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J M Tomas

Publications and source records attributed to J M Tomas.

18 recordsLinked to original sources
PubMed2005

Identification and characterization of putative virulence genes and gene clusters in Aeromonas hydrophila PPD134/91.

Aeromonas hydrophila is a gram-negative opportunistic pathogen of animals and humans. The pathogenesis of A. hydrophila is multifactorial. Genomic subtraction and markers of genomic islands (GIs) were used to identify putative virulence genes in A. hydrophila PPD134/91. Two rounds of genomic subtraction led to the identification of 22 unique DNA fragments encoding 19 putative virulence factors and seven new open reading frames, which are commonly present in the eight virulence strains examined. In addition, four GIs were found, including O-antigen, capsule, phage-associated, and type III secretion system (TTSS) gene clusters. These putative virulence genes and gene clusters were positioned on a physical map of A. hydrophila PPD134/91 to determine their genetic organization in this bacterium. Further in vivo study of insertion and deletion mutants showed that the TTSS may be one of the important virulence factors in A. hydrophila pathogenesis. Furthermore, deletions of multiple virulence factors such as S-layer, serine protease, and metalloprotease also increased the 50% lethal dose to the same level as the TTSS mutation (about 1 log) in a blue gourami infection model. This observation sheds light on the multifactorial and concerted nature of pathogenicity in A. hydrophila. The large number of putative virulence genes identified in this study will form the basis for further investigation of this emerging pathogen and help to develop effective vaccines, diagnostics, and novel therapeutics.

Aeromonas hydrophila↗
PubMed2004

A type III secretion system is required for Aeromonas hydrophila AH-1 pathogenesis.

Aeromonas hydrophila is a gram-negative opportunistic pathogen in fish and humans. Many bacterial pathogens of animals and plants have been shown to inject anti-host virulence determinants into the hosts via a type III secretion system (TTSS). Degenerate primers based on lcrD family genes that are present in every known TTSS allowed us to locate the TTSS gene cluster in A. hydrophila AH-1. A series of genome walking steps helped in the identification of 25 open reading frames that encode proteins homologous to those in TTSSs in other bacteria. PCR-based analysis showed the presence of lcrD homologs (ascV) in all of the 33 strains of A. hydrophila isolated from various sources. Insertional inactivation of two of the TTSS genes (aopB and aopD) led to decreased cytotoxicity in carp epithelial cells, increased phagocytosis, and reduced virulence in blue gourami. These results show that a TTSS is required for A. hydrophila pathogenesis. This is the first report of sequencing and characterization of TTSS gene clusters from A. hydrophila. The TTSS identified here may help in developing suitable vaccines as well as in further understanding of the pathogenesis of A. hydrophila.

Aeromonas hydrophila↗
PubMed1999

Surface antigen exposure by bismuth dimercaprol suppression of Klebsiella pneumoniae capsular polysaccharide.

The bacterial capsule is an important virulence determinant in animal and plant disease. Bacterial capsule and slime can be inhibited by bismuth compounds, especially when complexed with lipophilic thiol chelators. Bismuth dimercaprol (BisBAL) at 1 ppm of Bi3+ repressed Klebsiella pneumoniae capsule expression in defined medium by nearly 90%, which exposed subsurface structures. The phagocytic index for BisBAL-treated bacteria increased from <10 to 360 bacteria per 100 neutrophils in the presence of complement and anticapsular or anti-O antigen antiserum. BisBAL treatment also enhanced the reactivity of monoclonal antibodies (MAbs) specific for the O1-antigen lipopolysaccharide (LPS) or the LPS core in a dose-dependent manner as indicated by the results of enzyme-linked immunosorbent assays. When anti-O1 MAb was used, the reactivity increased significantly for fully encapsulated O1:K1 or O1:K2 cells but not for O1:K- cells. Deposition of C3b also increased significantly for BisBAL-treated O1:K1 or O1:K2 cells but not for O1:K- cells. Survival of a serum-sensitive strain was <0.1% when nonimmune human serum absorbed with O1:K1 cells was used and 107% when BisBAL-treated cells were used for absorption. Outer membrane proteins were also more accessible on the surface of K. pneumoniae after BisBAL treatment. Thus, at subinhibitory levels, BisBAL inhibited capsule expression, which promoted phagocytosis, enhanced the reactivity of specific antibodies for LPS O antigen, LPS core epitopes, or outer-membrane proteins, and enhanced complement interaction with encapsulated K. pneumoniae. By unmasking bacterial surface structures and enhancing the immune system reactivity to bacteria, bismuth thiols may prove useful as adjuncts for vaccination.

Animals↗
PubMed1997

Influence of osmolarity on lipopolysaccharides and virulence of Aeromonas hydrophila serotype O:34 strains grown at 37 degrees C.

Growth of Aeromonas hydrophila serotype O:34 strains at 37 degrees C at low and high osmolarity resulted in changes in the lipopolysaccharide (LPS) and virulence of the strains tested. We previously described the effect of growth temperature on LPS and virulence of these strains (S. Merino et al., Infect. Immun. 60:4343-4349, 1992). The effect of osmolarity can be observed when the cells grow at 37 degrees C but not when they grow at 20 degrees C. Purified LPS from cells cultivated at 37 degrees C and high osmolarity was smooth, while the LPS extracted from the cells cultivated at low osmolarity was rough. Furthermore, the strains were more virulent for fish and mice when they were grown at high osmolarity than when they were grown at low osmolarity and also showed increased extracellular activities when they were grown at high osmolarity. Finally, cells grown at high osmolarity showed better adhesion to HEp-2 cells than the same cells grown at low osmolarity, and furthermore the cells grown at high osmolarity were resistant to the bactericidal activity of nonimmune serum, while the same cells grown at low osmolarity were sensitive.

Aeromonas hydrophila↗
PubMed1996

Mesophilic Aeromonas sp. serogroup O:11 resistance to complement-mediated killing.

The complement activation by and resistance to complement-mediated killing of Aeromonas sp. strains from serogroup O:11 were investigated by using different wild-type strains (with an S-layer characteristic of this serogroup) and their isogenic mutants characterized for their surface components (S-layer and lipopolysaccharide [LPS]). All of the Aeromonas sp. serogroup O:11 wild-type strains are unable to activate complement, which suggested that the S-layer completely covered the LPS molecules. We found that the classical complement pathway is involved in serum killing of susceptible Aeromonas sp. mutant strains of serogroup O11, while the alternative complement pathway seems not to be involved, and that the complement activation seems to be independent of antibody. The smooth mutant strains devoid of the S-layer (S-layer isogenic mutants) or isogenic LPS mutant strains with a complete or rather complete LPS core (also without the S-layer) are able to activate complement but are resistant to complement-mediated killing. The reasons for this resistance are that C3b is rapidly degraded, and therefore the lytic membrane attack complex (C5b-9) is not formed. Isogenic LPS rough mutants with an incomplete LPS core are serum sensitive because they bind more C3b than the resistant strains, the C3b is not completely degraded, and therefore the lytic complex (C5b-9) is formed.

Aeromonas↗
PubMed1996

Cloning and characterization of two Serratia marcescens genes involved in core lipopolysaccharide biosynthesis.

Bacteriocin 28b from Serratia marcescens binds to Escherichia coli outer membrane proteins OmpA and OmpF and to lipopolysaccharide (LPS) core (J. Enfedaque, S. Ferrer, J. F. Guasch, J. Tomás, and M. Requé, Can. J. Microbiol. 42:19-26, 1996). A cosmid-based genomic library of S. marcescens was introduced into E. coli NM554, and clones were screened for bacteriocin 28b resistance phenotype. One clone conferring resistance to bacteriocin 28b and showing an altered LPS core mobility in polyacrylamide gel electrophoresis was found. Southern blot experiments using DNA fragments containing E. coli rfa genes as probes suggested that the recombinant cosmid contained S. marcescens genes involved in LPS core biosynthesis. Subcloning, isolation of subclones and Tn5tac1 insertion mutants, and sequencing allowed identification of two apparently cotranscribed genes. The deduced amino acid sequence from the upstream gene showed 80% amino acid identity to the KdtA protein from E. coli, suggesting that this gene codes for the 3-deoxy-manno-octulosonic acid transferase of S. marcescens. The downstream gene (kdtX) codes for a protein showing 20% amino acid identity to the Haemophilus influenzae kdtB gene product. The S. marcescens KdtX protein is unrelated to the KdtB protein of E. coli K-12. Expression of the kdtX gene from S. marcescens in E. coli confers resistance to bacteriocin 28b.

Amino Acid Sequence↗
PubMed1995

A survey of bacterial toxins involved in food poisoning: a suggestion for bacterial food poisoning toxin nomenclature.

There is at present no accepted nomenclature for bacterial protein toxins, although there have been several attempts at dividing them into groups by their mode of action. In this paper we will not try to describe all known bacterial protein toxins, but concentrate on the toxins involved in food poisoning. Although most of these toxins are enterotoxins (protein exotoxins with the site of action on the mucosal cells of the intestinal tract) there are also other toxins involved in food poisoning, like the neurotoxins. In Table 1 the most important food pathogens in Europe are listed. For most, but not all, of these food pathogens, toxins are virulence factors. Generally, we divide food poisoning into infections and intoxications, where Salmonella spp. and Shigella spp. are typical examples of infections and Clostridium botulinum and Staphylococcus aureus for intoxications. We consider it better to make four different groups of food pathogenic bacteria, according to Table 2. Today the first three groups are all defined as infections, although for both group 2 and 3 the bacterium itself does not harm the host directly. The bacterium in such locations is like an 'enterotoxin factory'. The bacteria belonging to group 3 do not even interact with the epithelial cells in the intestine, while the bacteria of group 2 must colonise the epithelial cells prior to enterotoxin production.

Bacillaceae↗
PubMed1995

Emerging pathogens: Aeromonas spp.

Aeromonas spp. are Gram-negative rods of the family Vibrionaceae. They are normal water inhabitants and are part of the regular flora of poiquilotherm and homeotherm animals. They can be isolated from many foodstuffs (green vegetables, raw milk, ice cream, meat and seafood). Mesophilic Aeromonas spp. have been classified following the AeroKey II system (Altwegg et al., 1990; Carnahan et al., 1991). The major human diseases caused by Aeromonas spp. can be classified in two major groups: septicemia (mainly by strains of A. veronii subsp. sobria and A. hydrophila), and gastroenteritis (any mesophilic Aeromonas spp. but principally A. hydrophila and A. veronii). Most epidemiological studies have shown Aeromonas spp. in stools to be more often associated with diarrhea than with the carrier state; an association with the consumption of untreated water was also conspicuous. Acute self-limited diarrhea is more frequent in young children, in older patients chronic enterocolitis may also be observed. Fever, vomiting, and fecal leukocytes or erythrocytes (colitis) may be present (Janda, 1991). The main putative virulence factors are: exotoxins, endotoxin (LPS), presence of S-layers, fimbriae or adhesins and the capacity to form capsules.

Aeromonas↗
PubMed1992

Modulation of surface antigen expression by Klebsiella pneumoniae in response to growth environment.

Growth in pooled human body fluids [urine, serum and peritoneal dialysate (HPD)] modulated the expression of cell envelope antigens in virulent (serotype O1:K1) and avirulent (serotype O1:K66) Klebsiella pneumoniae strains. Marked variations in the outer membrane protein (OMP) and lipopolysaccharide (LPS) profiles were noted when broth-grown cells were compared with those of bacteria cultured in body fluids. In particular, for the O1:K1 serotype strain, growth in the latter resulted in: (a) the expression of at least five iron-regulated OMPs in the 74-87 kDa range, the pattern of which was medium dependent; (b) alterations in the migration of the LPS core polysaccharide; and (c) the reversion of isogenic O-:K+ and O-:K- mutants to the O+ phenotype after growth in fresh serum but not in heat-inactivated serum, urine or HPD. Similar results were obtained for the O1:K66 serotype, although no variation in the migration of the LPS core was noted. For both O1:K1 and O1:K66 serotypes, neither the surface exposure of O1 serotype LPS nor the production of K-antigen (capsular polysaccharide) was affected by growth in body fluids. No reversion of K- mutants to the K+ phenotype was observed. These data illustrate the phenotype flexibility of this opportunistic pathogen and emphasise the crucial role of the O- rather than the K-antigen in protecting K. pneumoniae from complement-mediated serum killing.

Antigens, Bacterial↗
PubMed1991

National substance abuse epidemiology initiatives in the United States: what works for what.

Data concerning the epidemiology of drug abuse in the United States are presented in this paper. The paper includes an integrating approach from various data sources: national, state, and local level; surveillance systems; and outbreak investigations. An analysis of trends and patterns of drug abuse are discussed, both from the perspective of historical development and the current situation. Analyses focus on changes in levels of use for different drugs as well as changes in user characteristics and consequences of use. Current studies concerning the epidemiology of drug abuse in special populations and findings from recent field investigations are presented. Future research directions in the field of epidemiology from a national perspective are outlined.

Epidemiologic Methods↗
PubMed1990

Influence of lipopolysaccharide chemotype on the interaction between Klebsiella pneumoniae and human polymorphonuclear leucocytes.

A series of isogenic mutants lacking either the O1 (O-:K66) or K66 (O1:K-) antigens or both (O-:K-), some of which had additional defects in their LPS core polysaccharide was used to examine the interaction between polymorphonuclear leucocytes (PMNLs) and K. pneumoniae serotype O1:K66. In the absence of serum complement, only a O-:K- strain with a deep rough LPS chemotype elicited a PMNL-dependent chemiluminescent (CL) response. However, following opsonization of the non-capsulated strains by complement, the largest CL response was to the O1:K- mutant. This mutant also activated and bound more complement C3 than any of the other encapsulated or non-capsulated strains examined. Despite the surface exposure of smooth and rough LPS in the encapsulated parent and mutant strains, the K66 antigen reduced the binding of C3 and prevented PMNL activation. Both anti-LPS and anti-K66 antibodies, however, stimulated a PMNL-dependent CL response to the K66 bearing strains.

Antibodies, Bacterial↗
PubMed1990

Association between intravenous drug use and early misbehavior.

This investigation uses an epidemiologic strategy to examine a suspected association between misbehavior in early life and subsequent involvement in intravenous drug use. The 222 cases of i.v. drug use in this study were a subset of all persons recruited for a continuing study of drug use, HIV-1 infection, and the natural history of AIDS. It was possible to match these 222 i.v. drug users to 588 subjects from the NIMH Epidemiologic Catchment Area survey sample, for a total of 810 subjects in 60 matched sets. The matching factors were gender and census tract of residence. Conditional logistic regression models were used to estimate the degree of association between early misbehavior and i.v. drug use. The results showed a strong association between i.v. drug use and early misbehavior. For every unit increase in misbehavior score, the odds of being an i.v. drug user increased by a factor of 1.74 (P less than 0.001). Compared to subjects with low misbehavior scores, those with high scores were 24.67 times more likely to be i.v. drug users (P less than 0.001). Results of this study add to evidence now supporting field trials of public health strategies to modify the link between misbehavior in early life and later illicit drug use.

Adolescent↗
PubMed1989

Novel aerobactin receptor in Klebsiella pneumoniae.

Several Klebsiella pneumoniae strains which produced enterochelin but not aerobactin were nevertheless sensitive to cloacin DF13. In contrast, a strain of serotype K1:O1 which produced both siderophores was cloacin-resistant. Loss by mutation of the O1 but not K1 antigen rendered this strain cloacin-sensitive, indicating that the O1 antigen prevented access of cloacin to the cloacin/aerobactin receptor. Unlike the K1:O1 strain, the aerobactin-negative strains failed to hybridize in a colony blot assay with an aerobactin receptor gene probe prepared from pColV-K30. However, antisera raised against the 74 kDa pColV-K30 aerobactin receptor cross-reacted with a 76 kDa outer-membrane protein in each K. pneumoniae strain. In addition to the 76 kDa protein, the K1:O1 strain also produced a strongly cross-reacting 74 kDa protein. To determine whether these aerobactin-negative strains could use aerobactin, mutants unable to synthesize siderophores were isolated. Aerobactin promoted the growth of these mutants in iron-deficient media. The evidence presented suggests that some K. pneumoniae strains produce an aerobactin iron-uptake system without apparent production of aerobactin and which is probably based on a 76 kDa receptor, the gene for which does not hybridize with aerobactin receptor gene encoded on pColV-K30.

Bacterial Outer Membrane Proteins↗
PubMed1988

Surface exposure of the O-antigen in Klebsiella pneumoniae O1:K1 serotype strains.

Isogenic non-encapsulated (K-) mutants (O1:K-) were obtained from several different Klebsiella pneumoniae O1:K1 serotype strains. By employing K. pneumoniae bacteriophages FC3-1, FC3-2, and bacteriophage phi 1, the bacterial surface receptors of which, are the O1-antigen of lipopolysaccharide and the K1 capsular polysaccharide (K1) respectively, the K1 polysaccharide was found to completely cover the O1 lipopolysaccharide molecules in each of the O1:K1 strains examined. Exposure of the O-antigen at the cell surface was only observed after growth in the presence of sub-minimum inhibitory concentrations of antibiotics. The implications of these findings for the design of vaccines for the prevention of Klebsiella infections is discussed.

Antigens, Bacterial↗
PubMed1986

Influence of chelating agents on the toxicity, distribution and excretion of vanadium in mice.

The effects of the chelating agents Na2Ca-ethylendiaminetetraacetate (EDTA), Na3Ca-diethylentriaminepentaacetate (DTPA), L-cysteine, 4,5-dihydroxy-1,3-benzene-disulfonic acid (Tiron) and deferoxamine mesylate and the reducing agent ascorbic acid on the toxicity, excretion and distribution of i.p. injected vanadium were studied in male Swiss mice. Chelating and reducing agents were administered intraperitoneally at doses equal to one-fourth of their respective LD50. To determine the effects of the various chelators on the mortality of vanadium, various doses of NaVO3 (0.30-1.20 mmol kg-1 i.p.) were given, followed immediately by one of the chelating or reducing agents. Survival was recorded at the end of 14 days. Significant increases in survival were noted with ascorbic acid, Tiron and deferoxamine, with ascorbic acid being the most effective. Deferoxamine and Tiron were the most effective in increasing the excretion of vanadium and reducing the concentration of vanadium found in various tissues. However, ascorbic acid appears to be the most effective agent in the prevention of vanadium intoxication.

Animals↗
PubMed1985

Short-term toxicity studies of vanadium in rats.

Vanadium (in the form of NaVO3) was given in drinking water to groups of 10 male Sprague-Dawley rats over a period of 3 months at concentrations of 0, 5, 10 and 50 ppm. Vanadium accumulated dose-dependently in the kidneys and spleen. Appearance, behaviour, food and water consumption, growth and mortality of the treated rats of all groups were not affected during the 3-month period. Histopathological investigation showed only mild though dose-dependent lesions in kidneys and spleen. The plasma concentrations of urea and uric acid were increased in the highest exposure groups.

Animals↗