Livestock host composition rather than land use or climate explains spatial patterns in bluetongue disease in South India.

Culicoides-borne arboviruses of livestock impair animal health, livestock production and livelihoods worldwide. As these arboviruses are multi-host, multi-vector systems, predictions to improve targeting of disease control measures require frameworks that quantify the relative impacts of multiple abiotic and biotic factors on disease patterns. We develop such a framework to predict long term (1992-2009) average patterns in bluetongue (BT), caused by bluetongue virus (BTV), in sheep in southern India, where annual BT outbreaks constrain the livelihoods and production of small-holder farmers. In Bayesian spatial general linear mixed models, host factors outperformed landscape and climate factors as predictors of disease patterns, with more BT outbreaks occurring on average in districts with higher densities of susceptible sheep breeds and buffalo. Since buffalo are resistant to clinical signs of BT, this finding suggests they are a source of infection for sympatric susceptible sheep populations. Sero-monitoring is required to understand the role of buffalo in maintaining BTV transmission and whether they must be included in vaccination programs to protect sheep adequately. Landscape factors, namely the coverage of post-flooding, irrigated and rain-fed croplands, had weak positive effects on outbreaks. The intimate links between livestock host, vector composition and agricultural practices in India require further investigation at the landscape scale.

Genetic clustering of recent classical swine fever virus isolates from Karnataka, India revealed the emergence of subtype 2.2 replacing subtype 1.1.

The phylogenetic analysis of 11 CSFV isolates from Karnataka, India obtained during the year 2012-13 was undertaken to obtain the most reliable genetic typing of the CSFV isolates based on E2, NS5B and 5'UTR genomic regions. The study indicated that all the 11 CSFV isolates belonged to subgroup 2.2. The most reliable classification was obtained with sequence data from the NS5B region which separated all the isolates based on the history of outbreak and geographic origin. Analysis of full length E2 amino acid sequences revealed different genetic makeup of Indian 2.2 isolates compared to 2.2 isolates from different countries. The group 2.2 viruses are gradually spreading as confirmed by frequent detection/ isolation of group 2.2 viruses in the recent years and replacing the subgroup 1.1 viruses, which were hitherto predominantly involved in CSF outbreaks in India.

Diagnosis and control of peste des petits ruminants: a comprehensive review.

Peste des petits ruminants (PPR) is an acute, highly contagious, world organization for animal health (OIE) notifiable and economically important transboundary viral disease of sheep and goats associated with high morbidity and mortality and caused by PPR virus. PPR is considered as one of the main constraints in augmenting the productivity of small ruminants in developing countries and particularly severely affects poor farmer's economy. The disease is clinically manifested by pyrexia, oculo-nasal discharges, necrotizing and erosive stomatitis, gastroenteritis, diarrhoea and bronchopneumonia. The disease can be diagnosed from its clinical signs, pathological lesions, and specific detection of virus antigen/antibodies/genome in the clinical samples by various serological tests and molecular assays. PPR is the one of the priority animal diseases whose control is considered important for poverty alleviation in enzootic countries. Availability of effective and safe live attenuated cell culture PPR vaccines and diagnostics have boosted the recently launched centrally sponsored control programme in India and also in other countries. This review article primarily focus on the current scenario of PPR diagnosis and its control programme with advancement of research areas that have taken place in the recent years with future perspectives.

Prevalence of Peste-des-petits-ruminant virus antibodies in cattle, buffaloes, sheep and goats in India.

The present study describes the prevalence of Peste-des-petits-ruminant virus (PPRV) antibodies in cattle, buffaloes, sheep and goats carried out during the period 2011 using the serum samples randomly collected from different villages of five states of India. A total of 1,498 serum samples [n = 605 (cattle); n = 432 (buffaloes); n = 173 (sheep); n = 288 (goats)] were collected from 52 districts in five states (Andhra Pradesh, Gujarat, Jammu and Kashmir, Maharashtra and Rajasthan) of India and were screened for PPRV-specific antibodies by using PPR monoclonal antibody-based competitive ELISA kit. Analysis of 1,498 samples indicates that an overall seroprevalence of 21.83 % with 11.07 % in cattle, 16.20 % in buffaloes, 45.66 % in sheep and 38.54 % in goats. This report presents the results of PPRV-specific antibodies in situations where the subclinical, inapparent or nonlethal or recovery of infection was suspected in cattle, buffaloes, sheep and goats. The presence of PPRV antibodies demonstrate that bovines are exposed to PPRV infection and it implies the importance of cattle and buffaloes as subclinical hosts for the virus besides widespread presence of the disease in sheep and goats. Further, the study showed that the prevalence of PPRV antibodies in apparently healthy livestock under natural situation, 21.83 % of the animals were protected from PPRV re-infection. This inturn help in the implementation of disease control strategies such as vaccination in that particular geographical area.

Prevalence of peste des petits ruminants in goats in North-East India.

The present study describes prevalence of peste des petits ruminants (PPR) virus infection in goats in various parts of North-East (NE) India by screening of suspected serum samples collected during outbreak investigation and random samples during 2013-2014 survey. A total of 391 serum samples (318 random and 73 outbreak/suspected) were collected from 28 districts in 7 states (Meghalaya, Assam, Manipur, Nagaland, Arunachal Pradesh, Tripura and Mizoram) of NE India. Serum samples were screened for PPRV-specific antibodies by using PPR monoclonal-antibody based competitive ELISA. Analysis of 391 serum samples indicates that an overall seroprevalence of 17.90 % [CI 95 % 14.40-22.00] in goats {45.2 % in suspected [CI 95 % 34.32-56.58] and 11.63 % in random [CI 95 % 8.56-15.63] samples} in NE India. As expected prevalence was high in outbreaks vis-à-vis random samples. The random survey results (11.63 %) has specific implication in epidemiological perspectives, since it highlights the exact PPR prevalence under natural situations, where the subclinical, in apparent or nonlethal or recovery of infection was suspected in goats, as samples were collected from unvaccinated animals. It also warrants appropriate control measures against PPR in NE region to prevent spread of infection besides widespread presence of the disease in rest of India.

Prevalence of Neospora caninum antibodies in dairy cattle and water buffaloes and associated abortions in the plateau of Southern Peninsular India.

A seroprevalence study of bovine neosporosis was conducted among 1,927 dairy cattle and 341 water buffaloes from Karnataka and Andhra Pradesh states in plateau of southern peninsular India by employing competitive enzyme-linked immunosorbent assay. Overall, 12.61 and 9.97 % sera samples were found positive for the presence of Neospora caninum antibody, respectively, among cattle and water buffaloes. Out of 1,927 sera samples from cattle, 912 and 1,015 samples were collected from unorganized and organized herds, respectively. The cattle screened were of upgraded Holstein-Friesian and water buffaloes were of graded Surti breed. Significantly (p < 0.05) higher prevalence was found in the cattle in unorganized herds (16.66 %) in comparison to organized herds (8.96 %). The highest seroprevalence was recorded in the age group of 4 years and above in both type of cattle herds and water buffaloes. There was a significant variation of seroprevalence (p < 0.05) observed between different age groups of cattle. The rate of seroprevalence increased with the increment in the age of the animals suggesting a possibility of horizontal mode of transmission of the infection from the environment. The percentage of abortion history was more in seropositive group (51.65 %) in comparison to the seronegative group (5.84 %) and the seropositive cattle were 8.84 times more likely to experience abortion than the seronegative cattle. The occurrence of abortion among different age groups varied significantly (p < 0.05). The findings revealed the presence of neosporosis in the southern peninsular India among cattle and water buffaloes and a strong association between the seroprevalence and abortion.

Phylogenetic analysis of NS5B gene of classical swine fever virus isolates indicates plausible Chinese origin of Indian subgroup 2.2 viruses.

Twenty-three CSFV isolates recovered from field outbreaks in various parts of India during 2006-2009 were used for genetic analysis in the NS5B region (409 nts). Seventeen of these were studied earlier [16] in the 5'UTR region. Phylogenetic analysis indicated the continued dominance of subgroup 1.1 strains in the country. Detailed analysis of a subgroup 2.2 virus indicated the plausible Chinese origin of this subgroup in India and provided indirect evidence of routes of CSFV movement within South East Asia region.

Absence of heat intolerance (panting) syndrome in foot-and-mouth disease-affected Indian cattle (Bos indicus) is associated with intact thyroid gland function.

Foot-and-mouth disease (FMD) is a highly contagious and economically important viral disease with high morbidity and reduced productivity of affected animals. We studied the heat intolerance (HI) (panting) syndrome and the effect of FMD virus (FMDV) infection on thyroid gland function in Indian cattle (Bos indicus). Experimental infection with FMDV Asia 1 resulted in a mild form of disease with superficial lesions. Heat intolerance syndrome and its signs were not observed among the recovered animals. Subtle changes in the serum level of thyroid hormones, triiodothyronine (T3) and thyroxine (T4) were observed. However, there were no distinct histological changes in the thyroid gland, and FMDV antigens were not detected in the thyroid tissues. Our results thus suggest that the absence of panting syndrome in FMD-affected Bos indicus cattle may be associated with intact thyroid gland function.

PCR-based diagnosis of surra-targeting VSG gene: experimental studies in small laboratory rodents and buffalo.

Trypanosoma evansi, the causative organism of 'surra' expresses its variable surface glycoprotein (VSG) at early, middle and late stages of infection in animals. The variable antigenic nature of VSG caused by switching its expression type favours evasion from the host immune response and leads to chronic and persistent infection. Developing a polymerase chain reaction (PCR)-based diagnostic tool targeting the VSG gene is expected to be highly specific and sensitive for diagnosis of surra. Hence, in the present study, we have designed EXP3F/4R primer pair and amplified the 1.4 kb of VSG gene of T. evansi and studied the phylogenetic relationship by in silico analysis. The PCR method was standardised using another set of primer, DITRYF/R, and 400 bp was amplified from blood and tissue samples of experimentally infected animals. Applying the PCR method, we were able to detect as low as 0.15 trypanosomeml(-1). Considering the number of parasite-to-DNA concentration, the PCR method has a sensitivity of 0.015 pg ml(-1). The PCR could detect the presence of the parasite as early as 24h post-infection (p.i.) and 72 h p.i., respectively, in experimentally infected rats and buffalo. No amplification was observed with DNA of Babesia bigemina and Theileria annulata, indicating the primers are specific for T. evansi. The PCR method could detect the dog, lion and leopard isolates of T. evansi. Similarly, amplifying the DNA from the experimentally infected tissues was also found to be sensitive. Thus, the findings of this study favour the application of PCR over the parasitological methods for the detection of the early and/or chronic stage of surra in domestic and wild animals.

Genetic typing of recent classical swine fever isolates from India.

Seventeen classical swine fever virus (CSFV) isolates recovered during the period of 3 years (2006-2008) from India were subjected to nucleotide sequencing in the 5' untranslated region (UTR). For genetic typing, 150 nucleotides within this region were used. For better epizootiological understanding, 39 nucleotide sequences of the above region, including 13 Indian CSFV sequences, available either in the Genbank or published literature were also included in the study. Based on the phylogenetic analysis, the Indian isolates could be grouped in to two subgroups, viz., 1.1 and 2.2. The study also revealed predominance of subgroup 1.1 and involvement of viruses of more than one subgroup in an outbreak.

Kinetics of immune response to foot-and-mouth disease virus (type Asia 1) in experimental cattle.

Humoral and mucosal (secretory antibody)immune response to FMDV type Asia 1 in cattle was analyzed after vaccination and infection using virus neutralizing test (VNT). Vaccination (1/16th the usual dose) failed to protect cattle from generalized clinical disease following experimental FMDV Asia 1 infection. Our results showed that infection induced higher and prolonged serum antibody titres indicating antigen mass is important for optimal immune response. Experimental FMDV infection induced significant secretory antibody (mucosal) response in cattle. Though, there was no difference in the serum antibody response between the cattle that developed generalized infection (unprotected) and those with only localized infection (protected), secretory antibody response differed, wherein the unprotected cattle had higher secretory response than protected cattle. Thus, FMDV Asia 1 infection stimulates a similar serum antibody response and a unique secretory antibody response among the infected cattle.

Comparative study of experimental Foot-and-Mouth Disease in cattle (Bos indicus) and buffaloes (Bubalis bubalus).

Foot-and-mouth disease (FMD) is one of the most contagious diseases affecting wide range of host species with variable severity and decreased productivity. The present study was undertaken to compare the clinical and leucocytic changes in indigenous Indian cattle and buffaloes experimentally infected with FMD virus (FMDV) Asia 1. A mild type of disease was observed in the cattle, more so in buffaloes infected with FMDV. Difference in terms of type, site and healing of lesion was observed between cattle and buffaloes. Foot lesions were more common than tongue in buffaloes, which were mainly evident in bulb of the heel in contrast to interdigital foot lesions in cattle. Further, FMDV infection induced a transient moderate leucopenia with lymphopenia in both cattle and buffaloes, but monocyte levels diverged. Relationship between the raised body temperature, leucocytic changes and lesion development was observed. Microscopic changes were observed in the keratinized epithelium of tongue and foot. The findings of the present study indicated the need to investigate the early leucocytic changes in cattle and buffaloes in depth for better understanding of the disease process.

Seroepidemiology of peste des petits ruminants in sheep and goats of southern peninsular India.

This paper presents the results of a seroepidemiological study carried out between July 2006 and March 2007 to detect the presence of antibodies to peste des petits ruminants (PPR) virus in randomly collected serum samples from sheep and goats in southern peninsular India. The authors used a competitive enzyme-linked immunosorbent assay with a monoclonal antibody developed against a neutralising epitope of the haemagglutinin (HA) protein of the virus. A total of 1,492 sheep sera and 2,068 goat sera collected from the six southern Indian states were screened. It was determined that 41.35% of the sheep sera and 34.91% of the goat sera were positive for the presence of antibody. The study indicated an extensive endemicity of the disease in these states, which is attributed to the agro-climatic conditions and the migration of livestock.

Development and standardization of a piezo electric immunobiosensor for foot and mouth disease virus typing.

An immunobiosensor using a piezo electric (PZ) crystal was developed and standardized for foot and mouth disease (FMD) diagnosis and virus typing. A 6MHz quartz crystal was used as the frequency determining element. Foot and mouth disease virus (FMDV) type specific antibody raised in rabbits/monoclonal antibody was coated on the crystal surface and the resonance measured. One microlitre of the 10% aqueous suspension of the clinical sample (tongue or foot epithelium) was applied on both surfaces of the crystal and the resonance recorded. A difference in resonance of more than -2.5Hz was obtained in positive samples (homologous antigen and antibody). The test was standardized initially using various dilutions of FMD tissue culture antigen. Repeatability and sensitivity were also tested and it was found that the crystals could be washed and reused eight times. The test could be used for FMDV type specifically and no cross-reaction between FMDV types was observed. The shelf-life of the antibody-coated crystal stored at room temperature was 18 weeks. Application of the biosensor test to the FMDV clinical samples confirmed virus typing results when compared with enzyme-linked immunoabsorbent assay (ELISA) and it could also detect virus in ELISA negative samples and mixed virus infections.

E. coli expressed proteins as diagnostic reagents for typing of foot-and-mouth disease virus.

Truncated proteins corresponding to the C-terminal half of VP1 of four vaccine strains and two field variants of foot-and-mouth disease virus (FMDV) were expressed in E. coli. The expressed proteins were affinity purified and their type specific reactivity was confirmed by immunoprecipitation with anti-virus antibodies. Antibodies were raised against the purified proteins in guinea pigs and the type specificity of the anti peptide antibodies was confirmed by antigen capture reverse transcription polymerase chain reaction (Ag-RT/PCR) where the sera against a particular type captured the homologous virus. Antibodies were purified by immuno-affinity chromatography and tested for specificity by various serological tests. Using the purified proteins and the antibodies raised against them, tests like ELISA, Ag-RT/PCR, and latex agglutination test (LAT) were standardized. Application of the reagents in various tests was studied by screening a few field samples and by nucleotide sequencing. Specific reactivity of antibodies raised against expressed protein was seen with both vaccine virus and field samples. Thus E. coli expressed proteins and antibodies to them may form an alternative and cheap source of diagnostic reagents. The studies showed that antibodies against peptides were mono-specific and therefore may be used in LAT for rapid typing of FMDV and Ag-RT/PCR for typing ELISA negative field samples.

A note on outbreaks caused by mixed foot-and-mouth disease virus infections.

Two outbreaks of foot-and-mouth disease (FMD) in vaccinated cattle were investigated wherein a mixed infection due to FMD virus (FMDV) types O and Asia 1 was detected by sandwich enzyme-linked immunosorbent assay (ELISA) and confirmed by antigen capture polymerase chain reaction (PCR). The clinical picture and the epidemiological data on these outbreaks are presented. The isolated virus strains were compared to the respective vaccine strains by means of monoclonal antibody (MAb) profiling and nucleotide sequence analysis. The probable cause of the mixed FMDV infection and its significance in disease control are discussed.

Complexometric determination of mercury(II) with 2-imidazolidinethione as a selective masking agent.

A simple, rapid and selective complexometric method is proposed for the determination of mercury(II). Mercury(II) is first complexed with a known excess of EDTA and the surplus EDTA is back-titrated at pH 5.0-6.0 with lead nitrate, Xylenol Orange being used as indicator. 2-Imidazolidinethione is then added to displace EDTA from the Hg-EDTA complex quantitatively and the EDTA released is titrated with lead nitrate. Reproducible and accurate results are obtained for 2-75 mg of mercury, with a relative error of less than 0.3% and standard deviation of less than 0.04 mg.

Complexometric titration of thallium(III), by use of 4-amino-5-mercapto-3-propyl-1,2,4-triazole as replacing reagent.

A simple and selective EDTA method using a masking and demasking technique is proposed for the determination of thallium(III). The thallium is complexed with excess of EDTA, the surplus being back-titrated (pH 5-6, hexamine buffer) with zinc sulphate solution (Xylenol Orange as indicator). 4-Amino-5-mercapto-3-propyl-1,2,4-triazole is then added and the mixture heated on a water-bath for 5-10 min to displace EDTA from its thallium complex. The EDTA liberated is titrated with zinc sulphate solution. Reproducible and accurate results are obtained in the range 5-75 mg of thallium with both the relative error and coefficient of variation not exceeding 0.4%.

Complexometric determination of mercury(II) by use of 4-amino-5-mercapto-3-propyl-1,2,4-triazole as replacing reagent.

A simple, rapid, and selective complexometric method is proposed for the determination of mercury(II). Mercury(II) is first complexed with excess of EDTA and the surplus EDTA is back-titrated (pH 5-6) with zinc sulphate solution, with Xylenol Orange as indicator. 4-Amino-5-mercapto-3-propyl-1,2,4-triazole is then added to displace EDTA from the Hg-EDTA complex and the released EDTA is titrated with zinc sulphate solution. Reproducible and accurate results are obtained in the range 1-40 mg of mercury, with a relative error of approximately 0.4%.

4-Amino-3-methyl-5-mercapto-1,2,4-triazole as a gravimetric reagent for determination of silver in silver compounds, alloys and complexes.

A simple, convenient and accurate gravimetric method for the determination of silver is presented. 4-Amino-3-methyl-5-mercapto-1,2,4-triazole precipitates silver quantitatively from ammoniacal tartrate medium. The complex is weighed as AgC(3)H(5)N(4)S after drying at 120-30 degrees . Separation of silver from a large number of cations is described. Application of the method for quantitative analysis of alloys and complexes of silver is reported. The average relative error for the range 20-70 mg of silver is +/- 0.25% and the relative standard deviation at the 40-mg level is 0.2%.