No abstract

Aims-To detect and diVerentiate Mycobacterium tuberculosis simultaneously by polymerase chain reaction (PCR) in clinical samples prepared for histopathological analysis and for microscopic detection of acid fast bacteria. Methods-ParaYn wax embedded tissue samples and Ziehl-Neelsen (ZN) and auramine stained microscopic preparations from culture positive tuberculosis patients were subjected to DNA extraction and amplification by PCR. PCR was performed with primers specific for direct repeats and the product was detected by hybridisation to a set of 43 diVerent oligonucleotides, a procedure designated as "spoligotyping".

Detection of Shiga toxin-producing Escherichia coli (STEC) in The Netherlands is traditionally limited to serogroup O157. To assess the relative importance of STEC, including non-O157 serogroups, stool samples submitted nationwide for investigation of enteric pathogens or diarrhoea were screened with real-time PCR for the presence of the Shiga toxin genes. Patients were selected if their stool contained blood upon macroscopic examination, if they had a history of bloody diarrhoea, were diagnosed with haemolytic uraemic syndrome, or were aged <6 years (irrespective of the bloody aspect of the stool). PCR-positive stools were forwarded to a central laboratory for STEC isolation and typing. In total, 4069 stools were examined, with 68 (1.7%) positive PCR results. The highest prevalence was for stools containing macroscopic blood (3.5%), followed by stools from patients with a history of bloody diarrhoea (2.4%). Among young children, the prevalence (1.0%) was not significantly higher than among random, non-bloody, stool samples from diarrhoeal patients (1.4%). STEC strains were isolated from 25 (38%) PCR-positive stools. Eleven O-serogroups were detected, including five STEC O157 strains. As serogroup O157 represented only 20% of the STEC isolates, laboratories should be encouraged to use techniques enabling them to detect non-O157 serogroups, in parallel with culture, for isolation and subsequent characterisation of STEC strains for public health surveillance and detection of outbreaks.

EUCAST methodology resulted in higher co-amoxiclav E. coli resistance rates than CLSI methodology, but correlated better with clinical outcome. EUCAST-compliant microdilution and disc diffusion provided discrepant results.

Bacterial interactions in mixed infections may compromise antimicrobial therapy. The in vitro bactericidal activity of ampicillin and ampicillin-sulbactam against aerobic/anaerobic mixed cultures of Bacteroides fragilis, Escherichia coli and Enterococcus spp. was studied by means of broth dilution tests. The MBC of ampicillin for Enterococcus faecalis 6 was 0.25 mg/l when tested singly; in association with B. fragilis 1, however, the MBC for E. faecalis 6 was 16 mg/l. When tested singly E. coli 9 and B. fragilis 1 were both killed by ampicillin at a concentration of 4 mg/l in combination with 1 mg/l sulbactam. When both strains were associated, the MBC for B. fragilis 1 rose to > 256 mg/l. Results obtained indicate that the behaviour of bacteria as determined in pure cultures is not necessarily identical with the antibiotic susceptibility present in mixed cultures. It was shown that, besides beta-lactamase production by involved bacteria, other factors contribute to the alteration of bacterial susceptibility in mixed cultures. Synergistic and antagonistic effects between associated organisms were observed. For example, it was found that two moderately sensitive bacterial strains were resistant in mixed culture.