- Volume 18, Issue 1, 1984

Intraperitoneal injection of Campylobacter fetus ss. jejuni into HAM/1CR mice was lethal, but viable counts of bacteria from whole body homogenates, organs and blood indicated that death was not due to sustained bacterial multiplication. Heat-killed organisms (5 x 109 cfu) injected into 7-day-old mice caused death within 24 h and this was shown to be due to endotoxin. Both ferric iron and heterologous lipopolysaccharide enhanced virulence; the LD50 was lowered from 1.8 × 109 cfu to 2.7 × 107 cfu when both were used. Three-day-old or adult animals survived challenge with Campylobacter fetus without clinical symptoms when challenged orally or by intravenous or intraperitoneal routes.

Strains of six black-pigmented Bacteroides species and one un-named strain were examined for their ability to degrade the plasma proteins albumin, haemopexin, haptoglobin and transferrin. Strains of B. gingivalis were most effective, degrading all four plasma proteins at different rates. Strains of B. intermedins and B. asaccharolyticus showed intermediate activities, degrading different individual plasma proteins; strains of B. melaninogenicus, B. loeschei and B. denticola were least active, degrading only haemopexin. These findings are discussed in relation to the availability in tissue fluids of iron for bacterial growth.

The relationship between vaginal carriage and subsequent neonatal acquisition of Clostridium difficile was investigated. Vaginal carriage of C. difficile was detected in 11% of women attending the Department of Genital Medicine Clinic. C. difficile was isolated from the vagina in 18% of 50 mothers before delivery, and 8% after delivery; 62% of their babies had positive faecal cultures. Eight of nine of the babies whose mothers had positive cultures before delivery became colonised with C. difficile, while 23 of 41 babies whose mothers had negative cultures became colonised. This suggests that both the vagina and the environment may act as sources of neonatal acquisition of C. difficile. Broth enrichment culture proved a more sensitive method for isolating C. difficile from the vagina than direct plate culture and should be used in such investigations.

A total of 57 strains of Acinetobacter calcoaceticus was fingerprinted by SDS-PAGE of cellular protein. All strains were also examined by conventional, API and N/F-Tek methods, and antibiotic sensitivity patterns were determined. In general, using the API 20E and N/F-Tek methods, it was possible to assign isolates of A. calcoaceticus to the two accepted “biotypes” or “varieties”, A.c. anitratus and A.c. Iwoffi, but these methods did not offer useful subdivision of the biotypes. Gel electrophoresis permitted subdivision of the strains into clusters in a manner which suggests that the technique may be valuable in typing strains isolated during outbreaks of infection in hospital.

Phagocytosis of the luminous bacterium Vibrio cholerae var. albensis caused a similar decrease both in viable count and in the in-vivo luminescence. These effects of polymorphonuclear neutrophil leukocytes (PMN) were oxygen-dependent processes. Exposure of PMN to oxygen caused a prompt decrease in the luminescence of bacteria that had been ingested in anaerobic conditions. Cell-free supernates from active PMN suspensions caused a decrease in luminescence and as much as 10% of the killing could be attributed to extracellular killing. Similarly, bacteria entrapped on a membrane filter showed a decrease in luminescence upon addition of active PMN, even though they could not be ingested.

The adhesion of leptospires (Leptospira interrogans serovar. copenhageni L45) to mouse L-cells was studied by microscopic observations. Within 3 h of infection of monolayers many leptospires adhered to 95-100% of the cells, and intracellular leptospires were demonstrated by electron microscopy. No specific site of attachment on the cells or the leptospires was observed. Avirulent or dead leptospires adhered poorly but attachment of the saprophytic leptospire L. biflexa serovar. patoc occurred on cell and glass surfaces. After adhesion, microvilli on the cell surfaces disappeared within 6 h of infection and cell damage was observed after 12 h. The adhesion was greatly enhanced by the presence of specific antiserum at a subagglu-tinating concentration. No direct penetration by leptospires of the host cells was observed with transmission and scanning electron microscopy. It appears that (1) adhesion of leptospires to L-cells precedes cell damage, and (2) leptospires may enter cells either through damaged membranes, or by a phagocytosis-like mechanism.

Fimbriae have been demonstrated on strains of Bordetella pertussis that possess agglutinogen 2 (types 1, 2, 3 and 1, 2), but not on those that lack it (types 1,3 and 1). This correlation between fimbriation and the presence of agglutinogen 2 has been found with fresh isolates from children and with laboratory strains that are virulent for mice. If fimbriae enhance the attachment of bacteria to mucosal cells, these findings offer an explanation for the predominance of serotypes 1,2,3 and 1,2 in non-vaccinated communities. The findings also suggest that agglutinogen 3 is not a fimbrial antigen, and because this is an essential component of fully effective whole-cell vaccine, a subcellular vaccine prepared from fimbriae alone may be inadequate.

Trimethoprim-resistant urinary isolates of Escherichia coli, collected in 1982, were studied and the results compared with those obtained for isolates collected during the period 1979-1980. Ninety-eight (81%; a 10% increase) were resistant to trimethoprim 1024 µg/ml and 93 (95%) were also resistant to sulphamethoxazole 1024 µg/ml. The frequency distributions of plasmids were similar in both collections although there was a significant increase in the number of small plasmids (mol. wt ⩽ 20 × 106) in the 1982 collection. Transfer of resistance was associated with isolates that carried larger numbers of plasmids.

A significantly smaller proportion of isolates in this, than in the earlier series, transferred trimethoprim resistance to E. coli K12 suggesting continued transposition of trimethoprim resistance on to the bacterial chromosome. Fifteen different trimethoprim resistance plasmids were identified, of which five, including that found most frequently, were common to both collections. Plasmids which transferred trimethoprim resistance without sulphonamide resistance were more common in the 1982 series. Plasmids which transferred linked trimethoprim and streptomycin resistances, in particular those not carrying other resistance markers, were less common in the 1982 series of isolates. Although the majority of E. coli isolates highly resistant to trimethoprim remained resistant to sulphonamide, our results suggested changes in the genetic location and linkage of the resistance markers.

An improved understanding of the role of pili in adherence of Haemophilus influenzae type b to human epithelial cells (EC) would enhance knowledge of the pathogenesis of H. influenzae b infections. In this study a highly sensitive in-vitro assay allowed the quantitative assessment of H. influenzae b adherence to EC. The degree of adherence was influenced by incubation time, temperature, bacteria/EC ratio, EC type and the growth phase of the bacteria. Most serially subcultured (SC) capsular type-b strains originally isolated from cerebrospinal fluid, blood, nasopharynx or throat gave similar low degrees of adherence, as did representative single strains of capsular types a, c, d, e and f. SC non-capsulated H. influenzae strains adhered in significantly greater numbers than most SC capsulated strains (p< 0.001). One SC type-b strain isolated from a throat, with stable piliation, adhered in very high numbers despite capsulation. Filiated subpopulations selected from type-b capsulated strains adhered in greater numbers than did their parent strains. These data suggest that capsulation of H. influenzae is a deterrent to adherence of the bacteria to EC. However, the presence of pili may allow type-b organisms to overcome the effects of capsulation.

Forty-three strains of Campylobacter fetus ss. fetus isolated from sheep abortions in New Zealand, and reference strains of C. fetus ss. fetus (four), C. fetus ss. venerealis (two), C. jejuni (one) and C. coli (one) were examined by restriction endonuclease analysis with the enzymes BstE II and Xho I. DNA fragment patterns of C. fetus, C. jejuni and C. coli differed strikingly from each other, but there were many similarities in the patterns obtained for all strains of C. fetus ss. fetus (47) and C. fetus ss. venerealis (three). The 43 local strains of C. fetus ss. fetus were clearly divided into four types. Two of these types produced patterns nearly identical with those of overseas reference strains.

The iron concentration in the culture medium used to prepare the inocula influenced the virulence of Pseudomonas aeruginosa in a chronic pulmonary infection model in rats. Groups of rats were given transtracheal inocula of agar beads in which were embedded c. 104 cfu of P. aeruginosa strain PAO and the mutants of strain PAO, Fe5 and Fe18. When strain PAO was grown in low-iron medium before infection, it caused severe parenchymal changes including a dense mononuclear cell infiltration in the alveolar spaces, as well as intra- and peribronchial inflammation. When strain PAO was grown in high-iron medium, the pathological changes in lungs were restricted to intra- and peribronchial inflammation. Strain Fe5, in which the effect of iron on yields of elastase is deregulated, produced similar pathological changes regardless of whether it was grown in low- or high-iron media. All rats infected with strain Fel8, in which the effect of iron on yields of toxin A is deregulated, died within 48 h after infection. These data indicate that the iron concentration of the culture medium can influence the pathogenesis of P. aeruginosa in a chronic respiratory infection. These studies also suggest that the regulation of extracellular virulence factors by iron is important in the determination of P. aeruginosa virulence.