Pulsed-field gel electrophoresis fingerprinting of Listeria monocytogenes isolates recovered from foods of animal origin and fishes in North-Eastern India.

L isteria monocytogenes is a pathogen of great concern to the food industry. The present study was aimed to explore the clonal relationships amongst L. monocytogenes strains isolated from foods of animal origin (milk, beef, chevon (goat meat), pork and chicken) and fish. Forty-seven L. monocytogenes strains were characterized by pulsed-field gel electrophoresis (PFGE). The PFGE analysis using ApaI and AscI enzymes revealed 37 pulsotypes, with Simpson's discriminatory index of 0.987. This study demonstrated the presence of a few similar L. monocytogenes pulsotypes in different foods of animal origin in different places and years of isolation and this indicates that some L. monocytogenes subtypes may be ubiquitous which are acclimatizing and persisting in different foods of animal origin. This also emphasizes the importance of cross-contamination in local wet markets. Thus, the understanding of genetic diversity will contribute to the development of rational and workable strategies to control this important zoonotic infection.

Development of a novel and rapid polymerase spiral reaction (PSR) assay to detect Salmonella in pork and pork products.

The polymerase spiral reaction (PSR), a novel isothermal method for targeted DNA amplification, was effectively applied to detect Salmonella in artificially spiked pork. The specificity of the developed PSR was tested using 16 Salmonella and 15 non-Salmonella strains. The PSR assay was 10-fold more sensitive than conventional end-point PCR, having a sensitivity comparable to real-time PCR. The limit of detection of the developed assay was 4 × 103 per gram of pork without enrichment and 4 CFU per gram after a 6 h enrichment. The detection of 4 CFU per gram of pork was achieved within 8 h. The PSR assay was successful, and accurate in comparison to microbiological methods, in detecting Salmonella in 11 of 76 commercial pork samples. Therefore the positive predictive value, negative predictive value and accuracy rate of the developed assay were 100%. Considering its rapidity, user-friendliness, simplicity, cost-effectiveness and equipment-free nature, this PSR assay is a promising tool for the food industry for the detection of Salmonella and prevention of Salmonella outbreaks and recalls.

Biology, Predatory Potential, Life Table, and Field Evaluation of Propylea dissecta (Coleoptera: Coccinellidae), Against Lipaphis erysimi (Hemiptera: Aphididae) on Broccoli.

Propylea dissecta (Mulsant) (Coleoptera: Coccinellidae) is one of the most promising ladybird beetle against many sucking pests. Predation rates, developmental biology, life table, and field assessment of this ladybird were examined against mustard aphid, Lipaphis erysimi (Kalt.) (Hemiptera: Aphididae), on broccoli. Data on the life history were collected at 23 ± 1°C and 70 ± 1% RH and were evaluated using the two-sex, age-stage life table. Results showed that the two-sex, age-stage life table-based net reproductive rate (R0) was 11.264 ± 6.197 offspring. The adult females lived longer (33.8 ± 2.356 d) than the adult males (32.2 ± 0.841 d). The fourth instar consumed most of L. erysimi (113.97 ± 5.76) compared to the other larval stages of the predator. Male (1,821) and female (2,673) consumed more aphids than larvae. The net consumption rate was 741.78 ± 89.91 aphids. Other aphidophagous predators such as Coccinella septempunctata L., Micraspis discolor (F.), Coccinella transversalis (F.), and syrphid (Diptera: Syrphidae) were also noted in broccoli. Our research showed that inoculative release of 150 or 200 adults per 1,000 m2 for two times on broccoli achieved a significant decrease in aphids L. erysimi and Brevicoryne brassicae L. (Hemiptera: Aphididae) (>95%). The release rate of 150 adults per 1,000 m2 for two times may, therefore, be recommended to manage the aphid population on broccoli.

Characterization of Marek's disease virus and phylogenetic analyses of meq gene from an outbreak in poultry in Meghalaya of Northeast India.

The aim of the present study was to characterize the virus from the lesions and histopathology of organs associated with mortality in Kuroiler (dual purpose variety of poultry developed and marketed by Keggfarms Pvt. Ltd, India) birds suspected of Marek's disease. Among 1047 birds from two farms of different location with 5.5 and 34% mortality, two types of lesion were observed in post mortem examination; tumors in vital organs-liver, spleen, kidney, lung and ovaries and generalized small nodular tumour in the abdominal cavity. Molecular characterization based on detection of ICP4 gene showed the presence of Marek's disease virus (MDV) from tissues and cell culture adapted isolates in Madin Darby Canine Kidney cell lines. Histopathological examination revealed multinucleated immature lymphoid cells infiltration in the organs. Phylogenetic analysis of the isolates based on meq gene showed the isolates belongs to cluster I genotype of MDV. This is for the first time the MDV virus is characterized from an outbreak in the poultry flock in farmer's field affecting production in Meghalaya state of North east India.

Porcine circovirus 2 in the North Eastern region of India: Disease prevalence and genetic variation among the isolates from areas of intensive pig rearing.

Porcine Circovirus type-2 (PCV-2) is considered as a major threat to the piggery sector in India. To ascertain the epidemiological status and infection level of PCV2, a pilot study was undertaken to find out the prevalence of PCV2 in swine population by ELISA and PCR in the interior and border areas of Meghalaya which includes the area where accessibility and medical aid is a rare phenomenon. A total of 249 serum samples were collected from October 2014 to February 2016 from three divisions of Meghalaya: Khasi, Jaintia and Garo Hills Divisions. The mean positivity of PCV-2 antibodies in suspected sera was 83.93% whereas 62.25% of the suspected samples respectively were found to contain PCV2 as detected by PCR. Additional 190 tissue samples were collected during necropsy from both symptomatic and asymptomatic animals following reported outbreak in this region, which indicated a mean positivity of 18.94% (36/190); out of which 13 samples were subjected to sequencing to find out the genetic diversity of PCV2 amongst the field isolates. Molecular characterization and phylogenetic analysis of PCV2 isolates based on cap gene depicted genetic diversity among the strains in pig population of Meghalaya as the isolates belonged to PCV2a, PCV2b-1c and PCV2d genotypes; identification of the PCV2d genotype is probably the first report from Meghalaya. Four isolates forming an outlier group in the phylogenetic tree were arising out of natural inter-genotypic recombination between PCV2a and PCV2b. PCV2 being immunosuppressive in nature impairs the host immune response increasing the susceptibility to other co-infections leading to disease severity and high mortality in pig population. This baseline data gives a brief epidemiological status of PCV2 infection and circulating PCV2 genotype in this region which will be useful in the formulation of control and eradication programs in remotes areas of Meghalaya where accessibility is less and vaccination is a rare practice.

Recovery of Mycobacterium lentiflavum from bronchial lavage during follow-up of an extrapulmonary tuberculosis patient.

Initially diagnosed with cervical lymphadenitis, a 15-year-old boy was started with category I anti-tuberculosis (TB) drugs. Follow-up investigations led to isolation and identification of Mycobacterium lentiflavum by multiple diagnostic and identification approaches. Observation of this rare pathogen from human origin urges cautious diagnosis while attending TB cases.

Pan-genome analysis of Aeromonas hydrophila, Aeromonas veronii and Aeromonas caviae indicates phylogenomic diversity and greater pathogenic potential for Aeromonas hydrophila.

Aeromonas species are important pathogens of fishes and aquatic animals capable of infecting humans and other animals via food. Due to the paucity of pan-genomic studies on aeromonads, the present study was undertaken to analyse the pan-genome of three clinically important Aeromonas species (A. hydrophila, A. veronii, A. caviae). Results of pan-genome analysis revealed an open pan-genome for all three species with pan-genome sizes of 9181, 7214 and 6884 genes for A. hydrophila, A. veronii and A. caviae, respectively. Core-genome: pan-genome ratio (RCP) indicated greater genomic diversity for A. hydrophila and interestingly RCP emerged as an effective indicator to gauge genomic diversity which could possibly be extended to other organisms too. Phylogenomic network analysis highlighted the influence of homologous recombination and lateral gene transfer in the evolution of Aeromonas spp. Prediction of virulence factors indicated no significant difference among the three species though analysis of pathogenic potential and acquired antimicrobial resistance genes revealed greater hazards from A. hydrophila. In conclusion, the present study highlighted the usefulness of whole genome analyses to infer evolutionary cues for Aeromonas species which indicated considerable phylogenomic diversity for A. hydrophila and hitherto unknown genomic evidence for pathogenic potential of A. hydrophila compared to A. veronii and A. caviae.

Complete genome sequence of emerging porcine circovirus types 2a and 2b from India.

We report here the first characterized complete genome sequence of porcine circovirus types 2a and 2b from northeastern states of India. These isolates may serve as a potential reference for the Indian strains of porcine circovirus types 2a and 2b.

Complete genome sequence of classical Swine Fever virus subgenogroup 2.1 from assam, India.

We report the complete genome sequence of a classical swine fever virus (genogroup 2.1), isolated from an outbreak in Assam, India. This particular isolate showed a high degree of genetic variation within the subgenogroup 2.1 and may serve as a potential reference strain of the 2.1 genogroup of classical swine fever virus (CSFV) in the Indian subcontinent.

Detection of Pasteurella multocida isolates from local pigs of India by polymerase chain reaction and their antibiogram.

Pasteurella multocida has been recognized as an important veterinary pathogen for over a century. Conventional methods for diagnosis of pasteurellosis rely on the detection of the organism by microscopy and its isolation and identification. However, as far as pasteurellosis is concerned, it is not just sufficient to know the identity of the organism. To constitute effective control measures, it is important to know the serotype of the organism. A study was undertaken to characterize the Pasteurella isolates from local pigs in India with clinical respiratory disease by determination of their capsule types and presence or absence of toxin gene. Pasteurella could be isolated from 66.70% of pigs with clinical respiratory disease. All the isolates were confirmed through biochemical characterization and P. multocida-specific polymerase chain reaction. It has also been observed that all the isolates belonged to capsular type D. All the isolates were sensitive to chloramphenicol, chlortetracycline, doxycycline, and enrofloxacin, while the rest of the antibiotics were less effective. It has also been observed that all isolates were resistant to cephalexin, penicillin G, and sulphadiazine. The study revealed the detection of P. multocida serotype D from clinical respiratory diseases of local pigs of India, which could be one of the important respiratory tract pathogens responsible for mortality of local pigs in India.