Review of Salmonella detection and identification methods: Aspects of rapid emergency response and food safety

Salmonella has been recognized as a major and important foodborne pathogen for humans and animals over more than a century, causing human foodborne illness as well as high medical and economical cost. Accordingly, the effort to develop efficient and reliable Salmonella detection methods continues. This paper reviews and describes the development and application of commercially available Salmonella detection methods. These are categorized into several groups based on the principle applied: conventional culture methods, immunology-based assays, nucleic acid-based assays, miniaturized biochemical assays, and biosensors. Conventional culture methods serve as the basis in food testing laboratories despite rather laborious and time-consuming protocols. Considerable progress in rapid methods using emerging technologies yield faster answers and higher throughput of samples. This paper also shows and analyzes Salmonella test results and summarizes the features and limitations of the studies involving Salmonella detection methods developed for emergency response, mainly by food emergency response laboratories participating during recent fiscal years. The emergency response laboratories utilize Salmonella detection methods possessing properties that include simplicity, versatility, high sensitivity, good specificity, and cost efficiency. Collaboration of the food emergency response laboratories in the development of these technologies is important essentially to compare for the purpose of continually improving Salmonella detection methods.     

Quantification of Salmonella spp., Listeria monocytogenes and Escherichia coli O157:H7 in non-spiked food products and evaluation of real-time PCR as a diagnostic tool in routine food analysis

Escherichia coli O157:H7, Listeria monocytogenes and Salmonella spp. are foodborne pathogens frequently associated with foods such as poultry, ready-to-eat products, fruits and vegetables. PCR-based procedures are rapid, sensitive and accurate; in particular, real-time PCR (qPCR), which besides being an automated high-throughput technique, allows quantification of foodborne pathogens. In the present work, qPCR-based methods were applied for the quantitative detection of E. coli O157:H7, Salmonella spp. and L. monocytogenes in a total of 306 non-spiked food samples in a study carried out in two laboratories simultaneously. qPCR allowed the detection of the three pathogens in around 20% of the analyzed samples for each pathogen. Quantification results revealed the presence of the three pathogens mostly at levels between 10² and 10⁴ cells/g. Besides quantification, the qPCR results (presence/absence) were compared with those of the standard mini-VIDAS system. In order to determine which were the “true” positive samples, conventional PCR was carried out after the corresponding enrichment for each pathogen. These results were considered as the gold standard for further analysis. The statistical analysis of global data recording the presence of E. coli O157:H7 and L. monocytogenes, together with data previously obtained for Salmonella spp., revealed that qPCR outperformed the mini-VIDAS procedures, in terms of both time and accuracy. Thus, these results proved qPCR to be useful as a rapid diagnostic test for the direct detection of pathogens in food, without the need for enrichment steps.     

Rapid and simultaneous detection of Salmonella,Shigella, and Staphylococcus aureus in fresh pork using a multiplex real-time PCR assay based on immunomagnetic separation

The incidence of foodborne infections caused by Salmonella sp., Shigella sp. and Staphylococcus (S.) aureus in fresh pork is increasing each year, which poses a great potential threat to public health. In this study, a rapid and simultaneous detection for these three pathogens from fresh pork samples was developed by combining immunomagnetic separation (IMS) with multiplex real-time PCR (RT-PCR). Magnetic beads coated with specific antibodies were used to capture and purify the pathogens from 250 mL matrix prepared by both spiked and commercial samples, followed by DNA extraction. Then, multiplex RT-PCR was applied with three sets of specific primers and probes. The limit of detections were evaluated in 67 spiked pork samples and were 2.0 CFU/g for Salmonella, 6.8 CFU/g for Shigella, and 9.6 CFU/g for S. aureus. The sensitivity, specificity, and accuracy of IMS-multiplex RT-PCR method were 99.2%, 100%, and 99.5%, respectively. One hundred fifty-one samples were tested using the IMS-multiplex RT-PCR and culture methods, and a comparison of the results showed that the former was a potentially reliable method for rapid and effective detection of Salmonella sp., Shigella sp., and S. aureus in fresh pork.     

Bacterial assessment of phage magnetoelastic sensors for Salmonella enterica Typhimurium detection in chicken meat

Salmonella is one of the most common pathogens associated with foodborne illness in chickens. Food outbreaks from this pathogen haven’t declined in the past 15 years according to the data from Centers for Disease Control and Prevention. It is our goal to improve food safety monitoring in this area by developing a real time Salmonella detection sensor on food surfaces. Previously, we demonstrated the use of phage C4-22 immobilized onto a rapid magnetoelastic (ME) biosensor for use as a front-line detection ligand to detect all Salmonella enterica serotypes in Tris Buffer Saline (TBS). In this study, by using fluorescent imaging, the phage peptide binding to Salmonella enterica serotype Typhimurium cells is again confirmed. Moreover, we constructed two detection models to evaluate the detection of Salmonella on/in chicken meat using the phage coated ME sensors.In the chicken surface detection method, phage C4-22 sensors demonstrated more than 12 times higher Salmonella binding capacity than the control sensors with no phage for the Salmonella spiked at the concentration of 7.86 × 10⁵ cfu/mm². In the second model, phage sensors were placed at different depths inside the chicken breast (0.1 cm; 0.5 cm; 1.0 cm below the meat surface) after surface inoculation of Salmonella. The second detection system showed that 23.27%–33% of the inoculated Salmonella cells absorbed inside the chicken breast fillets below 0.1 cm of the surface.The data for direct detection on chicken showed that phage C4-22 ME biosensors bind ultimately when there are high concentrations of Salmonella on the chicken surface. The results also suggest that the phage sensors can detect Salmonella effectively when the bacterial contaminants are absorbed into the chicken, and are not detectable by the surface detection method.     

Thermal inactivation of Salmonella Typhimurium on dressed chicken skin previously exposed to acidified sodium chlorite or carvacrol

Salmonella is a leading cause of foodborne illness and live poultry is a main reservoir of this pathogen, worldwide. Cross-contamination and transportation of contaminated poultry meat act as an important vehicle of Salmonella infections in humans. In this study, we assessed the effect of two antimicrobials; acidified sodium chlorite (ASC) and carvacrol followed by thermal treatment to inactivate Salmonella Typhimurium on dressed chicken skin. D-values (time in min for the pathogen to decrease by 90%) of Salmonella Typhimurium at 56, 60 and 64 °C on dressed chicken skin in the control samples, determined by linear regression, were 6.17, 3.16, 1.32 min, respectively. Two D-values calculated using a logistic model, ranged from 6.28 (D_1, major population, plus T_L) and 11.66 (D_2, heat-resistant subpopulation, plus T_L) min at 56 °C to 1.08 (D₁ plus T_L), and 2.07 (D₂ plus T_L) min at 64 °C. Pre-dipping in 100–300 ppm ASC or 0.02–0.06% carvacrol rendered the pathogen more sensitive to the lethal effect of heat. Thus, combination of antimicrobials with thermal inactivation was more effective in reducing heat resistance of the pathogen on dressed chicken surface. The model developed will assist poultry processors in estimating the time required for specific log reductions of Salmonella Typhimurium on chicken skin and thus, will contribute in designing acceptance limits at critical control points for chicken skins at lower times and temperatures for cooking.     

Inactivation of foodborne pathogens on gala apples by application of antimicrobial waxes

There is a need for effective postharvest controls capable of inactivating bacterial foodborne pathogens, which can be applied to the organic and conventional apple industries. This study evaluated the efficacy of natural antimicrobials (essential oils or cultured dextrose) when incorporated into carnauba wax and applied to apples. Organic Gala apples were spot inoculated with a five strains/serovars cocktail (∼9 Log CFU/mL) of Listeria monocytogenes, Shiga-toxigenic E. coli O157:H7, or Salmonella. Inoculated apples were coated with organic carnauba wax containing a 1% and 2% (v/v) mixture of essential oils (EOs) in equal proportions (oregano, clove bud, cinnamon bark, and coriander seed) or commercially cultured dextrose at 0.5% (w/v). Apples coated with carnauba wax only and uncoated apples were used as control treatments. Apples were stored at 20 °C and sampled for up to 40 days. Five treatments with three biological replicates and three samples per replicate were used for each treatment (n = 9). A sensory discrimination test was used to identify differences between waxed apples for treatments with the greatest efficacy. Carnauba wax containing 2% EOs was the most effective treatment for inactivating target foodborne pathogens inoculated on apples for 40 days. This treatment showed 4.05, 1.38 and 2.81 Log CFU/apple reduction in L. monocytogenes, E. coli O157:H7, and Salmonella, respectively, compared to uncoated apples. Carnauba wax containing 2% EOs showed significantly (P ≤ 0.001) lower populations of L. monocytogenes and Salmonella compared with 1% EOs, 0.5% cultured dextrose, or wax-only control. In contrast, there were no significant differences (P > 0.05) for E. coli O157:H7. Sensory evaluation showed that panelists could detect differences for apples treated with wax containing 2% EOs, but not for 1% EOs compared to wax-only control. This study provides information to the organic and conventional apple industry about the efficacy of antimicrobial waxes to inactivate bacterial foodborne pathogens.     

Serovar diversity and antimicrobial resistance of non-typhoidal Salmonella enterica recovered from retail chicken carcasses for sale in different regions of China

Non-typhoidal Salmonella enterica (NTS), the etiological agents in foodborne salmonellosis, is a major public health concern. This study describes the serovar diversity and antimicrobial resistance phenotypes identified in NTS isolates from retail whole chicken carcasses across six provinces of China. From food samples tested, a total of 2210 Salmonella isolates were recovered and these were serotyped by conventional and molecular serotyping methods and tested for their susceptibility to a panel of antimicrobial compounds. Sixteen serogroups and 52 serovars were identified, with serogroups B, D1 and C1 common among Enteritidis, Indiana and Infantis isolates. The serovar distribution varied both geographically and seasonally. Most (80.18%) of these isolates were found to be resistant to at least one antimicrobial compound and 54.6% were multi-drug resistant (MDR). Resistance to nalidixic acid (NAL) was common (70.6%) among the 11 tested compounds and no isolate was found to be resistant to carbapenems. There were 119 antimicrobial resistance profiles identified in the study collection. Two-hundred eighty-four isolates, including 99 Salmonella enterica serovar Indiana (S. Indiana), were resistant to seven or eight classes of antimicrobial compound. One-hundred eighty-three S. Indiana isolates were found to be co-resistant to ciprofloxacin and cefotaxime and 179 of these were confirmed as extended-spectrum β-lactamase producers. These data begin to describe the serovar diversity and antimicrobial resistance of NTS isolates recovered from retail chicken carcasses in parts of China. The findings highlight the emergence of ciprofloxacin and cefotaxime co-resistant S. Indiana, a feature displaying serious antimicrobial resistance but not commonly reported in human infections of Salmonella until recently. The food safety implications of these findings are discussed.     

Prevalence,antimicrobial resistance and genetic diversity of Salmonella isolated from retail ready-to-eat foods in China

Salmonella is a leading cause of foodborne disease worldwide and ready-to-eat (RTE) foods are regarded as a major source of infection and outbreak. In recent years, the consumption of Chinese RTE foods has raised markedly with the pace of life increasing. However, the prevalence of Salmonella in these foods in China and its potential risk to public health has not been well determined and evaluated. This study was undertaken to investigate the prevalence of Salmonella in Chinese retail RTE food products and to determine serotype distribution, antimicrobial resistance, virulence, and genetic diversity of recovered Salmonella isolates. Out of the 539 RTE food products collected and tested from July 2011 to May 2014, 19 (3.5%) were positive for Salmonella. The contamination levels of Salmonella were mostly in the range of 0.3–10 most probable number (MPN)/g, with one sample exceeding 110 MPN/g. Among the 50 isolates identified, 37 isolates (74.0%) were resistant to at least one antimicrobial, while 21 isolates (42.0%) were resistant to more than three antimicrobials. High rates of resistance were observed for tetracycline (56.0%), ampicillin (38.0%), and streptomycin (34.0%). PCR analysis of 15 virulence genes showed that the avrA, ssaQ, mgtC, siiD, sopB, and bcfC genes were detected in all 50 isolates, whereas the genes located on plasmid and prophage varied significantly among the isolates. Ten distinct serovars were identified and S. Derby, S. Meleagridis, S. Enteritidis, and S. Senftenberg were the most prevalent serovars. A total of 11 sequence types by multilocus sequence typing and 20 profiles by pulsed-field gel electrophoresis were generated for the 20 selected isolates and the combination of these two methods presented a better knowledge of genetic diversity of Salmonella isolates. The study provided a systematical surveillance on prevalence of Salmonella in Chinese RTE foods and indicates its potential risk to public health.     

Quantitative detection of viable foodborne E. coli O157:H7, Listeria monocytogenes and Salmonella in fresh-cut vegetables combining propidium monoazide and real-time PCR

The increase of foodborne outbreaks associated with fresh vegetables has highlighted the importance of developing rapid and specific methods for the detection and quantification of foodborne pathogens. In this sense, real-time PCR (qPCR) fulfills these requirements although it may detect dead cells. Recently, a potential strategy to specifically detect viable cells has been proposed relying on the use of DNA binding molecules as sample pretreatment previous to the qPCR. In this study propidium monoazide (PMA) and reagent D, combined with qPCR, were evaluated for the detection and quantification of viable Escherichia coli O157:H7, Salmonella and Listeria monocytogenes. Initially, the optimal concentration of both reagents was determined for discrimination between viable and dead bacteria in cell suspensions. Although both reagents showed similar reductions for the three pathogens, reagent D was toxic to L. monocytogenes and Salmonella and therefore only PMA was used to evaluate the applicability of this technique on food samples. A final concentration of 50 μM PMA was assayed in artificially inoculated spinach and mixed salad. PMA-qPCR signal was negative for all dead cell concentrations tested except for mixed salad inoculated with L. monocytogenes at the highest concentration. These results demonstrate that PMA-qPCR is a suitable technique for the detection and quantification of viable pathogens in fresh-cut vegetables at the levels normally found in vegetable samples.     

Persistence of foodborne pathogens and their control in primary and secondary food production chains

This review highlights factors involved in the persistence of foodborne pathogens in selected food chains and covers aspects of the basis for persistence, the consequences of persistence in terms of food safety implications, and the strategies that can be employed to combat persistence. The examples selected are Escherichia coli O157 and Salmonella at primary production of cattle and pigs, respectively, Listeria monocytogenes and Cronobacter spp. at secondary production, while persistence of Campylobacter spp. represents both primary and secondary production.     

Assessment of foodborne pathogen presence in the peach supply chain and its potential risk to the end consumer

Peaches are popular, nutritious and widely consumed. Being a tree crop, it is considered a low risk fruit, with no direct water contact, and no previous foodborne disease outbreaks associated with its consumption. However, in 2014 the pioneer association between stone fruit and a foodborne illness was reported, linking Listeria monocytogenes to stone fruit. This highlights the need for better understanding of risk associated with contaminated fresh stone fruit, in order to implement adequate preventative measures. No information is available on the presence of foodborne pathogens on peaches in the supply chain. A case study approach was therefore followed to assess foodborne pathogen presence on the farm, focusing on the impact of irrigation water, facility sanitation and hygiene by collecting various fruit and environmental samples (n = 428). This study demonstrates the effectiveness of integrating basic microbial testing with safety management and risk assessment tools that can be collectively used to improve the food safety management system. No Salmonella Typhimurium was detected from samples, however, Escherichia coli O157:H7, Listeria spp. and Staphylococcus aureus were detected on fruit and environmental samples. Despite the GlobalG.A.P. certification status of the farm, livestock frequented water sources which lead to E. coli O157:H7 contamination. This conclusion was based on positive detection of foodborne pathogens from the water sources and subsequent removal of livestock which resulted in a definite decrease in pathogen detection. A number of E. coli O157:H7 and S. aureus were detected during the second year of monitoring from environmental samples and it was observed that the personal hygiene and facility sanitation was not adequately enforced. Based on feedback given to the farmer, enforcement was improved and a definite decrease in foodborne pathogens was observed in the following sampling cycle. Areas of risk that were still identified following the fourth year of monitoring included the water source used for irrigation and poor sanitation in the production and processing facilities. Limited foodborne pathogen prevalence on peaches over the full study period as well as the extended export supply chain at controlled temperatures resulted in low-to-medium calculated consumer risk. The correct and meticulous implementation of integrated and holistic pre- and post-harvest food safety management systems is therefore essential to prevent produce contamination, reduce the consumer risk and therefore ensure overall product safety.     

Consumption patterns,bacteriological quality and risk factors for Salmonella contamination in meat-based meals consumed outside the home in Kigali,Rwanda

Meat-based meals are consumed as a source of animal proteins and constitute one of the leading vehicles for food borne infections in humans. The main objective of this study was to determine the consumption pattern and the bacteriological quality of meat-based meals consumed outside households in Kigali. A survey on meat consumption patterns was carried out in 400 households by using a questionnaire, whereas different meat-based meals were sampled from 150 snack bars and restaurants. Enumeration of hygiene indicator bacteria (total mesophilic bacteria and Escherichia coli) and the qualitative detection of Salmonella were carried out by using conventional culture methods. The results indicated that goat was the type of meat that was consumed the most outside the home in Kigali and the meat intake varied significantly (p ≤ 0.05) with the social category of the household. The average levels of total aerobic bacteria and E. coli in meat-based meals were found to be 4.7 and 1.4 log cfu/g, respectively, whereas Salmonella was detected in 11.7% of all meat-based meals. Eight factors mostly linked to the cooking treatments and hygienic handling practices for cooked meals were found to be significantly (p ≤ 0.05) associated with the risk of Salmonella occurrence in meat-based meals consumed outside the home in Kigali. The findings from this study strongly suggest the need for proper cooking and/or improvements in hygiene in the establishments selling ready-to-eat meat-based meals in Kigali, particularly those located in rural localities.     

Simulation of decontamination and transmission of Escherichia coli O157:H7, Salmonella Enteritidis,and Listeria monocytogenes during handling of raw vegetables in domestic kitchens

Epidemiological data indicates that a large number of foodborne illnesses are attributed to cross-contamination during food preparation in the domestic kitchen. The objectives of this study were to evaluate the efficiency of household washing practices in removing Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella Enteritidis on artificially contaminated lettuce and to determine the transfer rate of these three foodborne pathogens from contaminated lettuce to wash water, tomato, cabbage, and cutting boards during washing and cutting processes. Washing under the running tap water with scrubbing for 60 s was the most effective method in reducing pathogen populations by 1.86–2.60 log₁₀ CFU/g. Also, final rinsing and scrubbing practices were found to enhance the efficiency of washing treatment. In this study, the transfer rates of S. Enteritidis, E. coli O157:H7, and L. monocytogenes from cutting board to cabbage and tomato via cutting process (17.5–31.7%) were higher (P < 0.05) than from wash water to cabbage and tomato (0.8–23.0%) during washing treatment. Overall, our findings suggest that wash water and cutting board can be potential vehicles in the dissemination of foodborne pathogens. Therefore, there is a need to promote consumer awareness for proper handling practices in the kitchen to minimise the risk of foodborne infection.     

Foodborne transmission of infectious intestinal disease in England and Wales, 1992–2003

This paper reports information on foodborne outbreaks of infectious intestinal disease (IID) in England and Wales during the period 1992–2003 collected as part of a major investigation into Breakdowns in Food Safety. Between 1992 and 2003, 7620 general outbreaks of IID were reported to the Communicable Disease Surveillance Centre and in 1729 (23%) the mode of transmission was described as foodborne. In total, 39,625 people were affected and 68 deaths reported.Salmonella spp. were implicated in over half of all foodborne outbreaks. During the surveillance period, the proportion of outbreaks attributed to Salmonella spp., Clostridium perfringens and Vero cytotoxin-producing Escherichia coli O157 decreased, whereas the proportion of outbreaks attributed to Campylobacters increased.     

Effects of blackberry juice on growth inhibition of foodborne pathogens and growth promotion of Lactobacillus

Berries such as blueberry, blackberry and raspberry possess several biological activities including antimicrobial and nutritional effects. In this study, the antimicrobial activities of blackberry (Rubus fruticosus) juice against foodborne pathogens including Listeria monocytogenes, Salmonella Typhimurium and Escherichia coli O157:H7 were investigated. Inhibition of growth of these foodborne pathogens was measured in broth (Luria–Bertani broth for E. coli O157:H7 and S. Typhimurium, and brain heart infusion broth for L. monocytogenes), skim milk and whole milk supplemented with 10% blackberry juice at different time points (0, 24, 48 and 72 h). The effects of blackberry juice on the growth of Lactobacillus casei, Lactobacillus plantarum and Lactobacillus rhamnosus were also investigated in Man–Rogosa–Sharpe (MRS) broth and skim and whole milk supplemented with blackberry juice. The growth of L. monocytogenes, S. Typhimurium and E. coli O157:H7 were significantly inhibited by blackberry juice by 1–3 logs in both milk and broth. We also observed that the growths of Lactobacillus strains were significantly stimulated (1–4 logs CFU/mL) by blackberry juice in both milk and MRS broth. These data clearly demonstrate that diluted blackberry juice can be used as a preservative in food processing and a preventive in foodborne infections as a natural antimicrobial.     

The microbiological quality of take-away raw salmon finger sushi sold in Hong Kong

The safety of ready-to-eat foods is an important issue. Improper storage and handling of ready-to-eat items may lead to foodborne disease outbreaks. In this study, raw salmon finger sushi (nigiri) was selected as a target ready-to-eat food for microbiological surveillance. The aim of this study was to evaluate the microbiological quality of take-away sushi sold in the licensed sushi shops in Hong Kong. Sushi samples were collected from 120 randomly selected licensed sushi shops in the 19 districts in Hong Kong from 1^st June to 30^th July 2014. They were tested for aerobic colony count (ACC), Escherichia coli and Staphylococcus aureus counts as well as the presence of Salmonella spp. to evaluate their overall hygienic quality. None of the samples was found to contain Salmonella spp. and 1.7% of the collected samples were classified as unsatisfactory for containing more than 100 CFU/g of E. coli indicating the overall hygienic quality of take-away sushi in Hong Kong was good. There was no significant difference between samples purchased from chain stores and those from self-hosted business, suggesting that microbiological quality of take-away sushi was not affected by these two types of business models. Based on the current findings, it was suggested that the government should have more frequent routine inspections and provide food hygiene education to the workers in the sushi shops in Hong Kong so as to minimize the risk of foodborne disease outbreaks.     

Food safety knowledge,attitudes and practices of street food vendors and consumers in Port-au-Prince,Haiti

This study had the major objective of determining the food safety knowledge, attitudes and practices of vendors and consumers of street food in Port-au-Prince, Haiti. Haiti currently has no food safety legislation in place. 160 consumers and 80 vendors from four different communes (Tabarre, Delmas, Pétion-ville and downtown Port-au-Prince) volunteered to participate in the study. In general, consumers and vendors exhibited average food safety knowledge and attitude levels. Gender, training, level of education and location did not have a significant effect (p < 0.05) on the level of food safety knowledge of the consumers. Vendors were determined to have higher levels of food safety knowledge than consumers, whilst trained vendors had better food safety knowledge and attitudes compared to untrained vendors. The majority of vendors and consumers were aware of the importance of washing hands and proper cleaning with regards to the prevention of foodborne diseases. However, some other aspects were of concern. Consumers and vendors did not know that Hepatitis A, Salmonella spp. and Staphylococcus spp. are pathogens responsible of foodborne diseases. They also had difficulties in identifying the groups at risk of foodborne diseases and most were unaware of the importance of reheating food to fight against foodborne diseases. In the observational part of the study, it was found that in 60% of the cases, flies and animals were evident around the stall and 65% did not have access to potable water. The majority served food with bare hands and did not wash their hands after handling money. Additionally, 70% of the vendors did not chill pre-cooked food. The conditions in which street food vendors operate in Port-au-Prince are largely unacceptable from a food safety point of view and an effort should be made to provide them with adequate infrastructure including potable water, toilets and waste disposal facilities. The results of this study should be used to generate part of the impetus towards the development of enforcement of appropriate food safety legislation in Haiti.     

Hygienic conditions and microbiological status of chilled Ready-To-Eat products served in Southern Spanish hospitals

Chilled Ready-To-Eat (RTE) foods are of concern in hospital foodservices because they can support microbial growth when subjecting to time/temperature abuses during processing and distribution together with poor handling practices. This study was conducted in five different hospitals (A–E) of Southern Spain during 2008–2009 to perform an evaluation of their sanitary conditions and microbiological quality of two RTE meals: lettuce salads and cooked ham. A checklist based on hygiene principles embedded in Food European legislation was developed and applied in each hospital. In parallel, microbiological analysis of food contact surfaces, air quality and time/temperature measurements along the distribution chain were carried out. RTE samples (n = 150) were examined for mesophilic aerobic bacteria (MAB), total coliforms, coagulase-positive Staphylococci (CPS), Escherichia coli, Listeria spp. and Salmonella spp. Differences were found between hospitals regarding handling practices and cleanliness of working surfaces. Cooked ham samples presented lower counts of MAB and total coliforms (<10³ and <10 cfu/g respectively) than lettuce salads (10⁴ to 10⁵ and 10 to 10⁴ cfu/g respectively), although concentration of CPS was higher in cooked ham samples reaching maximum levels close to 10³ cfu/g. Neither Listeria spp. nor Salmonella spp. were detected in any food sample. Prevalence of E. coli was low (3%). Surface counts and air quality presented high variability among the different hospitals evaluated. It was concluded that good manufacturing practices and HACCP principles should be followed together with special training of food handlers. This study can help risk managers to better define the control measures to be adopted in healthcare settings in order to prevent foodborne infections.     

Validation of real-time PCR for detection of six major pathogens in seafood products

Seafood can pose a public health concern to consumers. It is often consumed raw and may be contaminated with several foodborne pathogens. In order to guarantee the safety of seafood, real-time polymerase chain reaction (PCR) protocols may be used as these enable results to be provided within 24 h.The first goal of our work was to develop real-time PCR protocols enabling the detection of six foodborne pathogens that may be present in seafood products (Campylobacter jejuni, Campylobacter coli, enterohemorrhagic Escherichia coli, Salmonella spp., Vibrio parahaemolyticus, and Vibrio vulnificus). The corresponding gene targets were: 50S/VS1, rfbE, ttr, tlh, and vvp. A multiplex PCR was also developed to detect the virulence genes of V. parahaemolyticus: tdh and trh. A total of 420 bacterial strains belonging to four different genera/strains were used in this study. Sensitivity and specificity were always 100%, except in the case of Salmonella spp., where three strains were not detected by our PCR protocols.The second objective of our work was to assess the detection limit of our real-time PCR protocols on artificially contaminated seafood products (raw shrimps, cooked shrimps, and raw mussels), purchased in public stores. Six different levels of contamination were assayed in four replicates for each matrix. The real-time PCR protocols enabled a better level of detection than the ISO methods, except for Salmonella in raw shrimps and for V. vulnificus in shrimps (raw and cooked). The estimated level of detection was between 1 and 47 cfu/25 g sample for the ISO norms and between 1 and 315 cfu/25 g sample for the real-time PCR protocols tailored in our work.The real-time PCRs developed in our work allowed for good selectivity, sensitivity, and specificity. The sensitivity on seafood products was estimated at a level of 100%, except for Salmonella (97%). In the spiking assays, the levels of detection were lower with the real-time PCR protocol than those obtained with the ISO method. This was not the case for V. vulnificus in raw and cooked shrimps and for Salmonella in raw shrimps.These real-time PCR protocols appear to be good alternative methods for surveillance of seafood products to ensure the absence of foodborne pathogens.One additional conclusion is that laboratories have to use enrichment media that are compatible with those recommended by ISO standards. This may facilitate the isolation of the pathogen if the real-time PCR protocol gives a suspect positive signal during the first step of the seafood analysis.     

Occurrence and characterization of nontyphoidal Salmonella in retail table eggs in Kandy district of Sri Lanka

Salmonellosis in humans is typically a foodborne bacterial zoonotic disease. In this regard, eggs and egg products contaminated with nontyphoidal Salmonella represent a serious threat to consumers. Eggs are one of the most common sources of animal protein for Sri Lankans but minimum attention is paid to their quality assurance. Except for a few packaged eggs sold under brand names for higher prices, in general, grading, cleaning or cooling is not practised before sale posing a microbiological health hazard to the general public. Hence, the objective of this study was to identify the presence of Salmonella in raw table eggs available at retail outlets in one selected district of Sri Lanka. Eggs were purchased from 100 retail outlets situated in highly populated areas of the district. Samples were tested for the presence of Salmonella in eggshell washings and egg contents, separately. Isolation and identification of Salmonella was performed according to standard methods. The isolates were serotyped and their antimicrobial sensitivity was determined. Salmonella was isolated from the eggs purchased from 15 retail outlets, of which 12 yielded Salmonella from shell washings and three from egg contents. The serovars identified were S. Mbandaka, S. Braenderup, S. Corvallis, and S. Emek. Except one isolate showing resistance to nalidixic acid and another to a third generation cephalosporin, other isolates were sensitive for the tested antimicrobials. According to the results of this study, retail raw table eggs can be considered as an important source of nontyphoidal Salmonella to egg consumers in Sri Lanka.