Characterization of multiple antibiotic resistance of culturable microorganisms and metagenomic analysis of total microbial diversity of marine fish sold in retail shops in Mumbai, India.

Marine fish species were analyzed for culturable and total metagenomic microbial diversity, antibiotic resistance (AR) pattern, and horizontal gene transfer in culturable microorganisms. We observed a high AR microbial load of 3 to 4 log CFU g-1. Many fish pathogens like Providencia, Staphylococcus, Klebsiella pneumoniae, Enterobacter, Vagococcus, and Aeromonas veronii were isolated. Photobacterium and Vibrio were two major fish and human pathogens which were identified in the fish metagenome. Other pathogens that were identified were Shewanella, Acinetobacter, Psychrobacter, and Flavobacterium. Most of these pathogens were resistant to multiple antibiotics such as erythromycin, kanamycin, neomycin, streptomycin, penicillin, cefotaxime, bacitracin, rifampicin, trimethoprim, ciprofloxacin, and doxycycline with a high multiple antibiotic resistance index of 0.54-0.77. The fish microflora showed high prevalence of AR genes like bla TEM, Class I integron, tetA, aph(3')-IIIa, ermB, aadA, and sul1. Nineteen of 26 AR isolates harbored Class I integrons showing high co-resistance to trimethoprim, kanamycin, doxycycline, and cefotaxime. Mobile R-plasmids from 6 of the 12 AR pathogens were transferred to recipient E. coli after conjugation. The transconjugants harbored the same R-plasmid carrying bla CTX-M, dfr1, tetA, bla TEM, and cat genes. This study confirms that fish is a potential carrier of AR pathogens which can enter the human gut via food chain. To the best of our knowledge, this is the first study in the Indian subcontinent reporting a direct evidence of spread of AR pathogens to humans from specific marine fish consumption.

Species specific PCR based detection of Escherichia coli from Indian foods.

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.

Expression of virulence and stress response genes in Aeromonas hydrophila under various stress conditions.

Aeromonas hydrophila has emerged as an important human pathogen as it causes gastroenteritis and extra-intestinal infections. Information regarding the influence of environmental stresses on gene expression profile of A. hydrophila is lacking. The impact of nutrient replenishment, nutrient deprivation, acid stress, and cold shock on housekeeping, general stress-response, and virulence genes was studied using quantitative real-time PCR in two A. hydrophila strains, CECT 839T , and A331. These sub-lethal stresses invoked significant changes in the expression of these genes in a strain-dependent manner. Overall, nutrient replenishment and deprivation significantly induced the expression of housekeeping (rpoD), general stress regulators (uspA and rpoS), and virulence (aer) genes, indicating their importance in regulating the survival and virulence of A. hydrophila under these stress conditions. rpoS gene was significantly induced under cold shock; whereas, acid stress significantly induced the expression of uspA gene. This is the first study to investigate the effect of environmental parameters on the expression of stress-response and virulence genes in A. hydrophila strains.

Cellular and spectroscopic characterization of cancer stem cell-like cells derived from A549 lung carcinoma.

BACKGROUND: Cancer stem cells (CSCs) are increasingly being realized to play a significant role in the mechanism of chemo-, radio-resistance, and metastasis of cancer. However, studies for spectral markers of CSCs using Fourier transform infrared (FT-IR) and circular dichroism (CD) spectroscopy are limited in the literature. MATERIALS AND METHODS: In the present study, CSCs obtained from single cell assay of human lung adenocarcinoma (A549) cells were characterized using CD44+/CD24-/low phenotype expression, Hoechst 33342 dye efflux assay, and expression of stemness genes. Spectral changes in cancer cells and clones enriched with CSCs were studied by FT-IR and CD spectroscopy. RESULTS: The changes in FT-IR spectra of clones enriched with CSCs showed the difference in the secondary protein structure as compared to nonstem cancer cells. Moreover, A549 clone cells showed higher C-O band of carbohydrates and deoxyribose ring vibrations of Z-form of DNA. These results were further corroborated with CD spectroscopy that showed increased alpha helix proteins and difference in DNA conformation in clones enriched with CSCs. FT-IR studies also showed higher imidazole-metal interactions in clones enriched with CSCs. These results are in agreement with higher activity of one of the metalloproteins that is, superoxide dismutase in clones enriched with CSCs and their increased radioresistance. CONCLUSIONS AND GENERAL SIGNIFICANCE: Overall, these observations provide novel FT-IR and CD spectroscopy signatures in A549 clones enriched with CSCs, which may have implications in the quantifying magnitude of CSCs as prognostic markers in cancer therapy.

Diverse Profiles of N-acyl Homoserine L-Lactones, Biofilm, Virulence Genes and Integrons in Food-Borne Aeromonas Isolates.

Aeromonas are regarded as opportunistic as well as primary pathogens of humans and fish, and are associated with gastroenteritis and septicemia in humans. Production of N-acyl-homoserine lactone (AHL) signal molecules and biofilm was determined in 22 Aeromonas isolates, from different food products in India, using thin-layer chromatography (TLC) analysis and microtiter-plate assay, respectively. Overall, highly heterogeneous patterns of AHL production were observed, with the production of N-butanoyl homoserine lactone (C4-HSL) and N-hexanoyl homoserine lactone (C6-HSL) by the majority (81.8%) of Aeromonas food isolates. Moreover, putative N-pentanoyl homoserine lactone (C5-HSL), N-heptanoyl homoserine lactone (C7-HSL), and N-octanoyl homoserine lactone (C8-HSL) were produced by 72.7%, 27.3%, and 9.1% of isolates, respectively. This is the 1st report of production of C7-HSL by Aeromonas species. Aeromonas food isolates were highly variable in their biofilm forming abilities with majority of them as weak biofilm producers in 2 different media, TSB and M9 minimal medium supplemented with 0.4% glucose. The genes encoding for putative virulence factors, glycerophospholipid cholesterol acyltransferase (gcat), heat-labile cytotonic enterotoxin (alt), heat-stable cytotonic enterotoxin (ast), serine protease (ser), polar flagella (fla), and lateral flagella (lafA) were present in 95.5%, 59.1%, 22.7%, 81.8%, 77.3%, and 22.7% of the strains, respectively. Class 1 integrons (100 to 3000 bp) were found in 68.2% of food isolates; whereas, 50% isolates contained class 2 integrons (150 to 1600 bp). This study provides a baseline data on the diversity of AHLs, biofilm forming ability and presence of virulence genes and integrons in Aeromonas food isolates from India.

Radiation-resistant Macrococcus caseolyticus (A) isolated from radiation-processed semidried prawns.

A radiation-resistant bacterial isolate from gamma-radiation-processed (5 kGy) semidried prawns was identified as a new strain of Macrococcus caseolyticus and was designated as M. caseolyticus (A) on the basis of morphological and biochemical characterization and 16S rRNA sequencing. DNA-DNA hybridization studies with M. caseolyticus DSM 20597(T) further confirmed the isolate as M. caseolyticus. Major fatty acids present in M. caseolyticus (A) were C14:0, C16:1ω11c, and C18:1ω9c, whereas C15:0anteiso, C16:0iso, and C18:0iso were absent. The closest match for the isolate, as per fatty acid methyl ester analysis, was M. caseolyticus DSM 20597(T). However, the similarity index was significantly low (0.112), which indicates that the isolate could be a new strain of M. caseolyticus. The decimal reduction dose (D10) for M. caseolyticus (A), M. caseolyticus JCSC5402, and Staphylococcus aureus MTCC96 was 1.18, 0.607, and 0.19 kGy, respectively. This is the first report on radiation resistance of M. caseolyticus. Macrococcus caseolyticus (A) is more resistant to gamma and UV radiation stress than are M. caseolyticus JCSC5402 and S. aureus MTCC96; however, it is sensitive to heat as well as desiccation stress.

Induction, resuscitation and quantitative real-time polymerase chain reaction analyses of viable but nonculturable Vibrio vulnificus in artificial sea water.

Vibrio vulnificus, an important food-borne pathogen, is known to enter viable but nonculturable (VBNC) state under low temperature and low nutrition stress conditions. Present study examined the time required for induction of VBNC state and temperature which induces resuscitation of V. vulnificus YJ016. The change in cell morphology and gene expression during VBNC state and in resuscitated cells was also examined. V. vulnificus incubated in artificial sea water at 4 °C entered VBNC state after considerably extended time (70 days). An increase in temperature by 6 °C from the VBNC induction temperature (4 °C) resulted in resuscitation of VBNC cells; however, maximum resuscitation was observed when VBNC cells were held at 23 °C for 24 h. VBNC cells changed their morphology from comma shape to coccoid shape. Two rounds of induction of VBNC and resuscitation were possible with V. vulnificus cells; however, there was progressive reduction in number of resuscitated cells and after 190 days cells failed to resuscitate. Significant up-regulation of genes related to membrane proteins [porinH (10.4-fold), ompU (2.9-fold)], regulatory proteins [envZ (5.6-fold), toxR (4.5-fold), toxS (4.8-fold)], oxidative stress related protein katG (2.3-fold), cell division/maintenance proteins [ftsZ (4.3), mreB (6.5-fold)] and resuscitating promoter factor yeaZ (fourfold) was observed during resuscitation with respect to VBNC state indicating that these genes play a role during resuscitation. Gene expression data presented here would enhance our understanding of resuscitation of V. vulnificus from VBNC state. The results also highlight the importance of maintenance of low temperature during storage of seafood.

Comparative analysis of induction of osmotic-stress-dependent genes in Vibrio vulnificus exposed to hyper- and hypo-osmotic stress.

Vibrio vulnificus, a halophilic pathogenic bacterium of marine environments, encounters changes in salinity in its natural habitat and in the food-processing environment. The comparative response of V. vulnificus to hyperosmotic and hypoosmotic stress in terms of gene expression was investigated. Genes belonging to the proU operon for transport of compatible solutes and compatible solute synthesis were significantly upregulated (3- to 4.7-fold) under hyperosmotic stress. Under hypoosmotic stress, upregulation of genes coding for mechanosensitive channels of small conductance (mscS) was not observed. In hyperosmotic conditions a 2.3-fold decrease in the expression of aqpZ was observed. A 2-fold induction in gyrA was observed in V. vulnificus cells on exposure to hyperosmotic stress. groEL genes, VVA1659 (1.6-fold), and VV3106 (1-fold) were induced in hypoosmotic condition. Results of this study indicate that to manage hyperosmotic stress, V. vulnificus accumulated osmoprotectants through uptake or through endogenous synthesis of compatible solutes. Expression of mscS may not be necessary for immediate protection in cells exposed to hyper- and hypo-osmotic stress. Comparative analysis of important osmotic-stress-related genes showed up- or down-regulation of 14 genes in hyperosmotic stress as compared with up- or down-regulation of only 7 genes in hypoosmotic stress, indicating that the cells respond asymmetrically to hyper- and hypo-osmotic stress.

Characterization of Aeromonas strains isolated from Indian foods using rpoD gene sequencing and whole cell protein analysis.

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.

Prevalence, characterization, and antimicrobial resistance of Aeromonas strains from various retail food products in Mumbai, India.

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 ß-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.

Quantification of Salmonella in food samples from India using the MINI-MSRV MPN and modified MINI-MSRV MPN methods.

Salmonella is one of the most important food-borne pathogens. The MINI-MSRV MPN method was modified by replacing the isolation and confirmation of Salmonella on selective chromogenic agar with PCR/RT-PCR. This modification reduced the time for quantification by the MINI-MSRV MPN method by 24 h. Ninety-seven different food samples, comprising chicken, mutton, fish, and sprouts from different markets in Mumbai, India, were analyzed for quantification of Salmonella species by the MINI-MSRV MPN and modified MINI-MSRV MPN methods. Seventy-four percent of the chicken samples were found positive for Salmonella. However, Salmonella was found to be absent in fish, mutton, and sprouts samples. Salmonella load in the chicken sample was in the range of 1.30 to 120 MPN/g. This genotypic confirmation has advantage over variable phenotypic confirmation of pathogens.

Evaluation of the role of enzymatic and nonenzymatic antioxidant systems in the radiation resistance of Deinococcus.

Antioxidant enzymes and antioxidant metabolites appear to have different roles in the oxidative stress resistance responses of radiation-resistant bacteria belonging to the Deinococcus-Thermus group. Twelve distinct strains belonging to 7 Deinococcus species were characterized for their responses to hydrogen peroxide, ciprofloxacin, and ionizing radiation. The levels of catalase and peroxidase activities in these strains showed a positive correlation with resistance to hydrogen peroxide and ciprofloxacin. However, the levels of these enzymes and carotenoids did not appear to contribute significantly to radiation resistance. Our findings support the idea that enzymatic defense systems are not sufficient to account for the extreme radiation resistance of Deinococcus species. Consistent with previously published reports, the Deinococcus strains had high intracellular manganese/iron ratios. No significant correlation was found between intracellular manganese/iron ratios and radiation resistance within different Deinococcus species, suggesting that other components are involved in conferring radiation resistance.

Gamma radiation used as hygienization technique for foods does not induce viable but non-culturable state (VBNC) in Salmonella enterica subsp. enterica serovar Typhimurium.

Gamma radiation has been widely used for hygienization of food products. Whether gamma radiation stress induces VBNC state in Salmonella is of great concern. Therefore, the study was carried out to determine whether gamma radiation exposure induces VBNC state in Salmonella enterica subsp. enterica serovar Typhimurium (S. typhimurium). The parameters tested were culturability on agar medium, transcriptional activity by RT-PCR, cytoplasmic membrane integrity, and direct viable count using LIVE/DEAD BacLight bacterial viability kit. The LIVE/DEAD BacLight counts for S. typhimurium cells treated with 0.5 and 1.0 kGy radiation dose were 0.8 and 0.1% of the control, respectively. Plate counts for S. typhimurium cells treated with 0.5 and 1.0 kGy radiation dose were 0.7 and 0.05% of the control, respectively. No viable cells of S. typhimurium were detected by both plate count and LIVE/DEAD BacLight after radiation treatment with 2 kGy. No transcriptional activity was detected in cells treated with 2 kGy radiation dose. If there were VBNC cells present, then significant differences in the counts between the LIVE/DEAD BacLight microscopic counts and plate agar counts must be observed. No significant difference (P > 0.05) in the counts were observed. Thus, it can be concluded that treatment with 2 kGy results in complete killing and does not induce VBNC state in S. typhimurium.

Deinococcus piscis sp. nov., a radiation-resistant bacterium isolated from a marine fish.

A radiation-resistant, Gram-stain-positive, non-motile, non-sporulating, aerobic, coccoid bacterium, strain 3ax(T), was isolated from a marine fish (black pomfret, Parastromateus niger), with radiation as a selective pressure. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain 3ax(T) exhibited highest similarity (97.9 %) with Deinococcus proteolyticus DSM 20540(T). The DeltaT(m) for DNA-DNA hybridization of D. proteolyticus DSM 20540(T) and strain 3ax(T) was 15.3 degrees C, indicating that the novel strain was distinct from D. proteolyticus DSM 20540(T). The predominant respiratory quinone was MK-8. Strain 3ax(T) could grow at 20-42 degrees C; the optimum temperature for growth was 35 degrees C. The strain grew well at pH 6-9, with optimum growth at pH 7. The cell wall contained ornithine. The major fatty acids were 16 : 0, 16 : 1omega7c, 16 : 1omega9c and 18 : 1omega9c. Three phosphoglycolipids and one aminophospholipid were the major polar lipids. Based on the genotypic, phenotypic and chemotaxonomic characteristics, strain 3ax(T) was significantly different from D. proteolyticus DSM 20540(T) and, therefore, it was identified as representing a novel species of the genus Deinococcus, for which the name Deinococcus piscis sp. nov. is proposed. The type strain is 3ax(T) (=MTCC9123(T) =DSM 19767(T)).

Radiation resistance of Deinococcus radiodurans R1 with respect to growth phase.

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.

Rapid, sensitive, and validated method for detection of Salmonella in food by an enrichment broth culture - nested PCR combination assay.

A rapid nested PCR assay for detection of Salmonella from food was developed. The sensitivity of the assay developed was comparable to the traditional culture based methods with an advantage in reduction of assay time. The assay procedure with artificially contaminated samples was able to detect as low as 4CFU Salmonella/25g of food samples (sprout, carrot, cucumber and poultry meat). With two synthetic primers of 26 mer TS11 and 25 mer TS4, a 1.2kb fragment was amplified which served as a template for amplification of final 375bp product using TS11 and TS5 primers. No non-specific amplification from the native microbial flora of food samples was observed. The reaction generates a single band specific to Salmonella which allows the analyst to interpret data at ease and without any confusion. Enriched broth serves as template for the reaction which removes labour intensive DNA isolation procedures. In case of artificially contaminated samples, 6h enriched lactose broth can serve as template. However, for market samples where the organisms are under environmental stress, it is desirable to use template from Rappaport Vasiliadis medium. The assay also employes internal amplification control, which is amplified into a 300bp fragment and thus serves as positive control for the reaction and any possibility of false negative due to inhibitory action of food components on PCR reaction can be ruled out.

Distribution of Salmonella pathogenicity island (SPI)-8 and SPI-10 among different serotypes of Salmonella.

Many virulence phenotypes of Salmonella enterica are encoded by genes located on pathogenicity islands. Based on genome analysis, it is predicted that Salmonella pathogenicity island (SPI)-8 is restricted to Salmonella serovars Typhi and Paratyphi A, and SPI-10 to Salmonella serovars Typhi, Paratyphi, Enteritidis, Dublin and Gallinarum. This study was conducted to investigate the distribution of SPI-8 and SPI-10 among Salmonella isolates from sprouts, fish, water and blood. A total of 110 Salmonella isolates and 6 Salmonella serovars from the Microbial Type Culture Collection, Chandigarh, India, were screened. All isolates belonging to Salmonella serovars Washington, Enteritidis and Paratyphi A had both SPI-8 and SPI-10. All Salmonella serovar Typhi isolates from water and blood had both SPI-8 and SPI-10, whereas isolates from fish contained only SPI-8. SPI-8 and SPI-10 were also detected in only 3 out of 42 isolates belonging to Salmonella serovar Typhimurium. Both SPI-8 and SPI-10 were absent in Salmonella serovars Worthington, Dublin, Paratyphi B and Paratyphi C. These results contradict the predictions from Salmonella genome sequences available in GenBank and indicate that SPI-8 and SPI-10 are widely distributed among Salmonella serovars and that virulence factors other than those on SPI-8 and SPI-10 may be responsible for host specificity. This is the first report on the distribution of SPIs in Salmonella isolates from India.

Effectiveness of radiation processing for elimination of Salmonella Typhimurium from minimally processed pineapple (Ananas comosus Merr.).

The microbiological quality of market samples of minimally processed (MP) pineapple was examined. The effectiveness of radiation treatment in eliminating Salmonella Typhimurium from laboratory inoculated ready-to-eat pineapple slices was also studied. Microbiological quality of minimally processed pineapple samples from Mumbai market was poor; 8.8% of the samples were positive for Salmonella. D(10) (the radiation dose required to reduce bacterial population by 90%) value for S. Typhimurium inoculated in pineapple was 0.242 kGy. Inoculated pack studies in minimally processed pineapple showed that the treatment with a 2-kGy dose of gamma radiation could eliminate 5 log CFU/g of S. Typhimurium. The pathogen was not detected from radiation-processed samples up to 12 d during storage at 4 and 10 degrees C. The processing of market samples with 1 and 2 kGy was effective in improving the microbiological quality of these products.

Radiation processing for elimination of Salmonella typhimurium from inoculated seeds used for sprout making in India and effect of irradiation on germination of seeds.

The effect of radiation processing on the germination of the sprout seeds mung (Phaseolus aureus), matki (Phaseolus aconitifolius), chana (Cicer arietinum), and vatana (Pisum sativum) in terms of percent germination, germination yield, sprout length, vitamin C content, and texture was investigated. Gradual decreases in the percent germination, germination yield, and sprout length with increases in radiation dose (0.5 to 2.0 kGy) were observed. Vitamin C content and texture remained unaffected for the seeds treated with doses of up to 2 kGy. To determine the efficacy of radiation treatment in elimination of foodborne pathogens, seeds inoculated with 4 log CFU/g of Salmonella Typhimurium were treated with radiation doses of 1 and 2 kGy. A reduction in counts of Salmonella Typhimurium in inoculated seeds after radiation treatment was observed. A radiation dose of 2 kGy resulted in the complete elimination of 4 log CFU/g of Salmonella Typhimurium from the inoculated seeds. However, on sprouting for 48 h, the count of Salmonella Typhimurium reached 8 log CFU/g for the control seeds and the seeds treated with a 1-kGy radiation dose. The aerobic plate counts for seeds were 2.0 to 2.6 log CFU/g, which were reduced to 0.9 to 1.2 log CFU/g on treatment with a 2-kGy radiation dose. On sprouting for 48 h, the aerobic plate count reached 8 log CFU/g for both the control and radiation-treated seeds. The study demonstrates that irradiation can control bacterial levels on seeds but not contamination introduced during posttreatment handling. Therefore, radiation processing of the final product (sprouts) is recommended, rather than of the seeds.

Microbiological evaluation of sprouts marketed in Mumbai, India, and its suburbs.

A study was undertaken to assess the microbiological quality of sprouts marketed in Mumbai and its suburbs. A total of 124 sprout samples of four different legumes--mung (Phaseolus aureus), matki (Phaseolus aconitifolius), chana (Cicer arietinum), and vatana (Pisum sativum)--were analyzed over a period of 12 months for aerobic plate counts, coliforms, yeast and mold counts, staphylococci, Salmonella, Listeria monocytogenes, Escherichia coli, E. coli O157:H7, and coagulase-positive Staphylococcus aureus. Aerobic plate counts ranged from 7.6 to 8.9 log CFU/g, coliform counts ranged from 5.4 to 7.9 log CFU/g, yeast and mold counts ranged from 3.6 to 7.3 log CFU/g, and staphylococci counts ranged from 3.3 to 6.6 log CFU/ g. Nonpathogenic E. coli was detected in 13% of the mung, 26% of the matki, 40% of the chana, and 19% of the vatana samples. Salmonella Typhimurium was detected in 21% of the mung, 40% of the matki, and 4% of the chana samples. Salmonella Dublin was detected in 2% of the mung samples, and Salmonella Washington was detected in 4% of the matki samples. L. monocytogenes and E. coli O157:H7 were not detected in any of the samples examined. Coagulase-positive S. aureus was detected in 4% of the mung, 11% of the matki, and 4% of the chana samples. The results indicated that the marketed sprouts were of poor microbiological quality; therefore, further processing, such as radiation processing, is needed to ensure their safety.