Antimicrobial susceptibilities and epidemiological analysis of Salmonella enteritidis isolates in Korea by phage typing and pulsed-field gel electrophoresis

A total of 81 isolates of Salmonella Enteritidis were analyzed by antibiotic susceptibility, phage typing, and pulsed-field gel electrophoresis (PFGE). Thirty-two isolates came from broiler carcasses and pig feces, and 49 isolates were from humans in Seoul and suburbs of Seoul, Korea. Antibiotic resistance was most prevalent among human isolates. Of human isolates, 89.8% were resistant to more than two antibiotics, while 64.7% of poultry isolates and 13.3% of pig isolates showed multiple resistance to more than two antibiotics. The most common phage type (PT) was PT1, followed by PT30 or 33, PT21 and PT20a. The isolates showed six PFGE patterns with XbaI or SpeI digestion, and five PFGE patterns with NotI digestion. But a single pattern, PFGE X1, S1, or N1, was predominant and the rest of the PFGE patterns differed by only one or two bands. Results indicated the spread of a genetically related clone of Salmonella Enteritidis in foods and humans in Korea and that phage typing as well as PFGE may offer an improved level of discrimination for the epidemiological investigation of Salmonella Enteritidis.     

Typing of Lactobacillus sanfranciscensis isolates from traditional sourdoughs by combining conventional and multiplex RAPD-PCR profiles

In the present work, a rapid and reproducible molecular method, based on the combination of conventional and multiplex RAPD-PCR reactions, was developed for typing Lactobacillus sanfranciscensis isolates from traditional sourdoughs. At first, four random primers, two used singly and two combined with the primer RD1, were chosen on the basis of their differentiating capability and reproducibility. The four resulting profiles for each isolate were integrated into a unique profile to be statistically treated by cluster analysis. The method was validated on 58 L. sanfranciscensis isolates coming from three traditional Italian sourdoughs. This new RAPD method was useful for determining the genomic diversity within the L. sanfranciscensis species. In particular, the intraspecific diversity of this species seemed to be related to the sourdough origin.     

Comparison of antibiotic susceptibility profiles and molecular typing patterns of clinical and environmental Salmonella enterica serotype Newport

The genus Salmonella is composed of more than 2,400 serotypes, many of which cause enteric diseases in humans and animals. Several Salmonella serotypes are multidrug resistant, and there is evidence of the clonal spread of these strains from animals to humans. Salmonella enterica serotype Newport is one of the serotypes that increasingly present a multidrug-resistant phenotype. Source tracking and antibiotic resistance testing are important considerations for identifying the outbreak strain. The first goal of this study was to examine the antibiotic susceptibility patterns of clinical and environmental Salmonella Newport isolates from various geographic locations and to compare the discriminatory ability of two DNA fingerprinting techniques. The second goal was to determine whether the antibiotic resistance profiles and typing patterns correlated. Thirty Salmonella Newport isolates, including environmental and human clinical strains, were subjected to pulsed-field gel electrophoresis (PFGE), ribotyping, and antibiotic susceptibility testing. Eighty percent of the isolates showed total or intermediate resistance to one or more drugs; 75% of the isolates were multidrug resistant. Ribotyping with the EcoRI enzyme and PFGE with the XbaI enzyme each divided the isolates into 14 groups. Cluster analysis based on antibiotic susceptibility patterns generated 23 profiles. The susceptible and resistant isolates were not differentiated on the basis of either of the molecular typing techniques. Hence, no correlation was observed between the antibiotic resistance profiles and the DNA subtyping patterns. In conclusion, ribotyping is as discriminatory as PFGE and, when used in combination with antibiotic resistance profiles, provides a powerful tool for the source tracking of Salmonella Newport.     

Diversity of Salmonella isolates using serotyping and multilocus sequence typing

One hundred and twenty-one Salmonella isolates were obtained from food, feed, and live chicken samples derived from 13 countries or regions. In this study, their subtypes were evaluated by serotyping and multilocus sequence typing (MLST), and their genetic profiles were also characterized. It was demonstrated by serotyping on these isolates that 36 various serovars were obtained in this study, of which three serotypes S. Babelsberg, S. Fresno, and S. II were first found in mainland China. Based on Simpson’s index of diversity, the serotyping method had a 0.943 discriminatory power. Meanwhile, there were a total of 42 unique sequence types (STs) characterized by MLST, and the discriminatory power of MLST (D = 0.947) was close to that of the serotyping method. In MLST, hisD revealed the highest levels of nucleotide diversity. In addition, ST-92 was the most common ST represented by 16 Salmonella isolates, followed by ST-367 which was represented by 14 isolates. Seven new alleles were identified, which were associated with other alleles and resulted in the assignment of nine new STs. It was concluded from the results that MLST was generally associated with serotype, but not associated with the epidemiological source of the samples, and antimicrobial resistance patterns.     

Effects of plasmid curing on antibiotic susceptibility, phage type, lipopoly saccharide and outer membrane protein profiles in local Salmonella isolates

Five different Salmonella isolates (four Salmonella enteritidis and one Salmonella havana) of human, chicken or egg origin which all expressed smooth lipopolysaccharide were subjected to plasmid curing. Upon this treatment, four isolates lost at least one of their resident plasmids while all five isolates became rough in their LPS profiles. Two of the S. enteritidis isolates, CA13 and CE7, were found to convert phage types (PTs) as well. The phage type conversion in CA13 from PT6 to PT21 was found to be accompanied by the loss of a 19·0-MDa plasmid as well as the loss of sulbactam ampicillin and tetracycline resistance. The change in the PTs of the strain CE7 from PT20a to the PT20 variant was accompanied by the loss of two plasmids of 10·9 and 1·45 MDa. Outer membrane protein (OMP) type change occurred only in the isolate CA13, which lost an OMP of 27·2 kDa. The strains, CA13 and CE7, retained their high molecular weight plasmid (36 MDa), which has been very commonly found (77 of 82) among local S. enteritidis isolates.     

Rapid discrimination of Salmonella isolates by single-strand conformation polymorphism analysis

A molecular typing technique was developed for the differentiation of Salmonella isolates based on single-strand conformation polymorphism (SSCP) analysis of amplicons generated by PCR. Amplicons from parts of the fimA (both the 5' and 3' ends), mdh, invA, and atpD genes were generated separately from a panel of Salmonella strains representing Salmonella bongori, and four subspecies and 17 serovars of Salmonella enterica. These amplicons were subjected to SSCP analysis for differentiation of the salmonellae on the basis of different conformational forms arising due to nucleotide sequence variations in the target genes. Several distinct SSCP banding patterns (a maximum of 14 each for atpD and fimA 3' end) were observed with this panel of Salmonella strains for amplicons generated from each target gene. The best discrimination of Salmonella subspecies and serovar was achieved from the SSCP analysis of a combination of at least three gene targets: atpD, invA, and either mdh or fimA 3' end. This demonstrates the applicability of SSCP analysis as an important additional method to classical typing approaches for the differentiation of foodborne Salmonella isolates. SSCP is simple to perform and should be readily transferable to food microbiology laboratories with basic PCR capability.     

Survival of Salmonella enteritidis phage type 30 on inoculated almonds stored at -20, 4, 23, and 35 degrees C

To evaluate the survival of Salmonella on raw almond surfaces, whole almond kernels were inoculated with Salmonella Enteritidis phage type (PT) 30 collected from a 24-h broth culture or by scraping cells from an agar lawn. Kernels inoculated with lawn-collected cells to 8, 5, 3, and 1 log CFU per almond after a 24-h drying period were stored for 161 days at 23 +/- 3 degrees C. Calculated rates of reduction were similar for the four inoculum levels (0.22, 0.28, 0.29, and 0.22 log CFU/month, respectively). Kernels inoculated to 7.1 or 8.0 log CFU per almond after drying were stored for 171 or 550 days, respectively, at selected temperatures, including -20 +/- 2 degrees C, 4 +/- 2 degrees C, 23 +/- 3 degrees C, and 35 +/- 2 degrees C. No significant reductions of Salmonella were observed during storage at -20 and 4 degrees C over 550 days. At 35 degrees C, a biphasic survival curve was observed, with calculated reductions of 1.1 log CFU/month from days 0 to 59 and no significant reduction from days 59 to 171. At 23 degrees C, reductions of 0.18 and 0.30 log CFU/month were calculated for 171 and 550 days of storage, respectively. When combined with data from the study of inoculum levels, an overall average calculated reduction at 23 degrees C was 0.25 +/- 0.05 log CFU/month. Significantly greater reductions were observed during the 24-h drying period when broth-collected cells were used as the inoculum, suggesting that cells collected from agar lawns were more resistant to drying. However, after initial drying, the rates of reduction at 23 degrees C did not differ significantly between the inoculum preparation methods. Salmonella Enteritidis PT 30 survives for long periods on almond kernels under a variety of commonstorage conditions.     

Inactivation by ionizing radiation of Salmonella enteritidis,Salmonella infantis,and Vibrio parahaemolyticus in oysters (Crassostrea brasiliana)

Irradiation is considered one of the most efficient technological processes for the reduction of microorganisms in food. It can be used to improve the safety of food products, and to extend their shelf lives. Oysters are considered one of the most important vehicles for pathogenic bacteria because of their feeding characteristics. The aim of this study was to evaluate the influence of a gamma radiation process on high levels of Salmonella Enteritidis, Salmonella Infantis, and Vibrio parahaemolyticus incorporated by oysters (Crassostrea brasiliana), as well as the effects of the process on the survival of the oysters and on their sensory attributes. The oysters were exposed to gamma radiation (60Co) in doses ranging from 0.5 to 3.0 kGy. A dose of 3.0 kGy was generally sufficient to reduce the level of Salmonella serotypes by 5 to 6 log10 units. A dose of 1.0 kGy was sufficient to produce a 6-log10 reduction in the level of V. parahaemolyticus. The highest irradiation dose did not kill the oysters or affect their sensory attributes. Hence, a dose of 3.0 kGy can be considered effective in inactivating Salmonella and V. parahaemolyticus in oysters without changing their odor, flavor, or appearance.     

Characterization of Lactobacillus isolates from fermented olives and their bacteriocin gene profiles

Near one hundred isolates of Lactobacillus paraplantarum, Lactobacillus pentosus and Lactobacillus plantarum from table olives were studied. Strains were genotyped by rep-PCR. Although the technique failed to differentiate some isolates at the species level, it proved a robust and easy procedure that could be useful for distinguishing between related strains of L. paraplantarum, L. pentosus and L. plantarum from a large pool of unrelated strains of these species. A PCR-based screening revealed the presence of the plantaricin encoding genes plnA, plnB, plnC, plnD, plnE/F, plnF, plnI, plnJ, plnK, plnG and plnN in most isolates of the three species. Sequences of bacteriocin genes present in L. paraplantarum and L. pentosus were homologous to L. plantarum genes. Through a discriminating analysis of the bacteriocin gene profiles, it was possible to establish a relationship between the origin of isolation and the LAB isolates, regardless of species.     

Prevalence of Salmonella isolates and antimicrobial resistance patterns in chicken meat throughout Japan

We investigated the prevalence of Salmonella in chicken meat from northern, central, and southern Japan. Between 2006 and 2008, 821 samples from these three regions were collected and examined. Salmonella isolates were detected in 164 (20.0%) of these samples, with 15 (10.0%) of 150, 113 (27.5%) of 411, and 36 (13.8%) of 260 recovered from the northern, central, and southern regions, respectively. We recovered 452 Salmonella isolates. From the isolates, 27 serovars were identified; the predominant serovars isolated were Salmonella Infantis (n=81), Salmonella Kalamu (n=56), and Salmonella Schwarzengrund (n=43). Of the 452 isolates, 443 (98.0%) were resistant to one or more antibiotics, and 221 (48.9%) showed multiple-antibiotic resistance, thereby implying that multiple-antibiotic resistant Salmonella organisms are widespread in chicken meat in Japan. Resistance to oxytetracycline was most common (72.6%), followed by dihydrostreptomycin (69.2%) and bicozamycin (49.1%). This study, the first to report Salmonella prevalence in chicken meat throughout Japan, could provide valuable data for monitoring and controlling Salmonella infection in the future.     

Antimicrobial resistance and phage and molecular typing of Salmonella strains isolated from food for human consumption in Spain

The aims of this study were to ascertain the population structure and antimicrobial susceptibility of Salmonella enterica serovars isolated in 2002 from food in 16 Spanish regions. Serovars were characterized by serotyping, phage typing, antimicrobial susceptibility, and pulsed-field gel electrophoresis (PFGE) typing, and 264 nonrelated strains were selected for further analysis. The main sources were eggs and their derivatives (21.6% of strains), poultry and related products (16.6%), and seafood (16.3%). High serotype diversity was detected (51 serotypes); the most common were Enteritidis (n = 96, 36.3%) and Typhimurium (n = 53, 20.1%), followed by a miscellaneous group of 49 different serotypes (n = 115, 43.5%). A 15% increase in Salmonella Enteritidis isolation was observed. Common phage types for Salmonella Enteritidis were PT1 (41.6% of isolates), PT4 (9.4%), PT6 (9.4%), and PT6a (9.4%), and common types for Salmonella Typhimurium were DTU302 (18.8%), DT104 (15.1%), and DT104B (13.2%). Salmonella Enteritidis strains were categorized into eight PFGE types with a similarity of 81 to 96%, and 73.9% of the strains were grouped into just one cluster. Salmonella Typhimurium isolates were divided into 13 PFGE types with a similarity of 64 to 86%, and one predominant clone contained 41.5% of the strains. Resistance rates for Salmonella Enteritidis, Salmonella Typhimurium, and the miscellaneous group were, respectively, 8.3, 69.8, and 13.9% for ampicillin, 3.1, 52.8, and 59% for streptomycin, 40.6, 22.6, and 10.4% for nalidixic acid, 15.6, 71.7, and 31.1% for tetracycline, 7.3, 18.8, and 9.5% for trimethoprim-sulfamethoxazole, 0, 50.9, and 4.3% for chloramphenicol, and 6.2, 71.7, and 17.4% for multiple (at least four) antimicrobials. All the strains remained susceptible to other beta-lactams and fluoroquinolones. Surveillance of S. enterica isolated from food is strongly recommended to reduce community exposure to antimicrobial resistant strains.     

Variation in Salmonella resistance to poultry chemical decontaminants, based on serotype, phage type, and antibiotic resistance patterns

Chemical decontaminants are currently under review for final approval by the European Union authorities with the aim of reducing the number and/or prevalence of pathogenic microorganisms on poultry. The purpose of the research being reported here was to determine the association, if any, of decontaminant resistance with the serotype, phage type, and antibiotic resistance of Salmonella strains. Sixty poultry isolates of Salmonella enterica (serotypes Enteritidis: phage types 1, 4, 4b, 6a, 14b, and 35; Typhimurium; Newport; Infantis; Poona; Virchow; Agona; Derby; and Paratyphi B) showing resistance to none (sensitive), one (resistant), two, three, four, five, six, seven, or nine (multiresistant) antibiotics were screened for resistance to 1,000 ppm acidified sodium chlorite, 1.2% trisodium phosphate, or 25% citric acid. D-values (seconds required for 1-log reduction in the number of bacteria) in peptone water, using a linear regression, of Salmonella in the presence of acidified sodium chlorite varied widely with serotype (the highest resistance levels were shown by serotypes Typhimurium, Newport, and Derby) and antibiotic resistance pattern (average values of 8.37 +/- 1.69 s for multiresistant strains as compared with 5.96 +/- 0.54 s for sensitive, P < 0.05). A positive relationship (0.775, P < 0.001) was found between acidified sodium chlorite D-values and the number of antibiotics to which strains were resistant. Both serotype and antibiotic resistance had only a slight influence over Salmonella resistance to trisodium phosphate, with average D-values from 12.44 +/- 0.91 s (sensitive strains) to 13.28 +/- 0.77 s (multiresistant) (P < 0.05). Neither serotype nor antibiotic profile was associated with Salmonella resistance to citric acid (average D-value of 12.20 +/- 0.81 s). Minimal differences in resistance to decontaminants were found among Salmonella Enteritidis phage types. Results in the present study highlight the importance of selecting an adequate strain (serotype and antibiotic resistance pattern) when acidified sodium chlorite and trisodium phosphate are tested against Salmonella to ensure that concentrations capable of inactivating all strains are used.     

Evaluation of avian-specific probiotic and Salmonella enteritidis-, Salmonella typhimurium-, and Salmonella heidelberg-specific antibodies on cecal colonization and organ invasion of Salmonella enteritidis in broilers

Salmonella Enteritidis colonizes the intestinal tract of poultry and causes foodborne illness in humans. Reduction of Salmonella Enteritidis colonization in the intestinal tract of poultry reduces potential carcass contamination during slaughter. The purpose of this study was to determine the effect of an avian-specific probiotic combined with Salmonella Enteritidis-, Salmonella Typhimurium-, and Salmonella Heidelberg-specific antibodies on the cecal colonization and organ invasion of Salmonella Enteritidis in broiler as well as on body weights. The treatment group was defined as chicks spray-vaccinated with Avian Pac Plus at the hatchery and given Avian Pac Plus for the first 3 days after placement. An intermediate treatment was given at 10 and 14 days, 2 days prior to vaccination and 2 days postvaccination. All birds were vaccinated with Newcastle disease vaccine, La Sota virus (one drop/eye) at 12 days of age. A final treatment was given 3 days preslaughter. The control group was defined as chicks not given Avian Pac Plus at any time. Six hours after oral administration of the probiotic suspension (treatment group) or water (control group) at placement, the chicks were challenged with Salmonella Enteritidis. All chickens were orally inoculated with 0.25 ml of Salmonella Enteritidis that contained 4 x 10(7) CFU/1.0 ml. Cecal colonization and organ invasion were evaluated for Salmonella Enteritidis on days 0, 1, 3, 7, 10, 17, 24, 31, 38, and 41. The probiotic-treated group had a significantly lower concentration of Salmonella Enteritidis cecal colonization at days 3, 7, 10, 17, 24, 31, 38, and 41 when compared to the nontreated, control group (P < 0.05). Similarly, there was a significant difference (P < 0.05) in the isolation of Salmonella Enteritidis from the internal organs (liver and spleen) when probiotic-treated and nonprobiotic-treated groups were compared. There was no significant difference (P > 0.05) in the mean body weight between the two experimental groups at each collection period. These results indicated that a combination of Lactobacillus acidophilus, Streptococcus faecium, and Salmonella Enteritidis, Salmonella Typhimurium, and Salmonella Heidelberg-Specific antibodies have a beneficial effect in reducing the colonization of Salmonella Enteritidis in market-aged broilers.     

Effect of phage on survival of Salmonella enteritidis during manufacture and storage of cheddar cheese made from raw and pasteurized milk.

The ability of Salmonella Enteritidis to survive in the presence of phage, SJ2, during manufacture, ripening, and storage of Cheddar cheese produced from raw and pasteurized milk was investigated. Raw milk and pasteurized milk were inoculated to contain 10(4) CFU/ml of a luminescent strain of Salmonella Enteritidis (lux) and 10(8) PFU/ml SJ2 phage. The milks were processed into Cheddar cheese following standard procedures. Cheese samples were examined for Salmonella Enteritidis (lux), lactic acid bacteria, molds and yeasts, coliforms, and total counts, while moisture, fat, salt, and pH values were also measured. Salmonella Enteritidis (lux) was enumerated in duplicate samples by surface plating on MacConkey novobiocin agar. Bioluminescent colonies of Salmonella Enteritidis were identified in the NightOwl molecular imager. Samples were taken over a period of 99 days. Counts of Salmonella Enteritidis (lux) decreased by 1 to 2 log cycles in raw and pasteurized milk cheeses made from milk containing phage. In cheeses made from milks to which phage was not added, there was an increase in Salmonella counts of about 1 log cycle. Lower counts of Salmonella Enteritidis (lux) were observed after 24 h in pasteurized milk cheese containing phage compared to Salmonella counts in raw milk cheese with phage. Salmonella Enteritidis (lux) survived in raw milk and pasteurized milk cheese without phage, reaching a final concentration of 10(3) CFU/g after 99 days of storage at 8 degrees C. Salmonella did not survive in pasteurized milk cheese after 89 days in the presence of phage. However, Salmonella counts of approximately 50 CFU/g were observed in raw milk cheese containing phage even after 99 days of storage. In conclusion, this study demonstrates that the addition of phage may be a useful adjunct to reduce the ability of Salmonella to survive in Cheddar cheese made from both raw and pasteurized milk.     

Characterization and gene expression profiles of thermotolerant Saccharomyces cerevisiae isolates from Thai fruits

For industrial applications, fermentation of ethanol at high temperature offers advantages such as reduction in cooling costs, reduced risk of microbial contamination and higher efficiency of fermentation processes including saccharification and continuous ethanol stripping. Three thermotolerant Saccharomyces cerevisiae isolates (C3723, C3751 and C3867) from Thai fruits were capable of growing and producing 38 g/L ethanol up to 41°C. Based on genetic analyses, these isolates were prototrophic and homothallic, with dominant homothallic and thermotolerant phenotypes. After short-term (30 min) and long-term (12 h) exposure at 37°C, expression levels increased for the heat stress-response genes HSP26, SSA4, HSP82, and HSP104 encoding the heat shock proteins small HSP, HSP70, HSP90 and the HSP100 family, respectively. In isolates C3723 and C3867, expression was significantly higher than that in reference isolates W303 and TISTR5606 for TPS1 encoding trehalose-6-phosphate synthase, NTH1 encoding neutral trehalase and GSY1 encoding glycogen synthase. The results suggested that continuous high expression of heat stress-response genes was important for the long-term, heat stress tolerance of these thermotolerant isolates.     

Eggshell factors influencing eggshell penetration and whole egg contamination by different bacteria, including Salmonella enteritidis

Trans-shell infection routes and whole egg contamination of 7 selected bacterial strains; Staphylococcus warneri, Acinetobacter baumannii, Alcaligenes sp., Serratia marcescens, Carnobacterium sp., Pseudomonas sp. and Salmonella enteritidis, recovered from egg contents, were studied. The first objective was to correlate bacterial eggshell penetration with various eggshell characteristics and bacterial strains. An agar approach was used to assess the eggshell penetration. The second objective was to assess the contamination of whole eggs with the bacterial strains; whole intact eggs were used in this case. The intact shells of agar-filled and whole eggs were inoculated with 10(3) -10(4) cfu of the selected strains. During 3 weeks storage at 20 degrees C and 60% relative humidity, the bacterial eggshell penetration was regularly monitored. The whole egg contamination was only analyzed after 3 weeks. The eggshell characteristics such as area eggshell, shell thickness and number of pores did not influence the bacterial eggshell penetration. For each individual bacterial strain the mean cuticle deposition was lower for penetrated compared to non-penetrated eggshells. For the individual strain Carnobacterium sp. and for the global results of all strains this difference was statistical significantly. The whole egg contamination was not influenced by neither the area of the eggshell nor the porosity of the eggshell. The results of the agar approach indicate that the Gram-negative, motile and non-clustering bacteria penetrated the eggshell most frequently; Pseudomonas sp. (60%) and Alcaligenes sp. (58%) were primary invaders followed by S. enteritidis (43%). All selected strains were able to penetrate; penetration was observed most frequently after ca. 4-5 days. Particularly S. enteritidis was a primary invader of whole eggs: the membranes and/or the content of 32% of the whole eggs was contaminated. The remaining bacterial eggshell contamination with the selected strain was determined after 3 weeks storage. Penetrated eggshells and contaminated whole eggs showed a significantly higher bacterial contamination on the eggshell compared to non-penetrated eggshells and non-contaminated whole eggs respectively (global results of all strains). The influence of hen age on bacterial eggshell penetration and egg content contamination was not significant. While the agar approach is suitable to study the influence of the eggshell characteristics on the bacterial eggshell penetration, the intact egg approach gives an estimation of the penetration of the shell followed by the probability of survival and migration in whole eggs.     

Pulsed field,ribotype and integron analysis of multidrug resistant isolates of Salmonella enterica serovar Newport

Twenty-one isolates of Salmonella enterica serovar Newport were evaluated for their antimicrobial resistance, pulsed field gel electrophoresis (PFGE) profiles, ribotype profiles, and their integron profiles. Antimicrobial resistance profiles indicated that 20 of the 21 isolates were resistant to the following antibiotics: amoxicillin–clavulanic acid (AMOX/CA), ampicillin (AMPC), cefoxitin (CFOX), ceftiofur (TIO), cephalothin (CRIN), chloramphenicol (CHL), streptomycin (STR), tetracycline (TET), and sulfamethoxazole (SMX). Five isolates showed resistance to gentamycin (GEN) and kanamycin (KAN). Trimethoprim–sulfamethoxazole (SMX/TMP) resistance was observed in six isolates. Eight of the twenty one isolates showed intermediate resistance to ceftriaxone (CTRX), with one isolate exhibiting complete resistance. PFGE clearly resolved the Salmonella Newport isolates into nine distinct clusters, and a good congruence was observed between PFGE and antibiotic resistance patterns. Automated riboprinting clearly distinguished between antibiotic resistant and sensitive strains of Salmonella Newport, and resolved the isolates into two ribogroups. One group consisted of the multidrug resistant isolates, and the other grouping contained the sensitive isolate. Three different integrons (1.0, 1.2, and 1.8 kb) were observed in many of the isolates, and several isolates contained more than one integron. Restriction fragment length polymorphisms (RFLP) indicated that integrons of the same size were indistinguishable.When integron analysis and ribotype analysis were used in conjunction, four subtypes of multidrug resistant Salmonella Newport isolates were clearly defined. These results demonstrate the possibility of utilizing automated ribotyping and integron analysis to rapidly subtype multidrug resistant Salmonella Newport isolates.     

Growth of Salmonella enteritidis and Salmonella typhimurium in the presence of quorum sensing signalling compounds produced by spoilage and pathogenic bacteria

The effect of acylated homoserine lactones (AHLs) and autoinducer-2 (AI-2) signalling compounds present in the cell-free culture supernatants (CFS), of Pseudomonas aeruginosa, Yersinia enterocolitica-like GTE 112, Serratia proteamaculans 00612, Y. enterocolitica CITY650 and Y. enterocolitica CITY844, on the growth of two Salmonella Enteritidis and two S. Typhimurium strains was assessed though monitoring of changes in conductance of the medium. Detection times (T_det), area and slope of conductance curves were recorded. Except for P. aeruginosa 108928, which was not found to produce AI-2, all other strains produced both AHLs and AI-2. Thereafter, aliquots (20% in the final volume) of these CFS were transferred into NZ Amine broth inoculated with ca. 10³ CFU/ml of stationary phase cultures of each Salmonella strain. While the CFS of P. aeruginosa induced a shorter detection time, i.e. acceleration of the metabolic activity, the CFS of the other microorganisms increased the detection time of Salmonella strains compared to control samples (i.e. without CFS). Results indicate that the growth of Salmonella may be affected by the presence of Quorum sensing (QS) signalling compounds and/or other novel signals existing in CFS, produced by other bacterial species and confirm the complexity of bacterial communication.     

Inactivation of Salmonella enteritidis and Salmonella senftenberg in liquid whole egg using generally recognized as safe additives, ionizing radiation, and heat

The effect of combining irradiation and heat (i.e., irradiation followed by heat [IR-H]) on Salmonella Enteritidis and Salmonella Senftenberg inoculated into liquid whole egg (LWE) with added nisin, EDTA, sorbic acid, carvacrol, or combinations of these GRAS (generally recognized as safe) additives was investigated. Synergistic reductions of Salmonella populations were observed when LWE samples containing GRAS additives were treated by gamma radiation (0.3 and 1.0 kGy), heat (57 and 60 degrees C), or IR-H. The presence of additives reduced the initial radiation Dgamma -values (radiation doses required to eliminate 90% of the viable cells) by 1.2- to 1.5-fold, the thermal decimal reduction times (D,-values) by up to 3.5- and 1.8-fold at 57 and 60 degrees C, respectively, and the thermal D,-values after irradiation treatments by up to 3.4- and 1.5-fold at 57 and 60 degrees C, respectively, for both Salmonella serovars. Of all the additives investigated, nisin at a concentration of 100 IU/ml was the most effective at reducing the heat treatment times needed to obtain a 5-log reduction of Salmonella. Thus, while treatments of 21.6 min at 57 degrees C or of 5 min at 60 degrees C should be applied to achieve a 5-log reduction for Salmonella in LWE, only 5.5 min at 57 degrees C or 2.3 min at 60 degrees C after a 0.3-kGy radiation pretreatment was required when nisin at a concentration of 100 IU/ml was used. The synergistic reduction of Salmonella viability by IR-H treatments in the presence of GRAS additives could enable LWE producers to reduce the temperature or processing time of thermal treatments (current standards are 60'C for 3.5 min in the United States) or to increase the level of Salmonella inactivation.     

Comparison of antimicrobial resistance patterns and phage types of Salmonella Typhimurium isolated from pigs,pork and humans in Belgium between 2001 and 2006

Infections with non-typhoid Salmonella represent a major problem in industrialized countries.The emergence and spread of antimicrobial-resistant pathogens, among them Salmonella, has become a serious health hazard worldwide. One of the most commonly isolated non-typhoid Salmonella serovars in pigs, pork and humans is Salmonella Typhimurium. In this study the comparison of the incidences of resistance to nine antimicrobials, resistance patterns and phage types between S. Typhimurium isolated from pigs (n = 581), pork (n = 255) and humans (n = 1870) in Belgium in the period 2001 to 2006 was performed.Resistance to the antimicrobials ampicillin, chloramphenicol, streptomycin, sulfonamides and tetracycline was frequently observed and varied between 23.5% and 83.1%. Resistance ranged from 15.6% to 20.7% for the combination trimethoprim–sulfonamides and from 3.4% to 5.8% for nalidixic acid. Resistance to the critical important antimicrobials cephalosporins and fluoroquinolones was found sporadically (≤ 1.2%). Resistance to the different antimicrobials was observed to be similar in S. Typhimurium isolates from the various origins. Twenty-seven antimicrobial resistance patterns representing in total 75.2%, 89.0% and 89.6% of the isolates from pigs, pork and humans respectively were found to be common among the three groups and 73 combinations antimicrobial resistance pattern/phage type were found to be common among pork and human isolates, representing 70.1% of the pork isolates and 51.0% of the human isolates. The high percentage of isolates that have a common resistance pattern, and in a less pronounced way a common combination phage type/resistance pattern, are in agreement with the hypothesis of transfer of antimicrobial resistant Salmonella from pigs via the consumption of pork to humans as one of the possible pathways. The most prevalent combination in Belgium within both the pork isolates (7.4%) and the human isolates (13.2%) was S. Typhimurium DT104 resistant to ampicillin, chloramphenicol, streptomycine, sulfonamides and tetracycline.