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AIM: The aim of this study was to characterize beta-lactamase antimicrobial resistance in Klebsiella and Enterobacter species isolated from healthy and diarrheic dogs in Andhra Pradesh. MATERIALS AND METHODS: A total of 136 rectal swabs were collected from healthy (92) and diarrheic (44) dogs, bacteriological cultured for Klebsiella and Enterobacter growth and screened for beta-lactamase antimicrobial resistance phenotypically by disc diffusion method and genotypically by polymerase chain reaction targeting blaTEM, blaSHV, blaOXA, blaCTX-M Group 1, 2, blaAmpC, blaACC, and blaMOX genes. RESULTS: A total of 33 Klebsiella and 29 Enterobacter isolates were recovered. Phenotypic beta-lactamase resistance was detected in 66.6% and 25% of Klebsiella and Enterobacter isolates, respectively, from healthy dogs and 66.6% and 60% of Klebsiella and Enterobacter isolates, respectively, from diarrheic dogs. Overall, incidence of extended-spectrum beta-lactamase (ESBL) phenotype was found to be 21.2% (7/33) in Klebsiella isolates, whereas none of the Enterobacter isolates exhibited ESBL phenotype. Predominant beta-lactamase genes detected in Klebsiella species include blaSHV (84.8%), followed by blaTEM (33.3%), blaCTX-M Group 1 (15.1%), and blaOXA (6.1%) gene. Predominant beta-lactamase genes detected in Enterobacter species include blaSHV (48.2%), followed by blaTEM (24.1%), blaAmpC (13.7%), and blaOXA (10.3%) gene. CONCLUSION: The present study highlighted alarming beta-lactamase resistance in Klebsiella and Enterobacter species of canine origin in India with due emphasis as indicators of antimicrobial resistance.

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The field isolates of Trypanosoma evansi was collected from the infected cattle and it was propagated in rats. Trypanosoma evansi parasites were separated from the blood of infected rats by using diethylaminoethyl cellulose column chromatography. Whole cell lysate antigen (WCL) was prepared from purified trypanosomes by ultrasonication and centrifugation. The prepared WCL antigen was further purified by 50 % ammonium sulphate precipitation. Protein concentration of WCL antigen of T. evansi was 60 mg/ml. Protein concentration was adjusted to 1.0 mg/ml in PBS, pH 8.0 and stored at -20(0) C.   Polypeptide profiles of WCL antigen of T. evansi was determined by sodium dodecyl sulphate polyacrylamide gel electrophoresis. A total of eight polypeptide bands of the size ranging from 25 to 85 kDa in WCL antigen of T. evansi were obtained. Five prominent bands with molecular weight of 74, 60, 53, 42 and 37 kDa and three light bands with molecular weight of 85, 34 and 25 kDa were observed.

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A total of 56 foot swabs were collected from inter digital spaces of sheep with footrot lesions were screened for 16 rRNA of Dichelobacter nodosus by PCR. Out of the 56 samples, 38(67.85%) were found to be positive. All the positive samples were subjected to multiplex PCR targeting fimA gene for identification of serogroups of D. nodosus. Serogroup H was found along with serogroup B in 12 (55.26%) samples and with serogroup I in 8 (22.2%) samples. The serogroup H was identified for the first time from the Indian subcontinent. The phylogenetic analysis of the present sequence with the available serogroup H sequences of GenBank revealed to be in close association with the serotype H1.

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AIM: The loop mediated isothermal amplification (LAMP) was standardized for rapid detection of Clostridium perfringens. MATERIALS AND METHODS: A total of 120 fecal samples were collected from enterotoxemia suspected lambs were used for screening of C. perfringens cpa gene by LAMP. The specificity of the LAMP amplified products was tested by digesting with restriction enzyme XmnI for alpha toxin gene. RESULTS: Out of 120 samples screened 112 (93.3%) samples were positive by both LAMP and polymerase chain reaction (PCR) for detection of cpa gene which indicated the equal sensitivity of both the tests. The enzyme produced single cut in 162 base pair amplified product of alpha toxin gene at 81 base pair resulting in a single band in gel electrophoresis. CONCLUSION: Both LAMP and PCR for detection of cpa gene indicated the equal sensitivity of both the tests. Standardization of LAMP reaction for amplification of epsilon and beta toxin genes will help to identify the C. perfringens toxin types from the clinical samples. The test could be a suitable alternative to the PCR in detection of toxin types without the help of sophisticated machinery like thermal cycler. Considering its simplicity in operation and high sensitivity, there is the potential use of this technique in clinical diagnosis and surveillance of infectious diseases.

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The diagnosis of Trypanosoma evansi in animals with low parasitaemia is hampered by low diagnostic sensitivity of traditional detection methods. The present study was undertaken with an objective to improve the diagnostic tools for detection of antibodies against T. evansi infection using indirect enzyme-linked immunosorbent assay (ELISA) in bovines. The optimum concentration of antigen, test sera and conjugate were determined as 5 µg per well, 1:10 and 1: 6,000 dilutions, respectively. Among 320 cattle and 382 buffaloes examined in different parts of Rayalaseema region of Andhra Pradesh for T. evansi infection, 36.12 and 31.87 percent were found positive by indirect ELISA, respectively.

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AIM: Identification of different serogroups of Dichelobacter nodosus prevailing in the region and to understand the degree of genetic heterogeneities among the different isolates of D. nodosus. MATERIALS AND METHODS: A total of 150 exudate samples of footrot lesions with a lesion score of 2-4 were collected from naturally infected sheep. The samples were screened by polymerase chain reaction (PCR) targeting D. nodosus specific 16srRNA. Of 150 samples screened, 70 samples were found to be positive. The positive samples were attempted for isolation of D. nodosus, out of which 16 isolates were recovered. All the isolates were subjected to serogrouping by multiplex PCR targeting fimA gene using A-I serogroup specific primers. RESULTS: Of 16 isolates, 7 (43.75%) isolates were serogroup B, 4 (25.00%) isolates were serogroup A, 3 isolates (18.75%) were serogroup I and 2 (12.5%) isolates yielded both serogroup A and B. phylogenetic analysis was performed using neighbor-joining algorithm of the ClustelX2 software in order to study whether the serogroups isolated in the present investigation differed genetically from other published serogroups. The fimA gene sequence of present isolates of serogroups A, B, and I were segregated into three distinct groups with high bootstrap values. The serogroup B clustered with Australian isolate of serotype B1 suggesting high genetic similarity of the present isolate with serotype B1. CONCLUSIONS: The clinical samples were collected from suspected outbreaks of footrot and identified the prevalence of D. nodosus by PCR targeting 16srRNA gene. Identified serogroups A, B, and I from different districts of Andhra Pradesh. The phylogenetic analysis will help for the tentative identification of serotypes present in the serogroup and to understand the degree of genetic heterogeneities among the different isolates of D. nodosus.

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An outbreak of sheep associated malignant catarrhal fever in crossbred cattle in a village of Andhra Pradesh, southern India, affected thirteen adult cows and two calves from a population of forty animals. All the affected animals were died between December and January 2013-14. The clinical and gross postmortem findings were typical of MCF in Indian crossbred cattle. Migrating sheep flocks were suspected source of infection for the cattle. The diagnosis was confirmed by heminested PCR in all the affected cattle and the suspected sheep flock. The PCR provided evidence of ovine herpes virus type 2.

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The present communication records the first determination of the prevalence of footrot in the unexpected situation of the tropical climate of Andhra Pradesh and Tamil Nadu, two states in southern India where the maximum temperature rises to 42 degrees C. In total, 73 outbreaks of footrot in Nellore brown sheep were investigated in 11 districts of Andhra Pradesh and one district of Tamil Nadu during the period March 2009 to March 2011.The overall prevalence of ovine footrot was 15%, with severity scores of 2 to 4 (lesion severity scale 0 to 4). The outbreaks occurred mostly during the rainy season, which is usually from June to December. From a total of 1,050 samples of lesions in naturally infected sheep, 478 (45.5%) were positive for Dichelobacter nodosus. Serogrouping of the isolates revealed six serogroups: A, B, C, E, F and I. Among the positive samples, 448 (93.7%) were a single serogroup and 30 (6.3%) carried a mixed infection with two serogroups. Taking single and mixed infections together, serogroup B was most frequent at 50.4% and was found in all districts, followed by serogroup I in 29.3% of samples, A in 14%, F in 6.7% and C in 5.6%. Serogroup E was detected in only one sample. Serogroups A and F were detected for the first time in India. All of 58 D. nodosus isolates in a sub-sample representing different serogroups were found to be virulent, based on the production of thermostable proteases and the presence of the integrase A gene intA. Thus, the present paper reporting isolation and characterisation of D. nodosus confirms the occurrence of virulent footrot in the tropical climate of southern India.

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Rehabilitation of speech is tantamount to closure of defect in cases with velopharyngeal insufficiency. Often the importance of speech therapy is sidelined during the fabrication of obturators. Usually the speech part is taken up only at a later stage and is relegated entirely to a speech therapist without the active involvement of the prosthodontist. The article suggests a protocol for speech therapy in such cases to be done in unison with a prosthodontist.

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In this study we present the first report on partial amplification, sequencing and phylogenetic relationship of VP2 of the Indian isolate BTV-2. A PCR product of 1135 bp was amplified, cloned and sequenced. About 1063 bp of partial VP2 gene (1792-2854 bp region) of the Indian isolate was subjected to sequence analysis with already published sequences available in the genome database. The percent similarity of 85.2 was observed with Taiwan isolate and 59% with other isolates of BTV-2. However, 46.2% similarity with Australian BTV-1 and no significant similarity were noted with other serotypes. In-silico analysis and restriction enzyme digestion confirmed the presence of conserved SalI site at 2380 bp position in both Indian and Taiwan isolates. Phylogenetic analysis showed that all BTV-2 isolates formed one distinct group in which BTV-2 Indian and Taiwan isolate is more closely related and further demonstrated that BTV's of the same serotype from different geographical regions were closely related at nucleotide and amino acid level, respectively.

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The occurrence of bluetongue (BT) disease in India was initially confined to exotic breeds of sheep and subsequently became endemic in native breeds. BT virus (BTV) antibodies are common in cattle, buffaloes and goats although clinical disease has not been reported. Exotic breeds of sheep and their cross-breeds are more susceptible to disease than native breeds. Overall, morbidity, mortality and case fatality rates of 9.3%, 2.7% and 28.8%, respectively, have been reported in rural flocks; these rates are higher than in organised farms. The disease is mostly cyclical in occurrence. Outbreaks usually occur between June and December during the monsoon period when livestock biting midges greatly increase. BTVs have been isolated from native sheep, and sentinel herds have been used to demonstrate virus activity. A total of 21 serotypes of BTV have now been reported in the country. Major impediments to control the disease include the presence of multiple virus serotypes, the broad vertebrate host range of the virus and a lack of detailed knowledge of vectors. Inactivated vaccines prepared from local isolates are currently under field trials. BTV occurs in regions adjacent to India. An antibody prevalence of 48.4% has been reported in Pakistan with serotypes 3, 9, 15, 16 and 18 identified. BTV antibody, but not disease, has been reported in Bangladesh and Sri Lanka.

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