Occurrence of Foot and Mouth Disease ( FMD ) during 2014-2016 in cattle of Sirajganj district , Bangladesh

Foot and mouth disease (FMD) is a severe, highly contagious, and economically devastating viral disease which affects domestic cloven-hoofed cattle, pigs, sheep, goats as well as more than 70 species of mammals (Barnett & Cox, 1999; Brown, 2003; Grubman & Baxt, 2004; Du et al., 2007; Giasuddin et al., 2016). FMD is prevalent in many parts of the world and notorious for its ability to severely affect and indeed disrupt regional and international trade in animals and animal products. FMD is very common in many parts of the world, particularly in developing countries of Asia, Africa, the Middle East, and some parts of Europe. The occurrence of FMD in these countries results in severe economic losses. In FMD-endemic countries, usually the developing countries, the disease threatens food security and the livelihoods of smallholders. This disease causes an economic loss of US$60–150 million annually in Bangladesh (Momtaz et al., 2014).

The etiological agent of Foot and Mouth Disease Virus (FMDV) is a small non-enveloped virus and belongs to the genus Aphtho virus of the family Picornaviridae.FMDVs are classified into 7 serological types based on antigenic properties of the capsid proteins.The serotypes are O, A, C, Asia-1, SAT-1, SAT2 and SAT3 (Mittal et al., 2005).Presently, 6 serotypes of FMDV (O, A, Asia-1, SAT-1,-2, and -3) are circulating worldwide, and serotype C has not been recorded since 1995 (Biswal et al., 2012).Distribution of different serotypes of FMDV varies region to region.Serotypes O, A, and C mostly occur in Europe, South America, Africa, and Asia; SAT 1, SAT 2, and SAT 3 are normally limited to sub-Saharan Africa; and Asia1 occurs only in Asia (Ansell et al., 1994).
In a previous study, sequence analysis revealed that serotype O and Asia1 is more prevalent while serotype A has very low frequency in Bangladesh.It was also found that the VP1 sequences of FMDV of the study samples were quite similar to sequence of FMDV isolated from animals in India and other neighboring countries (Giasuddin et al., 2016).These outcomes imply the possibility of virus transmission during cross-country import of animals, which lacks strict policy for animal health check.
Moreover, some other environmental factors may play crucial role in transmission and progression of the disease in animal host.Despite vaccination, the disease appears every year across the country due to high evolutionary rates.Therefore, if Bangladesh needs to ensure food security, increase livelihood of rural people and get access to the lucrative markets for her livestock and livestock products, the control of FMD need to be addressed more effectively.Therefore, the present study was undertaken to identify environmental factors and host factors that influence the FMDV outbreak in Bangladesh.Besides we also aim to investigate molecular epidemiology, genotyping and employ phylogenetic analysis to determine the relationship of FMDV serotypes circulating in Bangladesh.Therefore this study was performed to detect the factors (both environmental and host) responsible for FMD outbreak and occurrence of FMD to highly susceptible area (Sirajganj District).
Total 68 samples were collected from clinically FMD-positive animals (cattle) from Sirajganj, Bangladesh.Samples preferably included mucosal epithelium (Tongue epithelium), tissue from the inter-digital space (Hoof tissue), saliva and fluid from the ruptured vesicles.Samples were collected in the viral transport medium (VTM) containing 0.04 M phosphate buffer (pH 7.2-7.6),1% phenol red, antibiotics (penicillin 100 U/ml and streptomycin 100 μg/ml), and equal volume of glycerol.All of the available information (such as date of sample collection, age, sex and breed of the cattle, disease history etc.) for every sample were recorded on the FMD sample submission form provided by Bangladesh Livestock Research Institute (BLRI).The collected samples were mixed with PBS 5 times of the volume of sample and centrifuged at 3000 rpm for 10 minutes maintaining temperature at 4°C.The supernatant was collected in 50ml Falcon tube, marked properly and stored at -80°C till further processing.
Total RNA was extracted from original epithelial suspensions using QIAamp® Viral RNA Extraction kit (Qiagen, Germany) following the recommendations of the suppliers.After elution in nuclease-free water, RNA samples were stored in aliquots at -80°C until required (Giasuddin et al., 2016).Assays has been played an important role in detection of FMDV in clinical samples (Laor et al., 1992;Meyer et al., 1991).After RNA extraction, the target sequence was amplified by using QIAGEN® one step RT-PCR kit (QIAGEN Inc., Netherlands).The oligonucleotide primer for the detection of FMDV and FMDV serotypes was used from the 1D, 2B and 5 ‫׳‬ UTR regions of the viral genome as previously published (Callens et al., 1998;Reid et al., 2000).More specification of these primers is listed in the Table 1.All oligonucleotide primers were synthesized by Sigma, USA.The amplified PCR products of the expected length were subjected to electrophoresis in a 2% agarose gel and visualized by straining with 0.6 mg/ml ethidium bromide and documented with Alpha Imager Mini System Protein Simple, USA (Giasuddin et al., 2016).The impact of seasonal variation on the outbreak of foot and mouth disease (FMD) in cattle was observed.During summer (March-June), monsoon (July-October), winter (November-February) 28, 18, 22 suspected samples were collected and among them 18, 16, 14 samples were found FMDV positive, respectively.In summer season the highest number 18 positive samples were identified.Animals were categorized into three agegroups and tested for the presence of FMD virus.Age groups were young, adult and old.FMD outbreaks were influenced by the age of host animals.Outbreak rate of FMD virus was varied among male and female cattle.Out of 31 male cattle, 20 were found positive, and 28 were positive in 37 female cattle, comprising 64.5% and 75.7% of total positive sample, respectively.Diagnosis of FMD outbreak also varied based on sample types.Results of RT-PCR detection technique varied between the sample types.Highest number of positive isolates was found from tissue sample that comprised 83.9% of total positive isolates.During the study period, out of 23 and 45 samples collected from indigenous and cross breed, 13 and 35 samples were found FMDV positive in respective groups.The susceptibility of various breed of cattle to FMD virus is shown in Table 3.It is found that, a significant variation of breed susceptibility was observed which affected mostly crossed breed cattle (72.9% of total positive sample) compared to indigenous cattle (27.1% of total positive sample).Susceptibility of the cattle to FMDV was variable and influenced by vaccination profile.
In this study out of 28 and 40 samples from vaccinated and non-vaccinated cattle respectively, 17 and 31 samples were found FMDV positive in respective groups.
Outbreak rate also fluctuated during different seasons and highest numbers of positive results were found in Monsoon (July-October).Female cattle were more susceptible than male cattle.Animals of young age were more vulnerable than other age groups.As vaccination could protect the emergence of foot and mouth disease in some cases, nonvaccinated cattle were more susceptible than vaccinated animals.Breeding patterns also influenced FMD outbreak, highest numbers of positive results were found in the animals having cross breed.The representative sequences of the positive isolates in this study and a previous study showed remarkable similarity with sequences of Bangladesh, Bhutan and India in 2012-2014 (Giasuddin et al., 2016).This has implied that FMD is a transboundary disease.Therefore, we have to establish animal health check during the importation of animals and animal products from surrounding countries to minimize the emergence of foot and mouth disease in animal population.
Present study has confirmed that the occurrence of FMD in cattle of Sirajganj district is high and its association with variation of the seasonal effect, age, sex and breed of cattle is significant.Most risk group for outbreak was also identified.The results of this study could help to control the FMDV outbreak from this area.