Lack of virulence factors in Escherichia coli strains of enteropathogenic serogroups isolated from water.

Thirty-eight Escherichia coli strains belonging to 14 human enteropathogenic serogroups were isolated from 33 of 208 water samples studied. No virulence factor or virulence-related gene sequences were found in any of the 38 strains analyzed. The results point out the importance of detecting specific virulence factors before incriminating water as a source of human diarrhea.     

Sewage dilution and loss of antibiotic resistance and virulence determinants in E. coli

The possession of potential virulence factors (serum resistance, aerobactin production, colicin production) and antibiotic resistance was evaluated in 418 E. coli strains isolated from river water. The strains were isolated from 11 points showing different levels of contamination. From the data obtained, it can be concluded that bacteria from less contaminated water present less antibiotic resistance and virulence factors than those isolated from highly contaminated water. This situation suggests that E. coli strains producing plasmid encoded antibiotic resistance and/or virulence factors, survive less well in aquatic environments without selective pressure) than the sensitive non-virulent ones.     

Evaluation of Virulence Test Procedures for Yersinia enterocolitica Recovered from Foods

Recently a heat stable toxin (ST) and the vw factors of the plague bacteria were identified in Yersinia enterocolitica recovered from human infections. The vw factors were reported to be associated with a plasmid of 42-46 M Daltons in size and were essential for the expression of virulence. With this knowledge virulence tests were developed which allowed us to assess the virulence potential of food-borne Y. enterocolitica regardless of biotypes or serotypes. The tests evaluated were: (1) rapid presumptive test for the virulence plasmid; (2) gel electrophoretic confirmation of the virulence plasmid; (3) Laird's qualitative oral feeding test with thirst stressed mice; (4) quantitative LD₅₀ determination by i.p. injection of the mouse lethal ( i.e. serotype 0:8) strains in saline; (5) quantitative LD₅₀ determination of mouse non-lethal ( i.e. serotype 0:3) strains by i.p. injection of these strains suspended in 1 ml of 10% iron dextran saline solution for virulence enhancement. These tests were evaluated with the serotypes 0:3 and 0:8 strains associated with human infections with and without the virulence plasmid with reproducible results. Then the virulence tests procedures were applied to 79 food isolates. The virulence plasmid was detected only in the Nilehn biotype 2, 3 and 4 strains, but it was absent in Nilehn biotype 1 or the atypical strains that ferment rhamnose. The virulence of food and clinical isolates of Y. enterocolitica can be assayed fairly accurately with the above tests.     

Influence of growth temperature on the production of extracellular virulence factors and pathogenicity of environmental and human strains of Aeromonas hydrophila

The biochemical properties, virulence for mice and trout, and the extracellular virulence factors at 28° and 37°C of 11 environmental and nine human strains of Aeromonas hydrophila were compared. All the environmental isolates and four of the human group were virulent for trout at 3 x 10⁷ cfu, but only human strains were able to cause death or lesions in mice by the intramuscular route. Extracellular virulence factors such as haemolysins, cytotoxins and proteases were also investigated in supernatant fluids of cultures grown at 28°C and 37°C. The production of haemolysins, caseinases, elastases and growth yields of environmental strains decreased sharply during cultivation at 37°C but cytotoxins were produced to the same extent, or slightly less, than at 28°C. The human strains differed from the environmental strains in response to growth temperatures: protease activity decreased at 37°C, although growth yield was not affected, but more haemolysins and cytotoxins were produced by the virulent strains at this temperature than at 28°C. Sodium caseinate SDS-PAGE of culture supernatant fluids of selected human strains revealed that temperature selectively inhibited the production of certain proteases.     

Host specificity of septicemic Escherichia coli: human and avian pathogens

Extraintestinal pathogenic Escherichia coli (ExPEC) strains are the cause of a diverse spectrum of invasive human and animal infections, often leading to septicemia. ExPEC strains contain virulence factors that enable them to survive in the host blood and tissues. Most of these virulence factors are distributed in ExPEC strains in a host-independent fashion. Genomic analyses of these strains provide evidence for numerous recombinational events and horizontal gene transfer, as well as for a high diversity of virulence factors. In studies of human and avian septicemic strains of serotypes O2 and O78 it appears that there is a positive correlation between virulence, invasiveness and clonal origin. Yet, it is clear that clonal division in these strains, as well as distribution of virulence factors, is independent of the host and closely related clones reside in different hosts. Although the possibility exists that ExPEC strains do have a certain degree of host specificity, which is not obvious from genomic studies, it is clear that the similarity of virulence factors presents a significant zoonotic risk.     

Distribution of Aeromonas hydrophila hybridization groups and their virulence properties in Australasian clinical and environmental strains

A total of 182 Aeromonas hydrophila strains isolated from environmental (food and water) and clinical (stool and other sources) samples taken in mainland Australia, Tasmania and New Zealand were assigned to one of three DNA/DNA hybridization groups (HGs) on the basis of biochemical characteristics, and tested with regard to their ability to produce virulence factors. Strains from HG2 were rarely isolated; strains from HG1 were most commonly isolated from clinical sources; and strains from HG3 formed the majority of environmental strains. There was no correlation of HG to geographic source. Strains from HG2 infrequently produced virulence factors. Strains from HG1 were more likely to produce virulence factors if they came from a clinical source. Overall, strains from mainland Australia produced virulence factors more frequently than those from Tasmania or New Zealand. Strains from HG1 may be of more clinical significance than strains from the other two HGs.     

Importance of flagella and enterotoxins for Aeromonas virulence in a mouse model

A genetic characterization of eight virulence factor genes, elastase, lipase, polar flagella (flaA/flaB, flaG), lateral flagella (lafA), and the enterotoxins alt, act, and ast, was performed using polymerase chain reaction with 55 drinking water and nine clinical isolates. When 16 Aeromonas hydrophila strains, seven Aeromonas veronii strains, and seven Aeromonas caviae strains exhibiting different combinations of virulence factor genes were tested in immunocompromised mice by intraperitoneal injection, only those strains that had one or more of the enterotoxins flaA, flaB, and either flaG or lafA showed signs of being virulent. The correlation was seen in 97% (29/30) of the strains, which included strains from drinking water. Thus, Aeromonas water isolates have the potential to be pathogenic in immunocompromised hosts.     

Chromosomal-encoded siderophores are required for mouse virulence of enteropathogenic Yersinia species

Abstract Yersinia enterocolitica and Y. pseudotuberculosis are enteropathogenic for humans. Essential virulence functions of these pathogens are determined by a 40-mDa plasmid. Plasmid-bearing Y. pseudotuberculosis strains and Y. enterocolitica strains of serotypes 0 : 8, 0 : 13, 0 : 20 and 0 : 40 are lethal for mice. In contrast, human pathogenic Y. enterocolitica strains of serotype 0 : 3, 0 : 9 and 0 : 5.27 are not mouse-lethal. Using a sensitive siderophore-indicator CAS-agar, we were able to detect siderophore production in all mouse-lethal Y. enterocolitica and Y. pseudotuberculosis strains mentioned above. By Tn5-transposon insertions into the chromosome of a serotype 0 : 8 strain we obtained two siderophore-deficient mutants. Introduction of the virulence plasmid did not render these mutants mouse-lethal, indicating that siderophore production is an essential virulence factor. The human nonpathogenic, aerobactin-producing strains of Y. intermedia, Y. kristensenii and Y. frederiksenii remained avirulent for mice after receiving the virulence plasmid of Y. enterocolitica . Obviously the siderophore aerobactin does not contribute to virulence in the genus Yersinia .     

The virulence of Staphylococcus aureus correlates with strain genotype in a chicken embryo model but not a nematode model

Staphylococcus aureus infections are of major importance in human and veterinary medicine. Studies of the virulence of this bacterium are complicated by inconsistent results obtained in different animal models. We searched for an uncomplicated and inexpensive model suitable to study virulence of poultry strains of S. aureus using a genome-wide approach. We determined that a useful model would clearly differentiate strains of high and low virulence, and that this would generally correlate with the genetic relatedness among strains. To this end Gallus gallus (chicken) embryo and Caenorhabditis elegans (nematode) models were selected, and their response to challenge by a set of well-characterized Staphylococcus strains was evaluated. The chicken embryo model allowed to determine variation in virulence among strains of poultry and human origin. The survival of embryos ranged from 0% to almost 100% for the various strains. In contrast, variation in virulence of most strains in the nematode model was comparable, regardless of their origin or genotype, demonstrating limited usefulness of this model. Most importantly, a clear correlation was found between the virulence level in the embryo model and the genotype of the tested poultry strains. Our findings indicate the potential usefulness of embryo model for future identification of host-specific adaptations and virulence factors in S. aureus.     

Characterization of Klebsiella Isolates from Natural Receiving Waters and Comparison with Human Isolates

Two hundred sixty-six strains of Klebsiella pneumoniae isolated from natural water sources in geographically diverse areas (Florida, Massachusetts, and Oregon) were analyzed to determine the serotype, biochemical, virulence, and antimicrobial susceptibility differences between these natural strains and human Klebsiella isolates. Sixty of 72 defined serotypes were found among 210 typable strains. Geographic patterns were present, but in general were not pronounced among serotypes. Reactions with 28 biochemical tests showed percentage responses which were very similar to the summaries of primarily human Klebsiella isolates (as reported by Edwards and Ewing, 1972) and that represented diverse geographic sampling. Virulence studies in representative strains showed no geographic variability and little difference from comparable hospital patient-obtained isolates. In contrast to human hospital isolates, strains demonstrated 90% or greater susceptibility to all antibiotics except ampicillin and carbenicillin; and in further contrast, there was little multiple antibiotic resistance beyond that with ampicillin and carbenicillin.     

Occurrence, characterisation and detection of potential virulence determinants of emerging aquatic bacterial pathogens from the Philippines and Thailand

Strains of Aeromonas spp., 'non-cholera vibrios' (NCVs) and Plesiomonas shigelloides isolated from aquatic environments, fish and human diarrhoeal cases in the Philippines and Thailand were characterised for potential virulence markers. Thus, the production of cytotoxin, cell-associated and cell-free haemolysin and their capacity to adhere to human intestinal (Henle 407) cells in vitro was investigated. In addition, the occurrence of tlh and tdh haemolysin genes and urease activity among V. parahaemolyticus strains was investigated. The results showed that strains recovered from clinical sources (human and fish) produced these virulence factors, whereas these are absent in environmental strains.     

Gastrointestinal Colonization Rates for Human Clinical Isolates of <Emphasis Type="Italic">Aeromonas Veronii</Emphasis> Using a Mouse Model

A variety of environment-associated gastrointestinal infections have been associated with the Aeromonas group of bacteria which contain both non-virulent strains as well as virulent strains within a particular species. This study monitors the colonization rates of colon tissue in a mouse-streptomycin dose/response model involving isolates of Aeromonas veronii biovar sobria obtained from human clinical specimens. The ability to successfully colonize mouse colon tissues by the human clinical isolates was then compared with the rates achieved in a previous study of Aeromonas isolates obtained from environmental drinking water samples. Results suggest that strains of Aeromonas isolated from drinking water environmental samples contain pathogenic and virulence capabilities similar to those seen in Aeromonas veronii clinical isolates from human infections.     

Lecithinase activity of Vibrio cholerae and of vibrios not agglutinated by O-cholera serum

The lecithinase activities of 187 V. cholerae strains differing in their virulence and 70 cultures of NAG vibrios of different origin varied in all experimental groups. The level of lecithinase activity in the groups of V. cholerae strains having low virulence, or no virulence at all, was considerably higher than in virulent strains. NAG vibrios isolated from diarrhea patients and from samples of lake and pond water did not differ in the activity of this enzyme.     

The role of capsular antigens in serum resistance and in vivo virulence of Escherichia coli

Abstract To identify critical microbial determinant(s) in serum resistance and in vivo virulence, 5 strains of Escherichia coli with different combinations of K and O types were examined along with their unencapsulated mutants. All 3 E. coli strains possessing the K1 capsule were considerably more resistant to normal human serum in vitro and more virulent in newborn rats than their isogenic mutants lacking the K1 capsule. In contrast, two K5 encapsulated strains and their unencapsulated mutants were essentially the same for their degree of serum resistance and in vivo virulence. These findings suggest that for the demonstration of serum resistance and in vivo virulence, a capsule is required for E. coli strains encapsulated with K1, whereas certain E. coli strains encapsulated with K5 may not need a capsule.     

Host- and tissue-specific pathogenic traits of Staphylococcus aureus

Comparative genomics were used to assess genetic differences between Staphylococcus aureus strains derived from infected animals versus colonized or infected humans. A total of 77 veterinary isolates were genetically characterized by high-throughput amplified fragment length polymorphism (AFLP). Bacterial genotypes were introduced in a large AFLP database containing similar information for 1,056 human S. aureus strains. All S. aureus strains isolated from animals in close contact with humans (e.g., pet animals) were predominantly classified in one of the five main clusters of the AFLP database (cluster I). In essence, mastitis-associated strains from animals were categorized separately (cluster IVa) and cosegregated with bacteremia-associated strains from humans. Distribution of only 2 out of 10 different virulence genes differed across the clusters. The gene encoding the toxic shock syndrome protein (tst) was more often encountered among veterinary strains (P < 0.0001) and even more in the mastitis-related strains (P<0.0001) compared to human isolate results. The gene encoding the collagen binding protein (cna) was rarely detected among invasive human strains. The virulence potential, as indicated by the number of virulence genes per strain, did not differ significantly between the human- and animal-related strains. Our data show that invasive infections in pets and humans are usually due to S. aureus strains with the same genetic background. Mastitis-associated S. aureus isolated in diverse farm animal species form a distinct genetic cluster, characterized by an overrepresentation of the toxic shock syndrome toxin superantigen-encoding gene.     

Niche-specific association of Aeromonas Ribotypes from human and environmental origin

A total of 88 Aeromonas isolates from distinct locations and sources (39 from extraintestinal infections, 31 from diarrhoeic, ten from non-diarrhoeic faeces, all human, and eight from fresh water) were subjected to phenospecies identification, serotyping, ribotyping and detection of some virulence markers. The strains belonged to four different phenospecies marked by 19 O serogroups and 38 ribotypes. No strong correlation between these parameters was found, and no group, as defined by the typing methods, could be characterized with a particular set of virulence markers. There was a clear association of ribotypes with the source of the strains. Cluster analysis allowed the identification of a complex of ribotypes belonging to distinct but related sources, including clinical and environmental isolates. These results suggest that ribotyping may be an epidemiological tool suitable for the study of Aeromonas infections.     

禽致病性大肠杆菌IMT5155 疑似毒力基因在人源及动物源大肠杆菌中的分布

[目的]检测禽致病性大肠杆菌IMT5155自分泌黏附素基因等具有代表性的疑似毒力基因在不同来源大肠杆菌中的分布,为进一步研究其致病机理提供依据.[方法]采用PCR和Dot blot,检测疑似毒力基因在不同地区(101株大肠杆菌中国分离株和121株大肠杆菌德国分离株)、不同来源(人源、禽源及猪源)大肠杆菌中的分布,并分析其和大肠杆菌系统进化分群的关系.[结果]自分泌黏附素基因B11等11个疑似毒力基因在禽致病性大肠杆菌中分布率较高,阳性率分别为:A1 36.4%(32/88)、A8 53.4%(47/88)、A1063.6%(56/88)、B1137.5%(33/88)、F3 59.1%(52/88)等,且疑似毒力基因主要存在于大肠杆菌B2进化群中.值得注意的是,D1、E9和F11基因片段在新生儿脑膜炎大肠杆菌中有较高的分布率,分别为60%(6/10)、80%(8/10)和90%(9/10),而在新生儿脑膜炎大肠杆菌中未检测到B11基因.[结论]自分泌黏附素B11等疑似毒力基因与禽致病性大肠杆菌关系密切,但疑似毒力基因D1、E9和F11与新生儿脑膜炎大肠杆菌密切相关,提示禽致病性大肠杆菌可能是新生儿脑膜炎大肠杆菌的毒力基因储库.     

Evidence for Coexistence of Distinct Escherichia coli Populations in Various Aquatic Environments and Their Survival in Estuary Water

Escherichia coli, a commensal bacterium from the intestinal tracts of humans and vertebrate animals, has been used as one of two bacterial indicators of fecal contamination, along with intestinal enterococci, to monitor the microbiological quality of water. However, water environments are now recognized as a secondary habitat where some strains can survive. We investigated the survival of E. coli isolates collected from bodies of water in France exhibiting distinct profiles of contamination, defined according to the following criteria: vicinity of the point sources of contamination, land use, hydrology, and physicochemical characteristics of the receiving water. We selected 88 E. coli strains among a collection of 352 strains to carry out a microcosm experiment in filtered estuarine water for 14 days at 10°C. The relationship between the survival of E. coli strains and genotypic and phenotypic characteristics was analyzed. This work showed that distinct E. coli survival types, able to survive from between 7 and 14 days to less than 2 days, coexisted in the water. E. coli isolates that rapidly lost their culturability were more frequently isolated in water recently contaminated by fecal bacteria of human origin, and most were multiresistant to antibiotics and harbored several virulence factors. In contrast, persistent strains able to survive from 4 to 14 days were more often found in water with low levels of fecal bacteria, belonged mainly to the B1 phylogroup, often harbored only one virulence factor, kspE or ompT, and were able to grow at 7°C.     

Intracellular vacuolation induced by culture filtrates of Plesiomonas shigelloides isolated from environmental sources

AIMS: Potential virulence factors produced by culture filtrates of Plesiomonas shigelloides isolated from water were investigated. METHODS AND RESULTS: Culture filtrates of P. shigelloides strains were assayed for cytotoxic activity in CHO (Chinese hamster ovary), Vero (African green monkey kidney), HeLa (human cervix), HT29 (human epithelial intestinal) and SK6 (swine epithelial kidney) cells. Microscopic analyses revealed intensive cytoplasmic vacuolation including cell rounding and swelling, with gradual destruction of the monolayer in filtrate-treated cells. Neutral red assays showed that CHO, HeLa and Vero cells were the most sensitive to the vacuolating activity, which was evident within 30 min of culture filtrate exposure. This activity was inactived by heating at 56 degrees C for 15 min and partially neutralized by antiserum to the cytotoxin of Aeromonas hydrophila. All P. shigelloides strains had a cell-associated haemolysin in the agar plate assay. Three isolates were found to produce a cell-free haemolytic activity at 37 degrees C. In the suckling mouse test, two P. shigelloides culture supernatants were positive for enterotoxic activity. CONCLUSIONS: P. shigelloides culture filtrates isolated from aquatic environment cause intracellular vacuolation on mammalian cells, and produce haemolytic and enterotoxic activities. SIGNIFICANCE AND IMPACT OF THE STUDY: This work revealed the presence of putative virulence factors that could be associated with human infections involving Plesiomonas strains.