PubMed Health⌕ Search

Biomedical subjects

J Hacker

Publications and source records attributed to J Hacker.

At least 217 records · Page 12Linked to original sources
PubMed1987

Comparison of the genetic determinant coding for the S-fimbrial adhesin (sfa) of Escherichia coli to other chromosomally encoded fimbrial determinants.

DNA probes specific for different regions of the S-fimbrial adhesin (sfa) determinant were constructed and hybridized with DNA sequences coding for P (F8 and F13), mannose-sensitive hemagglutinating type 1 (F1A), and F1C fimbriae. While the sfa and F1C DNA determinants exhibited homology along their entire lengths, the P-fimbrial and type 1-fimbrial determinants exhibited homology to regions of the sfa cluster responsible for the control of transcription and, to a minor extent, to regions coding for proteins involved in biogenesis and/or adhesion of the fimbriae and for the N-terminal part of the fimbrillin subunit.

Adhesins, Escherichia coli↗
PubMed1986

Influence of cloned Escherichia coli hemolysin genes, S-fimbriae and serum resistance on pathogenicity in different animal models.

The virulence of the uropathogenic E. coli strain 536 (O6:K15:H31) which produces the S-fimbrial adhesin (Sfa+), is serum-resistant (Sre+) and hemolytic (Hly+) and its derivatives were assessed in five different animal models. Cloned hemolysin (hly) determinants from the chromosomes of O6, O18 and O75 E. coli strains and from the plasmid pHly152 were introduced into the spontaneous Sfa-, Sre-, Hly- mutant 536-21 and its Sfa+, Sre+, Hly- variant 536-31. As already demonstrated for the 536-21 strains (Infect. Immun. 42: 57-63) the O18-hly determinant but not the plasmid-encoded hly determinant of pHly152 transformed into 536-31 contribute to lethality in a mouse peritonitis model. Similar results were obtained with both Hly- host strains and their Hly+ transformants in a chicken embryo test and in a mouse nephropathogenicity assay in which the renal bacterial counts were measured 15 min to 8 hours after i.v. infection. S-fimbriae and serum resistance had only a marginal influence in these three in vivo systems. In contrast all three factors, S-fimbriae, serum resistance and hemolysin, were necessary for full virulence in a respiratory mouse infection assay. In a subcutaneously-induced sepsis model in the mouse restoration of S-fimbriae and serum resistance and separately chromosomally-encoded hemolysis increased virulence to a level comparable to that of the parental 536 strain.

Animals↗
PubMed1986

Characterization of Escherichia coli wild-type strains by means of agglutination with antisera raised against cloned P-, S-, and MS-fimbriae antigens, hemagglutination, serotyping and hemolysin production.

E. coli strains isolated from patients with urinary tract infections (UTI) very often possess mannose-sensitive (MS) and mannose-resistant (MR) adherence factors (fimbriae). According to their receptor specificity the mannose-resistant adhesins can be divided into several types, P, S, M and X. We have cloned the determinants of three groups of UTI E. coli adhesins, MS, P and S, and prepared specific antisera against the fimbriae antigens. 189 hemagglutination (HA+)-positive strains, 96 fecal isolates and 93 strains isolated from UTI have been tested with these specific antisera and further characterized by receptor specific HA, HA patterns and further of the "common O serogroups" 01, 02, 04, 06, 07, 08, 018, 025, 075, most prevalent in UTI, and hemolysin production. 68 (73%) of the UTI strains and 50 (52%) of the fecal isolates showed P-receptor specificity; 16 (17%) of the uropathogenic bacteria and 33 (34%) of the fecal strains exhibited S, M or X-fimbriae antigens. 24% of the P-hemagglutinating (P+) strains reacted with P (F8)-specific antiserum. In contrast, more than three quarter of the S+-strains were agglutinated by S-specific antiserum. HA-pattern VI and 018 antigen were found to be associated with P-fimbriae strains, whereas HA-pattern V and VII and the O antigens 02 (M-type), 06 and 018 (S-type) occurred most frequently in P--strains. A high percentage of P-fimbriated strains showed mannose-sensitive hemagglutination and hemolysin production.

Adhesiveness↗
PubMed1986

Binding of Escherichia coli S fimbriae to human kidney epithelium.

Purified S fimbriae and an Escherichia coli strain carrying the recombinant plasmid pANN801-4 that encodes S fimbriae were tested for adhesion to frozen sections of human kidney. The fimbriae and the bacteria bound to the same tissue domains, and in both cases the binding was specifically inhibited by the receptor analog of S fimbria, sialyl(alpha 2-3)lactose. S fimbriae bound specifically to the epithelial elements in the kidneys; to the epithelial cells of proximal and distal tubules as well as of the collecting ducts and to the visceral and parietal glomerular epithelium. In addition, they bound to the vascular endothelium of glomeruli and of the renal interstitium. No binding to connective tissue elements was observed. The results suggest that the biological function of S fimbriae is to mediate the adhesion of E. coli to human epithelial and vascular endothelial cells.

Adenocarcinoma↗
PubMed1986

Analysis of the genetic determinants coding for the S-fimbrial adhesin (sfa) in different Escherichia coli strains causing meningitis or urinary tract infections.

Recently we have described the molecular cloning of the genetic determinant coding for the S-fimbrial adhesin (Sfa), a sialic acid-recognizing pilus frequently found among extraintestinal Escherichia coli isolates. Fimbriae from the resulting Sfa+ E. coli K-12 clone were isolated, and an Sfa-specific antiserum was prepared. Western blots indicate that S fimbriae isolated from different uropathogenic and meningitis-associated E. coli strains, including O83:K1 isolates, were serologically related. The Sfa-specific antibodies did not cross-react with P fimbriae, but did cross-react with F1C fimbriae. Furthermore the sfa+ recombinant DNAs and some cloned sfa-flanking regions were used as probes in Southern experiments. Chromosomal DNAs isolated from O18:K1 and O83:K1 meningitis strains with and without S fimbriae and from uropathogenic O6:K+ strains were hybridized against these sfa-specific probes. Only one copy of the sfa determinant was identified on the chromosome of these strains. No sfa-specific sequences were observed on the chromosome of E. coli K-12 strains and an O7:K1 isolate. With the exception of small alterations in the sfa-coding region the genetic determinants for S fimbriae were identical in uropathogenic O6:K+ and meningitis O18:K1 and O83:K1 strains. The sfa determinant was also detected on the chromosome of K1 isolates with an Sfa-negative phenotype, and specific cross-hybridization signals were visible after blotting against F1C-specific DNA. In addition homology among the different strains was observed in the sfa-flanking regions.

Adhesins, Escherichia coli↗
PubMed1986

Contribution of cloned virulence factors from uropathogenic Escherichia coli strains to nephropathogenicity in an experimental rat pyelonephritis model.

Escherichia coli 536 (O6:K15:H31), which was isolated from a case of urinary tract infection, determines high nephropathogenicity in a rat pyelonephritis system as measured by renal bacterial counts 7 days after infection. The loss of S fimbrial adhesin formation (Sfa-) (mannose-resistant hemagglutination [Mrh-] and fimbria production [Fim-]), serum resistance (Sre-), and hemolysin production (Hly-) in the mutant 536-21 led to a dramatic reduction of bacterial counts from almost 10(5) to only 40 cells per g of kidney. The reintroduction of the cloned S fimbrial adhesin determinant (sfa) increases the virulence of the avirulent mutant strain by a factor of 20; almost the same effect was observed after restoration of serum resistance by integration of an sfa+ recombinant cosmid into the chromosome. Additional reintroduction of the Hly+ phenotype by transformation of two hly determinants increased the virulence of the strains. Hemolysin production determined increased renal elimination of leukocytes and erythrocytes. Thus all three determinants investigated, S fimbriae, serum resistance, and hemolysin, contribute to the multifactorial phenomenon of E. coli nephropathogenicity.

Adhesins, Escherichia coli↗
PubMed1986

Role of Escherichia coli alpha-hemolysin and bacterial adherence in infection: requirement for release of inflammatory mediators from granulocytes and mast cells.

We investigated the role of bacterial mannose-resistant fimbriation of S fimbriae (Fim), mannose-resistant hemagglutination (S-Mrh), and hemolysin (Hly) production by an Escherichia coli parent and genetically cloned strains as regards their effect on histamine release from rat mast cells and generation of the chemiluminescence response, leukotriene, and enzyme release from human polymorphonuclear granulocytes. These mediators are involved in the induction of inflammatory disease processes and lead, e.g., to the enhancement of vascular permeability, chemotaxis, aggregation of granulocytes (leukotriene B4), lysosomal enzyme release, and smooth-muscle contraction (leukotrienes C4, D4, and E4). The content of azurophilic and specific granules in polymorphonuclear granulocytes consists of highly reactive enzymes which amplify inflammatory reactions. Washed bacteria (E. coli 764 Hly+/-, E. coli 21085 Hly+/- Fim+/- Mrh+/-), as well as their culture supernatants, were analyzed at various times during their growth cycle. No differences exist between parent and cloned or mutant strains with respect to their outer membrane proteins and lipopolysaccharide pattern. Washed bacteria [E. coli 764 and 21085(pANN202-312)] which produced hemolysin, unlike Hly- strains, induced high levels of histamine release from rat mast cells and led to a significant chemiluminescence response and enzyme and leukotriene release from human polymorphonuclear granulocytes. Bacterial culture supernatants from Hly+ and secreting strains showed similar results with the exception of E. coli 21085(pANN202-312), which is a hemolysin-producing but not a secretory strain. Our data suggest a potent role for hemolysin as a stimulus for noncytotoxic mediator release from various cells. Furthermore, we showed that the presence of Fim and S Mrh potentiates mediator release. The simultaneous presence of Mrh and Fim [E. coli 535/21(pANN801-4)] increased mediator release compared with Mrh+ Fim- strains [E. coli 536/21(pANN801-1)]. E. coli 536/21 (Msh- Mrh- Fim- Hly-) did not induce mediator release.

Arachidonic Acid↗
PubMed1986

Large, unstable inserts in the chromosome affect virulence properties of uropathogenic Escherichia coli O6 strain 536.

The hemolytic, uropathogenic Escherichia coli 536 (O6:K15:H31) contains two inserts in its chromosome (insert I and insert II), both of which carried hly genes, were rather unstable, and were deleted spontaneously with a frequency of 10(-3) to 10(-4). These inserts were not found in the chromosome of two nonhemolytic E. coli strains, whereas the chromosomal sequences adjacent to these inserts appeared to be again homologous in the uropathogenic and two other E. coli strains. Insert I was 75 kilobases in size and was flanked at both ends by 16 base pairs (bp) (TTCGACTCCTGTGATC) which were arranged in direct orientation. For insert I it was demonstrated that deletion occurred by recombination between the two 16-bp flanking sequences, since mutants lacking this insert still carried a single copy of the 16-bp sequence in the chromosome. Both inserts contained a functional hemolysin determinant. However, the loss of the inserts not only affected the hemolytic phenotype but led to a considerable reduction in serum resistance and the loss of mannose-resistant hemagglutination, caused by the presence of S-type fimbriae (sfa). It is shown that the Sfa-negative phenotype is due to a block in transcription of the sfa genes. Mutants of strain 536 which lacked both inserts were entirely avirulent when tested in several animal model systems.

Adhesins, Escherichia coli↗
PubMed1986

Effect of dextrose-crystalloid priming solution on fluid requirements and urine output during cardiopulmonary bypass.

We examined the influence of the addition of dextrose to crystalloid cardiopulmonary bypass priming solution. Ten patients received only lactated Ringer's solution during the perioperative period and as their cardiopulmonary bypass priming solution, while ten others, managed identically in all other respects, received only 5% dextrose in lactated Ringer's solution (D5LR). During cardiopulmonary bypass, patients who did not receive glucose required more supplementary fluid (20.0 vs 2.2 ml Kg(-1) hr(-1)) to maintain adequate flow rates, but made less urine (1.1 vs 2.8 ml Kg(-1)) than their glucose-treated cohorts. Postoperative hemoglobin was significantly lower in the group receiving lactated Ringer's solution (-14%) but not in the patients receiving D5LR (-6%). We conclude that the addition of dextrose to a crystalloid priming solution decreases intraoperative fluid requirements and helps restore precardiopulmonary bypass hemoglobin without the need for diuretics or blood products.

Journal Article↗
PubMed1985

Analysis of the flanking regions from different haemolysin determinants of Escherichia coli.

The haemolysin (hly) determinant of the plasmid pHly152 contains an IS2 element at 469 bp upstream of the hlyC gene. The sequence at the other (right-hand) end (RS) also shows multiple hybridization with the plasmid pHly152 and the chromosome of some Escherichia coli strains but the nucleotide sequence of this region does not reveal the typical properties of an IS element. Similar arrangements in the regions flanking the hly determinant are also found on various Hly plasmids from uropathogenic E. coli strains. Chromosomal hly determinants lack both flanking sequences (IS2 and RS) in the immediate vicinity of the hly genes. The sequences immediately upstream of the hlyC gene have been determined from several chromosomal hly determinants and compared with the corresponding sequence of the hly determinant of the plasmid pHly152. We show that these sequences, which contain one promoter (left promoter, phlyL) in all hly determinants tested, vary considerably although common sequence elements can still be identified. In contrast, only relatively few nucleotide exchanges have been detected in the adjacent structural hlyC genes. The A + T content of the 200 bp sequence upstream of hlyC is very high (72 mol% A + T) but even the structural hly genes show a considerably higher A + T content (about 60 mol%) than the E. coli chromosome on average (50 mol% A + T) suggesting that the hly determinant may not have originated in E. coli.

Amino Acid Sequence↗
PubMed1985

Cloning and characterization of genes involved in production of mannose-resistant, neuraminidase-susceptible (X) fimbriae from a uropathogenic O6:K15:H31 Escherichia coli strain.

The uropathogenic Escherichia coli strain 536 (O6:K15:H31) exhibits a mannose-resistant hemagglutination phenotype (Mrh) with bovine erythrocytes and delayed Mrh with human and guinea pig erythrocytes. Neuraminidase treatment of the erythrocytes abolishes mannose resistant hemagglutination, which is typical for X fimbriae. E. coli strain 536 synthesizes two different fimbriae (Fim phenotype) protein subunits, 16.5 and 22 kilodaltons in size. In addition the strain shows mannose-sensitive hemagglutination and common type I (F1) fimbriae. The cosmid clone E. coli K-12(pANN801) and another nine independently isolated Mrh+ cosmid clones derived from a cosmid gene bank of strain 536 express the 16.5-kilodalton protein band, but not the 22-kilodalton protein, indicating an association of the Mrh+ property with the "16.5-kilodalton fimbriae." All cosmid clones were fimbriated, and they reacted with antiserum produced against Mrh+ fimbriae of the E. coli strain HB101(pANN801) and lacked mannose-sensitive hemagglutination (F1) fimbriae. From the Mrh fim cosmid DNA pANN801, several subclones coding for hemagglutination and X fimbriae were constructed. Subclones that express both hemagglutination and fimbriae and subclones that only code for the hemagglutination antigen were isolated; subclones that only produce fimbriae were not detected. By transposon Tn5 mutagenesis we demonstrated that about 6.5 kilobases of DNA is required for the Mrh+ Fim+ phenotype, and the 1.5- to 2-kilobase DNA region coding for the structural protein of the fimbriae has been mapped adjacent to the region responsible for the Mrh+ phenotype. Two different regions can thus be distinguished in the adhesion determinant, one coding for hemagglutination and the other coding for fimbria formation. Transformation of plasmid DNA from these subclones into a Mrh- Fim- mutant of E. coli 536 and into a galE (rough) strain of Salmonella typhimurium yielded transformants that expressed both hemagglutination and fimbria production.

Animals↗
PubMed1985

Bacterial adherence and hemolysin production from Escherichia coli induces histamine and leukotriene release from various cells.

We investigated the role of bacterial adherence and hemolysin production from Escherichia coli parent and genetically cloned strains as to their effects on histamine release from rat mast cells and leukotriene generation from human polymorphonuclear granulocytes. These mediators were involved in the induction of inflammatory disease processes and led, for example, to enhancement of vascular permeability, chemotaxis (leukotriene B4 [LTB4]), chemoaggregation, lysosomal enzyme release, and smooth muscle contraction, (LTC4, LTD4, and LTE4). Washed bacteria (E. coli K-12 MS+ Hly +/-; E. coli 536 MS+ MR +/-) as well as their culture supernatants were analyzed. Washed E. coli K-12 (Hly+), unlike Hly- strains, induced high amounts of histamine release from rat mast cells and chemotactic activity from human polymorphonuclear granulocytes. Significant leukotriene release was obtained with washed E. coli K-12 Hly+ strains and their bacterial culture supernatants. Leukotriene induction was dependent on the amount of hemolysin activity present in the supernatant. However, additional soluble factors should also be considered. The presence of hemolysin appeared to accelerate and enhance the rate of phagocytosis of bacteria by neutrophils. When E. coli 536 (MS+ MR +/- Hly +/-) strains were analyzed, the simultaneous presence of MR+ pili and hemolysin production led to an increase in histamine release as compared with MR- Hly+ strains. The genetically cloned MR+ Hly+ E. coli 536 strain induced higher amounts of leukotrienes as compared with the wild-type strain. Our data suggest a potent role for adhesins and hemolysin as virulence factors in inducing the release of inflammatory mediators.

Adhesiveness↗
PubMed1984

Multiple copies of hemolysin genes and associated sequences in the chromosomes of uropathogenic Escherichia coli strains.

The O6 serogroup Escherichia coli strain 536 carries two hemolysin (hly) determinants integrated into the chromosome. The two hly determinants are not completely identical, either functionally or structurally, as demonstrated by spontaneous deletion mutants carrying only one of them and by cloning each of the two determinants separately into cosmid vectors. Each hly determinant is independently deleted at a frequency of 10(-4), leading to variants which exhibit similar levels of internal hemolysin but different amounts of secreted hemolysin. The two hly determinants were also identified in the O4 E. coli strain 519. The three E. coli strains 251, 764, and 768, which belong to the serogroup O18, and the O4 strain 367 harbor a single chromosomal hly determinant, as demonstrated by hybridization with hly-gene-specific probes. However, a hybridization probe derived from a sequence adjacent to the hlyC-proximal end of the plasmid pHly 152-encoded hly determinant hybridizes with several additional chromosomal bands in hemolytic O18 and O6 E. coli strains and even in E. coli K-12. The size of the probe causing the multiple hybridization suggests a 1,500- to 1,800-base pair sequence directly flanking hlyC. Spontaneous hemolysin-negative mutants were isolated from strains 764 and 768, which had lost the entire hly determinant but retained all copies of the hlyC-associated sequence.2+.

Base Sequence↗
PubMed1983

Hemolysin production as a virulence marker in symptomatic and asymptomatic urinary tract infections caused by Escherichia coli.

Potential virulence, as defined by combined levels of adhesion to urinary epithelial cells, serum resistance, and mouse toxicity, was assessed for Escherichia coli strains causing symptomatic and asymptomatic urinary tract infections in relation to the carriage of hemolysin and other suspected virulence determinants. Hemolysin production (Hly), associated with certain O (O4, O6, O18, and O75), K (5), and hemagglutination (VI and VII) antigenic types but not colicin V production (Cva), was evident in 83 and 60% of isolates in groups possessing high potential virulence and in only 11 and 6% of those with low virulence. Strains of particular O-types were not more virulent per se, but among the serotypes, specific combinations of virulence factors appeared decisive, e.g., O18 HAVI B/D/G Hly(+) K5(+/-) and O18 HAIII/IVB/V Hly(-) Cva(+/-) K1(+/-) strains were, respectively, of high and low potential virulence. Isolates with high potential virulence were found to a similar extent in symptomatic and asymptomatic infections.

Colicins↗
PubMed1983

Cloned hemolysin genes from Escherichia coli that cause urinary tract infection determine different levels of toxicity in mice.

After intraperitoneal injection of mice with Escherichia coli strains isolated from patients with urinary tract infections, the mortality due to hemolytic (Hly+) and nonhemolytic (Hly-) isolates was 77 and 40%, respectively. Deletion of the chromosomal hemolysin (hly) determinant in an E. coli O6:K15:H31 urinary tract infection strain led to a significant reduction in toxicity for mice, and its reintroduction on a recombinant plasmid partially restored the original toxicity. Although introduction of the cloned plasmid pHly152-encoded hly determinant into the Hly- E. coli O6 mutant strain increased toxicity by only a marginal degree, transformation with the cloned chromosomal hly determinants from two E. coli strains of serotypes O18ac:K5:H- and O75:K95:H? resulted in markedly greater toxicity, even exceeding that of the original Hly+ E. coli O6 wild-type strain.

Animals↗