Susceptibility of Cryptococcus neoformans by the NCCLS microdilution and Etest methods using five defined media.

The susceptibility of 12 isolates of Cryptococcus neoformans to amphotericin B, 5-fluorocytosine, fluconazole, itraconazole and ketoconazole was tested using the NCCLS and Etest methods with yeast nitrogen base (YNB) pH 5.6 and pH 7.0, RPMI MOPS pH 7.0 with and without added glucose (2%) and RPMI buffered with phosphate buffer to pH 7.0. Some isolates yielded poor growth in RPMI MOPS after 72 h. Tests indicated that YNB pH 5.6 was the best medium for 5-fluorocytosine but was unsuitable for ketoconazole. In conclusion, YNB pH 7.0 or RPMI MOPS with 2% glucose can be used with either method.

An outbreak of methicillin-resistant staphylococcus aureus (MRSA) in a dermatology day-care unit.

We describe an outbreak of methicillin-resistant Staphylococcus aureus (MRSA) in a dermatology day-care unit and the methods used to determine the mechanism of spread and control it. The epidemic strain had a characteristic sensitivity pattern and was typeable with phages 29, 80, 95, 47, 54 and 77, which was of considerable value in interpreting the epidemiological data. The method of spread was studied by examination of the medical and nursing records of patients who had acquired MRSA (to determine which members of staff they had encountered and which other MRSA-positive patients had been present in the department at the same time) and by the microbiological screening of all patients and staff. However, screening of all staff by nasal swabbing failed to identify carriage of the epidemic strain, while extensive swabbing of surfaces on the day-care unit also failed to show any evidence of MRSA in the environment. This suggests that the MRSA was most probably spread from patient to patient via the hands of staff, although there was also the possibility of direct transmission from patient to patient. Nine patients acquired the unique strain of MRSA and once acquired it proved difficult to eradicate, although in the majority, the infection did not appear to be clinically significant. However, in two patients MRSA contributed to a fatal outcome: these were the two most elderly patients and were the only two who were receiving systemic corticosteroids. The outbreak was brought under control with rigorous hygienic measures and the decision to discharge all patients with MRSA from the day-care unit. Repeat screening (swabs of nose, axilla and groin) of all day-care unit and in-patients 11 months after the last MRSA case showed no evidence of any residual MRSA infection in the day-care unit.

Comparison of a new insertion element, IS1407, with established molecular markers for the characterization of Mycobacterium celatum.

Genomic analyses of 18 Mycobacterium celatum strains obtained from different patients in three countries (United States, United Kingdom, and France) were performed; the methods used in this study were restriction fragment length polymorphism (RFLP) analysis, pulsed-field gel electrophoresis (PFGE) analysis, and PCR restriction analysis (PRA) of the hsp-65 gene. A new insertion sequence, IS1407 (GenBank accession no. X97307), belonging to the IS256 family, was identified in M. celatum type 1 strains and was characterized by sequencing. When a probe for Mycobacterium xenopi IS1395-like sequences was used, the RFLP analysis of M. celatum type 1 strains revealed that they contained three or four copies of IS1407 in identical genomic positions, while this element was absent in all M. celatum type 2 strains. PFGE performed with three different endonucleases revealed a unique large restriction fragment (LRF) pattern for M. celatum type 1 strains, whereas the LRF patterns obtained for M. celatum type 2 strains were polymorphic. Moreover, PFGE of nondigested genomic DNA revealed extrachromosomal elements in M. celatum type 2. The type strain of M. celatum type 3 could not be differentiated from M. celatum type 1 strains on the basis of the results of the RFLP analysis, the PFGE analysis, and the PRA of IS1407. In this study we confirmed that M. celatum types 1 and 2 represent distinct genomic clusters and that the molecular markers in M. celatum type 2 exhibit greater polymorphism than the molecular markers in M. celatum type 1.

Pilot studies of azithromycin, letrazuril and paromomycin in the treatment of cryptosporidiosis.

Pilot studies of the safety and efficacy of 3 drugs thought to have anticryptosporidial activity were carried out to determine whether any of them are suitable for large-scale clinical trials. Open studies of the use of azithromycin, letrazuril and paromomycin in patients with acquired immunodeficiency syndrome (AIDS) and confirmed cryptosporidial diarrhoea for at least a month. Azithromycin 500 mg daily was ineffective. Letrazuril 150-200 mg daily was associated with an improvement in symptoms in 40% of patients treated and cessation of excretion of cryptosporidial oocysts in the stool in 70%; however biopsies remained positive. Paromomycin therapy was associated with a complete resolution of diarrhoea in 60% of patients treated and some improvement in symptoms in a further 5% but it did not eliminate the infection. None of the drugs had any major toxicities. Dose escalation studies of azithromycin should be performed. Letrazuril should be further investigated for efficacy in double-blind placebo-controlled trials. Paromomycin appears to result in prolonged symptomatic remission of cryptosporidial diarrhoea, but has no effect on cryptosporidial cholangitis.

A new group (type 3) of Mycobacterium celatum isolated from AIDS patients in the London area.

We describe a new group (type 3) of the recently proposed species Mycobacterium celatum isolated from eight patients with AIDS in London, England. Sequences of genes coding for 16S rRNA (EMBL accession no. Z46664) showed a divergence of 17 bases from M. celatum type 2 reference isolates and a divergence of 7 bases from M. celatum type 1 reference isolates. A reference strain is available (NCTC 12882).

Rapid identification of mycobacteria from AIDS patients by capillary electrophoretic profiling of amplified SOD gene.

Aim-Rapid differentiation of mycobacterial species at the genomic level.Methods-The manganese superoxide dismutase (SOD) gene (464 bp) and 16SrRNA (353 bp) from 104 isolates (18 species) of mycobacteria were amplified using polymerase chain reaction (PCR). Products were sequenced and a phenogram of SOD sequences derived. PCR products of SOD gene were digested with HaeIII, and restriction fragment profiles visualised using capillary electrophoresis.Results-Novel SOD sequences were found for M szulgai, M marinum, M phlei, M smegmatis, M chelonei, M paratuberculosis, M malmoense, M intracellulare serotype 7, M intracellulare serotype 18, and M celatum types 1, 2, and 3. Phylogenetic analysis indicated that 18 of 19 species studied had 8-29% interspecies and <6% intraspecies sequence diversity in the SOD gene. No consistent differences were detected between AIDS and non-AIDS isolates. M paratuberculosis showed a unique SOD sequence with a 1.1% (SD 0.5%) diversity from M avium. Capillary electrophoresis profiles were able to differentiate 16 of 18 species within 24 hours.Conclusions-A phenogram of SOD sequences clearly delineated all mycobacterial species and showed two distinct clusters, fast growing species, and the M avium complex (MAC). Within the MAC, M avium (five types), M intracellulare (five types), M scrofulaceum (two types), and M paratuberculosis (one type) could be demonstrated. Phylogenetic diversity of M celatum from MAC, previously suggested by 16SrRNA data, was confirmed. This simple and rapid method for DNA extraction, in conjunction with capillary electrophoresis of SOD restriction fragments, allows rapid identification of mycobacterial isolates.

Long-term impact of HIV diagnosis on mood and substance use--St Stephen's cohort study.

Twenty HIV positive and 68 HIV negative subjects were assessed by the Hospital Anxiety and Depression Scale and by the Alcohol and Drugs Frequency Schedule immediately prior to notification of their HIV serostatus and 6 months after serodiagnosis. The 2 groups did not differ significantly in levels of anxiety or depression at baseline or follow-up. There were borderline levels of pathological anxiety prior to notification of HIV serostatus in both groups. The drop to normal levels of anxiety which had occurred by follow-up was significant in the HIV positive group. About a third of subjects in both groups were regularly making use of alcohol and/or drugs, both at baseline and follow-up. Mean levels of weekly alcohol intake for both groups ranged from about 20 to 30 units per week. The drugs most commonly used (in any frequency) were nitrates ('poppers') and cannabis.

Septicaemia in patients with and without AIDS at Westminster Hospital, London.

The types of organism causing septicaemia in patients with AIDS and without AIDS at Westminster Hospital were examined prospectively over a period of 2 years (1990-1991). Altogether 417 episodes of septicaemia were diagnosed, 148 (35%) of which were in patients with AIDS. Of septicaemias in patients with AIDS, 53 (36%) were caused by mycobacteria. Non-mycobacterial septicaemias were associated with IV access devices in 58 (61%) of patients with AIDS and in 50 (19%) of those without AIDS. Gram-negative organisms were responsible for septicaemia associated with IV access devices in 16 (28%) of 58 patients with AIDS and in 8 (16%) of 50 patients without AIDS. Non-typhoidal Salmonella species or Shigella species caused 13 (31%) of 42 episodes of septicaemia caused by Gram-negative organisms in patients with AIDS. These findings have influenced the strategy for empirical therapy of septicaemia in patients with AIDS at Westminster Hospital.

The clinical comparison of Oxoid Signal with Bactec blood culture systems for the detection of streptococcal and anaerobic bacteraemias.

Blood cultures were taken from 47 patients 1-2 min after dental extraction. These samples were tested by the radiometric Bactec 460 and Oxoid Signal systems for the detection of streptococcal and anaerobic bacteraemias. Streptococci were isolated from 19 (40%) patients and anaerobes from 15 (32%). In this study the Oxoid Signal system was significantly better for isolating oral anaerobes than the Bactec system; five isolates were obtained with the Bactec system and 14 with the Signal system. There was no significant difference in the isolation of streptococci between these two systems (Bactec 14, Oxoid Signal 10). The contamination rate was 4.1% for Bactec and 7.5% for the Oxoid Signal system.

Salmonella, Campylobacter and Shigella in HIV-seropositive patients.

OBJECTIVE: To study the incidence, clinical features, treatment and outcome of patients with Salmonella, Shigella or Campylobacter infection. DESIGN: Retrospective analysis. SETTING: Two dedicated HIV units within a London teaching hospital. METHODS: All patients with Salmonella, Shigella or Campylobacter infection were reviewed retrospectively by correlating the records of the gastrointestinal and microbiology departments with the computerized records of all HIV-positive patients attending the two clinics. RESULTS: Between July 1985 and June 1991, 56 episodes of Salmonella, 37 of Campylobacter and eight of Shigella infection were documented in HIV-seropositive patients. Shigella was most likely to occur early in HIV disease, whilst patients with Campylobacter or Salmonella were more likely to have had a previous AIDS diagnosis. Septicaemica was most common in patients with Salmonella and was especially likely to occur in individuals with an AIDS diagnosis. Relapse of infection was common in patients with Salmonella, especially in those with low CD4 lymphocyte counts, those with an initial septicaemic illness and those not treated with ciprofloxacin. CONCLUSIONS: Patients with Salmonella who have low CD4 lymphocytes counts and/or a septicaemic illness should be considered for life-long secondary prophylaxis with ciprofloxacin because of the high rate of relapse observed. Administration of zidovudine or cotrimoxazole as prophylaxis against Pneumocystis carinii pneumonia may prevent the development of salmonellosis: significantly fewer patients with this infection were taking these drugs than patients with Campylobacter.

The in-vitro bactericidal activities of combinations of antimicrobial agents against clinical isolates of Mycobacterium avium-intracellulare.

The in-vitro activities of five antimicrobial agents (rifabutin, clarithromycin, ethambutol, ciprofloxacin and amikacin), alone and in combination, were evaluated against 21 strains of Mycobacterium avium-intracellulare isolated from patients with AIDS. The combined activities of these agents were studied on solid medium by a full chequerboard method. Synergy was demonstrated most frequently (28-71% of isolates) with those combinations that included ethambutol. In killing curve experiments where double and triple combinations of agents were tested against two of the strains, 99% kill was achieved in seven days at concentrations well below those that are attainable in serum. However, an additive rather than a synergic effect was seen in most instances. Although ciprofloxacin alone had the greatest bactericidal activity against these two strains, its activity was antagonized in the presence of rifabutin; this antagonism became inapparent when a third agent was added. Demonstration of bactericidal activity in broth culture may be more relevant than the results of susceptibility testing on solid medium when choosing antimicrobial therapy for patients with this infection.

Cryptosporidiosis in HIV-seropositive patients.

The incidence of cryptosporidiosis in our unit has increased over the last 6 years, being diagnosed in approximately 5 per cent of all patients with HIV infection and in 21 per cent of those with AIDS, but a marked seasonal variation occurs. We have studied the course of the infection in 128 patients and identified four clinical patterns of disease: transient (28.7 per cent), chronic (59.7 per cent), fulminant (7.8 per cent) and asymptomatic (3.9 per cent). Transient disease occurred in patients with a wide range of CD4 lymphocyte counts, but was more common in less immunosuppressed patients. Fulminant disease, defined by the passage of more than 2 l of stool/day from the time of presentation, only occurred in patients with a CD4 count less than 50/mm3. This group had lost more than 7 kg in weight at presentation and more commonly had other intercurrent gastrointestinal infections. They survived for a median of only 5 weeks, compared with 20 weeks for those with chronic diarrhoea and 36 weeks for those with transient infection. The survival was unaffected by any treatment other than zidovudine. Cryptosporidiosis in HIV-infected individuals is a heterogeneous disease.