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PubMed1999

Comparison of CHROMagar Salmonella medium and hektoen enteric agar for isolation of salmonellae from stool samples.

CHROMagar Salmonella (CAS), a new chromogenic medium, was retrospectively compared to Hektoen enteric agar (HEA) with 501 Salmonella stock isolates and was then prospectively compared to HEA for the detection and presumptive identification of Salmonella spp. with 508 stool samples before and after enrichment. All stock cultures (100%), including cultures of H2S-negative isolates, yielded typical mauve colonies on CAS, while 497 (99%) isolates produced typical lactose-negative, black-centered colonies on HEA. Following overnight incubation at 37 degreesC, a total of 20 Salmonella strains were isolated from the 508 clinical samples. Sensitivities for primary plating and after enrichment were 95% (19 isolates) and 100% (20 isolates), respectively, for CAS and 80% (16 isolates) and 100% (20 isolates), respectively, for HEA. The specificity of CAS (88.9%) was significantly higher than that of HEA (78.5%; P < 0.0001). On the basis of its good sensitivity and specificity, CAS medium can be recommended for use for primary plating when human stool samples are screened for Salmonella spp.

Agar↗

PubMed2026

Deletion of the Salmonella pathogenicity island 2 gene, spiC, in attenuated Salmonella Typhimurium VNP20009 optimizes its potential for bacterial schwannoma therapy.

UNLABELLED: Recent advances in systems biology and immunotherapy have spurred the investigation of bacteria as therapeutic vehicles for cancer treatment. Currently, Bacillus Calmette-Guérin remains the only FDA-approved bacterial cancer therapy; it is a live attenuated mycobacterium that is indicated for the treatment and prophylaxis of carcinoma in situ of the urinary bladder and for the prophylaxis of primary or recurrent papillary tumors following transurethral resection. Although safety concerns have been raised, attenuated Salmonella Typhimurium strains such as VNP20009 have advanced to clinical trials targeting fast-growing human tumors. Notably, this strain induces robust immunological control of slow-growing tumors such as NF2-related schwannomatosis (NF2-SWN) in preclinical murine models. Here, we genetically characterize VNP20009 with the goal of constructing genetically defined attenuated strains that retain its promising therapeutic features while improving safety. Specifically, we investigated the contribution of the Salmonella pathogenicity island I (SPI-1) and SPI-2 type III secretion systems to antitumor efficacy and biosafety. Mutation of the SPI-1 gene sipB, a key structural component required for SPI-1 type III secretion system function, partially reduced tumor control in NF2-SWN murine schwannoma models, suggesting that bacterial invasion alone does not fully account for antitumor activity. In contrast, deletion of the SPI-2 gene spiC, a key effector required for intracellular survival, preserved robust tumor regression in NF2-SWN murine schwannoma models while improving safety and reducing systemic toxicity. To create a genetically defined and tractable platform, we generated two attenuated strains-AST101 and AST101-ΔspiC-which retain key mutations present in VNP20009 but lack ill-characterized background mutations. In the syngeneic NF2-SWN mouse schwannoma model, both strains significantly suppressed tumor growth compared to PBS. Collectively, these findings support the development of rationally engineered Salmonella Typhimurium strains with enhanced safety and preserved antitumor efficacy. IMPORTANCE: Given long-standing safety concerns surrounding the therapeutic use of live bacteria, we constructed a ΔspiC mutant of VNP20009 and demonstrated that it provides a markedly improved safety profile while retaining antitumor efficacy in NF2-related schwannomatosis mouse schwannoma models. In addition, we created two genetically defined Salmonella Typhimurium strains, AST01 and AST01-ΔspiC, which incorporate the key-targeted mutations found in VNP20009 and VNP20009-ΔspiC, respectively. These engineered strains offer a well-defined genetic background, enabling precise investigation of the bacterial traits responsible for Salmonella Typhimurium-mediated tumor control and thus further improvement of attenuated strains optimized for bacteriotherapy of neoplasms.

Salmonella typhimurium↗

PubMed2006

Epidemiology of salmonella infections in pig units and antimicrobial susceptibility profiles of the strains of Salmonella species isolated.

One hundred and thirteen finishing pig units and 74 sow units in Catalonia, Spain, were examined to determine the prevalence of salmonella infections and the factors that could be associated with them. Pooled faecal samples were taken from the finishing units, and samples of faeces were collected from individual sows. The Salmonella isolates were serotyped, phage typed and examined for their antimicrobial susceptibility to 18 common antimicrobial drugs. In addition, blood samples from pigs on 141 farms were analysed by ELISA. In both the bacteriological and serological surveys, a questionnaire with 84 questions was completed for each farm. Salmonella species were isolated from 20 per cent of the finishing units and 24 per cent of the sow units; 14 serotypes were detected in the finishing pigs and 11 in the sows. More than 30 per cent of the strains were resistant to tetracycline, sulphonamides, ampicillin or streptomycin, and 69 per cent of the strains were resistant to three or more agents up to 10 compounds. Seventy-seven per cent of the farms had at least one seropositive animal, and 26 per cent of these farms had an individual seroprevalence of 50 per cent or more. The factors associated (P<0.05) with the excretion of Salmonella species in the finishing units were the practice of raising livestock other than pigs (odds ratio [OR]=6.18), the herd size (OR=5.87), and a past history of clinical salmonellosis (OR=4.97). For the sows, the factors associated (P<0.05) with the excretion of Salmonella species were having open-flushed drainage of sewage (OR=34.48), a lack of rodent control measures (OR=0.05) and the number of sows in the unit (OR=9.26). Factors associated with seropositivity in the finishing units were a lack of bird-proof nets (OR=0.30) and the use of water from private wells (OR=3.64).

Animal Husbandry↗

PubMed2001

[Prevalence of the Salmonella plasmid virulence gene "spvD" in Salmonella strains from animals].

Strains of Salmonella isolated from animals in Germany (n = 878) were analysed for the presence of the spvD gene ("Salmonella plasmid virulence gene D") by DNA-DNA hybridization. The spvD gene was only detected in strains of serovars Typhimurium (93.3%), Enteritidis (97.1%), and Dublin (100%) as well as in two rough strains of Salmonella enterica. Salmonella isolates from mammals carried the gene more frequently (cattle 94.0%, horses 92.6%, pigs 73.7%) than those from birds (33.3%) or reptiles (4.5%). Due to its high prevalence in epidemiologically relevant salmonellae, the virulence factor spvD may represent a sensitive and specific target in various serovars for diagnostic or immunization strategies.

Animals↗

PubMed1996

[Inhibition phenomena between Salmonella strains--a new aspect of salmonella infection control in poultry].

Freshly hatched chickens show a very high susceptibility to Salmonella infections and control measures are therefore frequently focused on the period shortly after hatching. Experimental investigations using one strain against itself, differentiated by different antibiotic resistance markers, have shown that colonisation with Salmonella prevents the establishment of subsequently inoculated challenge organisms in the chicken gut. The inhibition effect lasts for several days and is detectable even when a challenge dose of 10(8) organisms is used. It is dependent of the breed of bird. Chickens colonised with Salmonella shed a subsequently inoculated challenge strain with significant lower numbers for several weeks than do non colonised control birds. The phenomenon is strain specific but not serovarspecific as has been shown in investigations using different strains of the same and other serovars for colonisation and challenge. The phenomenon shows a large variability between strains. Using other Enterobacteriaceae strains comparable inhibition against Salmonella was not observed. One important topic for further investigation is the capability of Salmonella live vaccines given orally to establish a protection effect, based on the inhibition phenomenon in the first few days of live, developing into a long-lasting immunity when birds reach immunological maturity.

Animals↗

PubMed2005

Distribution of Salmonella enterica serovars from humans, livestock and meat in Vietnam and the dominance of Salmonella Typhimurium phage type 90.

Epidemiologically unrelated non-typhoid Salmonella isolates from humans (n = 56) and animal origin (n = 241, from faeces, carcasses and meat) in Vietnam were investigated. Salmonella Typhimurium, S. Anatum, S. Weltevreden, S. Emek, and S. Rissen were the most prevalent serovars. S. Typhimurium phage type 90 was predominant among S. Typhimurium isolates. The serotype and phage type distribution of the Salmonella isolates was different from that in Europe and America. Many sero- and phage types found in humans were also found in cattle, pigs, and poultry suggesting that food producing animals are an important source of human non-typhoid Salmonella infection in Vietnam.

Animals↗

PubMed1996

Oral immunization with recombinant Salmonella typhimurium expressing surface protein antigen A (SpaA) of Streptococcus sobrinus: effects of the Salmonella virulence plasmid on the induction of protective and sustained humoral responses in rats.

Recombinant strains of Salmonella typhimurium have been studied as antigen delivery systems to determine their effectiveness as multivalent vaccines. Here we compare the efficacy of two strains of S. typhimurium. chi 4072 (pYA2905) and chi 3987 (pYA2905), expressing SpaA of Streptococcus sobrinus 6715 as oral vaccines for dental caries. Both strains are attenuated delta cya delta crp delta asd mutants with their Asd phenotypes complemented by the Asd+ plasmid pYA2905, which also encodes a peptide fragment of SpaA. S. typhimurium chi 3987 (pYA2905), unlike S. typhimurium chi 4072 (pYA2905), contains the 100 kb S. typhimurium virulence plasmid. Fischer rats were orally immunized with approximately 10(9) S. typhimurium chi 3987 (pYA2905) or chi 4072 (pYA2905) and then challenged with cariogenic S. sobrinus 6715. Rats orally immunized with either strain of recombinant Salmonella developed salivary IgA anti-SpaA responses and had lower levels of S. sobrinus-induced dental caries than nonimmunized, infected animals. In a second series of experiments, the kinetics and isotype of the serum and salivary antibody responses were determined in rats orally immunized with S. typhimurium chi 3987 (pYA2905) or chi 4072 (pYA2905) on weeks 0 and 8. IgG and IgM serum antibody responses to SpaA and S. typhimurium were detected after the primary and secondary immunizations, and the secondary immunization boosted serum IgG anti-Salmonella activity. In general, animals immunized with chi 3987 (pYA2905) had higher serum anti-SpaA, as well as serum and salivary anti-Salmonella, responses than animals immunized with chi 4072 (pYA2905). This study demonstrates the effective use of two recombinant S. typhimurium strains as oral vaccines for inducing protective and sustained immune responses against a mucosal pathogen and suggests that the recombinant Salmonella vaccine strain carrying the virulence plasmid induced similar or higher protective immune responses than the strain lacking the virulence plasmid.

Administration, Oral↗

PubMed1988

Development of a Salmonella-specific biotinylated DNA probe for rapid routine identification of Salmonella.

Classical microbiological techniques used in the detection and identification of Salmonella spp. in foods, drinking water and clinical samples are relatively lengthy. Immunoassays, on the other hand, have the major disadvantage of often generating false positives and false negatives. Recombinant DNA technology offers more efficient alternatives to the detection of a specific organism by employing cloned DNA sequences unique to the organism. Demonstration of a presence of complementary sequences among a heterogeneous population of molecules of DNA isolated from bacteria can be made by using a DNA-DNA hybridization technique. We have obtained a fragment of DNA from Salmonella typhimurium chromosomal DNA, cloned it in Escherichia coli plasmid and tested it in colony hybridization tests with 57 strains of Salmonella and other enterobacteriaceae. In all tests, the fragment was found to be Salmonella-specific in that it gave a positive reaction with all strains of Salmonella tested and was negative when tested against other Enterobacteriaceae.

Animals↗

PubMed2002

SifA, a type III secreted effector of Salmonella typhimurium, directs Salmonella-induced filament (Sif) formation along microtubules.

A unique feature of Salmonella enterica serovar typhimurium (S. typhimurium) is its ability to enter into (invade) epithelial cells and elongate the vacuole it occupies into tubular structures called Salmonella-induced filaments (Sifs). This phenotype is dependent on SifA, a Salmonella virulence factor that requires the SPI-2-encoded type III secretion system for delivery into host cells. Previous attempts to study SifA and other type III secreted proteins have been limited by a lack of suitable reagents. We examined SifA function by expressing SifA with two internal hemagglutinin epitope tags. By employing subcellular fractionation techniques, we determined that translocated SifA was membrane associated in infected HeLa cells. Confocal microscopy revealed that SifA associated with the Salmonella vacuole and with Sifs. Our analysis also revealed that microtubules serve as a scaffold for Sifs, and that SifA colocalizes with microtubules at sites of interaction between lysosomal glycoprotein-containing vesicles and Sifs. Treatment with the microtubule inhibitor nocodazole blocked Sif formation but did not prevent SifA translocation into the Salmonella vacuole. While polymerized actin has been observed on Sifs, this phenotype was transient and did not play a role in promoting or maintaining Sif formation. Our findings demonstrate the essential role of microtubule dynamics in the formation of Sifs and the utility of this epitope tagging strategy for the study of bacterial type III secreted proteins.

Actins↗

PubMed1993

UDP-sugar hydrolase isozymes in Salmonella enterica and Escherichia coli: silent alleles of ushA in related strains of group I Salmonella isolates, and of ushB in wild-type and K12 strains of E. coli, indicate recent and early silencing events, respectively.

Escherichia coli contains a single periplasmic UDP-glucose hydrolase (5'-nucleotidase) encoded by ushA. Salmonella enterica, serotype Typhimurium, also contains a single UDP-glucose hydrolase but, in contrast to E. coli, it is membrane-bound and is encoded by the non-homologous ushB gene; Salmonella enterica (Typhimurium) also contains a silent allele of the ushA gene (ushA0). In this report, we show that nearly all natural isolates of Salmonella contain both UDP-sugar hydrolases, i.e. they are UshA+ UshB+. The only exceptions are all from sub-group I (S. gallinarum, S. pullorum, and most Typhimurium strains), are UshA- UshB+, and several have been shown to contain an ushA0 allele. These data, together with the fact that these latter strains are closely related genetically, strongly suggests a recent silencing mutation(s). We also report the presence in E. coli K-12, and in natural isolates of E. coli, of a DNA sequence which is homologous to the ushB gene of Salmonella; since E. coli does not contain UshB activity, we tentatively refer to this sequence as ushB0. Since all E. coli strains investigated are UshB-, we conclude that the silencing mutation(s) occurred relatively early following the divergence of Escherichia coli and Salmonella from a common ancestor that was ushA+ ushB+.

Alleles↗

PubMed1998

Inoculum composition and Salmonella pathogenicity island 1 regulate M-cell invasion and epithelial destruction by Salmonella typhimurium.

In the mouse model of Salmonella typhimurium infection, the specialized antigen-sampling intestinal M cells are the primary route of Salmonella invasion during the early stages of infection. Under certain experimental conditions, M-cell invasion is accompanied by M-cell destruction and loss of adjacent regions of the follicle-associated epithelium (FAE), although the conditions responsible for expression of the cytotoxic phenotype in a proportion of previous studies have not been defined. In the present study, we have demonstrated that the cytotoxic effect exerted by wild-type S. typhimurium on mouse Peyer's patch FAE is dependent on the inoculum composition. We have also demonstrated that the extent of FAE destruction correlates with the extent of M-cell invasion. Bacteria inoculated in Luria-Bertani (LB) broth induce extensive FAE loss and exhibit efficient M-cell invasion, whereas bacteria inoculated in phosphate-buffered saline fail to induce significant FAE disruption and invade M cells at significantly lower levels. Similarly, inoculation in LB significantly enhances invasion of Madin-Darby canine kidney cells by wild-type S. typhimurium. Mutants defective for expression of invA, a component of Salmonella pathogenicity island 1 which is vital for efficient invasion of cultured cells, fail to induce FAE destruction and, when inoculated in LB, are attenuated for M-cell invasion. Variation in inv gene expression is, therefore, one possible mechanism by which inoculate composition may regulate the virulence of wild-type S. typhimurium. Our findings suggest that the composition of the gut luminal contents may be critical in determining the outcome of naturally acquired Salmonella infections and that both vaccine formulation and dietary status of vaccine recipients may significantly affect the efficacy and safety of live Salmonella oral vaccine delivery systems.

Animals↗

PubMed2000

Invasion genes are not required for Salmonella enterica serovar typhimurium to breach the intestinal epithelium: evidence that salmonella pathogenicity island 1 has alternative functions during infection.

Salmonella enterica serovar Typhimurium invasion genes are necessary for bacterial invasion of intestinal epithelial cells and are thought to allow salmonellae to enter and cross the intestinal epithelium during infection. Many invasion genes are encoded on Salmonella pathogenicity island 1 (SPI1), and their expression is activated by HilA, a transcription factor also encoded on SPI1. We have studied the role of Salmonella invasion genes during infection of mice following intragastric inoculation. We have found that strains containing a mutation in hilA or invG were recovered from the intestinal contents, intestinal tissues, and systemic tissues at a lower frequency than their parental wild-type strain. In contrast, a strain in which SPI1 is deleted was recovered from infected mice at a frequency similar to that of its parental wild-type strain. The DeltaSPI1 phenotype indicates that S. enterica does not require invasion genes to cross the intestinal epithelium and infect systemic tissues. This result has forced us to reconsider the long-held belief that invasion genes directly mediate bacterial infection of the intestinal mucosa and traversion of the intestinal barrier during infection. Instead, our results suggest that hilA is required for bacterial colonization of the host intestine. The seemingly contradictory phenotype of the DeltaSPI1 mutant suggests that deletion of another gene(s) encoded on SPI1 suppresses the hilA mutant defect. We propose a model for S. enterica pathogenesis in which hilA and invasion genes are required for salmonellae to overcome a host clearance response elicited by another SPI1 gene product(s).

Animals↗

PubMed1976

[Occurrence of Salmonella in healthy snakes and snake cadavers- isolation of a new Salmonella species belonging to the sub-genus IV (S. IV 18:Z36, Z38:-) (AUTHOR'S TRANSL)].

Forty-seven isolation attempts performed with 20 fecal samples of exotic snakes yielded 27 Salmonella species belonging to the sub-genus I-IV. The majority of isolates (78.7%), however, was identified as sub-genus III (Arizona). The snakes under study were hold captured for less than one year (one part) or for longer than one year. Bacteriological investigations of the 7 snakes studied, repeated 7 to 21 months thereafter, revealed uniformly other Salmonella species in comparison to those found at primary examination. Necropsy investigations were performed with materials from 3 snakes. One Salmonella species (Arizona) was isolated from the colon of one snake presenting with massive necrotic enteritis. The results with blood, bile and other colon sections were negative. Several Salmonella species were found in the colon of two other snakes presenting with infiltrative and necrotic changes in the organs, but without any pathologic finding in the intestinal tract. A new Salmonella species belonging to the sub-genus IV (S. IV 18:Z36, Z38:-) was recovered from fecal sample of a Black Mamba (Dendroaspis polylepis) and another Cascabel snake (Crotalus durissus terrificus).

Animals↗

PubMed1979

[Epidemiological studies on Salmonella in a particular area ('Walcheren Project'). IV. The incidence of Salmonella in the sewage system, in faeces of man and pets as well as in shops, kitchens and lavatories in the village of Aagtekerke (author's transl)].

As part of the epidemiological investigations on Salmonella on the island of Walcheren, the contamination of sewage water in the village of Aagtekerke was studied over a prolonged period. These studies showed that this sewage water was frequently contaminated by large numbers of Salmonella. In the present study, efforts were made to find an answer to the question of the origins of this contamination. For this purpose, the incidence of Salmonella in the sewage system, in faeces of man and pets as well as in shops, kitchens and lavatories in the village of Aagtekerke was studied for a period of three weeks in June 1977. In addition to the findings in the sewage system (including the inlet water and effluents of the sewage works) which were positive for Salmonella, this organism was only isolated from a few faecal samples of human and animal origin. Serotyping showed that the strains isolated from human faeces were similar to those found to be present in the sewage system. This fact, in conjunction with the results of the bacterial counts. suggests that the sewage system was only contaminated by the small number of carriers. The reduction of contamination observed in the sewage system during the period of investigation could be evidence of the fact that Salmonella organisms cannot survive by themselves in an environment of this type, at least not at the temperatures recorded on collecting samples from the sewage water. Growth of organisms under more favourable conditions cannot be ruled out. An inquiry on dietary habits and kitchen hygiene of the local population showed that contamination within households. Origination with the food, is a real possibility.

Adult↗

PubMed1979

[Diagnostic value of the acid production from galacturonate to differentiate strains of "Salmonella" sub-genus I and monophasic sub-genus III from the other "Salmonella" strains belonging to sub-genera II, diphasic III, IV, "Citrobacter" and "Hafnia alvei" (author's transl)].

Two methods were compared: (1) the ability to use sodium glacturonate as source of carbon and energy in defined medium; (2) the ability to produce acid by fermentation of this substrate in peptone water with phenol red. Results of growth on, acid formation from, galacturonate are identical. Salmonella strains of sub-genus I and monophasic serotypes of sub-genus III do not use galacturonate. Salmonella of sub-genera II and IV as well as diphasic serotypes of sub-genus III use galacturonate. Glacturonate-negative Salmonella are isolated mainly from man and warm-blooded animals whereas galacturonate-positive Salmonella are most often isolated from cold blooded animals and from the enrivonment. Citrobacter and Hafnia alvei which may be confused with Salmonella spp. are galacturonate-positive.

Citrobacter↗

PubMed1980

Epidemiological studies on Salmonella in a certain area ("Walcheren Project"). IV. The incidence of Salmonella in the sewage system, in the faeces of man and pets as well as in shops, kitchens and lavatories in the village of Aagtekerke.

As part of the epidemiological investigations on Salmonella ion the former island of Walcheren, the contamination of sewage water in the village of Aagtekerke was studied over a prolonged period of time. These studies showed that this sewage water was frequently contaminated by large numbers of Salmonella. In the present study efforts were made to find the source of this contamination and for this purpose the incidence of Salmonella in the sewage system, in the faeces of man and pets as well as in shops, kitchens and lavatories in the village of Aagtekerke was studied for a period of three weeks in June 1977. Salmonella was only isolated from a few samples of faecal material of human and animal origin, and from the sewage system (including the inlet water and effluents of the sewage works). Serotyping showed that the strains isolated from human faeces were similar to those found to be present in the sewage system. This fact taken in conjunction with the results of the bacterial counts, suggests that the sewage system was only contaminated by a small number of carriers. The reduction of contamination observed in the sewage system during the period of investigation could be evidence that Salmonella organisms cannot survive by themselves in an environment of this type, at least not at the temperatures recorded when collecting samples from the sewage water. Growth of organisms under more favourable conditions cannot be ruled out. A questionnaire about the dietary habits and kitchen hygiene of the local population showed that contamination within households, originating with the food, is a real possibility.

Adult↗

PubMed1981

Comparative efficiency of brilliant green, bismuth sulfite, Salmonella-Shigella, hektoen enteric, and xylose lysine desoxycholate agars for the recovery of Salmonella from foods: collaborative study.

The relative efficiency of brilliant green (BG), bismuth sulfite (BS), Salmonella-Shigella (SS), xylose lysine desoxycholate (XLD), and Hektoen enteric (HE) agars for the recovery of Salmonella from 5 foods was collaboratively studied in 11 laboratories. The analytical efficiency of various paired combinations of the 5 agars was statistically compared according to 3 parameters: (1) productivity or recovery of Salmonella, (2) rate of enumeration of cultures that were false positive for Salmonella, and (3) rate of enumeration of false-negative reactions. In descending order of productivity, the sequential ranking was BS, XLD, HE, BG, and SS agars. In ascending order, the rates of false-positive reactions based on a statistical analysis of paired agar combinations was HE, BS, BG and XLD (tie), and SS agars. Analogously, in ascending order, the sequence of false-negative reaction rates was BS, XLD, HE, BG, and SS agars. The combination of BS, XLD, and HE agars is more efficient for recovery of Salmonella from foods than is the present official combination of BG, BS, and SS agars. The revision of official final action method 46.054 to replace the combination of BG, BS, and SS agars with a combination of BS, XLD, and HE agars has been adopted official first action.

Agar↗

PubMed1996

Collaborative ring-trial of Dynabeads anti-Salmonella for immunomagnetic separation of stressed Salmonella cells from herbs and spices.

Eight laboratories participated in a Salmonella detection ring-trial which compared selective enrichment by conventional broths with immunomagnetic separation (IMS) using Dynabeads Anti-Salmonella. Laboratories analyzed six types of herbs and spices that were spiked with one of six freeze-dried Salmonella species. Each herb and spice analysis comprised of 12 samples (25 g each) which had been spiked at three different levels, plus a negative control and stored for one week prior to testing. Out of a total 468 samples analyzed, 195 (41.7%) were positive by both methods. Eighteen samples were positive only by IMS enrichment, in comparison with 19 positive samples by conventional enrichment broths and not IMS. These results confirm the potential use of IMS as an alternative to enrichment broths for Salmonella isolation.

Culture Media↗