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Detection of Salmonella from raw food samples using Dynabeads anti-Salmonella and a conventional reference method.

A Dynal core method has been established using Dynabeads anti-Salmonella to detect Salmonella from all categories of food samples. The protocol consists of the standard pre-enrichment of samples in buffered peptone water followed by immunomagnetic separation and subsequent selective enrichment of the bead-bacteria complexes in Rappaport-Vassiliadis Soya Peptone broth before plating onto Salmonella selective media. This modified IMS cultural method is intended to replace or augment traditional cultural methods used for Salmonella detection due to its specificity and increased sensitivity. The optional direct plating of bead-bacteria complexes onto solid media using a swab-streak technique is suitable for processed foods or samples known to have a history of very low resident flora. In an evaluation using 100 naturally contaminated samples, this IMS core method detected 39 of the 44 positive samples detected by all the methods combined, compared to 31 detected by the conventional ISO 6579 reference method. Furthermore, in ten different food matrices inoculated with low levels (1-5 cells/25 g) of twenty Salmonella serovariants, frozen for one month before being examined, the IMS core method, showed a 90% concordance with the ISO method and isolated two more Salmonella positive samples than the conventional ISO method.

Food Microbiology↗

Enzyme-linked immunosorbent assay for Salmonella typhimurium in food: feasibility of 1-day Salmonella detection.

A microtitration plate, antibody-capture, enzyme-linked immunosorbent assay was developed for detection of Salmonella typhimurium. The assay utilizes a monoclonal detector antibody which shows no cross-reactions with non-Salmonella species and only a slight cross-reaction with one other Salmonella serotype. By using only one cultural stage (in a nonselective, chemically defined medium) prior to the enzyme-linked immunosorbent assay, low numbers of cells in food (10 cells 25 g-1) were detected in 19 h. Non-Salmonella competing organisms did not interfere with detection of S. typhimurium even when present in the ratio of 10(6):1 (non-Salmonella/Salmonella spp.). The assay shows the feasibility of rapid, 1-day testing for Salmonella spp. with antibody technology.

Antibodies, Monoclonal↗

Effects of the evisceration and cooling processes on the incidence of Salmonella in fresh dressed turkeys grown under Salmonella-controlled and uncontrolled environments.

In July and December 1981, turkeys grown in a Salmonella-free environment were sampled at the end of the slaughter and evisceration processes to determine the effect of slaughter and evisceration on the incidence of Salmonella in raw, eviscerated turkeys. A total of 109 eviscerated turkeys from the Salmonella-free flocks and 79 control turkeys were sampled. A total of 75 environmental swab samples and 5 chiller water samples were collected. Less than 1% of the eviscerated Salmonella-free turkeys were Salmonella positive and 6.3% of the eviscerated control turkeys were Salmonella positive. The results of the swab samples indicated generally good processing equipment sanitation.

Abattoirs↗

Detection of Salmonella enteritidis in feces from poultry using booster polymerase chain reaction and oligonucleotide primers specific for all members of the genus Salmonella.

Salmonella enteritidis was identified in feces from hens using the polymerase chain reaction (PCR) and oligonucleotide primers specific for all members of the genus Salmonella. Feces from specific-pathogen-free Leghorn hens were determined to be negative for Salmonella by microbiological culture and by the PCR. Fecal samples were inoculated with known numbers of colony-forming units of S. enteritidis. The DNA was extracted from fecal samples and amplified by the PCR using genus-specific primers. Salmonella were detected in all samples known to be positive; the sensitivity of the assay extended to 1 cfu of S. enteritidis/g feces. Feces that were not inoculated with Salmonella were negative. Microbiological culture was less sensitive than the PCR assay; results of culture of feces with less than 10(2) cfu/g were negative. Although S. enteritidis was used in this study, the oligonucleotide primers used in this study have been previously demonstrated to be genus-specific for Salmonella.

Animals↗

Effect of calf age and Salmonella bacterin type on ability to produce immunoglobulins directed against Salmonella whole cells or lipopolysaccharide.

A commercially available Salmonella bacterin was administered to Holstein calves starting at 1 to 19 weeks of age. Serum samples were obtained before administering bacterin and at 2-week intervals thereafter. An ELISA with Salmonella dublin lipopolysaccharide (LPS) or S dublin whole cells as antigen, was used to measure specific IgG and IgM responses. Antibody responses to LPS were not detected from calves < 12 weeks old inoculated with killed bacterin. Immunoglobulin responses to whole-cell antigen were detected from all age groups of calves inoculated with the same killed Salmonella bacterin. Calves < 11 weeks old are able to produce immunoglobulins to some whole-cell antigens, but are unable to produce anti-LPS immunoglobulins when inoculated with killed Salmonella bacterin. This age-related response to killed Salmonella antigens may account, in part, for increased susceptibility to salmonellosis in calves < 12 weeks old. In comparison to the response for killed antigen, 8 calves given modified-live aromatic-dependent S dublin bacterin at 1 to 3 weeks of age had detectable anti-LPS immunoglobulins after immunization, although the response was not as rapid and was of a lesser magnitude than that of older calves given killed Salmonella bacterin.

Aging↗

Diagnostic potential of sefA DNA probes to Salmonella enteritidis and certain other O-serogroup D1 Salmonella serovars.

Salmonella enteritidis thin fimbriae, SEF14, were found to be restricted to S. dublin and the predominantly poultry-associated members of the Salmonella O-serogroup D1, S. enteritidis, S. berta, S. gallinarum and S. pullorum, when tested by Western and ELISA analysis from among 90 Salmonella isolates of 42 serovars, as well as from members of several related genera of the Enterobacteriaceae. These five serovars and a single isolate of S. typhi (D1) were also detected by hybridization of genomic DNA from 732 Salmonella isolates of 117 serogroups to gene probes derived from the S. enteritidis sefA (fimbrin gene), sefB (chaperone) or sefC (outer membrane protein) genes encoding proteins involved in SEF14 biosynthesis. None of 250 Enterobacteriaceae or 27 other eubacterial isolates tested hybridized to the sef probes. The sefA, sefB and sefC genes were amplified from these six Salmonella serovars by PCR using primer pairs designed from sefA, sefB or sefC of S. enteritidis. DNA sequencing of sefA genes from these five serovars indicated limited sequence variability among sefA genes and recognition of individual base pairs which could potentially differentiate certain strains of S. enteritidis, S. dublin and S. gallinarum.

Animals↗

Identification of Salmonella typhimurium genes responsible for interference with peptide presentation on MHC class I molecules: Deltayej Salmonella mutants induce superior CD8+ T-cell responses.

Salmonella-derived epitopes are presented on MHC molecules by antigen-presenting cells, and both CD4+ and CD8+ T cells participate in protective immunity to Salmonella. Therefore, mechanisms that allow Salmonella to escape specific immune recognition are likely to have evolved in this bacterial pathogen. To identify Salmonella genes, which potentially interfere with the MHC class I (MHC-I) presentation pathway, Tn10d transposon mutagenesis was performed. More than 3000 mutants, statistically covering half of the Salmonella genome, were individually screened for altered peptide presentation by infected macrophages. Two mutants undergoing enhanced antigen presentation by macrophages were identified, carrying a Tn10d insertion in the yej operon. This phenotype was validated by specific inactivation and complementation experiments. In accordance with their enhanced MHC-I presentation phenotype, we showed that (i) specific CD8+ T cells were elicited at a higher level in mice, in response to immunization with yej mutants compared to their parental strain in two different experimental settings; and (ii) yej mutants were superior vaccine carriers for heterologous antigens compared to the parental strain in a tumour model.

Animals↗

Genomic subtraction identifies Salmonella typhimurium prophages, F-related plasmid sequences, and a novel fimbrial operon, stf, which are absent in Salmonella typhi.

Salmonella typhimurium causes systemic and fatal infection in inbred mice, while the related serotype Salmonella typhi is avirulent for mammals other than humans. In order to identify genes from the virulent strain S. typhimurium ATCC 14028 that are absent in S. typhi Ty2, and therefore might be involved in S. typhimurium mouse virulence, a PCR-supported genomic subtractive hybridization procedure was employed. We have identified a novel putative fimbrial operon, stfACDEFG, located at centisome 5 of the S. typhimurium chromosome, which is absent in S. typhi, Salmonella arizonae, and Salmonella bongori but was detected in several other Salmonella serotypes. The fimbrial genes represent a genomic insertion in S. typhimurium compared to the respective region between fhuB and hemL in Escherichia coli K-12. In addition, the subtraction procedure yielded F plasmid-related sequences from the S. typhimurium virulence plasmid, a number of DNA fragments representing parts of lambdoid prophages and putative sugar transporters, and several fragments with unknown sequences. The majority of subtracted chromosomal sequences map to three distinct locations, around centisomes 5, 27, and 57.

Animals↗

The serologic response to Salmonella enteritidis and Salmonella typhimurium in experimentally infected chickens, followed by an indirect lipopolysaccharide enzyme-linked immunosorbent assay and bacteriologic examinations through a one-year period.

Three groups of 100 individually marked salmonella-free chickens were followed for a period of 53 wk. The chickens were infected as day olds by crop instillation of 10(8) colony-forming units: one group with Salmonella enteritidis and a second group with Salmonella typhimurium. A third group was kept uninfected as controls. The groups were monitored bacteriologically by examination of cloacal swabs and organs and serologically by examination of serum and egg yolk by a lipopolysaccharide enzyme-linked immunosorbent assay throughout the period. Within the first week, 100% of birds in both infected groups were excreting salmonella bacteria in the feces. However, the number of fecal excretors declined rapidly with time, down to 6% in 16 wk for S. typhimurium and down to a similar level within the first 8 wk for S. enteritidis. For the latter, relapses with up to 40% positive birds were observed at the onset of egg production. For both S. typhimurium and S. enteritidis, positive bacteriologic cultures were obtained by sampling from internal organs at the end of the experiment, more than 1 yr from the time of infection. At the age of 6-7 wk, 50% of the chickens in the two infected groups showed a measurable serologic response in serum samples. The response persisted throughout the study in both serum and egg yolk samples. The inclusion of serologic methods is a valuable additional tool in the detection of salmonella in poultry, but serology should be used in conjunction with bacteriologic methods in surveillance programs, in particular to detect flocks in early stages of infection before a measurable serologic response has been raised.

Animals↗

Salmonella-reactive synovial fluid T-cell clones in a patient with post-infectious Salmonella arthritis.

From a patient with reactive arthritis following Salmonella typhimurium enteritis, synovial fluid T-lymphocytes were cloned and expanded in vitro. Seven out of 74 clones showed a marked proliferative response to antigens of heat-killed Salmonella typhimurium with autologous T-cell-depleted peripheral blood mononuclear cells as antigen-presenting cells. The Salmonella-reactive clones were of the CD4+ phenotype, antigen-induced proliferation could be inhibited by a monoclonal antibody to HLA class II. One clone recognized both Salmonella and Campylobacter jejuni antigens in the proliferation assay. The multiclonality of Salmonella-reactive synovial fluid T-cells indicates that the microorganisms have been present, at least transiently, within the affected joint and thus recruited specific T-lymphocytes that might initiate synovitis.

Adult↗

[Significance of motility of Salmonella enteritidis and Salmonella typhimurium as a virulence factor and on the expression of the inhibition phenomenon in vitro and in vivo in SPF chickens].

Methods of immunoprophylaxis for poultry using live Salmonella vaccines are increasingly gaining in importance. Methods of a simple and reliable bacteriological as well as serological differentiation between vaccine and field strains will be of decisive importance for the acceptance and use of live Salmonella vaccines. The absence of motility in Salmonella strains may be a marker fulfilling these criteria. The studies described served to examine whether virulence and the ability to inhibit other Salmonella strains could be influenced by the absence of motility in Salmonella (S.) Enteritidis and (S.) Typhimurium. In a cell-culture model (IEC 6) under in vitro conditions, non-motile transposon mutants (TnphoA) of S. Enteritidis and S. Typhimurium exhibited a clearly reduced invasion potential in comparison with the respective motile parental strain. Under in vitro conditions (nutrient broth culture), the inhibitory potential of these non-motile mutants was also reduced compared to the motile original strains. In contrast, in vivo studies in a-few-days-old chickens revealed that there was no reduction of the virulence of non-motile mutants of S. Enteritidis and S. Typhimurium in comparison with the motile parental strain. In day-old chicks, the inhibitory potential of non-motile strains was significantly reduced and in some cases, had even become completely lost.

Animals↗

[Attempt to estimate the share of human Salmonella infections, which are attributable to Salmonella originating from swine].

On the basis of the data obtained in 1996 and 1997 in Germany on the frequency of detection of Salmonella and also of single Salmonella serovars in foods and infected humans, attempts were made to assess the relevance of Salmonella from swine as a source of human infection. Four assumptions considered as reasonable were made concerning the development of human salmonellosis. On the basis of these assumptions, a model for the estimation of the share of human Salmonella infections attributable to the swine population is proposed. According to this, ca. 20% of human cases of salmonellosis have been caused by Salmonella originating from swine.

Animals↗

Evaluation of cross-protection afforded by a Salmonella Choleraesuis vaccine against Salmonella infections in pigs under field conditions.

This field study investigated the efficacy of a Salmonella Choleraesuis live vaccine (Argus SC) to reduce the number of infections with Salmonella. Twelve groups of about 380 pigs each were randomly allocated to either vaccination (V) or no vaccination (C). The vaccine was applied orally at 3 and 16 weeks. Forty pigs per group were blood sampled at 3, 10, 16, and 24 weeks to detect possible antibodies against Salmonella. The prevalence of Salmonella in the lymph nodes was the major variable. In the V groups, only 0.6% of the lymph nodes were positive, whereas 7.2% were positive in the C groups (p < 0.001). The percentage of seropositive pigs at 24 weeks (cut-off OD > 10) was 26% and 9% in the V and C groups, respectively (p < 0.001). The present study documented that vaccination with a live modified S. Choleraesuis vaccine is a useful tool to lower the prevalence of Salmonella in swine herds.

Animals↗

Peroxidase-antiperoxidase and immunogold labeling of Salmonella typhimurium and Salmonella choleraesuis var kunzendorf in tissues of experimentally infected swine.

Peroxidase-antiperoxidase immunoenzymatic labeling and immunogold labeling techniques were evaluated for microscopic detection and location of Salmonella organisms in tissues of experimentally infected swine. Salmonella typhimurium and Salmonella choleraesuis var kunzendorf were labeled specifically by the peroxidase-antiperoxidase technique in paraffin-embedded tissues of infected swine for conventional light microscopy and by postembedding immunogold labeling on ultrathin sections for electron microscopy. Salmonella typhimurium had a low tendency to invade the enteric mucosa and did not reveal any predilection for a specific intestinal location. Salmonella choleraesuis var kunzendorf, however, was located preferentially in colon and on the luminal surface of ileal M cells of Peyer patches and had a tendency to invade the enteric mucosa there.

Animals↗

Distribution of Salmonella typhi and Salmonella paratyphi B phage types in Italy from 1974 through 1985.

The temporal changes in occurrence and distribution in Salmonella typhi and paratyphi B phage types in Italy during a twelve year period are considered. From 1974 to 1985, 2.075 isolates of Salmonella typhi and 830 isolates of Salmonella paratyphi B were phage typed at the National Centre for Enterobacteria in Rome and the three Regional Centres for Enterobacteria in Milan, Palermo and Pisa. The most prevalent types of Salmonella typhi were A, C1, D1, Vi degraded strains E1, I+IV and C4. Types A, C1 and D1 accounted for 59.1% of the total. The distribution of the phage types as A, B2, C1, D1, D4, D9, E1, F1, N, 28 and I+IV, Vi degraded and Vi negative strains in the three time periods (1974-77; 1978-81 and 1982-85) was quite similar; some types such as D8, E2, 0 and 43 were found sporadically and others like C9, D6, F4, H and 45 which were present in the first period, disappeared in the last. The most prevalent types of Salmonella paratyphi B were Tauton, Dundee, 1, and 3a. Type 2, frequently found in the 1974-77 period was however not seen during 1981-85.

Bacteriophage Typing↗

Defined salmonella antigens for detection of cellular and humoral immune responses in salmonella infected calves.

Intracutaneous injection of a crude supernatant fraction from homogenised Salmonella typhimurium SVA 44 (O 4, 5, 12) or S dublin SVA 47 (O 9, 12) elicited highly significant (P less than 0.005) double skin-fold thickness increases in calves spontaneously infected with salmonella and verified as excretors. The use of isolated structurally defined outer membrane components from salmonella bacteria established that the delayed skin reactions could be elicited by either the lipopolysaccharide which contains O-antigenic polysaccharide chains homologous to the infecting strain, or an outer membrane protein fraction (porin). The porin preparation gave rise to skin reactions regardless of which salmonella serotype the calf was infected with. Histological examination of biopsy material indicated a delayed skin reaction. No such reactions were seen in biopsies from control calves. The use of lipopolysaccharide permitted a salmonella serogroup specific skin test although the endotoxic side effects were marked in doses above 50 micrograms. Purified O-antigen specific polysaccharides devoid of lipid A from S typhimurium (O 4, 12) or S enteritidis (O 9, 12) failed however to elicit skin reactions. Infected calves had humoral antibody titres against the O antigen of the infecting strain which were significantly (P less than 0.005) higher than those found in control calves.

Animals↗

Detection of Salmonella C1, D and V1 antigens, by coagglutination, in blood cultures from patients with Salmonella infections.

Protein A-containing Staphylococcus aureus was coupled to Salmonella C1, D and Vi monovalent antisera to produce C1-, D- and Vi-COAG reagents. The reagents were used to detect their homologous Salmonella antigens in blood cultures (BC). The D and Vi antigens were detected in 79 of 239 BC from patients with suspected typhoid fever and Salmonella typhi was later isolated from the same 79 BC. The C1 antigen was detected in 8 BC from which only S. oranienburg was later isolated. The COAG test was generally positive at the same time the BC became culture positive. However, because of subculture and biochemical identification requirements the COAG test could be interpreted 1-2 days before culture results were available. The COAG test can be used to presumptively identify Salmonella typhi and Salmonella group C1 in blood cultures before the culture results are available.

Agglutination Tests↗

Control of Salmonella infections in animals and prevention of human foodborne Salmonella infections. WHO Consultation.

In many countries the incidence of human salmonella infections has markedly increased in recent years. To discuss recent developments and current understanding on the control of salmonella infections in animals, WHO organized a Consultation on the Control of Salmonella Infections in Animals: Prevention of Foodborne Salmonella Infections in Humans, held in Jena, Germany, on 21-26 November 1993. The present article summarizes the recommendations made by the participants on the pathoimmunogenesis, diagnosis, epidemiology, and control of salmonella infections and contaminations in animal production.

Animals↗