Haemagglutination of Bacteroides fragilis

Abstract The relationship between the ability to cause haemagglutination (HA) and the presence of capsules and/or pili was examined for 50 clinical isolates of Bacteroides fragilis . HA was tested using a slide technique, and bovine, porcine, guinea pig, rat, rabbit, horse, human, chicken and pigeon erythrocytes. Chicken and pigeon erythrocytes were the best indicators for HA with 43 (86%) of the strains tested causing HA and 39 (78%) with strong reactions. Capsule staining showed that the same 43 strains causing HA also produced a demonstrable capsule. No pili were found on either encapsulated or non-encapsulated strains using transmission electron microscopy. These results suggest that adherence of B. fragilis is related to the presence of capsular material, not pili.     

Pili mediated agglutination of Serratia marcescens in human urine

Of 51 strains of Serratia marcescens isolated from patients with urinary or respiratory tract infections, 35 agglutinated in human urine. The agglutinating strains possessed numerous pili which were morphologically distinct from common pili or type I pili. The diameter of the pili was 3 nm and the average length was 0.3 micrometer. Electron microscopic examination showed that 80% or more of the cells of the agglutinating strains and 0 to 8% of the cells of the nonagglutinating strains were piliated. When an agglutinating strain was heated at 55 C for 10 min, it lost its agglutinating capacity and concomitantly its pili. These results suggest that the agglutination might occur because of interactions between the pili and some factors in human urine. The urinary slime appears to contain these agglutinating factors.     

Maintenance of broad host range vector pKT230 in marine unicellular cyanobacteria

Clinical isolate of Vibrio mimicus were examined for production of cell-associated hemagglutinin (HA) and pili and for adherence to formalin-fixed human intestinal mucosa. V. mimicus grown on CFA agar for 3 h at 37 degrees C possessed HA and adhered better to the mucus layer than to the epithelial cell surface. A significant correlation was found between the HA titers and adherence ability to the epithelial cell surface of villi (P less than 0.05); adherence to the ileal lymphoid follicle-associated epithelium occurred at higher levels. In contrast, V. mimicus grown on CFA agar for 20 h at 37 degrees C exhibited lower levels of HA and reduced adherence ability. The production of pili was more pronounced after 20 h of incubation than after 3 h of incubation. In comparison with V. cholerae 01 and V. cholerae non-01 cultured under similar conditions, V. mimicus showed inferior adherence, but with similar HA production or piliation.     

Limitations of immunofluorescence tests in the diagnosis of infectious mononucleosis

The relative value of heterophil agglutinins (HA) and of specific EBV antibodies in the diagnosis of infectious mononucleosis (IM) was assessed in 108 cases of the disease and in 280 controls. Among the 108 cases 93 were HA-positive by sheep cells in at least one of their sera, while 15 were HA-negative by the same test. Among the 280 controls false-positive HA tests were not encountered except in eight cases with the horse cell microtitre tests. With one of the two slide tests at least two false-positive tests and 12 false-negative tests were also found but these sera had low titres in microtitre tests. The HA life-span was found to be unexpectedly long in a few cases, sheep cell HA lasting up to 8 to 10 months and horse cell HA up to 21 to 23 months.Many false-positive tests may therefore not be true false-positives and may result from the persistence of HA following unrecognized mononucleosis months before. Virtually all cases of IM had (or developed) antibodies to Epstein-Barr virus, viral capsid antigen (EBV-VCA), whereas only half of the controls were EBV-VCA-positive. The comparative analysis of nonspecific and specific test results in mononucleosis allows the following conclusions: (1) horse cell microtitre tests and the monospot test are more sensitive than sheep cell microtitre tests and the monotest; (2) false-negative results are occasionally seen with the latter tests but not with the former; (3) more false-positive results, however, are probably seen with the former tests; and (4) specific EBV-IgM and EBV-EA antibody tests are useful in the diagnosis of selected borderline cases of mononucleosis.     

Involvement of a sialic acid-binding lectin with hemagglutination and hydrophobicity of Flavobacterium psychrophilum

Strains of Flavobacterium psychrophilum were studied for their ability to adhere and cause agglutination of erythrocytes and yeast cells. Strains of the serotype Th showed low or no hemagglutinating (HA) properties toward human, avian, bovine, and rainbow trout erythrocytes, whereas strains of serotype Fd and Fp(T) exhibited distinct HA properties. None of the strains was able to cause agglutination of yeast cells. Greater adherence specificity toward rainbow trout blood cells was seen for the HA-positive strains. Growth at 5 degrees C, compared to that at 15 degrees C, induced an increase in the hemagglutination of some strains. HA activities of F. psychrophilum were inhibited only by sialic acid (N-acetyl-neuraminic acid), heat treatment at 65 degrees C, and proteinase K treatment and not by any of seven other carbohydrates, periodate oxidation, or treatment with trypsin. The supernatant from washed bacterial cells also showed some HA properties. All strains were shown to be highly hydrophobic by the hydrophobic interaction chromatography test, although some contradictions to the results of the salt aggregation test (showing some strains as less hydrophobic) were seen. These results indicate that the aggregation of F. psychrophilum and erythrocytes depend on a lectin present on the surface of HA-positive F. psychrophilum strains and absent on HA-negative strains. This lectin reacts specifically with sialic acid. The adhesion differences observed for F. psychrophilum strains do not appear to correlate with the virulence but still provide insights into the interaction of F. psychrophilum and rainbow trout.     

Expression and detection of different biotype-associated cell-bound haemagglutinins of Vibrio cholerae O1

The expression of two cell-bound haemagglutinins, one sensitive to L-fucose (FSHA) and the other to D-mannose (MSHA), on Vibrio cholerae O1 strains of both the classical and the El Tor biotypes was studied by (i) agglutination of chicken and human group O erythrocytes in the presence of L-fucose or D-mannose, (ii) binding of the bacteria to L-fucose- and D-mannose-coated agarose beads, and (iii) agglutination of the bacteria by 'biotype-specific' antisera. All of the 12 classical strains studied that were isolated before 1979 gave FSHA of human O erythrocytes whereas only 6 of 17 classical strains isolated during recent epidemics expressed FSHA; a few of the classical strains expressed MSHA in addition to FSHA. All the El Tor strains gave MSHA of chicken erythrocytes and one strain also expressed FSHA. Both the cell-bound HAs were optimally expressed during the exponential phase of growth; FSHA markedly decreased during the late exponential phase while the MSHA usually persisted into the stationary phase. The expression of FSHA and MSHA correlated very well with the direct binding of vibrios to fucose- and mannose-coated agarose beads, respectively. Antiserum 'specific' for classical vibrios agglutinated classical strains expressing FSHA and also the El Tor strain exhibiting FSHA. Similarly, the anti-El Tor serum agglutinated all El Tor strains and also classical strains expressing MSHA, suggesting that the 'biotype-specific' sera were specific for the biotype-associated cell-bound HAs.     

Serological method for identification of Vibrio parahaemolyticus from marine samples.

Use of agglutination with antiserum against lateral flagella (H-agglutination) for the identification of Vibrio parahaemolyticus was studied. Sucrose-negative bacteria were isolated from seawater, and their characterization was carried out by traditional biological tests and slide agglutination with antiserum specific to lateral flagella of V. parahaemolyticus. Of 135 strains isolated, 78 were identified as V. parahaemolyticus by biological tests and were agglutinated with the above serum. Fifty-five strains did not agglutinate with the serum, and their biological characteristics were different from those of V. parahaemolyticus. Two strains also differed from V. parahaemolyticus in some biological characteristics but agglutinated with the antiserum. All clinically isolated V. parahaemolyticus strains also agglutinated with the above serum. These results suggest that our serological method is useful for the identification of V. parahaemolyticus, especially for samples in which there are many organisms related to V. parahaemolyticus, because many biological tests can be omitted.     

Serological method for identification of Vibrio parahaemolyticus from marine samples

Use of agglutination with antiserum against lateral flagella (H-agglutination) for the identification of Vibrio parahaemolyticus was studied. Sucrose-negative bacteria were isolated from seawater, and their characterization was carried out by traditional biological tests and slide agglutination with antiserum specific to lateral flagella of V. parahaemolyticus. Of 135 strains isolated, 78 were identified as V. parahaemolyticus by biological tests and were agglutinated with the above serum. Fifty-five strains did not agglutinate with the serum, and their biological characteristics were different from those of V. parahaemolyticus. Two strains also differed from V. parahaemolyticus in some biological characteristics but agglutinated with the antiserum. All clinically isolated V. parahaemolyticus strains also agglutinated with the above serum. These results suggest that our serological method is useful for the identification of V. parahaemolyticus, especially for samples in which there are many organisms related to V. parahaemolyticus, because many biological tests can be omitted.     

Fimbrial hemagglutinins in Klebsiella pneumoniae

Eleven Klebsiella pneumoniae strains were isolated from urine specimens which were examined for their ability to produce hemagglutinins (HAs). Bacteria were grown under various culture conditions. Suspension of bacteria grown in broth or on Phosphate-buffered nutrient agar (PBA) were tested for agglutination in the presence and absence of 2% (w/v) D-mannose, on rockedtiles at 4 degrees C and ambient temperature with mangroup-O(M), fowl(F), ox(O), guinea-pig(G), horse (H), rabbit (R) and sheep (S) erythrocytes and tannic acid treated, but not fresh oxen erythrocytes. Each of the 11 strains was hemagglutinating. Ten strains (99%) producing two or three hemagglutinins (HAs), were multiple hemagglutinating. One strain produced only mannose-resistant, Klebsiella, the "Tanned ox hemagglutinin" (MR/K-HA). Solely mannose-sensitive hemagglutinin (MS/HA) was not produced by any of the strains. No mannose-sensitive hemagglutinating strains acted on sheep erythrocytes. Three main kinds of hemagglutinin (HA) were detected. These were: (a) a mannose-sensitive hemagglutinin (MS-HA); (b) a mannose resistant, Klebsiella, the "tanned ox hemagglutinin (MR/K-HA); (c) mannose-resistant, Proteus hemagglutinin (MR/P-HA). All strains (100%) produced MR/K-HA, 45% of MR/K-HA+ strains produced MR/P-HA at 37 degrees C and 99% of all strains produced MS/HA, MR/P-HA activity was never dependent on MR/K-HA: Electronmicroscopic examination of bacteria showed that all strains were fimbriate.     

Some Properties of the Pili of Corynebacterium renale

Some properties of the pili of the gram-positive bacteria Corynebacterium renale were described. A relationship was found between the morphological features of pili and the types of C. renale. Strains of types I and III usually possessed a small number of pili, whereas those of type II possessed numerous pili. Thick and long bundles of pili characteristic of C. renale were frequently observable in type II strains. Piliation of C. renale was stable under various cultural conditions. No ability to agglutinate red blood cells was demonstrated by piliated strains of C. renale. Pili were isolated from the cells of C. renale and studied serologically by immunodiffusion. The pili of a type II strain were serologically identical with the pili of another type II strain but not with those of the strains belonging to types I and III. The pili were serologically distinct from the cell wall. The pili were broken into short pieces by boiling, but their antigenicity was increased after boiling.     

Haemagglutinins and Fimbriae in Different Serotypes and Biotypes of Yersinia enterocolitica

When cultured in static broth at 20°C, 46 of 115 strains of Yersinia enterocolitica , diverse in biotype and serotype, produced a broad-spectrum mannose-resistant (MR) adhesin that agglutinated the erythrocytes of all of 10 animal species examined. The production of haemagglutinin (HA) was associated with the presence of fimbriae of S nm diameter. Culture of HA+ strains at 37° resulted in the disappearance of haemagglutinating ability and loss of fimbrial production. Strains of Y. enterocolitica with K1 antigen produced an MR adhesin that agglutinated only fowl erythrocytes and was associated with fimbriae of 4–4.5 nm diameter. None of 14 strains of Y. pseudotuberculosis was haemagglutinating.     

Involvement of a Sialic Acid-Binding Lectin with Hemagglutination and Hydrophobicity of Flavobacterium psychrophilum

Strains of Flavobacterium psychrophilum were studied for their ability to adhere and cause agglutination of erythrocytes and yeast cells. Strains of the serotype Th showed low or no hemagglutinating (HA) properties toward human, avian, bovine, and rainbow trout erythrocytes, whereas strains of serotype Fd and Fp^T exhibited distinct HA properties. None of the strains was able to cause agglutination of yeast cells. Greater adherence specificity toward rainbow trout blood cells was seen for the HA-positive strains. Growth at 5°C, compared to that at 15°C, induced an increase in the hemagglutination of some strains. HA activities of F. psychrophilum were inhibited only by sialic acid (N-acetyl-neuraminic acid), heat treatment at 65°C, and proteinase K treatment and not by any of seven other carbohydrates, periodate oxidation, or treatment with trypsin. The supernatant from washed bacterial cells also showed some HA properties. All strains were shown to be highly hydrophobic by the hydrophobic interaction chromatography test, although some contradictions to the results of the salt aggregation test (showing some strains as less hydrophobic) were seen. These results indicate that the aggregation of F. psychrophilum and erythrocytes depend on a lectin present on the surface of HA-positive F. psychrophilum strains and absent on HA-negative strains. This lectin reacts specifically with sialic acid. The adhesion differences observed for F. psychrophilum strains do not appear to correlate with the virulence but still provide insights into the interaction of F. psychrophilum and rainbow trout.     

Properties of the erythrocyte receptors for influenza C virus.

Properties of the receptor for influenza C virus were studied. Although the receptor for influenza C virus on chicken erythrocytes was destroyed by the homologous virion, neuraminidase activity could not be detected in any of the influenza C virus strains tested. The receptor activity of chicken erythrocytes for influenza C virus was diminished by formaldehyde treatment but not by periodate oxidation. There was a considerable variation in the pattern and the titer of hemagglutination of influenza C virus when human erythrocytes of different blood types were used; the virus agglutinated most type B erythrocytes but not type A erythrocytes. By using human type B erythrocytes, differences among strains of influenza C virus in the hemagglutinating activity were also demonstrated. These results showed that both the receptor for and the receptor-destroying activity of influenza C virus were completely different from those of influenza A or B virus and also that carbohydrates were not involved in the receptor for influenza C virus.     

RNA polymerase heterogeneity in Streptomyces aureofaciens: characterization by antibody-linked polymerase assay

The ability of Escherichia coli with different receptor specificities to interact with meconium was studied. E. coli strains expressing P-fimbriae, specific for Gal alpha 1-4Gal beta-containing receptors, were agglutinated by meconium at high titres. This reaction was inhibited by globotetraosylceramide. The attachment of P-fimbriated E. coli to human colonic epithelial cells of the HT-29 cell line was inhibited by meconium. Some type 1 fimbriated strains were agglutinated by meconium, but the agglutination was rarely blocked by methyl alpha-D-mannoside. The attachment by type 1 fimbriated strains to HT-29 cells was reduced by meconium only in some cases. These results suggest that meconium interacts with the P-fimbriae of E. coli, in a way that may influence bacterial colonization of the neonatal intestine.     

Electron Microscopic Examination of Factors Influencing the Expression of Filamentous Surface Structures on Clinical and Environmental Isolates of Aeromonas veronii biotype sobria

Strains of Aeromonas veronii biotype sobria isolated from clinical and environmental sources were examined for their expression of surface structures under a variety of culture conditions. When grown on solid media at 37 C, more than 95% of bacteria from the majority of strains isolated from human diarrheal feces and chicken carcasses were non-piliated or expressed only a few pili of long, flexible morphology per cell. Strains isolated from water or other foods were much more likely to express pili. Heavily piliated strains (all sources) possessed pili of several morphological types, including long, flexible pili of varying widths and rigid pili of varying lengths. Expression of pili was favored by growth at temperatures ca. 20 C and below and growth in liquid medium. Most fecal strains expressed some pili under these conditions. In addition, other surface structures (fibrillar aggregates, fibrillar networks, bundle-forming pili) were seen on some strains from most sources. These were also seen most frequently when bacteria were grown in liquid media at temperatures ca. 20 C and below. Pili expression was not dramatically influenced by growth under anaerobic conditions, or in iron-depleted media, or by combinations of the above conditions. The role of the above surface structures in Aeromonas pathogenicity remains to be elucidated.     

Variations in the expression of pili: the effect on adherence of Neisseria meningitidis to human epithelial and endothelial cells

The effect of variations in Neisseria meningitidis pili on bacterial interactions with three epithelial cell lines as well as human umbilical vein endothelial cells was studied using a panel of seven strains expressing Class I or Class II pili. Comparison of adherence of piliated and pilus-deficient variants of each strain to epithelial cells suggested that Class I pili may mediate bacterial adherence with all three epithelial cell lines. In contrast, Class II pili of the strains used did not increase bacterial adherence to Hep-2 larynx carcinoma cells, although an increase in adherence to Chang conjunctival and A549 lung carcinoma epithelial cells was observed in the Class II pili-expressing strains. In addition to these interclass functional variations, differences in adherence to epithelial cells were also observed among Class I and Class II strains. Functionally different pilin variants of one Class I strain, MC58, were obtained by single colony isolation. One piliated variant was identified which had concurrently lost the ability to adhere to both Chang and Hep-2 cells ('non-adherent' phenotype; adherence of less than 2 bacteria per cell). In addition, several adherent pilin variants were isolated from non-adherent Pil- and Pil+ bacteria by selection on Chang cells (adherence of 10-25 bacteria per cell). In contrast to epithelial cells, all variant pili, whether of Class I or Class II, adhered to endothelial cells in substantially larger numbers (greater than 50 bacteria per cell) and therefore implied the existence of distinct mechanisms in pilus-facilitated interactions of N. meningitidis with endothelial and epithelial cells.     

The hemagglutinating properties of Pseudomonas aeruginosa strains isolated from patients with nonspecific lung diseases]

To detect d-mannose-sensitive (MS) pili in 31 P. aeruginosa strains isolated from the respiratory tract of patients with inflammatory and purulent destructive pulmonary diseases, the hemagglutination (HA) test was used. The isolated Pseudomonas under study differed in the degree of manifestation of their MS adhesins. Among them microorganisms with pronounced HA activity (high HA titer) occurred, as well as those whose HA activity was less pronounced (low HA titer). P. aeruginosa strains with pronounced HA activity were more frequently isolated from patients with purulent destructive processes in the lungs. A correlation between the state of the patient at the moment of bacteriological examination and the degree of manifestation of MS pili in the P. aeruginosa strain isolated from this patients was established. The value of HA titer in the presence of d-mannose is indicative of the presence of MS adhesins in a P. aeruginosa strain.     

Hemagglutinating properties of Proteus mirabilis strains isolated from clinical specimens

Haemagglutinating properties of 345 P. mirabilis strains isolated from various clinical samples were determined. Red blood cells of different origin as human group 0, bovine, horse, sheep and rat were used for the study. For the detection of MS and MR/P haemagglutinins the haemagglutination reaction was run with and without D-mannose. On the other hand, for the detection of type MR/K haemagglutinins tanned human and bovine erythrocytes were used. The majority of tested strains (90.14%) was polyhaemagglutinating i.e. showed simultaneously the presence of two or three haemagglutinins. Only three strains of P. mirabilis (0.87%) did not agglutinate any of the erythrocytes used for the study. The majority of strains (95.83-100%) in specific groups of clinical materials showed the presence of MR/K+ while MR/P+ 45.45-93.75% of strains and MS+ 45.83-73.1% of tested strains. Out of P. mirabilis strains isolated from urine, faeces and blood the highest percentage possessed at the same time all three haemagglutinin types (MS+, MR/K+, MR/P+) or pattern MR/K+, MR/P+. Bronchial isolates had usually pattern MR/K+ (31.82%) and strains isolated from skin possessed haemagglutinins of pattern MR/K+, MR/P+ (50%) and MS+, MR/K+, MR/P+ (43.75%). Among strains expressing MR/P+ at 37 degrees C a great differentiation of spectrum activity against tested erythrocytes was seen. Undoubtedly, the majority of MR/P+ strains from specific groups of clinical materials (with the exception of urine) agglutinated sheep and horse erythrocytes with and without D-mannose. The majority of strains isolated from urine agglutinated sheep and bovine erythrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pili of Aeromonas hydrophila: purification, characterization, and biological role

Aeromonas hydrophila (Ae6) has 2 morphologically distinctive kinds of pili. One appeared rigid, channeled, and straight with a diameter of 9 nm (Ae6-R pili). The other looked flexible, wavy, and having helical structure with a diameter of 7 nm (Ae6-W pili). Ae6-R pili were purified and characterized. The pili consisted of a subunit protein with a molecular weight of 18 kDa as estimated by SDS-PAGE, and contained 42.3% hydrophobic amino acids and one cysteine residue. The pilus was solubilized to 18 kDa subunit protein by 2-mercaptoethanol, dithiothreitol, hydrochloric acid, or heating at 120 C for 5 min. The organism Ae6 was strongly adhesive to rabbit intestines as well as human intestines, and agglutinated erythrocytes. Anti-pili antibody (Fab fraction) did not block the adhesion. Purified Ae6-R pili did not adhere to the intestine or to the erythrocytes. However, the anti-pili Fab inhibited pellicle formation of the organisms cultured in broth, and also inhibited salt agglutination with ammonium sulfate. From these results, Ae6-R pili are not likely a colonization factor but probably play a role in the autoaggregation of the organisms.     

Purification and characterization of Vibrio cholerae O139 fimbriae

Abstract A Vibrio cholerae O139 (strain Al-1841) isolated from a patient with a cholera-like disease in Bangladesh predominantly produced new curved, wavy fimbriae (Al-1841 fimbriae) and small numbers of previously reported V. cholerae non-O1 S7-like pili. The former was purified and characterized. The molecular mass of the Al-1841 fimbrial subunit was less than 2.5 kDa, and it was immunologically different from that of V. cholerae non-O1 S7 pili. This novel fimbrial antigen was detected in all 182 Gram-negative strains from five genera tested but was absent from the Gram-positive bacteria tested. The purified Al-1841 fimbriae did not agglutinate human or rabbit erythrocytes.