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Avian nephritis virus (ANV), which belongs to the family Astroviridae , is associated with different clinical manifestations (enteric and kidney disorders) in poultry. Despite being a significant pathogen of the avian industry worldwide, information regarding genetic features of these viruses in India is scarce. In this study, 386 intestinal samples collected from 37 slaughterhouses in two north Indian states (Rajasthan and Haryana) were screened for ANV with reverse transcription PCR (RT-PCR) targeting the conserved ORF1b gene, followed by nucleotide sequencing of the amplified product. RT-PCR and sequencing confirmed the presence of ANV in 32 clinical samples (8.29%), with concurrent infections of infectious bronchitis virus, chicken astrovirus, and fowl adenoviruses observed in some clinical samples ( n = 4). Virus isolations were successful from four out of 12 ANV-positive clinical samples passaged via the yolk-sac route in specific-pathogen-free embryonated chicken eggs. Additionally, the near-complete genomes of two viruses were determined through sequencing. Phylogenetic analysis based on full-length capsid protein sequences classified the viruses into ANV genotype 4 (ANV4), and to the best of our knowledge this is the first report of ANV4 from India. This study revealed the presence and circulation of new strains of ANV in Indian poultry. Genetic profiling and isolation of the viruses in this study will not only aid in the development of diagnostic tools and vaccines for ANV but also offer valuable insights into its epidemiology.

Salmonella enterica is a foodborne pathogen associated with both typhoid and non-typhoid illness in humans and animals. This problem is further exacerbated by the emergence of antibiotic-resistant strains of Salmonella enterica . Therefore, to meet public health and safety, there is a need for an alternative strategy to tackle antibiotic-resistant bacteria. Bacteriophages or (bacterial viruses), due to their specificity, self-dosing, and antibiofilm activity, serve as a better approach to fighting against drug-resistant bacteria. In the current study, a broad-host range lytic phage phiSalP219 was isolated against multidrug-resistant Salmonella enterica serotypes Paratyphi from a pond water sample. Salmonella phage phiSalP219 was able to lyse 28/30 tested strains of Salmonella enterica . Salmonella phage phiSalP219 exhibits activity in acidic environments (pH3) and high temperatures (70°C). Electron microscopy and genome analysis revealed that phage phiSalP219 is a member of class Caudoviricetes . The genome of Salmonella phage phiSalP219 is 146Kb in size with 44.5% GC content. A total of 250 Coding Sequence (CDS) and 25 tRNAs were predicted in its genome. Predicted open reading frames (ORFs) were divided into five groups based on their annotation results: (1) nucleotide metabolism, (2) DNA replication and transcription, (3) structural proteins, (4) lysis protein, and (5) other proteins. The absence of lysogeny-related genes in their genome indicates that Salmonella phage phiSalP219 is lytic in nature. Phage phiSalP219 was also found to be microbiologically safe (due to the absence of toxin or virulence-related genes) in the control of Salmonella enterica serovar Typhimurium infections in the ready-to-eat meat and also able to eradicate biofilm formed by the same bacterium on the borosilicate glass surface., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Jaglan, Verma, Vashisth, Virmani, Bera, Vaid and Anand.)

In the current scenario of growing world population, limited cultivable land resources, plant diseases, and pandemics are some of the major factors responsible for declining global food security. Along with meeting the food demand, the maintenance of food quality is also required to ensure healthy consumption and marketing. In agricultural fields, pest infestations and bacterial diseases are common causes of crop damage, leading to massive yield losses. Conventionally, antibiotics and several pesticides have been used to manage and control these plant pathogens. However, the overuse of antibiotics and pesticides has led to the emergence of resistant strains of pathogenic bacteria. The bacteriophages are the natural predators of bacteria and are host-specific in their action. Therefore, the use of bacteriophages for the biocontrol of pathogenic bacteria is serving as a sustainable and green solution in crop protection and production. In this review, we have discussed the important plant pathogens and their impact on plant health and yield loss. Further, we have abridged the role of bacteriophages in the protection of crops from bacterial disease by discussing various greenhouse and field trials. Finally, we have discussed the impact of bacteriophages on the plant microbiome, phage resistance, and legal challenges in the registration and commercial production of bacteriophage-based biopesticides., Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-024-01204-x., Competing Interests: Conflict of interestThe authors declare no conflict of interest., (© Association of Microbiologists of India 2024. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)

Extensive and multiple drug resistance in P. aeruginosa combined with the formation of biofilms is responsible for its high persistence in nosocomial infections. A sequential method to devise a suitable phage cocktail with a broad host range and high lytic efficiency against a biofilm forming XDR P. aeruginosa strain is presented here. Out of a total thirteen phages isolated against P. aeruginosa , five were selected on the basis of their high lytic spectra assessed using spot assay and productivity by efficiency of plating assay. Phages, after selection, were tested individually and in combinations of two-, three-, four-, and five-phage cocktails using liquid infection model. Out of total 22 combinations tested, the cocktail comprising four phages viz. φPA170, φPA172, φPA177, and φPA180 significantly inhibited the bacterial growth in liquid infection model ( p < 0.0001). The minimal inhibitory dose of each phage in a cocktail was effectively reduced to >10 times than the individual dose in the inhibition of XDR P. aeruginosa host. Field emission-scanning electron microscopy was used to visualize phage cocktail mediated eradication of 4-day-old multi-layers of XDR P. aeruginosa biofilms from urinary catheters and glass cover slips, and was confirmed by absence of any viable cells. Differential bacterial inhibition was observed with different phage combinations where multiple phages were found to enhance the cocktail's lytic range, but the addition of too many phages reduced the overall inhibition. This study elaborates an effective and sequential method for the preparation of a phage cocktail and evaluates its antimicrobial potential against biofilm forming XDR strains of P. aeruginosa .

Trueperella pyogenes is a Gram-positive opportunistic pathogen that causes severe cases of mastitis, metritis, and pneumonia in a wide range of animals, resulting in significant economic losses. Although little is known about the virulence factors involved in the disease pathogenesis, a comprehensive comparative genome analysis of T. pyogenes genomes has not been performed till date. Hence, present investigation was carried out to characterize and compare 19 T. pyogenes genomes originating in different geographical origins including the draftgenome of the first Indian origin strain T. pyogenes Bu5. Additionally, candidate virulence determinants that could be crucial for their pathogenesis were also detected and analyzed by using various bioinformatics tools. The pan-genome calculations revealed an open pan-genome of T. pyogenes . In addition, an inventory of virulence related genes, 190 genomic islands, 31 prophage sequences, and 40 antibiotic resistance genes that could play a significant role in organism's pathogenicity were detected. The core-genome based phylogeny of T. pyogenes demonstrates a polyphyletic, host-associated group with a high degree of genomic diversity. The identified core-genome can be further used for screening of drug and vaccine targets. The investigation has provided unique insights into pan-genome, virulome, mobiliome, and resistome of T. pyogenes genomes and laid the foundation for future investigations.

Emergence of multiple drug resistant (MDR) strains of Acinetobacter baumannii and a withering drug discovery pipeline necessitates the search for effective alternatives to replace or synergize with currently used antibiotics. In this report, we have described the synergy assessment of a virulent Acinetobacter baumannii phage φAB182 with a wide range of antibiotics. Myophage φAB182 was isolated from sewage against MDR A. baumannii and exhibited maximum stability at 25 °C and pH 7. It also had a short latent period of 9 min with a large burst size of 287. The phylogenetic analysis of its major capsid protein gene indicated an 84.15% similarity to the lytic  A. baumannii phage Acj9. In the presence of antibiotics, phage φAB182 showed the highest synergy ( p  < 0.0001) with colistin, followed by polymixin B, ceftazidime and cefotaxime and this synergistic effect was further validated by time kill kinetics. The combined action of phage φAB182 with colistin, polymixin B, ceftazidime and cefotaxime was also synergistic for the eradication of biofilms formed by A. baumannii as measured by MBEC combination /MBEC antibiotic values (<0.25). We thus propose bacteriophage φAB182 as a potential antibacterial candidate in combination therapy. The findings from this study strongly support the use of phage antibiotic synergy for the successful treatment of biofilm forming MDR A. baumannii infections.

In the present scenario, the challenge of emerging antimicrobial resistance is affecting human health globally. The increasing incidences of multidrug-resistant infections have become harder to treat, causing high morbidity, and mortality, and are posing extensive financial loss. Limited discovery of new antibiotic molecules has further complicated the situation and has forced researchers to think and explore alternatives to antibiotics. This has led to the resurgence of the bacteriophages as an effective alternative as they have a proven history in the Eastern world where lytic bacteriophages have been used since their first implementation over a century ago. To help researchers and clinicians towards strengthening bacteriophages as a more effective, safe, and economical therapeutic alternative, the present review provides an elaborate narrative about the important aspects of bacteriophages. It abridges the prerequisite essential requirements of phage therapy, the role of phage biobank, and the details of immune responses reported while using bacteriophages in the clinical trials/compassionate grounds by examining the up-to-date case reports and their effects on the human gut microbiome. This review also discusses the potential of bacteriophages as a biocontrol agent against food-borne diseases in the food industry and aquaculture, in addition to clinical therapy. It finishes with a discussion of the major challenges, as well as phage therapy and phage-mediated biocontrols future prospects., (Copyright © 2022 Jaglan, Anand, Verma, Vashisth, Virmani, Bera, Vaid and Tripathi.)

Combination therapy of bacteriophages and antibiotics requires careful selection of specific antibiotics as it is crucial towards determining the success of phage therapy to treat multiple drug-resistant bacterial infections. So, we examined how different antibiotics can affect phage lytic activity when used in combination against targeted bacteria. Various antibiotics targeting bacterial protein synthesis pathways were tested for their bactericidal action in combination with bacteriophages of Acinetobacter baumannii (φAB145, φAB182), Staphylococcus aureus (φSA115, φSA116) and Salmonella Typhimurium (φST143, φST188). The phages displayed highly significant antagonism with most of the protein/ribosomal machinery targeting antibiotics: φSA115 (13/13); φSA116 (13/13); φST143 (11/13); φAB145 (11/13); φST188 (9/13); φAB182 (7/13). To validate this antagonistic effect, synergy assessment of these phages with gentamicin (GEN) and tetracycline (TE) was performed using time kill curve assays and counting the remaining viable bacterial cells at the end of the experiment. An increase in bacterial turbidity in phage-antibiotic combination groups was observed as compared to the treatment with phages individually. Also, GEN exhibited 4.22, 5.90, 2.02, 3.15, 2.68, and 2.60 log proliferation in viable cell count, respectively, for φSA115, φSA116, φST145, φAB182, φST143 and φAB188 in combination group in comparison to their individual actions. TE supplementation also led to 2.40, 4.90, 1.61, 2.73, 3.93, and 1.81 log increments in viable bacterial count when combined with φSA115, φSA116, φST145, φAB182, φST143 and φAB188, respectively. This study concludes that antibiotics targeting the bacterial protein biosynthetic machinery may lead to a reduction in the lytic activity of bacteriophages, thus lowering their therapeutic potential. Hence, such compounds must be carefully screened before their employment in combination treatment regimens., Competing Interests: Declaration of Competing Interest The authors declare they have no conflicts of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Trypanosomes are the hemoflagellate kinetoplastid protozoan parasites affecting a wide range of vertebrate hosts having insufficient host specificity. Climatic change, deforestation, globalization, trade agreements, close association and genetic selection in links with environmental, vector, reservoir and potential susceptible hosts' parameters have led to emergence of atypical human trypanosomosis (a-HT). Poor recording of such neglected tropical disease, low awareness in health professions and farming community has approached a serious intimidation for mankind. Reports of animal Trypanosoma species are now gradually increasing in humans, and lack of any compiled literature has diluted the issue. In the present review, global reports of livestock and rodent trypanosomes reported from human beings are assembled and discrepancies with the available literature are discussed along with morphological features of Trypanosoma species. We have described 21 human cases from the published information. Majority of cases 10 (47%) are due to T. lewisi, followed by 5 (24%) cases of T. evansi, 4 (19%) cases of T. brucei and 1 (5%) case each of T. vivax and T. congolense. Indian subcontinent witnessed 13 cases of a-HT, of which 9 cases are reported from India, which includes 7 cases of T. lewisi and 2 cases of T. evansi. Apart from, a-HT case reports, epidemiological investigation and treatment aspects are also discussed. An attempt has been made to provide an overview of the current situation of atypical human trypanosomosis caused by salivarian animal Trypanosoma globally. The probable role of Trypanosoma lytic factors (TLF) present in normal human serum (NHS) in providing innate immunity against salivarian animal Trypanosoma species and the existing paradox in medical science after the finding on intact functional apolipoprotein L1 (ApoL1) in Vietnam T. evansi Type A case is also discussed to provide an update on all aspects of a-HT. Insufficient data and poor reporting in Asian and African countries are the major hurdle resulting in under-reporting of a-HT, which is a potential emerging threat. Therefore, concerted efforts must be directed to address attentiveness, preparedness and regular surveillance in suspected areas with training of field technicians, medical health professionals and veterinarians. Enhancing a one health approach is specifically important in case of trypanosomosis., (© 2022 Wiley-VCH GmbH.)

Salmonella enterica serovar Gallinarum biovar Pullorum (bvP) and biovar Gallinarum (bvG) are the etiological agents of pullorum disease (PD) and fowl typhoid (FT) respectively, which cause huge economic losses to poultry industry especially in developing countries including India. Vaccination and biosecurity measures are currently being employed to control and reduce the S. Gallinarum infections. High endemicity, poor implementation of hygiene and lack of effective vaccines pose challenges in prevention and control of disease in intensively maintained poultry flocks. Comparative genome analysis unravels similarities and dissimilarities thus facilitating identification of genomic features that aids in pathogenesis, niche adaptation and in tracing of evolutionary history. The present investigation was carried out to assess the genotypic differences amongst S.enterica serovar Gallinarum strains including Indian strain S. Gallinarum Sal40 VTCCBAA614. The comparative genome analysis revealed an open pan-genome consisting of 5091 coding sequence (CDS) with 3270 CDS belonging to core-genome, 1254 CDS to dispensable genome and strain specific genes i.e. singletons ranging from 3 to 102 amongst the analyzed strains. Moreover, the investigated strains exhibited diversity in genomic features such as virulence factors, genomic islands, prophage regions, toxin-antitoxin cassettes, and acquired antimicrobial resistance genes. Core genome identified in the study can give important leads in the direction of design of rapid and reliable diagnostics, and vaccine design for effective infection control as well as eradication. Additionally, the identified genetic differences among the S. enterica serovar Gallinarum strains could be used for bacterial typing, structure based inhibitor development by future experimental investigations on the data generated., Competing Interests: The authors have declared that no competing interests exist.

Phage display is one of the important and effective molecular biology techniques and has remained indispensable for research community since its discovery in the year 1985. As a large number of nucleotide fragments may be cloned into the phage genome, a phage library may harbour millions or sometimes billions of unique and distinctive displayed peptide ligands. The ligand-receptor interactions forming the basis of phage display have been well utilized in epitope mapping and antigen presentation on the surface of bacteriophages for screening novel vaccine candidates by using affinity selection-based strategy called biopanning. This versatile technique has been modified tremendously over last three decades, leading to generation of different platforms for combinatorial peptide display. The translation of new diagnostic tools thus developed has been used in situations arising due to pathogenic microbes, including bacteria and deadly viruses, such as Zika, Ebola, Hendra, Nipah, Hanta, MERS and SARS. In the current situation of pandemic of Coronavirus disease (COVID-19), a search for neutralizing antibodies is motivating the researchers to find therapeutic candidates against novel SARS-CoV-2. As phage display is an important technique for antibody selection, this review presents a concise summary of the very recent applications of phage display technique with a special reference to progress in diagnostics and therapeutics for coronavirus diseases. Hopefully, this technique can complement studies on host-pathogen interactions and assist novel strategies of drug discovery for coronaviruses.

Objectives: Klebsiella pneumoniae is an important emerging pathogen of humans and animals leading to serious clinical consequences. Increased antibiotic use has promoted the emergence of carbapenem-resistant and extended-spectrum β-lactamase (ESBL)-producing K. pneumoniae strains. Recently, phage therapy has gained momentum as a possible alternative against emerging antimicrobial resistance. This study was performed to assess the therapeutic effects of a novel lytic phage (VTCCBPA43) in a pneumonic mouse model in order to explore the efficacy of phage therapy against virulent K. pneumoniae infection., Methods: The tailed phage VTCCBPA43 was assessed for its growth kinetics, in vitro host range, and temperature and pH sensitivity. Protein constituents were analysed by SDS-PAGE and nLC-MS/MS. Therapeutic efficacy was observed 2 h post-challenge with virulent K. pneumoniae in a BALB/c mouse model., Results: Phage VTCCBPA43 was found to be highly temperature-tolerant (up to 80 °C). It was most active at pH 5, had a burst size of 172 PFU/mL and exhibited a narrow host range. It was identified as a KP36-like phage by shotgun proteomics. Following intranasal application of a single dose (2 × 10 9 PFU/mouse) post-challenge with virulent K. pneumoniae, the presence of biologically active phage in vivo and a significant reduction in the lung bacterial load at all time points was observed. A reduction in lesion severity suggested overall beneficial effects of VTCCBPA43 phage therapy in the pneumonic mouse model., Conclusion: This research represents the first in vivo evidence of effective phage therapy against K. pneumoniae infection by the intranasal route., (Copyright © 2019 International Society for Antimicrobial Chemotherapy. Published by Elsevier Ltd. All rights reserved.)

A virulent Aeromonas veronii biovar sobria and the corresponding novel, lytic bacteriophage (VTCCBPA5) were isolated from village pond water. The phage was found to belong to family Podoviridae. PCR analysis of major capsid protein gene confirmed its classification to T7-like genus. The protein profiling by SDS-PAGE indicated the major structural protein to be ~ 45 kDa. The phage (VTCCBPA5) is host specific and is stable over a range of pH (6-10) and temperatures (4-45 °C). On the basis of restriction endonuclease analysis combined with prediction mapping, it was observed to vary significantly from previously reported podophages of Aeromonas sp., viz. phiAS7 and Ahp1. The phylogenetic analysis on the basis of PCR-amplified segment of DNA polymerase gene of phage revealed it being an outgroup from podophages of Klebsiella sp. and Pseudomonas sp. though a small internal fragment (359 bp) showed the highest identity (77%) with Vibrio sp. phages. Thus, this is the first report of a novel Podoviridae phage against A. veronii. It expands the assemblage of podophages against Aeromonas sp. and BPA5 could be potentially useful in biocontrol of environmentally acquired Aeromonas veronii infections.

Bordetella bronchiseptica is a well-known Gram-negative bacterial pathogen causing a plethora of diseases in different animals. Although its infection has been reported from pigs and dogs in India, no report of B. bronchiseptica from horses is described. We report for the first time, isolation, identification and characterization of strains of B. bronchiseptica from respiratory infection in horses from different states in India. The antimicrobial susceptibility testing showed resistance to penicillins, ceftazidime, and chloramphanicol. The virulence capability of the strains was confirmed by sequencing genes such as adenylate cyclase toxin (cyaA), bordetella virulence gene (bvgA) and by PCR detection of flagellin gene (fla). We demonstrate the involvement of B. bronchiseptica strains in respiratory tract infection in horses in India.

The ecosystem is continuously exposed to a wide variety of antimicrobials through waste effluents, agricultural run-offs and animal-related and anthropogenic activities, which contribute to the spread of antibiotic resistance genes (ARGs). The contamination of ecosystems with ARGs may create increased opportunities for their transfer to naive microbes and eventually lead to entry into the human food chain. Transduction is a significant mechanism of horizontal gene transfer in natural environments, which has traditionally been underestimated as compared to transformation. We explored the presence of ARGs in environmental bacteriophages in order to recognize their contribution in the spread of ARGs in environmental settings. Bacteriophages were isolated against environmental bacterial isolates, purified and bulk cultured. They were characterized, and detection of ARG and intI genes including blaTEM, blaOXA-2, intI1, intI2, intI3, tetA and tetW was carried out by PCR. This study revealed the presence of various genes [tetA (12.7 %), intI1 (10.9 %), intI2 (10.9 %), intI3 (9.1 %), tetW (9.1 %) and blaOXA-2 (3.6 %)] and blaTEM in a significantly higher proportion (30.9 %). blaSHV, blaOXA-1, tetO, tetB, tetG, tetM and tetS were not detected in any of the phages. Soil phages were the most versatile in terms of ARG carriage. Also, the relative abundance of tetA differed significantly vis-à-vis source. The phages from organized farms showed varied ARGs as compared to the unorganized sector, although blaTEM ARG incidences did not differ significantly. The study reflects on the role of phages in dissemination of ARGs in environmental reservoirs, which may provide an early warning system for future clinically relevant resistance mechanisms.

Swinepox virus (SWPV), a member of the genus Suipoxvirus causes generalized pock-like lesions on the body of domestic and wild pigs. Although outbreak has been reported in India since 1987, virus isolation and genetic characterization remained elusive. In September 2013, an outbreak of acute skin infection occurred in piglets in a commercial piggery unit at Rohtak district in Haryana, India. The presence of SWPV in scab samples collected from piglets succumbed to infection was confirmed by virus isolation, PCR amplification of SWPV-specific gene segments and nucleotide sequencing. Phylogenetic analysis of host-range genes of the SWPV revealed that the Indian isolate is genetically closely related to reference isolate SWPV/pig/U.S.A/1999/Nebraska. To the best of our knowledge this is the first report on isolation and genetic characterization of SWPV from pigs in India., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

A bacteriophage (VTCCBPA6) against a pathogenic strain of Aeromonas hydrophila was isolated from the sewage of an organized equine breeding farm. On the basis of TEM analysis, phage belonged to family Myoviridae. PCR amplification and sequence analysis of gp23 gene (encoding for major capsid protein) revealed phylogenetic resemblance to T4 like virus genus. Protein profiling by SDS-PAGE also indicated its resemblance to T4 like phage group. However, the comparison of its gp23 gene sequence with previously reported phages showed similarity with T4-like phages infecting Enterobacteriaceae instead of Aeromonas spp. Thus, to our knowledge, this report points toward the fact that a novel/evolved phage might exist in equine environment against A. hydrophila, which can be potentially used as a biocontrol agent., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Equine influenza viruses (EIV)-H3N8 continue to circulate in equine population throughout the world. They evolve by the process of antigenic drift that leads to substantial change in the antigenicity of the virus, thereby necessitating substitution of virus strain in the vaccines. This requires frequent testing of the new vaccines in the in vivo system; however, lack of an appropriate laboratory animal challenge model for testing protective efficacy of equine influenza vaccine candidates hinders the screening of new vaccines and other therapeutic approaches. In the present investigation, BALB/c mouse were explored for suitability for conducting pathogenecity studies for EIV. The BALB/c mice were inoculated intranasally @ 2×106.24 EID50 with EIV (H3N8) belonging to Clade 2 of Florida sublineage and monitored for setting up of infection and associated parameters. All mice inoculated with EIV exhibited clinical signs viz. loss in body weights, lethargy, dyspnea, etc, between 3 and 5 days which commensurate with lesions observed in the respiratory tract including rhinitis, tracheitis, bronchitis, bronchiolitis, alveolitis and diffuse interstitial pneumonia. Transmission electron microscopy, immunohistochemistry, virus quantification through titration and qRT-PCR demonstrated active viral infection in the upper and lower respiratory tract. Serology revealed rise in serum lactate dehydrogenase levels along with sero-conversion. The pattern of disease progression, pathological lesions and virus recovery from nasal washings and lungs in the present investigations in mice were comparable to natural and experimental EIV infection in equines. The findings establish BALB/c mice as small animal model for studying EIV (H3N8) infection and will have immense potential for dissecting viral pathogenesis, vaccine efficacy studies, preliminary screening of vaccine candidates and antiviral therapeutics against EIV.

Salmonella enterica subsp. enterica serovar Gallinarum biovar Gallinarum causes fowl typhoid (FT), which results in huge economic losses to poultry farmers in India. We report the draft genome sequence of Salmonella biovar Gallinarum strain VTCCBAA614, isolated from a chicken in an FT affected broiler flock., (Copyright © 2015 Vaid et al.)

Phage therapy has been previously tried for treatment of diarrhoea in calves, pigs and lambs but those trials were conducted without any detailed information of used phages. Here, we report isolation of a broad-spectrum phage which showed bactericidal activity against 47.3 % of calf diarrhoeal isolates of Escherichia coli, in vitro. The isolated phage resembled the characteristics of Myoviridae family and showed ~97 % similarity with earlier reported bacteriophages of sub family-Tevenvirinae, genus-T4-like virus, based on nucleotide sequence of major head protein-gp23 gene. The phage exhibits the potential to be used as drug substitute tool against E. coli causing diarrhoea in cattle in farm environments.

Camelpox virus (CMLV), a close variant of variola virus (VARV) infects camels worldwide. The zoonotic infections reported from India signify the need to study the host-range genes-responsible for host tropism. We report sequence and phylogenetic analysis of five host-range genes: cytokine response modifier B (crmB), chemokine binding protein (ckbp), viral schlafen-like (v-slfn), myxomavirus T4-like (M-T4-like) and b5r of CMLVs isolated from outbreaks in India. Comparative analysis revealed that these genes are conserved among CMLVs and shared 94.5-100 % identity at both nucleotide (nt) and amino acid (aa) levels. All genes showed identity (59.3-98.4 %) with cowpox virus (CPXV) while three genes-crmB, ckbp and b5r showed similarity (92-96.5 %) with VARVs at both nt and aa levels. Interestingly, three consecutive serine residue insertions were observed in CKBP protein of CMLV-Delhi09 isolate which was similar to CPXV-BR and VACVs, besides five point mutations (K53Q, N67I, F84S, A127T and E182G) were also similar to zoonotic OPXVs. Further, few inconsistent point mutation(s) were also observed in other gene(s) among Indian CMLVs. These indicate that different strains of CMLVs are circulating in India and these mutations could play an important role in adaptation of CMLVs in humans. The phylogeny revealed clustering of all CMLVs together except CMLV-Delhi09 which grouped separately due to the presence of specific point mutations. However, the topology of the concatenated phylogeny showed close evolutionary relationship of CMLV with VARV and TATV followed by CPXV-RatGer09/1 from Germany. The availability of this genetic information will be useful in unveiling new strategies to control emerging zoonotic poxvirus infections.

The Pasteurella multocida subsp. multocida B:2 serotype causes hemorrhagic septicemia in bubalines with high morbidity and mortality in the Indian subcontinent. We report the draft genome sequence of Pasteurella multocida strain VTCCBAA264 isolated from the small-intestine of a buffalo calf that died of high fever., (Copyright © 2014 Vaid et al.)

The neuraminidase (NA) gene sequences of four Indian equine influenza viruses (EIVs) isolated from epizootic in 2008 and 2009 were analyzed. The phylogenetic relationship and selection pressure of NA genes were established in comparison to other EIVs circulating worldwide along with the domains and motifs of the encoded protein to find out the significance of mutational changes. Among Indian isolates, two amino acid (aa) changes each in Mysore/12/08 (Asn67Tyr & Asp396Gly), Gopeshwar/1/09 (Ile49Val & Asp396Gly), and Uttarkashi/1/09 (Ile49Val & Asp396Gly) isolates were observed in respect to Jammu-Katra/06/08 isolate. Amino acid (aa) sequence analysis also revealed five consistent aa residue changes viz, Gly/Arg40Glu, Tyr66His, Val191Ile, Val209Ile and Asp235Asn in Asian including Indian isolates, Spain/07 and Spain/09 isolates in comparison to other EIVs circulating worldwide. The topology of the phylogenetic tree revealed that the Indian, Chinese, Mongolian and Kazakhstan isolates together formed a subgroup with Yokohama/10 isolate. Spain/07 & Spain/09 isolates showed closest clustering with Asian isolates. This indicates that non-synonymous mutations in Asian isolates with temporal pattern originating from Spain/07, led to the subgroup of the Asian isolates within Florida clade 2 sublineage. The analysis of the predicted secondary structure has not shown any significant difference in the NA proteins of all Indian isolates. Fixed-effects likelihood (FEL) analysis of the selection pressure revealed three codons (43, 355 & 434) under positive selection pressure. The overall evolutionary changes (ω value) of 3.4 indicates NA gene to be under strong selection pressure. Further, seven putative N-glycosylation sites were observed in the NA protein. The mapping of specific aa changes, their mutational and functional analysis need to be carried out to ascertain their role in pathogenecity of the virus.

Buffalopox virus (BPXV), a close variant of vaccinia virus (VACV) has emerged as a zoonotic pathogen. The host tropism of poxviruses is governed by host-range genes. Among the host-range genes: E3L, K3L, and C7L are essential for virus replication by preventing interferon resistance, whereas B5R is essential for spread of the virus and evasion from the host's immune response as in VACV. We report sequence analysis of host-range genes: E3L, K3L, C7L, and membrane protein gene (B5R) of BPXVs from buffalo, cattle, and human from recent outbreaks in India-their phylogenetic relationship with reference strain (BP4) and other Orthopoxviruses. BPXVs revealed a sequence homology with VACVs including zoonotic Brazilian VACV-like viruses. The aa sequences of E3L and K3L genes were 100 % similar in buffalo, cattle, and human isolates. However, four significant point mutations (I11K; N12K and S36F in C7L gene and D249G in B5R gene) were observed specific to buffalo isolate only. This signifies that different strains of BPXV were circulated during the outbreak. The mutations in C7L and B5R could play an important role in adaptation of BPXV in human and cattle which needs further functional studies. The strain of BPXV isolated from buffalo may not be adopted in human and cow. Various point mutations were observed in the host-range genes of reference strain (BPXV-BP4) which may be due to several passages of virus in cell culture. The phylogeny constructed based on concatenated gene sequences revealed that BPXVs are not as closely related to vaccine strain (Lister and Lister-derived strain-LC16m8), as hypothesized earlier, rather they are more closely related to reference strain (BPXV-BP4) and other vaccinia and vaccinia-like viruses such as Passatempo and Aracatuba viruses. The availability of information regarding host tropism determinants would allow us to understand molecular mechanism of species tropism of poxviruses which would be useful in unveiling new strategies to control zoonotic poxviral infections.

Glanders, a bacterial disease of equines caused by Burkholderia mallei, is a fatal infectious disease of equines and has zoonotic significance. The disease has been eradicated from many countries by statutory testing, elimination of infected animals and import restrictions. However, it is still endemic in parts of Africa, Asia, the Middle East and Central and South America. In India, major glanders outbreaks were reported from different parts of the country between 1976 and 1982. Later, sporadic cases of the disease were reported in 1988, 1990 and 1998. The country remained free of glanders for about eight years until the recent outbreaks occurred in eight States from 2006 to 2007. Recurrent episodes have occurred in Himachal Pradesh and Uttar Pradesh, whereas fresh outbreaks occurred in Chhattisgarh from 2009 to 2010. A total of 164 equines were declared positive; a majority of the positive cases (n=77) were from Uttar Pradesh, followed by Maharashtra (n=23), Uttarakhand (n=21) and Andhra Pradesh (n=16). Under the provision of Prevention and Control of Infectious and Contagious Disease in Animals Act, 2009, all the infected animals were euthanised and bio-security measures were implemented to curb the further spread of the disease.

India faced an epizootic of equine influenza in 2008-2009. The isolated viruses were typed as H3N8 and grouped with the clade 2 viruses of Florida sublineage on the basis of haemagglutinin (HA) gene sequence analysis. This report describes the genetic analysis and selection pressure of matrix (M) and non-structural 1 (NS1) genes of the Indian isolates. All isolates shared 98.41% and 99.54% homology with other clade 2 viruses of Asian origin for M1 and M2 amino acid (aa) sequences, respectively. There were 3 and 4 unique aa residue changes respectively in M1 and M2 proteins in all Asian isolates. Phylogenetic analysis revealed clustering of Indian and Chinese isolates in a separate group designated here as Asian clade for M gene. Indian and Chinese isolates shared homology ranging from 98.17% to 99.08% at aa level. The M and NS1 genes were under negative selection pressure with estimated magnitude of pressure (ω) 0.054, 0.581 and 0.30 for M1, M2 and NS1, respectively., (Copyright © 2011 Elsevier B.V. All rights reserved.)

This study reports the first conclusive evidence of zoonotic camelpox virus (CMLV) infection in humans associated with outbreaks in dromedarian camels (Camelus dromedaries) in northwest region of India during 2009. CMLV infection is usually restricted to camels and causes localised skin lesions but occasionally leads to generalised form of disease. However, the present outbreak involved camel handlers and attendants with clinical manifestations such as papules, vesicles, ulceration and finally scabs over fingers and hands. In camels, the pock-like lesions were distributed over the hairless parts of the body. On the basis of clinical and epidemiological features coupled with serological tests and molecular characterization of the causative agent, CMLV zoonosis was confirmed in three human cases. Clinical samples such as skin scabs/swabs and blood collected from affected animals and humans were analysed initially, for the presence of CMLV-specific antigen and antibodies by counter immunoelectrophoresis (CIE); serum neutralization test (SNT); plaque-reduction neutralization test (PRNT) and indirect immunoperoxidase test which was later confirmed by amplification of CMLV-specific ankyrin repeat protein (C18L) gene. Virus isolation was successful only from samples collected from camels. Further, sequence analyses based on three full-length envelope protein genes (A27L, H3L and D8L) revealed 95.2-99.8% and 93.1-99.3% homology with other Orthopoxviruses at nucleotide and amino acid levels, respectively. Phylogram of the three genes revealed a close relationship of CMLV with Variola virus (VARV). Considering the emerging and re-emerging nature of the virus, its genetic relatedness to VARV, zoonotic potential and productivity losses in camels; the control measures are imperative in curtailing economic and public health impact of the disease. This is the first instance of laboratory confirmed camelpox zoonosis in India., (Copyright © 2011 Elsevier B.V. All rights reserved.)

An outbreak of equine influenza (EI) was reported in India in June, 2008 after a gap of two decades. The outbreak started from Jammu and Kashmir (Katra), northern state of India and spread to the other parts of the country affecting equines in 11 states. The virus (H3N8) was isolated from nasal swabs obtained from clinical cases in various locations in the country including Katra (Jammu and Kashmir), Mysore (Karnataka) and Ahmedabad (Gujarat) using embryonated chicken eggs. The virus isolates were identified as H3N8 by haemagglutination inhibition (HI) test titration with standard serum and by sequencing of full-length haemagglutinin (HA) gene and partial sequence of neuraminidase (NA) gene. Paired serum samples (n=271) showing more than fourfold rise in antibody titres tested from 11 states confirmed equine influenza. Serum samples (n=2517) of equines from 13 states of the country screened by HI test revealed 687 (26.85%) samples positive for antibodies to EI (H3N8). Phylogenetic analysis of the haemagglutinin (HA) gene confirmed the virus to be closely related to Clade 2 of the Florida sublineage in American lineage. Comparison of deduced amino acid sequence of HA gene with EIV isolates from various lineages showed substitutions in the antigenic regions C and D. HA1 gene sequence had highest amino acid identity to A/eq/Gansu/7/08 and A/eq/Hubei/6/08 isolates from China and Inner-Mongolia isolate, while the complete HA gene sequence was closest to A/eq/A/eq/Newmarket/5/03, A/eq/Bari/05 and A/eq/Kentucky/05/02 isolates. Recent outbreaks of Mongolia, China and India by clade 2 EI viruses imply their predominance in Asia in addition to Europe., ((c) 2009 Elsevier B.V. All rights reserved.)

The design, synthesis, and biological characterization of an orally active prodrug (3) of gemcitabine are described. Additionally, the identification of a novel co-crystal solid form of the compound is presented. Valproate amide 3 is orally bioavailable and releases gemcitabine into the systemic circulation after passing through the intestinal mucosa. The compound has entered clinical trials and is being evaluated as a potential new anticancer agent.