- Volume 51, Issue 5, 2002

Streptococcus mutans and S. sobrinus are associated with the development of dental caries. These bacteria were detected by PCR and then their presence was compared with the incidence of dental caries in 77 Japanese pre-school children. Plaque samples were collected from all erupted tooth sites in the subjects, aged 3–5 years old and each with primary dentition, with a sterile toothbrush. A dental examination was performed for dmft (decayed, missing, filled, total) with the WHO caries diagnostic criteria. In all subjects, the prevalence of S. mutans and S. sobrinus was 72.8% and 61.1%, respectively; 19 (24.7%) were positive for S. mutans alone, 10 (13.0%) were positive for S. sobrinus alone, 37 (48.1%) were positive for both S. mutans and S. sobrinus, and 11 (14.3%) were negative for both S. mutans and S. sobrinus. The dmft scores of children positive for both S. mutans and S. sobrinus were significantly higher than those positive for S. mutans alone. These results indicate that children harbouring both S. mutans and S. sobrinus have a significantly higher incidence of dental caries than those with S. mutans alone.

The bacterial composition of human faeces can vary greatly with factors such as age and disease, although relatively few studies have monitored these events, particularly at species level. In this investigation, bacteria were isolated from faecal samples from healthy young adults and elderly subjects, and elderly patients with Clostridium difficile-associated diarrhoea (CDAD). The organisms were identified to species level on the basis of their cellular fatty acid profiles with the MIDI system. In some groups of bacteria, species diversity was found to change with age despite the overall numbers of organisms being similar at genus level. Bacteroides thetaiotaomicron, B. ovatus and Prevotella tannerae were common gram-negative anaerobes isolated from young adults. Bacteroides species diversity increased in the faeces of healthy elderly people. Bifidobacterial species diversity decreased with age, with Bifidobacterium adolescentis and Bif. angulatum being the most common isolates. CDAD patients were characterised by greater diversity of facultative species, lactobacilli and clostridia, but greatly reduced numbers of bacteroides, prevotella and bifidobacteria. Such bacterial population changes in the normal microbiota could result in metabolic conditions favourable for the establishment of pathogenic micro-organisms, such as clostridia, and would have considerable effects on the biochemical capacity of the large intestine as a whole. Alterations in the community structure of bifidobacteria and lactobacilli have relevance for dietary and therapeutic interventions such as the use of pre- or probiotics that aim to modify the composition or metabolic activities of the intestinal microflora in a beneficial way, particularly in elderly people or individuals at risk of CDAD.

Adhesion of Mycoplasma pneumoniae and the closely related M. genitalium to HEp-2 cells was investigated. The main surface proteins known to be involved in adhesion are P1 of M. pneumoniae and its homologue, MgPa, of M. genitalium. Both proteins are also immunodominant proteins. Protein P116 is another immunodominant protein of M. pneumoniae. These immunogenic proteins were investigated for their surface exposure and involvement in adhesion to host epithelial cells. Immunofluorescence microscopy (IFM) was used to detect M. pneumoniae and M. genitalium adhering to HEp-2 cells. Monospecific antibodies were produced against fragments of the surface proteins lacking tryptophan stop codons and were used for adhesion detection, surface exposure and adhesion inhibition IFM assays. Three monospecific antibodies were made against MgPa covering regions in the N-terminal, the middle and the C- terminal part; two monospecific antibodies were produced against P1 covering regions of the N- and the C-terminal part and one monospecific antibody was made against most of P116. Only the C-terminal parts of P1 and MgPa were surface exposed and blocking of these regions with the monospecific antibody resulted in inhibition of cytadsorption. Protein P116 was shown to be surface exposed and an essential protein involved in adhesion because the anti-P116 antibody prevented attachment of M. pneumoniae to the HEp-2 cells independently of P1. This study adds to the understanding of the molecular biology of M. genitalium and M. pneumoniae and presents a method to study the proteins involved in adhesion of these mycoplasmas.

DNA-PCR and reverse transcription (RT)-PCR for the 18-kDa protein of Mycobacterium leprae were used to examine the efficacy of multi-drug therapy (MDT) in leprosy. MDT was administered for 0–24 months. Fourteen (63.6%) of 22 patients showed positive PCR results after treatment for 12 months and the positive results decreased to 30% after 24 months of MDT. These results did not correlate with the bacterial index (BI) or the IgM antibody titre for the phenolic glycolipid (PGL)-1. One-dimensional densitometric analysis of agarose gels from PCR from the longitudinal study showed a gradual reduction of the 360-bp band after 12–24 months of MDT. RT-PCR for mRNA of the 18-kDa protein successfully tracked bacterial RNA changes in the biopsies and confirmed a decrease in the RNA of M. leprae in patients after MDT for 12 months. Thus, DNA- and RT-PCR for the 18-kDa protein of M. leprae are effective in assessing the efficacy of MDT for leprosy.

Vibrio cholerae is again the subject of attention on account of the current increase in the world-wide incidence of cholera. In this study, 200 clinical isolates of V. cholerae serotypes O1 and non-O1, non-O139, were collected from different provinces in Iran. The isolates were subjected to biochemical analysis, antibiogram, PCR of toxin genes, plasmid profile, ribotyping and pulsed-field gel electrophoresis (PFGE). The analysis of plasmid content showed that 33–96% of V. cholerae isolated from different provinces carry a large plasmid. PCR analysis of V. cholerae O1 showed that the genes encoding cholera toxin (ctx), toxin co-regulated pilus (tcp), accessory cholera enterotoxin (ace) and zonula occludens toxin (zot) were present in 55–97% of isolates in different provinces. Restriction fragment length polymorphism (RFLP) of BglI-digested DNA probed with five oligonucleotides revealed three different ribotype patterns in isolates of V. cholerae O1. The ribotype pattern B21 of V. cholerae O1 El Tor was found to be the predominant pattern in the isolates studied. V. cholerae non-O1, non-O139 isolates showed a single ribotype pattern. PFGE analysis also showed 10 different patterns amongst the isolates, 9 of which were in V. cholerae O1. Overall, the analysis of polymorphism of ribotypes and PFGE patterns of the isolates showed that the provinces in Iran were affected by a limited number of clones of V. cholerae O1 and non-O1, non-O139 strains.

Clinically important fungi such as Candida albicans and Cryptococcus neoformans are known to undergo phenotypic changes after repeated subculture or passages in vivo. However, there are no reports describing this phenomenon in Trichosporon species. This study investigated whether in-vivo passages of environmental isolates of Trichosporon asahii in mice changes their phenotype; three environmental isolates and 14 clinical isolates (from deep-seated infections) were used. The shape of the colony and cell type were observed, and the titre of glucuronoxylomannan (GXM) antigen and concentration of (1←3)-β-d-glucan were measured for each isolate. Changes in these features were also examined after three passages of the environmental isolates in mice. The shape of colonies and cell types were clearly different in environmental and clinical isolates. Furthermore, the clinical isolates released significantly higher levels of GXM antigen than environmental isolates (titre: log2 9.4 SD 0.7 versus log2 5.4 SD 1.4). The phenotype of passaged isolates was significantly different from the original environmental isolates with respect to the morphology of colonies and cell type and GXM release (titre: log2 10.0 SD 0.7 versus log2 5.4 SD 1.4). These results suggest that the phenotypic changes in T. asahii occur as a result of in-vivo passages. This process may allow a proportion of the fungal population to escape eradication by the host immune system, as GXM antigen is considered to protect the fungi against phagocytosis by polymorphonuclear leucocytes and monocytes in vivo.