Occurrence of canine parvovirus type 2c in diarrhoeic faeces of dogs in Kolkata, India.

Canine parvovirus-2(CPV-2) causes a highly contagious disease of dogs characterised by acute hemorrhagic gastroenteritis, lethargy, vomiting, fever and usually bloody or mucoid diarrhoea. In the present study, 41 faecal samples collected from dogs exhibiting the signs of fever, vomition, bloody or mucoid diarrhoea in Kolkata, India were screened by haemagglutination test and PCR for detection of capsid protein coding VP2 gene. The viral genotype was detected by multiplex PCR and analysis of partial VP2 gene nucleotide sequences of selected PCR products with bioinformatics tool. Thirteen (31.71%) samples were found positive with HA titre ≥ 32 whereas 28 (68.29%) samples were positive by PCR of VP2 gene indicating higher sensitivity of PCR. Highest occurrence of CPV-2 was observed in the age group of 1-6 months (80.65%) and non-descript breeds with no history of vaccination (85%). Three samples were antigenic type CPV-2a, rest were CPV-2b/CPV 2c. Six CPV sequences were found to be highly similar to published CPV 2c sequences in BLAST analysis revealing a maximum identity of 99-100% with other CPV-2c strains and clustered together with CPV-2c strains of India and other countries in phylogenetic analysis. The present study highlights the need for continuous monitoring of samples to detect gradual changes in circulating CPV-2 genotypes in India.

Multi-drug resistant, biofilm-producing high-risk clonal lineage of Klebsiella in companion and household animals.

Antimicrobial resistance is a global emergency which needs one health approach to address. The present study was conducted to detect the prevalence of beta-lactamase and biofilm-producing Klebsiella strains in rectal swabs (n = 624) collected from healthy dogs, cats, sheep and goats reared as companion or household animals in India. The dogs and cats were frequently exposed to third- or fourth-generation cephalosporins for therapy. The sheep and goats were occasionally exposed to antibiotics and had environmental exposure. Phenotypical ESBL (n = 93) and ACBL (n = 88)-producing Klebsiella were isolated significantly more (P < 0·05) from companion animals than household animals. Majority of the Klebsiella possessed blaCTX-M-15 . The sequences blaCTX-M-15.2 , blaCTX-M-197 and blaCTX-M-225 are reported first time from the companion animals. All ACBL-producing isolates possessed blaAmpC . The present study detected 65·8% of Klebsiella strains as biofilm producers possessing the studied biofilm associated genes. The isolates showed phenotypical resistance against chloramphenicol, tetracycline, doxycycline, co-trimoxazole, ampicillin, cefotaxime/clavulanic acid. The present study showed that companion and household animals (dogs, cats, sheep, goats) may act as a carrier of ESBL/biofilm-producing, multi-drug resistant, high-risk clonal lineage of Klebsiella.

Characterization of beta-lactamase and biofilm producing Enterobacteriaceae isolated from organized and backyard farm ducks.

This study was undertaken to detect the occurrence of beta-lactamase and biofilm producing Enterobacteriaceae in healthy ducks. A total 202 cloacal swabs were collected from ducks kept in organized (n = 92) and backyard (n = 110) farms in West Bengal (India). The ducks had no history of antibiotic intake. Among the 87 phenotypically beta-lactamase producing Escherichia coli, 19 (17·43%), 6 (5·05%) and 15 (13·76%) isolates possessed blaTEM , blaSHV and blaCTX-M respectively. Whereas, 5 (38·46%) Salmonella isolates were found to harbour blaCTX-M . In K. pneumoniae 10 (33·33%), 3 (13·33%), 4 (13·33%) isolates possessed blaTEM , blaSHV and blaCTX-M respectively. The sequences of selected PCR products were found 98% cognate with blaCTX-M-9, blaSHV-12 and blaTEM-1 . Beta-lactamase producing E. coli isolates belonged to 14 different serogroups such as O1, O2, O3, O5, O7, O8, O35, O83, O84, O88, O119, O128, O145 and O157. Moreover, 87 E. coli (79·82%), six Samonella (46·15%) and 13 K. pneumoniae (43·33%) isolates were detected as AmpC producers possessing blaAmpC . Majority of E. coli (46·79%), Salmonella (46·15%) and K. pneumoniae (70%) isolates were detected as biofilm producers and possessed the associated genes (csgA, sdiA, rcsA, rpoS). Significantly higher occurrence of beta-lactamase and biofilm producing Enterobacteriaceae isolates was detected in backyard ducks than organized farms. SIGNIFICANCE AND IMPACT OF THE STUDY: Consumption of antibiotic through feed or during therapy is considered as potential reason for generation of antimicrobial resistant bacteria in birds. This study provides valuable evidence that exposure to contaminated environment may be an additional source for generation of antimicrobial resistant bacteria in backyard ducks. The backyard ducks are reared by marginal farmers in India who cannot offer antibiotics to them either through feed or during therapy due to high cost. The study also reveals a significant correlation between biofilm formation and possession of antimicrobial resistance genes in the bacterial isolates from the ducks.

Detection and characterization of pathogenic Pseudomonas aeruginosa from bovine subclinical mastitis in West Bengal, India.

AIM: Subclinical mastitis in bovines is mainly responsible for the huge economic loss of the dairy farmers, of which Pseudomonas aeruginosa is one of the causative agents. The study was aimed at a screening of suspected milk samples from different cattle farms of West Bengal for detection and confirmation of P. aeruginosa strains followed by their characterization. MATERIALS AND METHODS: Around 422 milk samples were screened from different dairy farms primarily by on-spot bromothymol blue (BTB) test and then in the lab by somatic cell counts (SCC) to finally consider 352 samples for detection of P. aeruginosa. Selective isolation and confirmation of the isolates were done using selective media, viz., cetrimide and Pseudomonas agar followed by confirmation by fluorescent technique. Molecular characterization of the strains was done by polymerase chain reaction for the presence of toxA (enterotoxin A, 352 bp) and exoS (exoenzyme S, 504 bp) genes. RESULTS: Approximately, 371 (87.9%) samples were positive in on-spot BTB test among which 352 (94.8%) samples revealed high SCC values (more than 3 lakh cells/ml) showing infection when screened. Among these, 23 (6.5%) samples yielded typical Pseudomonas sp. isolates out of which only 19 (5.4%) isolates were confirmed to be P. aeruginosa which showed characteristic blue-green fluorescence due to the presence of pigment pyoverdin under ultraviolet light. Out of these 19 isolates, 11 isolates were positive for toxA, 6 isolates for exoS, and 2 for both these pathogenic genes. CONCLUSION: Approximately, 5.4% cases of bovine subclinical mastitis infections in South Bengal were associated with P. aeruginosa which possess pathogenic genes such as toxA (63.2%) and exoS (36.8%).

Isolation, molecular characterization and antibiotic resistance of Shiga Toxin-Producing Escherichia coli (STEC) from buffalo in India.

In total, 363 Escherichia coli were isolated from 165 faecal samples of healthy buffaloes in West Bengal, India. Twenty-four of these isolates (6·61%) were found to carry at least one gene characteristic for Shiga toxin-producing Escherichia coli (STEC). These STEC strains belonged to 13 different O-serogroups. The stx1 gene was present in 23 (95·8%) of total STEC isolates, whereas 20 (83·3%) STEC isolates carried the gene stx2. Twelve strains of E. coli (50% of total STEC isolates) possessed enterohaemolysin (ehxA) gene in combination with others. Fourteen (58·33%) isolates found to possess saa gene. However, no E. coli was detected harbouring gene for intimin protein (eaeA). Of 23 stx1 -positive isolates, seven (30·43%) were positive for genes of the stx1C subtype. Of the 20 isolates with the stx2 gene, 25% (5/20) possessed stx2C and 10% (2/20) possessed stx2d gene. The phylogenetic analysis after RAPD of STEC strains revealed six major clusters. The isolated STEC strains were resistant most frequently to erythromycin (95·83%), cephalothin (62·5%), amikacin (54·17%), kanamycin (45·83%) and gentamicin (41·67%) group of antibiotics. No ESBL-producing (blaCTXM , blaTEM , blaSHV ) or quinolone resistance gene (qnrA) was detected in the STEC isolates.

Escheriosomes entrapped DNA vaccine co-expressing Cu-Zn superoxide dismutase and IL-18 confers protection against Brucella abortus.

In the present study, we evaluated prophylactic prospective of liposome based DNA vaccine co-expressing Cu-Zn superoxide dismutase (SOD) along with interleukin-18 (IL-18) against experimental murine brucellosis. The immunization schedule involves liposome-mediated delivery of pVsod (encoding SOD of Brucella abortus) and pVIL18-sod (encoding IL-18 of mouse and SOD of B. abortus) DNA constructs. The data highlight potential of Escherichia coli lipid liposome (escheriosome) based DNA delivery vehicle to induce SOD specific humoral and cellular immune responses in the immunized mice. The co-expression of SOD along with IL-18 ensued in higher lymphoproliferative response and IFN-gamma production in comparison to the group of animals that were immunized with free form of SOD-DNA. Antibody response developed upon immunization with both DNA vaccines was of IgG2a type mainly. The results of the present study show that co-expression of IL-18 along with SOD polarized the antigen specific immune responses toward Th-1 direction, a desirable feature to control intracellular pathogens.