Aims:  To study the occurrence and diversity of Salmonella serovars in urban water supply systems of Nepal. Methods and Results:  Occurrence of Salmonella was detected in 42 out of 300 water samples by enrichment culture technique in selenite F broth followed by plating on Salmonella Shigella agar. A total of 54 isolates identified to genus level by standard tests were subsequently confirmed by serotyping, phage typing and PCR detection of virulence genes (inv A and spv C). The predominant serotype was Salmonella Typhimurium, followed by Salm. Typhi, Salm. Paratyphi A and Salmonella Enteritidis. Most of the Salm. Typhi isolates were E1 phage type followed by UVS4, A and UVS1. All isolates of Salm. Paratyphi A and Salm. Enteritidis were an untypable (UT) phage type. The majority of isolates were multi‐drug resistant as revealed by Kirby–Bauer disc diffusion technique. Ceftriaxone resistant isolates of Salm. Enteritidis indicated the presence of one of the ESBL genes, blaSHV, whereas the genes blaTEM and blaCTX were absent. Conclusions:  The microbiological quality of the urban water supply is poor and indicates possibility of fatal outbreaks of enteric fever and related infections in Nepal. Significance and Impact of the Study:  The present study will be useful in water borne disease control and prevention strategy formulation in Nepal and in the global context.

Isolation of five ionizing radiation (IR)-resistant bacteria by screening of isolates from various habitats classified as common and stressed is reported. IR-resistant isolates exhibited varying degrees of resistance to gamma-radiation and were classified as highly and moderately radiation resistant. Resistance to ultraviolet (UV) radiation correlated well with gamma-radiation resistance, whereas a comparable desiccation resistance for all the highly and moderately radiation-resistant isolates was observed. However, salt tolerance failed to correlate with IR resistance, indicating a divergent evolution of the salt tolerance and radiation resistance. Characterization of isolates by the amplified rDNA restriction analysis profiling attested to the clustering of these isolates with their stress phenotype. 16S rRNA gene-based analysis of the isolates showed that the bacteria with similar-resistance physiologies clustered together and belonged to related genera. Hydrogen peroxide resistance and mitomycin survival patterns of the isolates indicated the roles of oxidative-stress tolerance in desiccation survival and recombination repair in higher radiation resistance, respectively.

A total of 154 food samples (chicken, fish, and ready-to-eat sprouts) from various retail outlets in Mumbai, India, were analyzed for the presence of Aeromonas spp. over a period of 2 y (January 2006 to March 2008). Twenty-two Aeromonas isolates belonging to 7 different species were isolated from 18 (11.7%) food samples. The highest percentages of isolation were from chicken (28.6%) followed by fish (20%) and sprout (2.5%) samples. Aeromonas caviae, A. veronii bv. sobria, and A. salmonicida were the most frequently isolated species from sprouts, chicken, and fish samples, respectively. The genes encoding for putative virulence factors, cytotoxic enterotoxin (act), hemolysin (hly), aerolysin (aer), elastase (ahyB), and lipase (lip) were detected using polymerase chain reaction method in 59.1%, 40.9%, 22.7%, 54.5%, and 31.8% of the strains, respectively. The isolated Aeromonas strains were found to be positive for virulence factors, that is, amylase, DNase, gelatinase, protease, and lipase production. More than 60% isolates were also positive for β-hemolytic activity. All these food isolates were found to be resistant to ampicillin and bacitracin, and sensitive to gentamicin, 3rd-generation cephalosporins (ceftazidime, cephotaxime, ceftriaxone), and chloramphenicol. Seventeen (77.2%) isolates harbored single and/or multiple plasmids (approximately 5 to >16 kb). The XbaI digestion patterns of chromosomal DNA of these isolates, using pulsed field gel electrophoresis, showed high genetic diversity among these isolates. Our results demonstrate the presence of various Aeromonas spp. with virulence potential and antimicrobial resistance in different food products marketed in Mumbai, India. The potential health risks posed by consumption of these raw or undercooked food products should not be underestimated.

Aeromonas are responsible for causing gastroenteritis and extra-intestinal infections in humans. Twenty-two Aeromonas strains isolated from different food sources were re-identified up to species level using rpoD gene sequence analysis. Biochemical tests and 16S rRNA gene sequencing were insufficient to identify Aeromonas till species level. However, incorporation of additional biochemical tests lead to correct identification of 95.5 % strains up to species level. The 16S rRNA gene sequencing was useful to identify Aeromonas isolates at the genus level only. Sequences of the rpoD gene showed greater discriminatory power than 16S rRNA gene and provided conclusive discrimination of the strains for which the phenotypic species identification was uncertain. All these 22 strains were accurately identified up to species level by rpoD gene as A. salmonicida (6), A. veronii bv. veronii (4), A. caviae (3), A. hydrophila (2), A. veronii bv. sobria (2), A. jandaei (1), A. trota (1), A. sobria (1), A. allosaccharophila (1) and A. bivalvium (1). All these strains were also characterized using whole cell protein (WCP) analysis by gradient SDS-PAGE and showed different whole cell protein (WCP) profile [22-28 polypeptide bands (~10 to >97 kDa)], indicating high genetic diversity. The present work emphasizes the use of molecular methods such as rpoD gene sequencing along with comprehensive biochemical tests for the rapid and accurate identification of Aeromonas isolates till species level. The WCP profile can be subsequently used to characterize Aeromonas isolates below species level.

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Deinococcus species exhibit an extraordinary ability to withstand ionizing radiation (IR). Most of the studies on radiation resistance have been carried out with exponential phase cells. The studies on radiation resistance of Deinococcus radiodurans R1 with respect to different phases of growth showed that late stationary phase cells of D. radiodurans R1 were fourfold more sensitive to IR and heat as compared with exponential or early stationary phase cells. The increased sensitivity of D. radiodurans R1 to IR in the late stationary phase was not due to a decrease in the intracellular Mn/Fe ratio or an increase in the level of oxidative protein damage. The resistance to IR was restored when late stationary phase cells were incubated for 15 min in fresh medium before irradiation, indicating that replenishment of exhausted nutrients restored the metabolic capability of the cells to repair DNA damage. These observations suggest that stress tolerance mechanisms in D. radiodurans R1 differ from established paradigms.

Escherichia coli is a faecal indicator and certain virotypes are known as pathogens. Therefore, detection and prevention of E. coli in food is very important. The existing rapid methods concentrate on detecting the pathogenic E. coli instead of total E. coli population. Present study evaluates the use of two molecular markers (uidA and flanking region of uspA) specific for the E. coli in combination with microbiological method for confirmation. Majority of the isolates (77%) were positive for both the genes tested. However, 22% of the E. coli isolates were positive for any one of the two primer sets [uidA (9%) and flanking region of uspA (13%)]. High levels of E. coli incidences (92% samples) were observed in beef while low occurrence (19% samples) was found in sprouts. Low percentage (7.3%) of E. coli isolates was positive for virulence genes tested (lt, ipaH, aggR, eaeA, stx1 and stx2). Two isolates were positive for stx genes. However, none of the isolates including stx positive isolates were E. coli 0157:H7. Maximum number of the E. coli (44%) isolates was characterized under phylogenetic group B2. This phylogenetic group comprises of extra intestinal and virulent E. coli strains.