Yersinia Yop-specific IgA antibodies in Hungarian blood donors.

Sera of 112 healthy Hungarian blood donors were tested for the presence of Yersinia enterocolitica and Y. pseudotuberculosis-specific agglutinins by tube agglutination, and for that of yersinia outer membrane protein (Yop)-specific IgA antibodies by ELISA. The positive results of this latter assay were confirmed by immunoblot. Only one sample gave a positive agglutination reaction with Y. enterocolitica antigen (group 03) and four exhibited an equivocal reaction with Y. pseudotuberculosis antigens (groups II and IV). Contrary to the low incidence of agglutinins, 15.1% of the samples showed a positive Yop-specific IgA reaction, while further 5.3% samples fell into the equivocal range by ELISA (17 and 6 specimens, respectively). Eleven of these samples (9.8% of all specimens tested) were also positive by immunoblot for the presence of Yop-specific IgA antibodies. These data suggest a higher incidence of yersinia infections than the 1.0-1.4 per 10(5) population predicted on the basis of stool culture results.

Specific binding of lactoferrin to Escherichia coli isolated from human intestinal infections.

The degrees of human lactoferrin (HLf) and bovine lactoferrin (BLf) binding in 169 Escherichia coli strains isolated from human intestinal infections, and in an additional 68 strains isolated from healthy individuals, were examined in a 125I-labelled protein binding assay. The binding was expressed as a percentage calculated from the total labelled ligand added to bacteria. The HLf and BLf binding to E. coli was in the range 3.7 to 73.4% and 4.8 to 61.6%, respectively. Enterotoxigenic strains demonstrated a significantly higher HLf binding (median = 19%) than enteropathogenic, enteroinvasive, enterohaemorrhagic strains or normal intestinal E. coli isolates (medians 6 to 9). Enteropathogenic strains belonging to serotypes O44 and O127 demonstrated significantly higher HLf binding compared to O26, O55, O111, O119 and O126. No significant differences in the degree of HLf or BLf binding were found between aerobactin-producing and non-producing strains. The interaction was further characterized in a high Lf-binding EPEC strain, E34663 (serotype O127). The binding was stable in the pH range 4.0 to 7.5, did not dissociate in the presence of 2M NaCl or 2M urea, and reached saturation within two h. Unlabelled HLf and BLf displaced the 125I-HLf binding to E34663 in a dose-dependent manner. Apo- and iron-saturated forms of Lf demonstrated similar binding to E34663. Among various unlabelled subepithelial matrix proteins and carbohydrates tested (in 10(4)-fold excess) only fibronectin and fibrinogen caused a moderate inhibition of 125I-HLf binding. According to Scatchard plot analysis, 5,400 HLf-binding sites/cell, with an affinity constant (Ka) of 1.4 x 10(-7) M, were estimated in strain E34663. These data establish the presence of a specific Lf-binding mechanism in E. coli.

Value of surveillance cultures in a bone marrow transplantation unit.

Because of the increased risk of infection with the associated diagnostic and therapeutic problems in bone marrow transplantation (BMT) patients, the usefulness of surveillance cultures (SC) at the BMT department of the National Institute of Haematology, Blood Transfusion, Transplantation and Immunology, Budapest, was reviewed. Between January 1992 and May 1995, 26 BMT operations were performed; 13 patients had 23 febrile espisodes. In 12 of these episodes infection was clinically documented; however, SC of these patients yielded bacteria identical with those in the blood culture in only two episodes (1 and 6 days before their blood cultures became positive, respectively). Out of a total of 1187 samples from these patients, potentially pathogenic bacteria were isolated from 145 SC and 43 blood cultures (drawn on 31 different days). Suppression of the gastrointestinal flora could be achieved by the department's decontamination regimen; however, overgrowth by gram-positive organisms (mainly coagulase-negative staphylococci) occurred in the intestine and at other body sites. On the basis of these results, SC are of limited value in predicting infection or identifying the causative organisms of fever. On the other hand, SC are useful in confirming the efficiency of suppression of the body flora by antimicrobial agents. Specific treatment was based on suitably sampled materials, and close contact between physicians, infectious disease specialists and microbiologists was essential.

Comparison of rapid methods for detection of heat-labile (LT) and heat-stable (ST) enterotoxin in Escherichia coli.

Oxoid VET-RPLA, ST-EIA and Pharmacia Phadebact ETEC-LT enterotoxin tests were compared to find a simple but reliable method for detecting enterotoxigenic Escherichia coli (ETEC) in Hungary. In the Oxoid tests, all six reference LT- or ST-producing strains, except one ST-producer, gave positive results. Of 11 reference porcine enterotoxigenic strains, all four LT-producers gave positive reactions for LT but three of 10 ST-producers gave negative reactions for ST. Thirteen of 50 strains from culture collections of H. Steinrück (Germany) were LT+ and nine of 33 were ST+. When 31 isolates were tested simultaneously with the Oxoid and the Pharmacia LT tests, 12 strains were LT+ by the Oxoid LT test but by the Phadebact LT test only seven of these strains were LT+ and, of the remainder, three gave uncertain results and two gave negative results. Of 69 porcine strains, seven were LT+ and three ST+. Of 901 human strains isolated in Hungary, 10 were LT+ and one of 24 tested was ST+. In two cases, ETEC strains were isolated from contacts of travellers returning from Mongolia and Bangladesh. Results of comparative studies with reference strains corresponded well to those of the classical toxin detection tests. The Oxoid test was rapid, sensitive, specific and easy to perform and is recommended for use in screening ETEC isolates.

[Significance of phenotypical properties in extra-intestinal E. coli infections]. / Az Escherichia coli fenotípusos tulajdonságainak jelentösége az extraintestinalis fertözésekben.

Virulence factors (serogroup, haemolysis, haemagglutination, antigens K1, K5, colicinogenicity) and their association with diseases of 3334 Escherichia coli strains isolated from different clinical specimens between 1979-1990 were analysed. Strains, that were isolated from cerebrospinal fluid of newborns under one month were characterized by certain serogroups (O7, O18, O45, O78, O83), possession of antigen K1 and production of colicin. On the basis of their LD50 they belonged to the pathogenic group (i.e. < 10 10(6)) significantly more frequently than those isolated from other materials. Strains originating from blood cultures belonged frequently to serogroup O4, O6, O18 and were haemolytic. Their pathogenicity was proved by mouse lung toxicity test. Properties of strains isolated from wound and throat swabs, urinary samples resembled to that of strains originating from blood cultures in many respects, expressing the fact that bacteria settle in different organs before sepsis is developing. Frequent occurrence of strains with antigen K1, haemolysin and haemagglutination positivity in vaginal swabs expose newborns to danger. Knowledge of virulence markers and prevention of infections are associated.

Association of human enteric pathogenicity and mouse lung toxicity of Escherichia coli.

Mouse lung toxicity of 439 strains (431 Escherichia coli, 1 Shigella dysenteriae 1, 1 Enterobacter cloacae, 5 Vibrio sp., 1 Klebsiella) was compared to other pathogenicity tests (mouse virulence, enterotoxicity, guinea pig eye test), to serogroup distribution, loss of virulence on storage, origin and haemolytic activity. Mouse lethality was highest in serogroup O4 (p < 0.001), O18a,c (p < 0.001); serogroups O6, O20, O75, O115, O147 were next in order. E. coli serogroups O19, O26, O28a,b, O32, O51, O53, O55, O73, O78, O79, O83, O105, O111, O112, O114, O117, O119, O124, O129, O136, O142 failed to show lung toxicity. Strains O4 and O18 isolated at different periods of time did not differ significantly in the lung test (p = 0.05, p = 0.01, p > 0.1, p = 0.05, p > 0.1). There was no significant difference between strains isolated from the stools of patients with enteritis and of healthy individuals (p = 0.1, p > 0.99) and between those isolated from all faecal specimens and from extraintestinal samples (p = 0.05, p > 0.3). There was no correlation between lung toxicity and other pathogenicity tests. Since strains isolated from healthy individuals were also toxic for mice, a positive lung test cannot be considered a criterion of the aetiological role of the agent.

Virulence factors of Escherichia coli. II. Antigens O4, O6 and O18, haemolysin production and mannose resistant haemagglutinating capacity are closely associated.

Escherichia coli strains isolated from a variety of human samples were examined for alpha haemolysin (Hly) production. A total of 1156 strains was compared for incidence of Hly positivity, and the capacity of mannose resistant haemagglutinating activity of human erythrocytes (MRHA). Incidence of Hly production in serogroups O4, O6 and O18ac was significantly higher than in other ones (70.1% vs. 18.7%), independently of the origin of strains; 78% of Hly+ strains belonging to serogroups O4, O6, O18 was MRHA+, too. The marked correlation between Hly positivity, MRHA activity and these serogroups suggested that in serogroup O4, O18 and in a lesser degree O6, genetic informations concerning O antigen, MRHA and Hly are linked in the chromosome.

Bacteriological proficiency testing in Hungarian clinical microbiology laboratories.

In course of a proficiency testing programme carried out in 1989, a total of 47 clinical microbiology laboratories of public health stations and of hospitals received freeze-dried cultures for isolation, identification and antimicrobial susceptibility testing. The specimens contained bacteria that occur in everyday work, including those that require improved methods of cultivation and identification. Nine public health laboratories and one hospital laboratory achieved excellent results. Good results were attained by 11 public health and 6 hospital laboratories. Four public health and 10 hospital laboratories were on the medium level and 4 hospital laboratories did not reach even this degree. The main failures were due to an insufficient anaerobic cultivation, unreliable identification and negligation of controls for drug susceptibility tests.

Computerized complex typing of Escherichia coli strains from different clinical materials.

A multivariate analysis of 3334 Escherichia coli strains originating from different clinical materials revealed that 50.2% of isolates belonged to the most common 12 (O1, O2, O4, O6, O7, O8, O15, O18, O45, O75, O78, O83) out of 133 serogroups. Haemolysin (Hly) production, mannose resistant haemagglutinating activity for human erythrocytes (MRHA) and colicinogenicity (Col) were recorded in 30, 30 and 36%, respectively. Antigens K1 and K5 were present in 11% and 6.6%, respectively. Association were found among certain serotypes and virulence markers (O1, H-, H7, K1, MRHA, Col; O2, H-, Kl, Col; O4, H-, H5, MRHA, Hly; O6, H-, H1, MRHA, Hly; O6, K5, MRHA, Col; O7, H-, H4, K1, MRHA, Col; O18ac, H7, K1, Col; O18ac, H-, K5, MRHA, Hly; O78, H-, Col (V-type); O83, H-, K1, Col). There were associations among clinical specimens, age of patients, nosocomial group of diseases, serogroups and virulence markers, too (cerebrospinal fluid-CSF-O7, O18ac, O45, O83-K1-newborn meningitis; O78-ColV-meningitis, sepsis, inflammations diseases of premature babies; CFS-O6, MRHA, Hly-adult-meningitis, sepsis, urinary tract infection-UTI-, pneumonia, other inflammatory diseases; blood-O2, O4, O6, O18ac, ONT, K5, MRHA, Hly-sepsis, UTI, hepatic diseases; urine-O1, O2, O4, O6, O18ac, O75, virulence markers fall to differ among upper and lower UTI; faeces-O1, O4, O6, O18ac, O78, virulence markers rare). Associations were also found among animal pathogenicity tests, specimens, serogroups and virulence factors: highly virulent group strains (i.e. LD50 below 10(6)) belonged to serogroups O2, O6, O18ac, possessed antigen K1 (less frequently the presence of MRHA, Hly, K5) and originated mainly from CSF. With mouse lung toxicity test correlations of serogroups (O4, O6, O18ac), antigen K5, MRHA, Hly and specimens (blood) were also shown. However, association was found between the lack of virulence factors and phage insensitivity and also between K5 positivity and sensitivity to phages 16, 17, there were no correlations between serogroups and phage patterns. On the basis of the above-described associations one can find correlations among virulence markers, serotype, and nosological group of diseases. Animal pathogenicity tests give additional data in understanding the pathomechanism of diseases. Correlations between phage patterns and serogroups reveal certain epidemiological relatedness and also virulence of strains.

Typing of coagulase-negative staphylococci isolated from immunocompromised patients.

A total of 3121 coagulase-negative staphylococcal strains sourced from clinical samples were characterized during a 4-year period. Biotype, antibiotic resistance pattern, phage pattern and slime production was determined. Plasmid profile analysis was performed on related isolates. Thirty percent of strains originated from the Bone Marrow Transplantation Unit, The National Institute of Haematology, Blood Transfusion and Immunology (NIHBTI), Budapest. Staphylococcus epidermidis occurred most frequently (48.8% in total, 58.2% source from NIHBTI). Total bacteriophage typability was 75.9%, and 603 phage patterns were observed. NIHBTI isolates differed in the incidence of multiply resistance, slime positivity and average frequency of phage patterns from the total suggesting spread of a selected hospital population. Statistical analysis of data obtained by typing showed no predominance of any endemic clone: the strains colonizing the immunocompromised patients and isolated from staff and inanimated environment differed from each other in biotype, phage pattern, antibiotic susceptibility, slime production and/or plasmid profile.

[Determining the pattern of outer membrane proteins of gram-negative bacteria as a contribution to complex typing]. / Die Bestimmung des Musters der Proteine der äusseren Membran gramnegativer Bakterien als Beitrag zur komplexen Typisierung.

The aim of these investigations was to study relations between the serotype of E. coli strains and the pattern of their outer membrane proteins ("OMP") in sodium dodecylsulfate polyacrylamide gel electrophoresis. Three groups of strains being well characterized at least serologically (01, 02, 018ac containing different K, H, and in part F antigens) were submitted to this analysis. In all cases a nearly complete paralellity between OMP pattern and O:K:H(F:) type was observed, provided that the strains were epidemiologically related. The possibility is discussed that the OMP type could be used as a guide marker for the complex typing of E. coli strains.

Incidence of enteritis of Enterobacteriaceae isolates possessing human colonization factor antigen. New human colonization factors.

Of 462 Enterobacteriaceae strains including 435 Escherichia coli isolated from 250 patients, 298 haemagglutinating (HA) cultures were classified into 36 different HA groups. Sixteen of them belonged to Evan's I or II groups, although none possessed CF I or CF II antigen detectable by slide agglutination. Seventy-seven strains showed 4+ mannose resistant (MR) HA with human (53), bovine (2), chicken (6), guinea pig (7) or human and guinea pig (9) erythrocytes. These strains were significantly more frequent in patients under one year of age. Eighty-eight percent of the typable strains belonged to E. coli serogroups O1, O2, O4, O6, O18. HA positivity and fimbrial structures were correlated in 2 isolates (15/1, O18a, c:-K77: H-; 12/2/1 O1: K1: H .). Fimbriae of the two strains exhibited adhesive properties. Their fimbrial antigens differed serologically from each other and from those of the reference strains H 10407 and PB 176. Forty-nine of 4+ human MRHA strains showed variable reactions in the two sera for the new fimbrial antigens.

O:K:H:F serotypes of fimbriated Escherichia coli strains isolated from infants with diarrhea.

Bacterial agglutination and crossed immunoelectrophoresis were compared as techniques for the subdivision of mannose-resistant hemagglutinating Escherichia coli fimbrial antigens. A total of 22 fimbrial strains, 15 of which had earlier been grouped into two fimbrial agglutination groups, were examined for the presence of fimbrial antigens F7 go F12 by crossed immunoelectrophoresis. Most of the strains were isolated from infants with diarrhea; they were neither enteropathogenic nor enterotoxigenic serotypes. Of the 10 strains in agglutination group 1, 4 had fimbrial antigens F7 and 1C, 4 had antigens F8 and 1C, and in 1 strain, only antigen 1C was demonstrated. The tenth strain did not, by the crossed immunoelectrophoresis test, fit into agglutination group 1. Agglutination group 2 comprised five strains. Four of these had antigens in common with the still unnumbered fimbrial antigens of an O4:K12:H5 strain. In the fifth strain, no known F antigens were demonstrated. The previously found correlation between some O:K:H serotypes and fimbrial antigens in strains from urinary tract infections was confirmed in this study in strains from diarrhea cases. We concluded that, although the agglutination test can indicate the presence of certain fimbriae, it cannot be used presently for an exact demonstration of these antigens because each strain often produces several different fimbriae.

Virulence factors of Escherichia coli. IV. Association in Escherichia coli of LD50 with haemolysin production, haemagglutinating capacity, antigens K1, K5 colicinogenicity and pathogenicity.

Haemolysin production (HLY), mannose resistant haemagglutinating activity (MRA), presence of antigens K1 and K5 and colicinogenicity (Col) were compared with LD50 for mice in 663 Escherichia coli strains, including 281 faecal, 129 urinary and 253 other extraintestinal isolates. Those isolates that LD50 value fell into less than or equal to 10(6) LD50 category were arbitrarily termed highly virulent (HV) and those which belonged to greater than or equal to 10(7) LD50 category were considered avirulent (AV). HV isolates occurred significantly more frequently (58%) among strains from different extraintestinal samples than from faeces (14%) or urine (16%). The incidence of HV strains was significantly higher in patients with sepsis (43%) or meningitis (100%) than in patients with enteritis (20%), urinary tract infections (UTI, 16%) or in healthy subjects (28%). The incidence of HV strains in the most frequent (O1, O2, O4, O6, O7, O18, O75) serogroups was significantly higher (60%) than in others (10%). Strains with different virulence markers (HLY, MRA, K1, K5, Col) belonged significantly more frequently to the HV group than those which failed to have these markers (44 vs 27%, 51 vs 25%, 83 vs 17%, 78 vs 27%, 52 vs 16%, respectively). Important role of antigen K1 playing in pathomechanism of meningitis was confirmed by data of analysis according to which significant difference was revealed in the incidence of HV strains between groups of isolates with MRA+K1+ (71%) and MRA+K1- (44%, p less than 0.02), or between groups of isolates with MRA+K1- and MRA-K1+ (91%, p less than 0.001). Moreover there were significant differences in the incidences of HV strains in K1+Col- (73%) and K1-Col+ (29%, p less than 0.001), and in K1+Col+ (86%) and K1-Col+ (29%, p less than 0.001) groups. Further evidence was given by those data that there were no significant differences between groups of HV strains with MRA+K1+ and MRA-K1+ (p greater than 0.05) or with K1+Col+ and K1+Col- (p greater than 0.1) properties. Isolates that possessed simultaneously two of MRA, HLY, Col markers were more pathogenic in LD50 assay than those that had one or the other of these markers alone. Strains in serogroup O18 killed mice significantly more frequently than those of other serogroups independently of having any virulence factor, suggesting that bacteria in serogroups O18 must have some special virulence other than K1, Col, MRA or K5. MRA+HLY+ HV strains occurred frequently in extraintestinal diseases (42%) supporting the preconception that these properties play an important role also in the pathomechanism of extraintestinal infections other than UTI.(ABSTRACT TRUNCATED AT 400 WORDS)

Escherichia coli Col V plasmids and their role in pathogenicity.

Out of 1474 Escherichia coli strains belonging to 70 serogroups and 247 serologically not identified ones, colicin V producers were found in serogroups O78, O1, O7, O18 and also among the not identified strains. The molecular weight of Col V1 plasmid from the standard strain was 70 Mdal. The Col V plasmids carried by E. coli O78 strains isolated from a hospital outbreak in Hungary had a molecular weight of 78 Mdal and also 78 Mdal was the molecular weight of the Col V plasmid carried by Rivier's strain (designated 23), which had caused meningitis in Switzerland. The molecular weight of Col V plasmids of O1: K1, O21, O161 and serologically not identified E. coli strains isolated from sporadic cases was of 94 to 119 Mdal. In the case of three strains the increase of LD50 values, which means the decrease of virulence, resulted in the loss of colicin V production and the loss of Col V plasmid. It was demonstrated by introduction of a transposon, inactivating colicin V production, into a wild type E. coli strain that the production of colicin was not essential for the increase of virulence controlled by Col V plasmid. In the case of one strain the loss of both R and Col V plasmid resulted in a decrease of virulence. No plasmid other than R and Col V was carried by this strain. The virulence determining gene could be eliminated together with both plasmids, which means that this gene could be attached equally to Col V and R plasmid.

Virulence factors of Escherichia coli. III. Correlation with Escherichia coli pathogenicity of haemolysin production, haemagglutinating capacity, antigens K1, K5, and colicinogenicity.

A total of 1156 Escherichia coli strains including 489 faecal, 384 urinary, 283 other extraintestinal isolates was compared for haemolysin production (Hly), mannose resistant haemagglutinating activity (MRA), presence of antigens K1 and K5 and colicinogenicity (Col). K1 capsule which was demonstrated only in a few serogroups (O1, O2, O7, O18) occurred more frequently among extraintestinal (32.1%) than among faecal (4.3%) or urinary (7.3%) isolates. In the incidence of antigen K5 there was no difference between faecal and urinary (3.3%; 3.1%) or between urinary and other extraintestinal (5.3%) isolates belonging mainly to serogroups O2, O6, O18 and O75. Col+ isolates occurred frequently in all samples (23.5% of faecal, 31.7% of urinary and 43.4% of other extraintestinal strains), they being significantly more frequent in serogroups O1, O2, O7, O18 than in others. A close association existed between K1+ and Col+ properties, mainly (24.4%) among strains isolated from extraintestinal sources other than urine. The frequent coexistence of K1+ and Col+ in serogroups O1, O2, O7, O18 offers a further explanation for the extraintestinal pathogenicity of these serogroups. Neither Hly+ and K1+, nor Hly+ and Col+ were associated. MRA+ and K1+ correlated mainly in serogroups O1, O2 but never occurred simultaneously in serogroup O18. Connection between MRA+ and Hly+ was not associated with other virulence factors (K1, Col). The results showing a close connection among certain serogroups (O1, O2, O4, O6, O7, O18) and certain markers of pathogenicity (MRA, Hly, K1, Col) support the concept that E. coli strains have a clonal connection.

Characterization of Escherichia coli serogroups causing meningitis, sepsis and enteritis. I. Serological properties and animal pathogenicity of O18, O78 and O83 isolates.

Escherichia coli O78: K80 strains isolated from an outbreak among premature and newborn infants with meningitis, sepsis and enteritis, from sporadic cases of enteritis and from healthy carriers were compared with one another and with different E. coli serogroups. The O78: K80 cultures uniformly failed to give the rabbit intestinal loop test and the guinea pig eye reaction and none of them contained L1 antigen. After intraperitoneal injection into mice, the organisms multiplied in the peritoneal cavity and caused bacteriaemia lasting at least 2 weeks. E. coli strains originating from septicaemia (O78: K80, O18a,c: K?, O83: K?) showed significantly lower LD50 values for mice (9 x 10(3)--7 x 10(5)) than did E. coli serogroups associated with infantile enteritis only (3 x 10(8)--7 x 10(8)). It is assumed that the isolates differ in pathogenicity not only from E. coli strains associated with "cholera-like" disease and with "dysenteriform" infection, but also from L1 antigen-containing cultures described in neonatal meningitis, and constitute a separate group characterized by an ability to cause meningitis, sepsis and enteritis within the same outbreak.

Characterization of Escherichia coli serogroups causing meningitis, sepsis and enteritis. II. Classification of Escherichia coli O78 strains by phage sensitivity, colicin type and antibiotic resistance.

Escherichia coli O78: K80 strains isolated from an outbreak of meningitis, sepsis and enteritis in infants, were compared with O78: K80 strains from sporadic cases of enteritis, healthy carriers and animals. The strains were uniform in antigenic structure and phage pattern but differed in colicinogenicity. The epidemic strains and calf-pathogenic cultures produced colicin V, the remaining isolates were characterized by other types of colicin or were not colicinogenic. Col V+ strains multiplied in the mouse peritoneal cavity more readily and killed the animals at significantly lower doses than did col V- strains. One half of antibiotic resistant O78: K80 strains carried R factor. The spread of R factor could be followed by phage restriction experiments.