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Results for “Virulence profiling”

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PubMed2001

Host microarray analysis reveals a role for the Salmonella response regulator phoP in human macrophage cell death.

Bacterial pathogens manipulate host cells to promote pathogen survival and dissemination. We used a 22,571 human cDNA microarray to identify host pathways that are affected by the Salmonella enterica subspecies typhimurium phoP gene, a transcription factor required for virulence, by comparing the expression profiles of human monocytic tissue culture cells infected with either the wild-type bacteria or a phoPTn10 mutant strain. Both wild-type and phoPTn10 bacteria induced a common set of genes, many of which are proinflammatory. Differentially expressed genes included those that affect host cell death, suggesting that the phoP regulatory system controls bacterial genes that alter macrophage survival. Subsequent experiments showed that the phoPTn10 mutant strain is defective for killing both cultured and primary human macrophages but is able to replicate intracellularly. These experiments indicate that phoP plays a role in Salmonella-induced human macrophage cell death.

Bacterial Proteins↗

PubMed2000

Biodistribution and genetic stability of the novel antitumor agent VNP20009, a genetically modified strain of Salmonella typhimurium.

VNP20009 is a genetically modified strain of Salmonella typhimurium possessing an excellent safety profile, including genetically stable attenuated virulence (a deletion in the purI gene), reduction of septic shock potential (a deletion in the msbB gene), and antibiotic susceptibility. VNP20009 is genetically stable after multiple generations in vitro and in vivo. In mice, VNP20009 is rapidly cleared from the blood from a peak level of 1x104 cfu/mL to undetectable levels in 24 h. In tumor-bearing mice, VNP20009 accumulates preferentially in tumors over livers at a ratio of 1000&rcolon;1. In nonhuman primates, VNP20009 was also rapidly cleared from the blood, from a peak level of 1.0x106 cfu/mL to undetectable levels in 24 h. VNP20009 was detected in the liver, spleen, and bone marrow of monkeys; the amount decreased over time, and VNP20009 was cleared from all organs by day 41; no VNP20009 could be detected in the urine or feces of the monkeys. VNP20009 is genetically stable after many generations of growth (>140) both in vitro and in vivo.

Animals↗

PubMed1996

Stationary phase-associated protein expression in Mycobacterium tuberculosis: function of the mycobacterial alpha-crystallin homolog.

The majority of active tuberculosis cases arise as a result of reactivation of latent organisms which are quiescent within the host. The ability of mycobacteria to survive extended periods without active replication is a complex process whose details await elucidation. We used two-dimensional gel electrophoresis to examine both steady-state protein composition and time-dependent protein synthetic profiles in aging cultures of virulent Mycobacterium tuberculosis. At least seven proteins were maximally synthesized 1 to 2 weeks following the end of log-phase growth. One of these proteins accumulated to become a predominant stationary-phase protein. N-terminal amino acid sequencing and immunoreactivity identified this protein as the 16-kDa alpha-crystallin-like small heat shock protein. The gene for this protein was shown to be limited to the slowly growing M. tuberculosis complex of organisms as assessed by Southern blotting. Overexpression of this protein in wild-type M. tuberculosis resulted in a slower decline in viability following the end of log-phase growth. Accumulation of this protein was observed in log-phase cultures following a shift to oxygen-limiting conditions but not by other external stimuli. The protein was purified to homogeneity from overexpressing M. smegmatis in two steps and shown to have a significant ability to suppress the thermal denaturation of alcohol dehydrogenase. Collectively, these results suggest that the mycobacterial alpha-crystallin protein may play a role in enhancing long-term protein stability and therefore long-term survival of M. tuberculosis.

Antigens, Bacterial↗

PubMed1996

Variant colony surface antigenic phenotypes within mycoplasma strain populations: implications for species identification and strain standardization.

Immunobinding assays with mycoplasma colonies on agar plates (immunofluorescence and immunoperoxidase techniques) or with imprints of colonies transferred to solid supports (colony immunoblotting) are widely used as standard diagnostic tests for serological species identification of mycoplasma isolates. However, in light of the high rate of variability of surface antigens in many mycoplasmas, diagnostic data obtained with these techniques require a more critical evaluation. In this report, we demonstrate with some examples that mycoplasma surface variability based on alterations in expression, in size, and in surface presentation of integral and peripheral membrane proteins may lead to misinterpretation of colony immunostaining reactions obtained by using specific monoclonal antibodies as well as conventional diagnostic hyperimmune sera. To more easily identify phenotypically mixed isolates or samples which contain more than one species, we have introduced some minor modifications of the colony immunoblot technique which provide sharp signals of positive as well as negative reactions and enable identification of cryptic epitopes. It is further demonstrated that because of the variability in colony surface antigenic phenotype, mycoplasma strains, including well-established reference and other prototype strains which are used under the same designation in many laboratories, can differ markedly in their antigen profiles and their potentially virulence-related surface properties, since they are usually purified by filter cloning and often propagated by subcultivation of randomly selected agar-grown subpopulations. We conclude from this study that because of this surface variability, the establishment of criteria for standardization of mycoplasma strains and diagnostic antisera is urgently required in order to obtain reproducible results in different laboratories.

Antibodies, Bacterial↗

PubMed2026

In-host adaptation of Staphylococcus aureus during recurrent prosthetic joint infections: a retrospective longitudinal study.

UNLABELLED: The aim of this study was to characterize the in vivo evolution of Staphylococcus aureus strains involved in recurrent prosthetic joint infections (PJIs) both phenotypically and genomically. We conducted a monocentric retrospective study in a 1,437-bed French teaching hospital between 2013 and 2021. All patients presenting a recurrent S. aureus-related PJI-defined as at least two strains isolated from distinct clinical samples more than 90 days apart-of the knee, hip, or shoulder were included. Clinical data were reviewed, and all isolates underwent phenotypic characterization, including antimicrobial susceptibility testing, growth rate determination, biofilm production assays, metabolic profiling (API 50 CH), and virulence evaluation using the Galleria mellonella infection model. Whole-genome sequencing (WGS) was performed for all strains, followed by analyses of core-genome multilocus sequence typing (cgMLST), resistome, virulome, and mobilome composition, and single-nucleotide polymorphisms (SNPs). Thirteen patients met inclusion criteria, yielding 55 S. aureus isolates. Eight patients experienced recurrent infections caused by genetically closely related strains throughout the clinical course (median: three strains per patient; range: 2-6), whereas five patients were infected by genetically distinct strains. At baseline, isolates were genetically diverse and susceptible to methicillin and rifampicin; two showed fluoroquinolone resistance due to grlA and/or gyrA mutations. In one patient (patient C), a recurrent isolate acquired an rpoB S486L mutation, conferring rifampicin resistance after rifampicin exposure. Due to the limited sample size, it is difficult to draw definitive conclusions from the phenotypic analyses. This study highlights the adaptive evolution of S. aureus during chronic PJIs and underscores the need for further research to better understand intra-host dynamics in long-standing infections. IMPORTANCE: This study conducted in a 1,437-bed French teaching hospital analyzed the genomic and phenotypic evolution of 55 Staphylococcus aureus strains recovered in recurrent PJIs from 13 patients. The first strains showed high genotypic diversity across 12 different sequence types. Among the 13 patients, only eight experienced a true recurrence with the same strain, while five were contaminated with a different strain of S. aureus, indicating a new infection. Moreover, this study underscores the complex within-host evolution of S. aureus and highlights the phenotypical and genotypical adaptation during chronic infection.

Staphylococcus aureus↗

PubMed1996

Molecular analysis of genetic differences between Mycobacterium bovis BCG and virulent M. bovis.

The live attenuated bacillus Calmette-Guérin (BCG) vaccine for the prevention of disease associated with Mycobacterium tuberculosis was derived from the closely related virulent tubercle bacillus, Mycobacterium bovis. Although the BCG vaccine has been one of the most widely used vaccines in the world for over 40 years, the genetic basis of BCG's attenuation has never been elucidated. We employed subtractive genomic hybridization to identify genetic differences between virulent M. bovis and M. tuberculosis and avirulent BCG. Three distinct genomic regions of difference (designated RD1 to RD3) were found to be deleted from BCG, and the precise junctions and DNA sequence of each deletion were determined. RD3, a 9.3-kb genomic segment present in virulent laboratory strains of M. bovis and M. tuberculosis, was absent from BCG and 84% of virulent clinical isolates. RD2, a 10.7-kb DNA segment containing a novel repetitive element and the previously identified mpt-64 gene, was conserved in all virulent laboratory and clinical tubercle bacilli tested and was deleted only from substrains derived from the original BCG Pasteur strain after 1925. Thus, the RD2 deletion occurred after the original derivation of BCG. RD1, a 9.5-kb DNA segment found to be deleted from all BCG substrains, was conserved in all virulent laboratory and clinical isolates of M. bovis and M. tuberculosis tested. The reintroduction of RD1 into BCG repressed the expression of at least 10 proteins and resulted in a protein expression profile almost identical to that of virulent M. bovis and M. tuberculosis, as determined by two-dimensional gel electrophoresis. These data indicate a role for RD1 in the regulation of multiple genetic loci, suggesting that the loss of virulence by BCG is due to a regulatory mutation. These findings may be applicable to the rational design of a new attenuated tuberculosis vaccine and the development of new diagnostic tests to distinguish BCG vaccination from tuberculosis infection.

BCG Vaccine↗

PubMed1995

Evidence for the existence of distinct populations of Vibrio anguillarum serogroup O1 based on plasmid contents and ribotypes.

A total of 103 Vibrio anguillarum serogroup O1 strains displaying 15 different plasmid profiles were characterized with respect to biochemical properties and ribotypes. The results confirmed that V. anguillarum O1 is a biochemically homogeneous group. The 103 strains could be allocated to three main clusters with high similarity coefficients. None of the biochemical properties were connected with the presence of plasmids. In total, 12 different ribotypes were demonstrated, with HindIII being used as the restriction enzyme. Forty of the strains were isolated from the same Danish fish farm, some from the kidneys of diseased fish and some from the environment, and some strains were isolated from the mucus, gills, and feces of healthy fish. Nineteen of these isolates possessed the 67-kb virulence plasmid alone or in combination with other plasmids, while 21 had no plasmids. All strains isolated from the kidneys of diseased fish on this farm had plasmids. Irrespective of their origin (kidneys, gills, or mucus), all 19 strains carrying the 67-kb virulence plasmid had the same ribotype, profile 1, while isolates without plasmids belonged to five different profiles, all different from profile 1. These results suggest that pathogenic V. anguillarum O1 strains possessing a virulence plasmid and nonpathogenic strains without plasmids from a small geographical area and even from the same fish may constitute two essentially distinct populations. Thus, it may be suggested that an exchange of virulence plasmids among strains is unlikely to occur in vivo.

Bacterial Typing Techniques↗

PubMed2006

Proteomic profiling of cell envelope-associated proteins from Staphylococcus aureus.

The emergence of highly virulent community acquired Staphylococcus aureus and continued progression of resistance to multiple antimicrobials, including methicillin and vancomycin, marks the reemergence of S. aureus as a serious health care threat. Investigation of proteins localized to the cell surface could help to elucidate mechanisms of virulence and antibiotic resistance in S. aureus. In this study, proteomic profiling methods were developed to solubilize, display, and evaluate abundance levels of proteins present in the supernatants of the lysostaphin-digested cell envelope from cultured vancomycin-intermediate S. aureus (VISA) cells. Combining approaches of 2-DE or chromatographic separation of proteins with MS analyses resulted in the identification of 144 proteins of particular interest. Of these proteins, 48 contained predicted cell wall localization or export signal motifs, including 14 with distinct covalent peptidoglycan-anchor sites, four of which are uncharacterized to date. One of the two most abundant cell envelope proteins, which showed remarkably high variations in MW and pI in the 2-DE gel display, was the S. aureus surface protein G. The display of numerous secreted proteins that are not covalently cell wall-anchored, suggests that, in the exponential growth phase, secreted proteins can be retained physiologically in the cell envelope and may interact with cell wall-anchored proteins and carbohydrate structures in a manner yet to be determined. The remaining 96 proteins, devoid of recognizable motifs, were repeatedly profiled in the VISA cell envelope fractions. We describe a novel semiquantitative method to determine abundance factors of such proteins in 2-DE gels of cell envelope fractions relative to whole cell lysates and discuss these data in the context of true cell envelope localization versus experimentally caused cell lysis.

Amino Acid Sequence↗

PubMed1994

Ovine Haemophilus somnus: experimental intracisternal infection and antigenic comparison with bovine Haemophilus somnus.

Experimental infection was produced by two of four isolates of ovine Haemophilus somnus given by intracisternal inoculation into two to three-month-old lambs. Isolate 2041 (originally obtained from a septicemic lamb in Alberta) caused lethal infection in eight of nine lambs, isolate 67p from the prepuce of a normal lamb produced less acute disease in four of nine lambs, and the other two isolates (93p and 1190) caused no detectable disease. Significant lesions were limited to the brain and spinal cord. Purulent meningitis was characteristic but vasculitis or septicemia were not detected, perhaps due to the route of inoculation. Since a difference in virulence was noted among strains, we analyzed surface proteins thought to be virulence factors of bovine H. somnus. Protein profiles of bovine and ovine H. somnus done by sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed similar patterns for virulent bovine isolates and ovine septicemic isolates. Preputial isolates showed a lower molecular mass major outer membrane protein than septicemic isolates. Antigenic analysis revealed that outer membrane proteins p270, p78, p76, p40, and p39 were detected in both ovine and bovine isolates except for 1190, which was probably not a true H. somnus isolate. Thus the preputial and septicemic isolates of ovine H. somnus were similar to bovine H. somnus in pathogenicity and in surface antigens.

Animals↗

PubMed1992

[Studies on genetic differentiation of Salmonella enteritidis isolates by plasmid profile].

Since 1989, the number of salmonellosis cases caused by S. Enteritidis has increased considerably in Japan. Genetic differentiation of 385 strains isolated from January 1982 to December 1988 and January 1989 to April 1991 were used for plasmid profiles. Plasmids were found in 377 out of 385 strains; therefore, only 8 strains carried no plasmid. Among 377 strains, 15 different plasmid profile types (OP-1 to OP-15) were classified. The most common plasmid profile types from 1982 to 1988 were OP-7 (70 kbp) and OP-8 (70 kbp and 2 kbp). On the other hand, the most common plasmid profile types from 1989 to 1991 were OP-1 (60 kbp) and OP-2 (60 kbp and 54 kbp). Serovar-specific virulence 60 kbp plasmids of S. Enteritidis were identified in 7 plasmid profile types (A total of 200 strains). In the other plasmid profile types, 70 kbp plasmids were found in 5 plasmid profile types (A total of 173 strains). In restriction enzyme analysis of 70 kbp plasmid DNAs obtained from 5 plasmid profile types of S. Enteritidis, we found that these plasmid DNAs shared both 60 kbp and 10 kbp fragments. These results indicate that these plasmid profile types also carried serovar-specific virulence plasmids of S. Enteritidis. The strains of plasmid profile type OP-2 were SA (sulfisoxazole) and SM (streptomycin) resistance, and the 54 kbp plasmids in the strains of OP-2 were transferred by bacterial conjugation into the E. coli strains. All transconjugants acquired SM resistance.

Humans↗

PubMed2001

Molecular and antigenic characterization of Bangladeshi isolates of infectious bursal disease virus demonstrate their similarities with recent European, Asian and African very virulent strains.

Three isolates of infectious bursal disease virus (IBDV), obtained from chickens in Bangladesh in 1999 and designated as BD 1/99, BD 2/99 and BD 3/99, were characterized. In an antigen capture enzyme-linked immunosorbent assay using a panel of VP2-specific neutralizing monoclonal antibodies (mAb), all three isolates showed a mAb-binding profile similar to that of very virulent IBDV (vvIBDV) strains. In contrast to the classical virulent strains, they did not react with mAb 3 and mAb 4. Molecular characterization was performed by direct sequencing of a 677-base pair cDNA corresponding to the VP2 variable domain of the polyprotein gene, synthesized by a reverse transcription-polymerase chain reaction. In comparison to the classical virulent strains, the Bangladeshi isolates were found to have five amino acid substitutions in this region. Four of these changes, Pro222Ala, Val256Ile, Leu294Ile and Asn299Ser, were also observed in other vvIBDV strains. The fifth substitution, Glu300Ala, was similar to that in some African strains of IBDV. The results support the observation that antigenically and genetically similar vvIBDV strains, first observed in Europe in the late 1980s, have spread to most parts of the world in a short period of time.

Africa↗

PubMed1995

The Salmonella dublin virulence plasmid mediates systemic but not enteric phases of salmonellosis in cattle.

Plasmid-bearing and plasmid-free isolates and a plasmid-cured strain of Salmonella dublin were compared for virulence in calves. The plasmid-bearing strains were highly virulent, causing severe enteric and systemic disease with high mortality. In contrast, the plasmid-free strains caused diarrhea but only low mortality. The infection kinetics of a wild-type and a derivative plasmid-cured strain were compared. Both strains were isolated in high numbers from intestinal sites at 3 and 6 days after oral challenge and were isolated at comparable frequencies from systemic sites at 3 days, but not at 6 days, when the wild-type strain was predominant. The strains were equally invasive in intestinal epithelia with and without Peyer's patch and elicited comparable secretory and inflammatory responses and intestinal pathology in ligated ileal loops. The effect of the virulence plasmid on growth kinetics and on the outer membrane protein profile was assessed in an in vivo growth chamber. The virulence plasmid did not influence either extracellular growth or the expression of major outer membrane proteins. These observations demonstrate that the virulence plasmid is not involved in either the enteric phase of infection or the systemic dissemination of S. dublin but probably mediates the persistence of S. dublin at systemic sites.

Animals↗

PubMed2001

Theileria annulata: attenuation of a schizont-infected cell line by prolonged in vitro culture is not caused by the preferential growth of particular host cell types.

Flow cytometry and monoclonal antibodies to bovine leucocyte surface antigens were used to identify the types of host cells that the sporozoites of Theileria annulata infect in cattle, to determine whether virulent schizont-infected cell lines (lines) differed phenotypically from avirulent lines, and to establish whether attenuation in vitro was accompanied by the preferential growth of particular host cell types. The surface antigens of four pairs of T. annulata (Ta) (Hisar) lines derived ex vivo and in vitro, including the virulent ex vivo-derived Ta Hisar S45 line, were consistent with a myeloid origin for all lines, irrespective of their derivation. The profiles of lines derived from cattle inoculated with a virulent line showed that the schizonts liberated from inoculated cells had transferred to myeloid cells. A number of other lines infected with different stocks of T. annulata expressed myeloid markers; a single line expressed CD21, a B cell marker. During prolonged in vitro culture, the parasites in the ex vivo (virulent)- and in vitro (avirulent)-derived Ta Hisar S45 myeloid lines became clonal, as defined by glucose phosphate isomerase (GPI) polymorphism, and the virulent line became attenuated. The two lines retained phenotypic profiles indicative of a myeloid origin but coexpressed some lymphoid antigens (CD2, CD4, CD8), although not CD3. Cloned schizont-infected lines, representing the three parasite GPI isotypes which constituted the virulent line, expressed similar patterns of myeloid and lymphoid markers to the virulent parent line. Some schizont-infected clones failed to establish as lines during the early weeks of culture because the cells died as the parasites differentiated into merozoites at 37 degrees C, the temperature at which schizont-infected cells normally grow exponentially. These results provided no evidence that prolonged culture induces preferential growth or loss of particular host cell types. However, a number of the alterations in host cell surface antigens induced by prolonged culture were shown to be linked to permanent changes in the parasite genome.

Animals↗

PubMed2002

Virulence control in group A Streptococcus by a two-component gene regulatory system: global expression profiling and in vivo infection modeling.

Two-component gene regulatory systems composed of a membrane-bound sensor and cytoplasmic response regulator are important mechanisms used by bacteria to sense and respond to environmental stimuli. Group A Streptococcus, the causative agent of mild infections and life-threatening invasive diseases, produces many virulence factors that promote survival in humans. A two-component regulatory system, designated covRS (cov, control of virulence; csrRS), negatively controls expression of five proven or putative virulence factors (capsule, cysteine protease, streptokinase, streptolysin S, and streptodornase). Inactivation of covRS results in enhanced virulence in mouse models of invasive disease. Using DNA microarrays and quantitative RT-PCR, we found that CovR influences transcription of 15% (n = 271) of all chromosomal genes, including many that encode surface and secreted proteins mediating host-pathogen interactions. CovR also plays a central role in gene regulatory networks by influencing expression of genes encoding transcriptional regulators, including other two-component systems. Differential transcription of genes influenced by covR also was identified in mouse soft-tissue infection. This analysis provides a genome-scale overview of a virulence gene network in an important human pathogen and adds insight into the molecular mechanisms used by group A Streptococcus to interact with the host, promote survival, and cause disease.

Animals↗

PubMed2002

[Evaluation of western blot methods for serologic diagnosis of Helicobacter pylori infections].

Recent studies have demonstrated that immunoblot method is more sensitive and specific than enzyme immunoassays (EIA) to diagnose Helicobacter pylori infections, and it also permits the detection of antibody profiles against different bacterial antigens including the virulence factors. The aim of this study was to evaluate the diagnostic value of Western Blot (WB) method in the serological diagnosis of H. pylori infections. For this purpose, H. pylori IgG and IgA antibodies have been searched qualitatively by a commercial WB test (Euroimmun GmbH, Germany), in the sera of 23 patients (9 female, 14 male) ages between 6-70 years old (13 children, 10 adults), who were diagnosed as H. pylori infection with the positive results of at least three of the conventional methods (culture and polymerase chain reaction of endoscopic biopsy specimens, 13C urea breath test, serum IgG-EIA methods). All of the patients (100%) were found positive by WB-IgG, and 9 (39%) were positive by WB-IgA tests. H.pylori IgG and IgA antibodies showed reactivity with the different combinations of p120 (CagA), p95 (VacA), p29 (UreA) and p66 (UreB) antigens. The number of positive samples for CagA-IgG, VacA-IgG, UreA-IgG, CagA-IgA and UreA-IgA were 19, 17, 21, 8 and 4, respectively, while there were no positive sample for VacA-IgA. The antigenic distribution did not show any difference between age and sex of the patients. In our study the sensitivities of WB-IgG and WB-IgA methods were estimated as 100% and 39%, respectively, which were in accordance with the studies in literature. As a result, WB-IgG method appears to be useful and reliable for the detection of antibody profiles to H. pylori antigens and virulence factors, rather than routine and screening purposes, however the diagnostic value of WB-IgA method is low. It was thought that the results of our preliminary study should be confirmed by other studies including larger patient populations and control groups, in our country.

Adolescent↗

PubMed1999

Comparative physical and genetic maps of the virulence plasmids of Salmonella enterica serovars typhimurium, enteritidis, choleraesuis, and dublin.

Using fragment profiling, PCR, and Southern hybridization, we found that Salmonella enterica serovar Choleraesuis harbored virulence plasmids of various sizes, whereas serovars Typhimurium, Enteritidis, and Dublin carried a plasmid of a unique size. Also, the virulence plasmid of Typhimurium contained genes in the same order detected in the other three plasmids, all of which contained deletions.

Base Sequence↗

PubMed1995

Identification of virulent Rhodococcus equi by amplification of gene coding for 15- to 17-kilodalton antigens.

During a survey of the prevalence of virulent Rhodococcus equi at horse-breeding farms by plasmid and protein profiles, cryptic plasmids of various sizes were found in 66 (3.8%) of 1,725 isolates from feces of horses and 129 (5.9%) of 2,200 isolates from soil. Twenty-two isolates, which contained cryptic plasmids of different sizes, were found by plasmid profiles, and their protein profiles and mouse pathogenicities were examined. Of the 22 isolates, 7 were virulent R. equi, contained both virulence and cryptic plasmids, and expressed 15- to 17-kDa antigens. The remaining 15 isolates were avirulent and did not express the antigens: 6 strains contained cryptic plasmids of two different sizes and 9 strains contained cryptic plasmids of various sizes. A PCR assay was developed for the rapid identification of virulence plasmids of R. equi. Oligonucleotide primers, derived from the sequence of a gene coding for the 15- to 17-kDa virulence-associated antigens of R. equi, amplified a 564-bp product from all the tested isolates harboring a virulence plasmid. This PCR product hybridized with virulence plasmid DNA in the Southern hybridization assay. Virulence plasmid-cured derivatives and all of the tested isolates harboring cryptic plasmids only were negative. The PCR is a rapid, sensitive, and specific test for the identification of virulent R. equi from environmental isolates compared with standard techniques, such as plasmid and protein profiles and the mouse pathogenicity test, and is considered to be a useful tool for epidemiological studies.

Animals↗

PubMed1990

Outbreak of Salmonella typhimurium infection traced to contaminated chocolate and caused by a strain lacking the 60-megadalton virulence plasmid.

We describe an outbreak of Salmonella typhimurium infection, caused by contaminated chocolate produced by one Norwegian company, which occurred in Norway and Finland in 1987. A total of 349 bacteriologically verified cases were recorded in Norway, and 12 cases were recorded in Finland. There was a predominance of young children among the patients (median age, 6 years), many of whom developed acute hemorrhagic diarrhea. The outbreak strain exhibited a rare phage lysis pattern and a characteristic plasmid profile lacking the 60-MDa virulence-associated plasmid. DNA hybridization failed to demonstrate any DNA sequence homology between the outbreak strain and the virulence plasmid. The outbreak strain was nonlethal for orally infected mice. The finding of only less than or equal to 10 S. typhimurium cells per 100 g of chocolate in about 90% of the positive samples obtained from retail outlets suggested that an inoculum of fewer than 10 organisms may have been sufficient to cause symptomatic disease.

Cacao↗