- Volume 41, Issue 3, 1994

An isolate of Streptococcus intermedius from a brain abscess showed neuraminidase (sialidase), β-D-galactosidase, N-acetyl-β-D-glucosaminidase and N-acetyl-β-D-galactosaminidase activities. The optimal pH values of these enzymes were 5.5–6.0, 5.5–6.0, 5.0–5.5 and 5.0–5.5, respectively. The km of the enzymes varied according to whether the type of substrate was chromogenic or fluorogenic; sialidase was most active at the lowest substrate concentrations, with a km of 0.01 mM. In semi-defined medium, with porcine gastric mucin—a model glycoprotein—as the sole source of fermentable carbohydrate, levels of the glycosidases were significantly increased. Addition of glucose to the mucin-containing medium, or growth of cells in media supplemented with glucose alone, repressed glycosidic activities and the majority of these were cell-associated. S. intermedius cells from cultures grown with mucin were able, simultaneously, to transport via sugar: phosphoenolpyruvate phosphotransferase (PTS) systems, monosaccharides which are constituents of carbohydrate side chains of glycoproteins. These cells also possessed significant levels of neuraminate-pyruvate lyase, involved in the intracellular catabolism of neuraminic acid; this was absent from cells grown with glucose. These mechanisms, collectively, may facilitate the persistence and growth of S. intermedius in vivo.

A study of Chlamydia trachomatis infection was conducted in two stages on 15656 subjects at urogenital clinics of the Faculty of Medicine and Surgery at La Sapienza University in Rome, the S. Anna Hospital in Turin, and the Niguarda Hospital in Milan. The overall incidence of the disease was 6.4% in patients examined throughout the whole study period. The rate of positive cases was 5.8 % for the 5270 patients examined up to 1990, and 6.7 % for the 10386 patients examined from 1990 to 1992, showing an increasing trend. There was a much higher positivity rate in men (9.8%) than in women (6.0%); the difference was statistically significant. Of all patients, 60%, were asymptomatic. In symptomatic patients, C. trachomatis was present in 18.5 % of cases of non-gonococcal urethritis and in 12.8 % of cases of salpingitis. The highest incidence of C. trachomatis infection was in women who had begun sexual activity at an early age, (under 25 years in age), had several sexual partners and used intra-uterine contraceptive devices or spermicides or both.

Considerable biochemical diversity and polynucleotide sequence variation have been reported amongst strains of Citrobacter spp. However, sequence heterogeneity has not been investigated at gene loci of clinical relevance. In this study, sequence heterogeneity in the β-lactamase structural gene, ampC, amongst 91 clinical isolates of Citrobacter spp. that showed resistance to various third-generation cephalosporins was investigated. Variation was examined by high-stringency polymerase chain reactions (PCR) with primers homologous to the known ampC sequences of C. freundii strains OS60 and I113, and C. diversus NF85. If an isolate contained an ampC gene homologous to one of these three characterised ampC genes, a single PCR band of a predictable size was generated with the appropriate primer set; 50 (60 %) of isolates gave a PCR product of the expected size with the OS60 primer set and nine (10%) gave a product with the I113 primer set. All these 59 isolates were identified as C. freundii by API-20E strips. Six isolates (7 %) gave a product with the C. diversus NF85 primer set but only four of these were identified as C. diversus in API-20E tests; the other two isolates were identified as C.freundii. Of the 91 isolates, 28 (31 %), were identified as either C.freundii or C. diversus, but gave no PCR product with any primer set tested. Five of these showed no homology to any of the reference strain ampC PCR products in hybridisation tests. Nevertheless, all showed β-lactamase activity. Overall, this method allowed the identification of novel ampC gene loci, which may serve as a basis for the identification of Citrobacter spp. rapidly at a molecular level.

A seroreactive protein (TB66) was purified from culture filtrate (CF) and cell sonicate (CS) of Mycobacterium tuberculosis H37Rv by immobilised metal affinity chromatography (IMAC) on a Ni-nitrilotriacetic acid (NTA) column. The TB66 preparations obtained by IMAC contained predominantly a 66-kDa protein with a pI of c. 5.5 as determined by two-dimensional electrophoresis. TB66 was detected in the CF as early as 1 week of growth of H37Rv. The NH2-terminal amino-acid sequence showed 85 % homology with the N-terminal sequence of bovine serum albumin (BSA) and 80 % homology with human serum albumin. Amino-acid analysis indicated a difference in the amino-acid content of TB66 when compared to BSA, with an abundance of acidic amino acids. A monoclonal antibody (MAb) OD4AG3, raised in this laboratory against an M. avium complex (MAC 101) sonicate cross-reacting with H37Rv sonicate, recognised a heat-stable and trypsin-sensitive epitope of this protein. TB66 was also recognised by MAbs IT1 and IT20 which also react with the 14-kDa antigen of the M. tuberculosis complex. Antibodies against TB66 were present in the sera of 62 of 64 patients with tuberculosis; sera from normal healthy individuals showed no significant reactivity. TB66 appears to be a predominant secretory protein of M. tuberculosis and could play an important role in the pathogenesis of this organism.

Isolates of Fusarium species from 18 patients with keratomycosis were examined for their C-29 and C-31 sterol content and for their capacity to synthesise mycotoxins. All isolates were resistant to azole antifungal agents in vitro and the sterol contents were indistinguishable. In-vitro toxin production was monitored by gas chromatography–mass spectrometry; 13 isolates produced nivalenol, six produced deoxynivalenol, nine gave T-2 toxin and two showed the presence of diacetoxyscirpenol at different time intervals. However, neither sterol content nor toxin production in vitro appeared to be related to the severity of infections observed in patients.