Characterization of Escherichia coli strains from long-term urinary tract infections by combined typing techniques.
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Biomedical subjects
Publications and source records attributed to D C Old.
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Fifty-two strains from seven species of Enterobacter, grown under a variety of conditions, were examined in rocked-tile tests for production of haemagglutinins and with the electron microscope for fimbriae. Thirteen non-haemagglutinating strains were non-fimbriate. Most (33) of the 39 haemagglutinating strains produced only one kind of haemagglutinin, either the mannose-sensitive haemagglutinin associated with type -1 Fimbriae or, the mannose-resistant, Klebsiella-like haemagglutinin associated with type-3 fimbriae. Multiply haemagglutinating strains were most common in E. aerogenes, in which species a third kind of haemagglutinin, also mannose-resistant, was found. The findings are discussed briefly in the light of the current taxonomy of Enterobacter.
When fimbriate (Fim+) strains of Salmonella typhimurium were grown in static broth, many bacteria were in the fimbriate phase and bore fimbrial mannose-sensitive haemagglutinin (MSHA) that enabled them to adhere to guinea-pig and other erythrocytes and to agglutinate them in rocked tile and static settling tests. When either Fim+ or Fim- strains were grown on phosphate-buffered nutrient agar, the bacteria formed a diffusible, mannose-resistant haemagglutinin (MRHA) that gave dispersed sediments with sheep and pig erythrocytes in static settling tests, but without evidence of bacterial adhesion to the erythrocytes. On exposure, from above or from below, to cultured HEp2 and HeLa cells for 30 or 90 min at 37 degrees C, motile MSHA-rich, MRHA-negative broth-grown bacteria adhered to the cells in large numbers (e.g., 20-100/cell), but motile MSHA-negative, MRHA-negative broth-grown bacteria and non-motile MSHA-negative, MRHA-rich agar-grown bacteria adhered in only small numbers (usually less than 1/cell). Thus, strong adhesiveness of bacteria for cultured cells in vitro appears to depend upon the presence of MSHA, not MRHA, and as Fim- (MSHA-negative) strains of S. typhimurium are known to be highly infective in animals, a strong reaction in the in-vitro model does not reflect a property of the bacteria essential for infectivity in vivo.
In strains of Proteus mirabilis of urinary origin, no correlation was found between proticine types, reflecting relative affinity for the urinary tract, and the production of haemagglutinins and presence of fimbriae, a measure of adhesiveness.
For 13 strains of Yersinia enterocolitica, there was a good correlation between the production of the broad-spectrum, mannose-resistant Yersinia haemagglutinin (MR/Y HA), the presence of fimbriae and high surface hydrophobicity. Each of these characters was expressed in cultures grown at low (less than 32 degrees C) but not at high (Greater than 35 degrees C) temperatures.
Strains of Klebsiella of the species (or 'patho-bio-sero-geogroups') Kl. atlantae, Kl. edwardsii and Kl. rhinoscleromatis produced neither haemagglutinins (HAs) nor fimbriae; strains of Kl. ozaenae were HA- but some produced type-6 fimbriae; and strains of Kl. pneumoniae (sensu stricto) and Kl. aerogenes that produced the mannose-sensitive HA (MS-HA) formed type-1 fimbriae. Most strains of Kl. aerogenes produced, in addition, one or both of the mannose-resistant HAs, MR/K-HA or MR/P-HA. The former, associated with type-3 fimbriae, was produced by 95%, and the latter by 57%, of the Kl. aerogenes strains. Some of the properties of the MR/P-HA, apparently a non-fimbrial HA not previously recognised in Klebsiella, are described.
For each of the isomers of tartaric acid, meso- or d- or l-, a pair of strains of Salmonella typhimurium was obtained, the one, a naturally occurring, non-fermenting strain and the other a spontaneous, tartrate-fermenting mutant derived from it. For each of the pairs of strains, competitive mixed cultures were grown from inocula of the tartrate-fermenting and tartrate-non-fermenting strains in peptone medium without or with the appropriate tartrate isomer. In each experiment, small numbers of tartrate-fermenting bacteria outgrew small or large numbers of tartrate-non-fermenting bacteria in 24 hours in tartrate-containing but not in tartrate-free peptone medium. The results of the experiments are discussed with reference to the choice of the definitive time of reading for tartrate-utilisation tests in the biotyping of S. typhimurium.
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An international collection of 419 isolates of Salmonella agona was phage typed, biotyped and colicine typed. Of 16 recognized phage types, 15 were represented. Three phage types (I, V and XVI) accounted for 84% of all isolates, were widely distributed and may be interconvertible. Biotyping afforded little type differentiation; thus 92.6% of the isolates belonged to biotype 1 a. A rhamnose non-fermenting variant line (of biotype 5a) became established in Zaire from 1979 to 1980. A maltose late-fermenting line of biotype 1 a, isolated in Scotland in 1974, did not thereafter become established. Two Col+ lines (producing colicine I b) accounted for 45 of 68 colicinogenic isolates. The implication of type diversification and the phylogenetic significance of these findings are discussed.
A two-tier system for biotyping Escherichia coli gave a fine and reliable differentiation of strains and is capable of future extension by the addition of new types and new tests. Strains were allocated to a primary biotype (1-16) by their reactions in four primary tests in culture media containing raffinose, sorbose, ornithine or dulcitol. Subtypes were distinguished within the primary biotypes by reactions in six secondary tests for rhamnose fermentation, lysine decarboxylation, aesculin hydrolysis, motility, type-1 fimbriation and prototrophy. Full biotypes were designated by letters indicating subtype reactions appended to primary type numbers. A series of 599 strains (1242 cultures) of E. coli from diverse sources was classified into 16 primary biotypes and into 213 full biotypes. The biotype characters of a strain were generally stable during its spread in the natural environment and in non-selective media used for storage.
One hundred and thirteen strains of Morganella, Proteus and Providencia, grown in different cultural conditions, were examined for their ability to produce haemagglutinins (HAs). Three main kinds of HA (MS, MR/K and MR/P) were detected, and 89% of the 112 HA+ strains were capable of producing two or three of the different HAs in the same or different cultures. The properties of the three HAs were partly defined and the difficulties of identifying their separate HA activities when present together are discussed. Electronmicroscopic examination of bacteria from HA+ cultures showed at least six distinct fimbrial types, the properties of which are described. We tried, with limited success, to correlate the presence of the different HAs with that of the different fimbrial types. The significance of our findings is reviewed in the light of recent taxonomic changes for this group of enterobacteria. The distribution of HAs and fimbriae in the species of Morganella, Proteus and Providencia is more complex than that so far described for other genera of Enterobacteriaceae.
Strains of Serratia marcescens, Serratia liquefaciens, Serratia marinorubra, and Serratia plymuthica produced one or more of the following hemagglutinins (HAs): mannose-sensitive HA and mannose-resistant K-HA (MR/K-HA) and P-HA (MR/P-HA) (J. P. Duguid and D. C. Old, in E. H. Beachey (ed.), Bacterial adherence, vol. 6., p. 185-217, 1980). Most strains (82%) were multiply hemagglutinating. The properties of the three HAs are described. Each HA was associated with a distinct type of fimbria: mannose-sensitive HA with type 1 fimbriae. MR/K-HA with type 3 fimbriae, and MR/P-HA with a new type of thin fimbriae provisionally called MR/P fimbriae. This is the first report of the production of MR/P-HA and MR/P fimbriae by Serratia species. The range of Serratia HAs, which may reflect in vivo colonization potential, is more complex than previously reported.
A biotyping scheme for Enterobacter cloacae is proposed. Tests with seven substrates that gave reliable, reproducible results with 110 isolates of E cloacae formed the basis of the biotyping scheme which would allow recognition of 128 potential biotypes.
When 1420 isolates of Shigella sonnei recovered in Dundee, Scotland between 1971 and 1976 were examined with indicator strains of known immune status, four new colicin types, not previously described among the 26 types of Horák (8), were observed. The types were: 11 variant (colicins B, M), 4 variant (Ib, B, M), SH1 (E3) and SH3 (E5). Only type 4 variant strains were of epidemiological significance in the Dundee area.
An epidemiological study of Sonne dysentery in Dundee during the years 1971-6 was made by examining, in respect of 1420 isolates of Shigella sonnei, the discriminating power of colicine typing, antibiogram testing, biotyping and resistotyping and the stabilty of the markers they provided. Colicine typing identified nine colicine types, including four not previously described. However, because types 4 and 4 var., determined by col Ib, and type U, producing no colicines, accounted for 96% of the isolates, discrimination with colicine typing was poor. In antibiotic sensitivity test, 13 different antibiogram patterns were noted. Less than 1% of the isolates were sensitive to all of the eight antibiotics tested; most were multiply drug-resistant. Resistance to kanamycin, neomycin and paromomycin (KNP) was apparently due to a single resistance determinant, widely distributed in a majority (53%) of the isolates. When definitive times were chosen for reading each biotyping test, only maltose and rhamnose of the 13 "sugars' tested differentiated isolates into prompt- and late-fermenting types. Though the ability to ferment rhamnose was a stable property, it discriminated only 1.5% of the minority, late-fermenting type. Resistotyping with six chemicals discriminated eight epidemiologically valid resistotypes, including three new types. However, 93 of the isolates belonged to only three resistotypes. Analysis of the data for isolated from 286 epidemiologically distinct episodes showed that the variability of colicine and antibiogram characters, found among isolates within, respectively, 40 and 28% of the episodes, was generally associated with loss or gain of a plasmid ("col Ib-KNP') which determined production of colicine Ib and KNP resistance. These characters varied both in vivo and in vitro. Variability of resistotype characters, on the other hand, was observed in only 28 (9%) episodes, 14 of which possibly represented examples of mixed or sequential infections. For accurate epidemiological tracing of strains of Sh. sonnei in a community, resistotyping, the technique showing the greatest discrimination and least variability of the four tested, should be included as the principal typing method.
Different strains of Shigella sonnei present in Dundee from 1971 to 1976 were identified by a "multiple typing' method in which resistotyping, used as the main method of differentiation, was supplemented by colicine typing, antibiogram testing and biotyping. At least 19 different "multiple types' (MTs) were identified by combining information from the four typing techniques. The relation of the different types and their possible derivation from each other are discussed. The practical value of multiple typing was demonstrated in a study of 247 isolates, of three distinct MTs, recovered from 178 persons involved in an extended outbreak centred primarily on day nurseries. A few episodes that yielded isolates of different resistotypes were analysed to determine whether the cultures were: isolates of the same strain different in resistotype as a result of in vivo or in vitro variation of resistotype characters, or isolates of distinct strains of different resistotypes. The multiple typing approach clarified the way in which different MTs emerged, persisted, disappeared or co-existed in the community during the 6 years of the study.
Twenty-three strains of Escherichia coli with mannose-resistant and eluting (MRE) haemagglutinins with eight different patterns of substrate specificity were examined by a variety of electronmicroscope methods. In 12 strains, the presence of MRE haemagglutinins with broad-spectrum patterns 1, 3 and 4 was correlated with that of fimbriae. In 11 strains with MRE haemagglutinins with the less common patterns 6, 7 var., 8, 9 and 10, fimbriae were not found on bacteria in MRE+ cultures, indicating that the latter haemagglutinins are non-fimbrial.
Strains of Yersinia enterocolitica from different O serogroups were tested for their ability to adhere to cultured epithelial cells (H.Ep 2) after growth of bacteria in broth at 22 C or 37 C. Strains that adhered to H.Ep 2 cells generally did so better after growth at 22 C than at 37 C. The data indicate that the ability to produce either of the mannose-resistant (MR) hemagglutinins, MR/Y or K1, each associated with fimbriae, does not correlate with ability to adhere to epithelial cells in this particular in vitro model.