Heterogeneity within apparently pure cultures of Escherichia coli freshly isolated from significant bacteriuria.

Pyrolysis mass spectrometry (PyMS), a highly discriminatory method for comparison of isolates, was used to assess the homogeneity of colonies taken from apparently pure cultures of Escherichia coli in 10 urine specimens. Ten randomly selected colonies were subcultured from each urine for comparison. For six urines, the set of 10 single-colony isolates proved indistinguishable by PyMS. However, for four urines there was clear heterogeneity. For two urines, one of the isolates was distinct, and the remaining nine indistinguishable. One urine yielded seven indistinguishable isolates, and three further individually distinct isolates. One urine yielded two clusters of indistinguishable isolates, one comprising seven and the other three isolates. Thus, significant heterogeneity (as measured by PyMS) occurs in apparently pure cultures of E. coli from urinary tract infection. The nature of this heterogeneity remains to be established, as does the significance, if any, of this phenomenon in urinary tract infection.

Crystal violet reactions of coagulase negative staphylococci.

Twenty four reference strains and 112 clinical isolates of coagulase negative staphylococci (CNS) were examined for their reactions in the crystal violet test. Some species gave a white reaction and others a purple reaction. Results were consistent and reproducible and each species gave only one pattern of crystal violet reaction. Within the limited variety of species represented in the clinical isolates, Staphylococcus saprophyticus and S haemolyticus gave crystal violet purple reactions, in contrast to S epidermidis, which always gave a white reaction. Investigations suggested that the mechanism of the crystal violet test in S haemolyticus may be similar to that previously described in S aureus. Further work is needed to characterise the ability of crystal violet to modify S epidermidis and other central nervous system species. The crystal violet reaction, which has strong associations with invasiveness, phage group susceptibilities, colonisation persistence abilities, and nosocomial origin in S aureus may also be useful in studies of CNS disease.

Crystal violet reactions of Staphylococcus aureus strains colonizing infants in the first six weeks.

Nasal colonization with Staphylococcus aureus occurred in 18% of babies leaving a maternity unit and had risen to 40% by 6 weeks after birth. S. aureus was first acquired by 34.5% of babies after discharge. Female infants were more likely to be colonized than males. Colonization was not significantly different between babies receiving standard postnatal care and those nursed on the Special Care Baby Unit. Crystal violet (CV) tests showed that purple-reacting isolates accounted for approximately 60% of strains, whether first detected at hospital discharge or subsequently acquired. Purple-reacting strains, once acquired, were significantly better able to persist than non purple-reacting strains and formed a cumulatively higher proportion of the strains isolated at 6 weeks after birth than at hospital discharge. CV purple-reactions were significantly associated with lysis by phages of groups III and I and non-purple-reactions were significantly associated with lysis by phages of group II and/or 94/96. Maternity units remain a significant route whereby strains of S. aureus with some characteristics associated with a hospital origin gain access to the community.

A study into the mechanism of the Crystal Violet reaction in Staphylococcus aureus.

The mechanism of the Crystal Violet (CV) reaction, a trait which has been related to biotype, source and pathogenicity in Staphylococcus aureus, was investigated in agar and broth studies. White reactions could be converted to purple and vice versa by altering the incubation conditions on agar. Broth reactions examined macroscopically and by spectrophotometry revealed that both white and purple human biotype strains take up CV but the former then progressively modify the dye more quickly than the latter. A cell-associated product of CV was detected in white and purple strains by reverse-phase thin-layer chromatography of methanol extracts. White strains appear to produce a second additional product from CV. The white reaction was not inhibited by chloramphenicol or azide but did depend on viable cells with a nutrient source. CV MICs and MBCs for 10 white and 10 purple reactors showed no gross differences in susceptibility, while a standardized assay for the rate of CV modifying activity (52 strains) demonstrated that the two categories comprise discrete populations which alter CV at different rates. Although most white strains belong to either or both of phage-typing groups V and II, purple strains with this pattern of susceptibility and white strains without it both occur. The capacity to modify CV slowly or rapidly appears to subdivide human biotype strains independently of their phage group and is associated in the former case with their capacity to produce hospital-acquired and invasive infections.

Rapid inter-strain comparison by pyrolysis mass spectrometry in nosocomial infection with Xanthomonas maltophilia.

Seventeen strains of Xanthomonas maltophilia and one strain of Pseudomonas cepacia were examined by pyrolysis mass spectrometry (PYMS). The Xanthomonas strains comprised 11 clinical and environmental isolates from a suspected outbreak of colonization and infection on a heart-lung transplant intensive care unit, two strains from patients elsewhere in the same hospital and four strains from a national reference collection. The single isolate of Pseudomonas cepacia was from a sink in the same affected intensive care unit. A series of discriminant analyses performed on the PYMS-derived data showed that, whereas six strains of Xanthomonas from the respiratory tract, blood and ventilatory equipment of one of the affected patients were indistinguishable, all the other isolates were distinct. The results of PYMS rapid inter-strain comparison were in accord with those of an epidemiological investigation which suggested that the episode was due to unauthorized reuse of disposable nebulizers and not to cross-infection between patients. Pyrolysis mass spectrometry with rapid data analysis is a potentially useful technique for the investigation of nosocomial infections due to organisms such as X. maltophilia.

Crystal violet reactions of fresh clinical isolates of Staphylococcus aureus from two British hospitals.

When 168 fresh clinical isolates of Staphylococcus aureus were examined for their reactions on a medium containing 1 part in 100,000 crystal violet 50.6% of strains produced a purple appearance, 39.3% produced a white appearance and 10.1% produced a yellow appearance. Purple-reacting isolates were significantly associated with both invasive infections (P less than 0.01) and hospital origin (P less than 0.001). There were no significant associations between the crystal violet reactions and either animal contact or other properties previously reported to be characteristic of white and yellow-reacting strains (beta haemolysin and bovine coagulase production). The results of phage typing showed associations between susceptibility to group III phages and purple-reacting strains and between phage group II susceptibility and white and yellow-reacting strains. There was also a highly significant association between white reactions on crystal violet agar and susceptibility to lysis by a combination of all three groups (that is, I + II + III) and white-reacting strains were significantly more susceptible to lysis by phages 94 and/or 96, whether as a restricted pattern or as part of a broader pattern. The purple reaction on crystal violet medium may be a reliable marker of the 'hospital staphylococcus'.