerging Infectious Disease Journal. Volume 15:1-5

4. Veterinary Science Team(2009). Global Animal Health–International Disease Monitoring Preliminary Outbreak Assessment Reference (http://archive.defra.gov.uk/foodfarm /farmanimal /diseases/monitoring/documents/fmd-meupdate-: VITT/1200 FMD in Middle East Date

5. Foot and Mouth Disease. Washington State Department of Health. March 2002. Archived from the original on (2007)(ttp://www. doh.wa .gov/ehp/ts/zoo/foot-and- mouth-disease)

6. Canadian Food Inspection Agency (2011).(http://www.inspection. gc.ca/english/anima /heasan/disemala/fmdfie/questionse.shtm.

7. Knowles NJ, Samuel AR(2003). Molecular epidemiology of foot-and-mouth disease virus. Virus Res ;91:65–80).

8. Martinez-Salas E, Saiz M, Sobrino F (2008). "Foot-and-Mouth Disease Virus". Animal Viruses: Molecular Biology. Caister Academic Press. pp. 1–38

9. Sharma PK. and Das SK (1984). Occurrence of foot-and-mouth disease and distribution of virus type in the hill states of North Eastern region of India, Indian Journal of Animal Sciences, vol. 4, pp. 117–118.

10. Suryanarayana VV., Pradeep B , Reddy GR and Misra LD (1999). Serotyping of footand-mouth disease virus from aerosols in the infected area Indian Veterinary Research Institute, Hebbal, Bangalore 560 024, India. pp1-4 online

11. Tamilselvan RP, Sanyal A De, and Pattnaik B. (2009). Genetic transitions of Indian serotype O Foot and Mouth Disease Virus isolates responsible for field outbreaks during 2001–2009: a brief note: OIE/FAO Reference laboratories network meeting: New Delhi, India, pp. 11-13

12. Woodbury EL (1995). A review of the possible mechanisms for the persistence of foot-and-mouth disease virus,? Epidemiology and Infection, vol. 114, no. 1, pp. 1–

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Dr. Pramod Kumar Manjhi joined Editor-in-Chief since July 2021 onwards

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Awards, Research and Publication incentive Schemes by IJCRR

Best Article Award: 

One article from every issue is selected for the ‘Best Article Award’. Authors of selected ‘Best Article’ are rewarded with a certificate. IJCRR Editorial Board members select one ‘Best Article’ from the published issue based on originality, novelty, social usefulness of the work. The corresponding author of selected ‘Best Article Award’ is communicated and information of award is displayed on IJCRR’s website. Drop a mail to editor@ijcrr.com for more details.

Women Researcher Award:

This award is instituted to encourage women researchers to publish her work in IJCRR. Women researcher, who intends to publish her research work in IJCRR as the first author is eligible to apply for this award. Editorial Board members decide on the selection of women researchers based on the originality, novelty, and social contribution of the research work. The corresponding author of the selected manuscript is communicated and information is displayed on IJCRR’s website. Under this award selected women, the author is eligible for publication incentives. Drop a mail to editor@ijcrr.com for more details.

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‘Emerging Researcher Award’ is instituted to encourage student researchers to publish their work in IJCRR. Student researchers, who intend to publish their research or review work in IJCRR as the first author are eligible to apply for this award. Editorial Board members decide on the selection of student researchers for the said award based on originality, novelty, and social applicability of the research work. Under this award selected student researcher is eligible for publication incentives. Drop a mail to editor@ijcrr.com for more details.


Best Article Award

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A Study by M. Muthu Uma Maheswari et al. entitled "A Study on C-reactive Protein and Liver Function Tests in Laboratory RT-PCR Positive Covid-19 Patients in a Tertiary Care Centre – A Retrospective Study" is awarded Best Article of Vol 13 issue 06 Special issue Modern approaches for diagnosis of COVID-19 and current status of awareness
A Study by Gainneos PD et al. entitled "A Comparative Evaluation of the Levels of Salivary IgA in HIV Affected Children and the Children of the General Population within the Age Group of 9 – 12 Years – A Cross-Sectional Study" is awarded Best Article of Vol 13 issue 05 Special issue on Recent Advances in Dentistry for better Oral Health
A Study by Alkhansa Mahmoud et al. entitled "mRNA Expression of Somatostatin Receptors (1-5) in MCF7 and MDA-MB231 Breast Cancer Cells" is awarded Best Article of Vol 13 issue 06
A Study by Chen YY and Ghazali SRB entitled "Lifetime Trauma, posttraumatic stress disorder Symptoms and Early Adolescence Risk Factors for Poor Physical Health Outcome Among Malaysian Adolescents" is awarded Best Article of Vol 13 issue 04 Special issue on Current Updates in Plant Biology to Medicine to Healthcare Awareness in Malaysia
A Study by Kumari PM et al. entitled "Study to Evaluate the Adverse Drug Reactions in a Tertiary Care Teaching Hospital in Tamilnadu - A Cross-Sectional Study" is awarded Best Article for Vol 13 issue 05
A Study by Anu et al. entitled "Effectiveness of Cytological Scoring Systems for Evaluation of Breast Lesion Cytology with its Histopathological Correlation" is awarded Best Article of Vol 13 issue 04
A Study by Sharipov R. Kh. et al. entitled "Interaction of Correction of Lipid Peroxidation Disorders with Oxibral" is awarded Best Article of Vol 13 issue 03
A Study by Tarek Elwakil et al. entitled "Led Light Photobiomodulation Effect on Wound Healing Combined with Phenytoin in Mice Model" is awarded Best Article of Vol 13 issue 02
A Study by Mohita Ray et al. entitled "Accuracy of Intra-Operative Frozen Section Consultation of Gastrointestinal Biopsy Samples in Correlation with the Final Histopathological Diagnosis" is awarded Best Article for Vol 13 issue 01
A Study by Badritdinova MN et al. entitled "Peculiarities of a Pain in Patients with Ischemic Heart Disease in the Presence of Individual Combines of the Metabolic Syndrome" is awarded Best Article for Vol 12 issue 24
A Study by Sindhu Priya E S et al. entitled "Neuroprotective activity of Pyrazolone Derivatives Against Paraquat-induced Oxidative Stress and Locomotor Impairment in Drosophila melanogaster" is awarded Best Article for Vol 12 issue 23
A Study by Habiba Suhail et al. entitled "Effect of Majoon Murmakki in Dysmenorrhoea (Usre Tams): A Standard Controlled Clinical Study" is awarded Best Article for Vol 12 issue 22
A Study by Ghaffar UB et al. entitled "Correlation between Height and Foot Length in Saudi Population in Majmaah, Saudi Arabia" is awarded Best Article for Vol 12 issue 21
A Study by Siti Sarah Binti Maidin entitled "Sleep Well: Mobile Application to Address Sleeping Problems" is awarded Best Article for Vol 12 issue 20
A Study by Avijit Singh"Comparison of Post Operative Clinical Outcomes Between “Made in India” TTK Chitra Mechanical Heart Valve Versus St Jude Mechanical Heart Valve in Valve Replacement Surgery" is awarded Best Article for Vol 12 issue 19
A Study by Sonali Banerjee and Mary Mathews N. entitled "Exploring Quality of Life and Perceived Experiences Among Couples Undergoing Fertility Treatment in Western India: A Mixed Methodology" is awarded Best Article for Vol 12 issue 18
A Study by Jabbar Desai et al. entitled "Prevalence of Obstructive Airway Disease in Patients with Ischemic Heart Disease and Hypertension" is awarded Best Article for Vol 12 issue 17
A Study by Juna Byun et al. entitled "Study on Difference in Coronavirus-19 Related Anxiety between Face-to-face and Non-face-to-face Classes among University Students in South Korea" is awarded Best Article for Vol 12 issue 16
A Study by Sudha Ramachandra & Vinay Chavan entitled "Enhanced-Hybrid-Age Layered Population Structure (E-Hybrid-ALPS): A Genetic Algorithm with Adaptive Crossover for Molecular Docking Studies of Drug Discovery Process" is awarded Best article for Vol 12 issue 15
A Study by Varsha M. Shindhe et al. entitled "A Study on Effect of Smokeless Tobacco on Pulmonary Function Tests in Class IV Workers of USM-KLE (Universiti Sains Malaysia-Karnataka Lingayat Education Society) International Medical Programme, Belagavi" is awarded Best article of Vol 12 issue 14, July 2020
A study by Amruta Choudhary et al. entitled "Family Planning Knowledge, Attitude and Practice Among Women of Reproductive Age from Rural Area of Central India" is awarded Best Article for special issue "Modern Therapeutics Applications"
A study by Raunak Das entitled "Study of Cardiovascular Dysfunctions in Interstitial Lung Diseas epatients by Correlating the Levels of Serum NT PRO BNP and Microalbuminuria (Biomarkers of Cardiovascular Dysfunction) with Echocardiographic, Bronchoscopic and HighResolution Computed Tomography Findings of These ILD Patients" is awarded Best Article of Vol 12 issue 13 
A Study by Kannamani Ramasamy et al. entitled "COVID-19 Situation at Chennai City – Forecasting for the Better Pandemic Management" is awarded best article for  Vol 12 issue 12
A Study by Muhammet Lutfi SELCUK and Fatma entitled "Distinction of Gray and White Matter for Some Histological Staining Methods in New Zealand Rabbit's Brain" is awarded best article for  Vol 12 issue 11
A Study by Anamul Haq et al. entitled "Etiology of Abnormal Uterine Bleeding in Adolescents – Emphasis Upon Polycystic Ovarian Syndrome" is awarded best article for  Vol 12 issue 10
A Study by entitled "Estimation of Reference Interval of Serum Progesterone During Three Trimesters of Normal Pregnancy in a Tertiary Care Hospital of Kolkata" is awarded best article for  Vol 12 issue 09
A Study by Ilona Gracie De Souza & Pavan Kumar G. entitled "Effect of Releasing Myofascial Chain in Patients with Patellofemoral Pain Syndrome - A Randomized Clinical Trial" is awarded best article for  Vol 12 issue 08
A Study by Virendra Atam et. al. entitled "Clinical Profile and Short - Term Mortality Predictors in Acute Stroke with Emphasis on Stress Hyperglycemia and THRIVE Score : An Observational Study" is awarded best article for  Vol 12 issue 07
A Study by K. Krupashree et. al. entitled "Protective Effects of Picrorhizakurroa Against Fumonisin B1 Induced Hepatotoxicity in Mice" is awarded best article for issue Vol 10 issue 20
A study by Mithun K.P. et al "Larvicidal Activity of Crude Solanum Nigrum Leaf and Berries Extract Against Dengue Vector-Aedesaegypti" is awarded Best Article for Vol 10 issue 14 of IJCRR
A study by Asha Menon "Women in Child Care and Early Education: Truly Nontraditional Work" is awarded Best Article for Vol 10 issue 13
A study by Deep J. M. "Prevalence of Molar-Incisor Hypomineralization in 7-13 Years Old Children of Biratnagar, Nepal: A Cross Sectional Study" is awarded Best Article for Vol 10 issue 11 of IJCRR
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A study by Karanpreet et al "Pregnancy Induced Hypertension: A Study on Its Multisystem Involvement" is given Best Paper Award for Vol 10 issue 09

List of Awardees

A Study by Ese Anibor et al. "Evaluation of Temporomandibular Joint Disorders Among Delta State University Students in Abraka, Nigeria" from Vol 13 issue 16 received Emerging Researcher Award


A Study by Alkhansa Mahmoud et al. entitled "mRNA Expression of Somatostatin Receptors (1-5) in MCF7 and MDA-MB231 Breast Cancer Cells" from Vol 13 issue 06 received Emerging Researcher Award


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International Journal of Current Research and Review (IJCRR) provides platform for researchers to publish and discuss their original research and review work. IJCRR can not be held responsible for views, opinions and written statements of researchers published in this journal

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