In-silico insights of ESBL variants and tracking the probable sources of ESBL-producing Escherichia coli in a small-scale poultry farm.

Commercial broiler farms faces challenges of extended spectrum beta-lactamase (ESBL)-producing Escherichia coli transmitted from both vertical and horizontal routes. Understanding the dynamics of ESBL-E. coli transmission in compromised biosecurity settings of small-scale rural poultry farms is essential. This study aimed to elucidate the probable transmission pathways of ESBL-E. coli in such settings, employing phylogenetic analysis and molecular docking simulations to explore the catalytic properties of ß-lactamase variants. Sampling was conducted on a small-scale poultry farm in West Bengal, India, collecting 120 samples at three intervals during the broiler production cycle. E. coli isolates underwent resistance testing against eight antimicrobials, with confirmation of ESBL production. Genotypic analysis of ESBL genes and sequencing were performed, alongside molecular docking analyses and phylogenetic comparisons with publicly available sequences. Among 173 E. coli isolates, varying resistance profiles were observed, with complete resistance to cefixime and high resistance to amoxicillin and tetracycline. The incidence of ESBL-E. coli fluctuated over the production cycle, with dynamic changes in the prevalence of blaCTX-M-Type and blaSHV-Type genes. Phylogenetic analysis indicated partial clonal relationships with human clinical strains and poultry strains from the Indian subcontinent. Molecular docking confirmed the catalytic efficiencies of these ESBL variants. The study highlights probable vertical transmission of ESBL-E. coli and emphasizes drinking water as a potential source of horizontal transmission in small-scale poultry farms. Strict biosecurity measures could prevent the spread of antimicrobial-resistant bacteria in birds and their products in a small scale poultry farm.

An indigenous tubular ceramic membrane integrated bioreactor system for biodegradation of phthalates mixture from contaminated wastewater.

Endocrine-disrupting phthalates (EDPs) are widely used as plasticizers for the manufacture of different plastics and polyvinyl chloride by providing flexibility and mechanical strength. On the other hand, they are categorized under priority pollutants list due to their threat to human health and the environment. This study examined biodegradation of a mixture of dimethyl, diethyl, dibutyl, benzyl butyl, di-2-ethylhexyl, and di-n-octyl phthalates using a CSTB (continuous stirred tank bioreactor) operated under batch, fed-batch, continuous, and continuous with biomass recycle operation modes. For operating the CSTB under biomass recycle mode, microfiltration using an indigenous tubular ceramic membrane was employed. Ecotoxicity assessment of the treated water was carried out to evaluate the toxicity removal efficiency by the integrated bioreactor system. From the batch experiments, the EDPs cumulative degradation values were 90 and 75% at 1250 and 1500 mg/L total initial concentration of the mixture, respectively, whereas complete degradation was achieved at 750 mg/L. In the fed-batch study, 93% degradation was achieved at 1500 mg/L total initial concentration of the mixture. In continuous operation mode, 94 and 85% degradation efficiency values were achieved at 43.72 and 52.08 mg/L⋅h inlet loading rate of phthalate mixture. However, continuous feeding with 100% biomass recycle revealed complete degradation at 41.67 mg/L⋅h inlet loading rate within the 84 h operation period. High seed germination index and low mortality percentage of brine shrimps observed with phthalate degraded water from the integrated bioreactor system revealed its excellent potential in the treatment and toxicity removal of phthalates contaminated environment.

Phytochemical Analysis and Demonstration of Antioxidant, Antibacterial, and Antibiofilm Activities of Ethnomedicinal Plants of North East India.

Ethnomedicinal plants are a rich reservoir of active compounds with potent pharmacological properties. Therefore, plants could serve as a source for the discovery of active antimicrobial and antioxidant agents and are focused because of their low toxicity, economic viability, easy availability, etc. In this regard, phytochemical analyses, viz. ß-carotene, total sugar, reducing sugar, vitamin C, total carotenoids, protein, total phenolic content (TPC), and total flavonoid content (TFC) of 20 ethnomedicinal plants of North East India (NEI) were evaluated in this study. The antibacterial activity against human pathogens and antioxidant potential of plant extracts was also demonstrated. The minimum inhibitory concentration (MIC80), minimum bactericidal concentration (MBC), and total antibacterial activity (TAA) of the active extracts were evaluated against Pseudomonas aeruginosa and Chromobacterium violaceum. The active extracts were also examined for antibiofilm as well as anti-pyocyanin activities against P. aeruginosa and anti-QS activity against C. violaceum at sub-MICs. The study demonstrated variable concentration of phytochemicals of the extracts, viz. ß-carotene (0.29-8.91 mg g-1), total sugar (2.92-30.6 mM), reducing sugar (0.44-14.5 mM), vitamin C (8.41-31.3 mg g-1), total carotenoids (14.9-267.0 mg g-1), protein (5.65-283 mg g-1), TPC (5.32-31.0 mg GAE/g DW), and TFC (1.74-68.2 mg QE/g DW). The plant extracts also exhibited potent antioxidant and antibacterial activities against both Gram-positive and Gram-negative bacteria. Some of the extracts also demonstrated significant biofilm inhibition and eradication, anti-pyocyanin, and anti-QS activities at sub-MICs. The selected ethnomedicinal plants are rich in phytochemicals and demonstrated potent antioxidant, antibacterial, and antibiofilm activities, thus could serve as the important source of novel antioxidant and antimicrobial agents.

Docking analysis of circulating CTX-M variants in multi-drug resistant, beta-lactamase and biofilm-producing E. coli isolated from pet animals and backyard livestock.

The generation of antimicrobial-resistant bacteria largely depends on the use of antimicrobials not only in humans but also in pet animals and livestock. The present study was conducted to detect the occurrence of beta-lactamase and biofilm-producing- E.coli in healthy pet and backyard livestock. The study also intended on molecular docking experiments to confirm the nature of the catalytic mechanism in ß-lactamase enzymes, encoded by the various blaCTX-M genotypes and phylogenetic analysis to reveal clonal relationship of the animal origin E. coli isolates with human clinical strains. The rectal swabs were collected from healthy dogs (n = 254), cats (n = 108), sheep (n = 119) and goats (n = 143) in India. In total 247 (76.47%) E. coli strains were identified as ESBL producers. The possession of ESBL-producers was significantly more (p < 0.05) in pets than in the backyard livestock. Most of the strains possessed blaCTX-M-15 like clones. E. coli strains possessing blaCTX-M-15.2, blaCTX-M-157, blaCTX-M-181 and blaCTX-M-218 like clones, isolated from pets were not reported earlier. The study detected 56.65% of E. coli strains as moderate or strong biofilm producers possessing biofilm-associated genes (csgA, rcsA, rpoS, sdiA). ESBL-producing E. coli showed phenotypical resistance to tetracycline (93.1%), azithromycin (89.8%), ampicillin (84.2%), cefotaxime (80.9%), doxycycline (82.5%), co-trimoxazole (80.9%), ampicillin/cloxacillin (76.9%). The CTX-M variants obtained in this study were modelled by the SWISS-MODEL and verified. Ligand having minimum binding energy, show the highest affinity of ß-lactamases for cefotaxime and cefpodoxime. The Gibbs free energy release for all 14 different complex ranges between -6.9 (CTX-M-15.2+cefpodoxime) to -5.3 (CTX-M-218+cefpodoxime) Kcal/mol. Phylogenetic analysis of the animal origin ESBL-E. coli strains revealed a partial clonal relationship with the clinical isolates of local human patients. The present study described the significant presence of biofilm and ß-lactamase producing, multi-drug resistant E. coli in pet animals having public health importance.

Ecofriendly phytofabrication of silver nanoparticles using aqueous extract of Cuphea carthagenensis and their antioxidant potential and antibacterial activity against clinically important human pathogens.

The green synthesis of nanoparticles (NPs) is the safest, ecofriendly, cost-effective, and non-hazardous approach of nanotechnology. In the current study, we described the green synthesis of silver nanoparticles (AgNPs) using Cuphea carthagenensis aqueous leaf extract as a reducing, capping, and stabilizing agent. The study aims at the synthesis, characterization, optimization, and determination of the antibacterial activity of Cc-AgNPs against clinically important human pathogens. Coating of cotton fabrics with Cc-AgNPs and their efficacy against skin infection causing organisms was also evaluated. Furthermore, antioxidant activity, growth assay and time kill assay of Cc-AgNPs were also performed in the study. The biosynthesized Cc-AgNPs were characterized by UV-visible spectrometry, energy-dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), transmission electron microscopy (TEM), and Fourier transform infrared spectroscopy (FTIR). The spectroscopic and microscopic analysis demonstrated biosynthesis of face-centered cubic (fcc) crystalline spherical Cc-AgNPs with an average particle size of 10.65 ± 0.1 nm. Optimized peak synthesis of Cc-AgNPs was reported at pH7, 55 °C, 4 mM silver nitrate, and 5:45 (plant extract: silver nitrate). Cc-AgNPs exhibited potent antioxidant effect and antibacterial activity against both Gram-positive and Gram-negative bacteria. The lowest MIC (15 µg/ml) and MBC (25 µg/ml) values were reported against S. typhimurium. The Cc-AgNPs coated fabrics demonstrated potent antibacterial activity against tested strains. This application could be helpful in wound healing management. Furthermore, the hemolytic analysis demonstrated that Cc-AgNPs exhibit non-toxic nature against Red Blood Cells (RBCs) at the tested concentrations. In conclusion, the investigation demonstrated a fast, stable, and eco-friendly approach to the biosynthesis of Cc-AgNPs along with their antibacterial and antioxidant properties.

Exploring broilers and native fowls of Andaman and Nicobar Islands as a source of ß-lactamase-producing Enterobacteriaceae even with limited anthropogenic activities and docking-based identification of catalytic domains in novel ß-lactamase variants.

Objectives: The present study was conducted to detect the occurrence of ß-lactamase and biofilm-producing Escherichia coli, Salmonella, and Klebsiella in broilers and native fowl reared in the Andaman and Nicobar Islands, India. The study also included molecular docking experiments to confirm the nature of the catalytic domains found in the ß-lactamase variants obtained and to reveal the clonal relationship of the isolates with human clinical strains from the database. Materials and methods: A total of 199 cloacal swabs were collected from five poultry breeds/varieties (broiler, Vanraja, Desi, Nicobari, and layer) in three districts of the Andaman and Nicobar Islands. E. coli, Salmonella enterica, and Klebsiella pneumoniae were isolated by standard techniques and confirmed by PCR. Phenotypical ß-lactamase producers were identified by a double-disc test. The genes (bla CTX, bla SHV, bla TEM , and bla AmpC) were screened, and selected sequences of ß-lactamase variants were submitted to DDBJ. Homology modeling, model validation, and active site identification of different ß-lactamase variants were done by the SWISS-MODEL. Molecular docking was performed to identify the catalytic domains of the ß-lactamase variants. The selected ß-lactamase sequences were compared with the Indian ESBL sequences from human clinical strains in NCBI-GenBank. Results: In total, 425 Enterobacteriaceae strains were isolated from the collected samples. Klebsiella pneumoniae (42.58%) was found to be the most prevalent, followed by Salmonella enterica (30.82%) and E. coli (26.58%). The phenotypical antibiogram of all 425 isolates showed the highest resistance against oxytetracycline (61-76%) and the lowest against gentamicin (15-20%). Phenotypical production of ß-lactamase enzymes was observed in 141 (33.38%) isolates. The isolation rate of ß-lactamase producing E. coli, Salmonella enterica, and Klebsiella pneumoniae was significantly higher (p < 0.05) in the birds reared in the South Andaman district (25.6, 17.5, and 18.7%, respectively) than in Nicobar (11.5, 7.6, 7.1%, respectively). Genotyping of the ß-lactamase-producing isolates revealed the maximum possession of bla TEM, followed by bla SHV and bla CTX - M. The nucleotide sequences were found to be similar with bla CTX - M-15, bla SHV - 11, bla SHV - 27, bla SHV - 228, bla TEM - 1, and bla AmpC in BLAST search. Distribution of studied biofilm-associated genes in Enterobacteriaceae strains from different varieties of the birds revealed that the layer birds had the maximum possession, followed by Vanraja, Desi, broilers, and Nicobari fowls. The phylogenetic analysis of selected sequences revealed a partial clonal relationship with human clinical strains of the Indian subcontinent. Molecular docking depicted the Gibbs free energy release for 10 different macromolecules (proteins) and ligand (antibiotic) complexes, ranging from -8.1 (SHV-27 + cefotaxime) to -7 (TEM-1 + cefotaxime) kcal/mol. Conclusion and relevance: The study revealed ß-lactamase variants circulating in the fowl population of the Andaman and Nicobar Islands (India), even in remote places with low anthropogenic activity. Most of the strains possessed bla TEM - 1, followed by bla CTX - M-15. Possession of bla SHV - 11, bla SHV - 27, and bla SHV - 228 in poultry Enterobacteriaceae strains was not reported earlier from any part of the world. The phylogenetic analysis revealed a partial clonal relationship of ß-lactamase sequences with the human clinical strains isolated from the Indian subcontinent.

Effect of dietary phytobiotic mixture on growth performance, nutrient utilization, and immunity in weaned piglets.

This study investigated the effects of dietary phytobiotic mixture on growth performance, blood profiles, immune response, and fecal microorganisms in weaned piglets. Twenty four weaned crossbred piglets were equally divided into four groups in a completely randomized design. The animals in 4 groups were fed a basal diet added with (1) no antibiotics and phytobiotics (CON), (2) bacitracin (0.5 g/kg; AB), (3) a blend of Cinnamomum zeylanicum and Trachyspermum copticum essential oils (0.3 g/kg and 0.4 g/kg, respectively; EO), and (4) plant extracts (PEO) of Mikania micrantha and Garcinia lanceifolia (2.8 g/kg and 1.4 g/kg, respectively) and C. zeylanicum and T. copticum essential oils (0.3 g/kg and 0.4 g/kg, respectively). Inclusion of AB, EO, and PEO did not affect final body weight, average daily gain, feed intake, feed efficiency, and nutrient digestibility. Compared with the CON, serum protein profiles were not affected, but a few lipid profiles were improved, particularly cholesterol, low-density lipoprotein, and high-density lipoprotein in the EO and PEO groups. Lymphocyte proliferation index and concentrations of IgG and IgA and TNF-α were not affected by any treatments. The concentrations of IgM increased (P = 0.04) at 28 days and tended to increase (P = 0.10) at 56 days in the EO group. Serum IL-1ß levels decreased on days 28 and 56 in the EO and PEO groups. Fecal Lactobacilli population generally increased (P < 0.01) in the AB, EO, and PEO groups compared with the CON. Fecal enterobacterial numbers were always greater for AB than for CON, EO, or PEO, but enterobacterial populations were sometimes lower in the EO group than the CON group. In conclusion, dietary EO or PEO has no effect on the growth performance, but it may improve a few lipid profiles, immune responses, and fecal microbial populations in piglets.

Isolation and molecular characterization of GP5 glycoprotein gene of Betaarterivirus suid 2 from Mizoram, India.

Porcine reproductive and respiratory syndrome (PRRS) is a serious swine disease causing great economic impact worldwide. The emergence of highly pathogenic strains in Asian countries is associated with large scale mortality in all age groups of pigs besides the classical presentation of severe respiratory distress, pneumonia, and a series of reproductive disorders in sows, like late-term abortion, premature farrowing, and an increased number of stillborn piglets. The present study was designed with the aim of isolation and characterization of the Betaarterivirus suid 2 from outbreaks in Mizoram in primary porcine alveolar macrophage and subsequently characterized the GP5 gene sequence of the isolate in terms of phylogenetic analysis and deduce amino acid sequence comparison. Virus propagation was performed in the porcine alveolar macrophage (PAM) primary cell culture and confirmed by immunoperoxidase test, FAT, and nested RT-PCR. The full-length GP5 gene (603nt) was amplified from the isolate and subsequently cloned and sequenced (MN928985). Phylogenetic analysis and sequence comparison of the present isolate was found to have similarity 98.7-98.8% with Myanmar HP-PRRS strains, 98-98.5% with Vietnam strains, 98.2-98.3% with China strains, indicating a close lineage with highly pathogenic PRRS strains. In deduced amino acid sequence analysis, one mutation was found in the primary neutralizing epitope (PNE) at position 39L → I39 and one more mutation was also found in the decoy epitope (DCE) at position 30 N → D30. The amino acid at this position is an N-linked glycosylation site, and mutation of the N-linked glycosylation is an immune escaped strategy adopted by this virus causing a persistent infection in the natural host.

Characterisation of Porcine Enteropathogenic Escherichia Coli Isolated in Northeastern India.

INTRODUCTION: Enteropathogenic Escherichia coli (EPEC) is one of the main pathotypes causing gastroenteritis, particularly in young immunocompromised hosts. The study reports the prevalence, characterisation, and molecular epidemiology of EPEC from piglets in northeastern India. MATERIAL AND METHODS: A total of 457 faecal samples were collected, from which 1,286 E. coli strains were isolated and screened by PCR. The resultant EPEC strains were serotyped and phenotypically characterised for resistance against 15 antimicrobials. Also, the phylogenetic sequence was analysed for 11 selected strains. RESULTS: A total of 42 strains (3.26%) belonged to atypical EPEC, of which, 15 (35.71%, and 2.29% of the 654 strains from this farm type) were isolated from organised and 27 (64.29%, and 4.27% of the 632 strains from this farm type) from unorganised farms; further, 5 (11.90% of the EPEC strains and 1.51% of the 330 strains from this breed) were isolated from the indigenous breeds and 37 (88.10%, and 3.87% of the 956 strains from this breed) from crossbred piglets. Serogroups O111 (11.9%) and O118 (7.14%) were the most prevalent of the 10 present. Sequence analysis of a length of the eaeA gene of 11 isolates of the region showed them to have 100% homology with each other and their identity ranged from 99.4% to 99.7% with GenBank reference sequences. All the EPEC isolates were multi-drug resistant, showing the highest resistance to amoxicillin (80.9%) and cephalexin (76.19%). CONCLUSION: The study highlighted the association of EPEC with piglet's diarrhoea in northeastern India. EPEC isolates belonged to many serotypes and phenotypically all were multi-drug resistant with close genetic homology.

Systematic evaluation of species-independent serum pre-fractionation strategies revealed cost-effective methods to reduce proteome complexity.

In this study, the efficiency of one commercial (ProteoMiner™ -PM) and five simple and cost-effective laboratory chemicals (Acetone, TCA/acetone, DTT, ACN and DTT-ACN) based serum protein pre-fractionation strategies was compared in pig model by label-free quantitation based mass spectrometric approach to find out the most suitable strategy for reducing the complexity of serum proteome for subsequent proteomic studies. The highest serum protein depletion percentage and highest depletion of albumin, the most abundant serum protein, was observed in DTT-ACN method. The maximum number of serum proteins was identified in ACN followed by DTT-ACN method and importantly, detection of more number of low-abundant proteins (LAPs) could also be achieved by these two methods. Although PM method resulted into lowest dynamic range of protein abundance, quite a less number of proteins were identified by this method. Overall, sequential depletion using DTT-ACN and ACN methods provided advantage of simultaneous detection of more number of proteins along with LAPs with a reasonably high dynamic range of protein abundances over other methods and thus emerged as cheaper and effective alternatives to the commercial methods. Further, these methods are species-independent and hence can be applied in human and in any livestock species to simplify the serum proteome.

An outbreak of Goatpox virus infection in Wild Red Serow (Capricornis rubidus) in Mizoram, India.

Goatpox virus (GTPV) belongs to the genus Capripoxvirus associated with characteristic clinical lesions in fully susceptible breeds of sheep and goats. To date, there is no report of outbreaks of GTPV infection in any wild animals. This study reports the outbreak of GTPV infection in wild Red Serow (Capricornis rubidus.) in Mizoram, India. A total of 113 wild Serow carcasses were recovered from seven districts of Mizoram between May 2015 to October 2016. A postmortem revealed presumptive pox-like lesions. Clinical specimens (lung, skin, and trachea) were examined for the aetiological agents. GTPV could be isolated in PLT cells and confirmed in PCR assays by targeting RPO30 and P32 genes. The genetic and phylogenetic analysis reveled that over 99.8% sequence identity with GTPV from India and other parts of the world. To the authors' knowledge, this is the first report of GTPV infection in wild ruminants.

Multidrug-resistant Staphylococcus pettenkoferi isolated from cat in India.

BACKGROUND AND AIM: Coagulase-negative staphylococci (CoNS) are considered to be one of the emerging pathogens in human and animals in recent times. Staphylococcus pettenkoferi, a novel pathogen under CoNS, is discovered in 2002 in humans with multiple clinical manifestations in various patients. To date, the pathogens have not yet been reported from any animals. The present study reported the first ever isolation, identification, and characterization of multidrug-resistant S. pettenkoferi from a cat with peritonitis in India. MATERIALS AND METHODS: Peritoneal fluid was collected aseptically from 3 years old cat processed for bacteriological culture by standard techniques. Isolates were confirmed by BD Phoenix™ automated bacterial identification system and were subjected to plate and tube coagulase tests. All the isolates were tested for antimicrobial sensitivity profile by disc diffusion assay, extended-spectrum ß-lactamase production by double disc diffusion assay, in vitro biofilm production ability by microtiter plate assay, and detection of virulence genes and mecA gene by polymerase chain reaction assay. RESULTS: A total of five clonally expanded isolates of S. pettenkoferi were isolated from peritoneal fluid of the affected cat. All the isolates were resistant against 36 antimicrobial agents and were also methicillin-resistant staphylococci. Phenotypically, all the isolates were negative for biofilm production but were carrying multiple biofilm-producing genes (icaA, IS257, nuc, and mecA). CONCLUSION: Although S. pettenkoferi was previously reported once from animal (cat) environment, this is probably the first ever report of isolation of the organism directly from any animals. This is also probably the first report from any species in India.

The emergence of porcine circovirus 2 infections in the Northeastern part of India: A retrospective study from 2011 to 2017.

Porcine circovirus (PCV) infection has emerged as an alarming threat to the pig population of India, especially in the Northeastern region (NER) over the last 10 years. The present study is a comprehensive report of the seroepidemiology of PCV2 and its incidences in the pig population from organized and unorganized farms of the entire NER of India from 2011 to 2017. A total of 5697 serum samples were screened by ELISA and the mean positivity of PCV2 antibodies in suspected sera was 31.27%. A total of 22 confirmed cases of PCV2 infection were recorded during the years 2014-2017. Seroprevalence of PCV2 infection in sows causing reproductive disorders in NER suggested its higher incidence in organized farms (65.7%) as compared to unorganized farms (17.6%). A detailed pathological and histopathological examination of the tissue samples collected from the affected animals indicated the presence of PCV2. Molecular characterization and phylogenetic analysis of four PCV2 isolates depicted the circulation of PCV2d genotype in the states of Meghalaya and Assam.

Molecular evidence and phylogenetic analysis of orf virus isolates from outbreaks in Tripura state of North-East India.

This study describes the first confirmed report of contagious ecthyma in Black Bengal goats from Tripura state, a North-Eastern state of India situated at the Indo-Bangladesh border. Outbreaks were characterized by the high rates of morbidity (58-67%), low mortality (8-10%) and case fatality (11-15%). The etiology of the outbreaks was confirmed as orf virus (ORFV) by standard virological/serological and molecular techniques including sequence analysis of B2L, a major envelop protein gene of genus Parapoxvirus. Sequence and phylogenetic analysis based on B2L gene of ORFV isolates from Tripura revealed that they were closely related to each other and also to other Indian isolates, in particular to ORFV-Shahjahanpur 82/04 isolate from North India. They revealed several specific nucleotide/amino acid substitutions, namely G299A (G100D), G660A, C705T, C795T (N267D) and G872A (R291H) which may be of notable epidemiological significance. This report necessitates the systematic investigation of orf outbreaks in susceptible populations including wild species particularly at transboundary regions by use of rapid diagnostics to control the infection by deploying an effective vaccine/therapeutics and better managemental practices.

Prevalence of CTX-M-Producing Klebsiella spp. in Broiler, Kuroiler, and Indigenous Poultry in West Bengal State, India.

This study was undertaken to detect the prevalence of CTX-M-producing Klebsiella spp. in healthy broiler, indigenous, and kuroiler birds reared in West Bengal (India) during November 2014-February 2015. In addition to CTX-M gene, the study was also conducted to reveal the occurrence of other ß-lactamase and class I integron genes in Klebsiella spp. isolates along with their clonal relationship. A total of 321 cloacal swabs from healthy broiler, indigenous, and kuroiler birds were collected from different places of West Bengal, India. Klebsiella spp. isolation rate varies among different types of poultry birds (43.8-72.3%). In total, 33 (10.7%) Klebsiella spp. isolates were detected phenotypically as CTX-M producers and all the isolates possessed blaCTX-M in polymerase chain reaction. Whereas 17 (51.5%) and 16 (48.5%) Klebsiella spp. isolates possessed blaSHV, and blaTEM with blaCTX-M, respectively. None of the CTX-M-producing Klebsiella spp. isolates in this study possessed class I integron gene. Randomly amplified polymorphic DNA-based phylogenetic tree revealed the presence of clonal relationship among the CTX-M-producing Klebsiella spp. isolates, recovered from broilers and indigenous birds. This study identified broilers and indigenous game birds as a potential reservoir of CTX-M-producing Klebsiella spp., which could be transmitted to the human food chain directly or indirectly.

Prevalence of multi drug resistant enteropathogenic and enteroinvasive Escherichia coli isolated from children with and without diarrhea in Northeast Indian population.

BACKGROUND: Diarrheagenic Escherichia coli are associated with infantile diarrhea in the developing countries. The present study was conducted to determine the occurrence and antimicrobial resistance pattern of enteropathogenic and enteroinvasive E. coli associated with diarrhoea among the paediatric patients. METHODS: A total of 262 stool samples were collected from children with and without diarrhea from Mizoram, Northeast India. E. coli were isolated and subjected to multiplex PCR to detect virulent genes of EPEC (eaeA and bfpA) and EIEC (ial). Isolates were subjected to antimicrobial sensitivity assay using disc diffusion method. Selected eaeA genes were sequenced for identification and genetic relationship. RESULTS: A total of 334 E. coli was isolated, of which 17.37% were carrying at least one virulent gene. Altogether, 14.97 and 2.40% isolates were categorized as EPEC and EIEC, respectively. Among the DEC isolates, 4.79% were EPEC and 7.78% were EIEC. A total of 8 (2.40%) isolates were EIEC (ial+), of which 6 (1.80%) and 2 (0.60%) were from diarrhoeic and non-diarrhoeic patients, respectively. A total of 24 (41.40%) DEC isolates were MDR (resistance against ≥5 antimicrobials). CONCLUSIONS: A high frequency of EPEC pathotypes associated with paediatric diarrhea was observed in Mizoram, Northeast India and majority of the isolates are resistant to antibiotics with a high frequency of MDR, which is a matter of concern to the public health. This also raises an alarm to the world communities to monitor the resistance pattern and analyse in a global scale to combat the problems of resistance development.

Co-infection of methicillin-resistant Staphylococcus epidermidis, methicillin-resistant Staphylococcus aureus and extended spectrum ß-lactamase producing Escherichia coli in bovine mastitis--three cases reported from India.

Emergence of antimicrobial resistance among bovine mastitis pathogens is the major cause of frequent therapeutic failure and a cause of concern for veterinary practitioners. This study describes intra-mammary infection of methicillin-resistant Staphylococcus epidermidis (MRSE), methicillin-resistant Staphylococcus aureus (MRSA) and extended spectrum ß-lactamase (ESBL) producing Escherichia coli in two Holstein Friesian crossbred cows with subclinical mastitis and one non-descript cow with clinical mastitis in two different districts of West Bengal, India. In total, three MRSE, one MRSA and three ESBL producing E. coli were isolated from these cases. Both the crossbreds were detected with MRSE (HFSE1 and HFSE2) and ESBL producing E. coli (HFEC1 and HFEC2), whereas, simultaneous infection of three pathogens viz. MRSA (NDSA1), MRSE (NDSE1) and ESBL producing E. coli (NDEC1) was found in the non-descript cow. The methicillin-resistant isolates possessed mecA gene and exhibited resistance to various antibiotics such as amikacin, tetracycline and glycopeptides. The ESBL producers were positive for blaCTX-M and blaTEM genes; in addition, HFEC1 and HFEC2 were positive for blaSHV and possessed the genes for class I integron (int1), sulphonamide resistance (sul1), quinolone resistance (qnrS) and other virulence factors (papC, iucD and ESTA1). All the ESBL producers exhibited resistance to a variety of antibiotics tested including third- and fourth-generation cephalosporins and were also intermediately resistant to carbapenems. This is the first ever report on simultaneous occurrence of MRSE, MRSA and ESBL producing E. coli in bovine mastitis indicating a major concern for dairy industry and public health as well.

ESBL-producing Shiga-toxigenic E. coli (STEC) associated with piglet diarrhoea in India.

The present study was conducted to investigate the prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli carrying other virulence genes associated with piglet diarrhoea. Faecal samples from the piglets with history of diarrhoea were processed for isolation of E. coli and were tested for their ability of ESBL production. ESBL encoding genes and other virulence genes were detected by specific PCR, and amplicons were sequenced. Ability of transferring the ESBL genes between the enteric bacteria was tested in in vitro HGT. A total of 170 E. coli was isolated, of which 43 (25.29 %) were confirmed as ESBL producer by double disc synergy test (DDST). Altogether, 6.47 and 2.94 % isolates were positive for bla TEM and bla CTX-M-15 genes, respectively, of which 2.35 % isolates were positive for both the genes and only 0.6 % isolate was positive for the bla CTX-M gene alone. The resistance traits could not be transferred to the recipient host. Based on PCR, 2 (1.18 %) and 1 (0.59 %) isolates were recorded as Shiga-toxin-producing E. coli (STEC) and enteropathogenic E. coli (EPEC), respectively. Both the STEC (one isolate positive for stx 2 and another positive for stx 2 and hlyA) isolates belonging to serogroup O2 and NT were also positive for the bla CTX-M-15 gene. This is the first report of ESBL-producing E. coli isolate possessing the STEC gene associated with piglet diarrhoea.

Seroprevalence of brucellosis in yaks (Poephagus grunniens) in India and evaluation of protective immunity to S19 vaccine.

The present study was carried out to explore the seroprevalence of brucellosis in yaks of North-Eastern hilly yak tracts of Arunachal Pradesh, India. Of 374 animals tested, 23.79, 21.11 and 18.98% were found positive for brucellosis using avidin-biotin ELISA (AB-ELISA), Rose-Bengal plate test (RBPT) and standard tube-agglutination test (STAT), respectively. The relative sensitivity and specificity for STAT were 79.77 and 100%, respectively and the same for RBPT were 88.76 and 100%, respectively in comparison to AB-ELISA. The alarming prevalence as recorded was highest among the yak cows (31.42%) followed by heifers (23.85%) and bulls (8.88%). The immune response in yaks following standard dose of calfhood vaccination with Brucella abortus strain 19 vaccine showed that protective antibody level persisted up to 210 days. This is the first report from India on prevalence of brucellosis and immunization with B abortus strain 19 vaccine in yaks. The present investigation would be a valuable guideline for future control measure and eradication programme of brucellosis in yaks.