Phenotypic switching and genetic diversity of Cryptococcus neoformans.

Niger seed agar was used as a primary plating medium for the isolation of Cryptococcus neoformans from cerebrospinal fluid specimens from AIDS patients with untreated primary cryptococcosis. The medium was used as the primary means to detect variations in the colony morphology of the yeast. To search for phenotypic and genetic variations, nine patients individually harboring two or three types of colony morphology were studied. Intraindividual isolates from nine patients had minor variations in the API 20C profile, and the MICs of one or more antifungal agents (amphotericin B, fluconazole, and itraconazole) for isolates from three patients were significantly different. Intraindividual isolates from three patients had minor karyotype differences, and one showed a dramatic chromosomal length polymorphism. In addition, three serial isolates from a patient with two episodes of infection showed similar karyotypes, confirming persistent infection by the same strain. Random amplified polymorphic DNA products were identical for all isolates (including three isolates from a relapse case). Our results provided evidence suggesting that (i) in humans, C. neoformans may undergo phenotypic and genetic changes during early infection prior to antifungal agent administration; (ii) dramatic variations in electrophoretic karyotypes and in phenotypes, as demonstrated during the early infection of one patient, may be due to infection by different strains; and (iii) the use of niger seed agar as a primary plating medium is useful for studying antifungal susceptibility, phenotypic switching, genetic diversity, and multiple-strain infections.

Antilipopolysaccharide II: an antibody protective against fatal melioidosis.

This was a study of IgG antibody responses to two S-type lipopolysaccharides (LPS I and LPS II) and flagellin of Burkholderia pseudomallei in patients with melioidosis. The specificity of these antibodies was 91.7%, 90.3%, and 93.8%, respectively, when compared to responses in a population where the organism is not endemic. Only the level of antibody to LPS II (anti-LPS II) was significantly higher in patients who survived than in those who died, as well as in patients with nonsepticemic vs. septicemic melioidosis. Results of logistic regression analysis, controlled for confounding factors such as duration of illness before treatment and bacteremic status, confirmed that a high level of anti-LPS II was a significant factor protective against fatal melioidosis. Thus, LPS II of B. pseudomallei would be a potentially useful component of a vaccine developed against fatal melioidosis. Further studies are in progress to determine the level of this antibody among those with asymptomatic infection in areas where melioidosis is endemic.

Recovery of Cryptococcus neoformans from the nasopharynx of AIDS patients.

Nasopharyngeal swabbings, obtained from AIDS patients, were plated onto Niger seed agar containing antibiotics Cryptococcus neoformans was isolated from 35 out of 84 patients (41.7%) diagnosed as primary cryptococcal cases before antifungal administration, and 8 out of 86 (9.3%) cryptococcosis patients on antifungal therapy. The fungus could not be isolated from any of 447 samples from 194 AIDS patients not diagnosed with cryptococcosis. These findings are novel in that the presence of C. neoformans in AIDS patients at this site has never been looked at previously.

Pathogenicity of basidiospores of Filobasidiella neoformans var. neoformans.

Basidiospores of Filobasidiella neoformans var. neoformans (progeny of Cryptococcus neoformans MT 100.1 x VR 45980) were able to induce cryptococcosis in Swiss albino mice if inoculated by intraperitoneal injection, nasal instillation or nasal spraying. The latter method, with the aid of a jet nebulizer, was first adopted to imitate the natural entrance of infectious particles. Using this method the small number of basidiospores (7 x 10(3)) could induce cryptococcosis in mice, while the higher number of the parental laboratory-grown yeast cells (1.5 x 10(6)) did not produce infections. By nasal instillation Cyclophosphamide (Cy)-treated mice were more susceptible to the basidiospores, showing 80% cryptococcosis (eight of 10). Seven of the eight infected mice had disseminated cryptococcosis. Immunocompetent mice were more resistant to basidiospore infection than Cy-treated mice, as 40% of that group developed only pulmonary cryptococcosis; none had disseminated infection. Thus, we propose that basidiospores are one form of the infectious propagules of F. neoformans var. neoformans which can cause cryptococcosis, particularly in immunocompromised people.

Streptococcus suis toxic-shock syndrome and meningitis.

Three cases with S. suis bacteremia and meningitis were reported. The first case was a 23-year-old butcher who was a regular drinker of alcohol for two years and developed streptococcal toxic-shock syndrome. The organism was transmitted to him through a minor cut in his right arm. The second cases was a 49-year-old female laborer who had been consuming locally produced alcohol for 20 years and developed fever and meningitis. Unfortunately, she succumbed in seven days despite intensive supportive and cefotaxime treatments. The third case was a 45-year-old regular alcoholic drinker and car painter who was seen at a private hospital due to contusion at his left lateral chest wall. However, fever and confusion due to meningitis was detected upon admission. Irreversible deafness developed within 48 hours of ceftriaxone therapy for meningitis. He finally recovered with deafness. S. suis was isolated from blood and cerebrospinal fluid cultures in all three cases though initially reported to be viridans group of streptococci.

Serotypes of Cryptococcus neoformans isolated from patients prior to and during the AIDS era in Thailand.

One hundred and eighty-seven strains of Cryptococcus neoformans isolated from patients in Thailand were characterized by biochemical varieties relating to serogroups. Canavanine-glycine-bromothymol blue (CGB) agar was used for differentiating the varieties of C. neoformans. Slide agglutination tests were performed with Crypto Check (Iatron, Inc., Tokyo) to determine their serotypes. Fifty-five percent (10 out of 18) of the pre-AIDS isolates were serotype B, 28% were serotype A, 5% were serotype D, and an unexpected 11% (2 out of 18) were serotype C. These are the first to be recorded in Asia. In contrast, among the 169 clinical isolates obtained between January 1993 and March 1995 (AIDS epidemic), serotype A was outstandingly predominant--93% (157 out of 169), serotype B was relatively low (3.6%) and both serotypes D and AD were 1.8%. The pattern of serotypes of the 59 isolates from known HIV-positive patients was closely similar to the total isolates during the AIDS epidemic. In determining the varieties of C. neoformans by CGB, only 1 of the 187 isolates gave a false reaction. On the basis of our findings, we believe that in the pre-AIDS era either C. neoformans var. gattii serotype B or serotype C were the common causative agents of cryptococcosis in Thailand. The advent of AIDS changed the pattern of serotypes with serotype A becoming predominant as has been reported world wide.

Occurrence of Penicillium marneffei infections among wild bamboo rats in Thailand.

Penicilliosis marneffei has emerged as an endemic systemic mycosis in Southeast Asia among humans and wild bamboo rats. To gain an insight into the epidemiology of this life-threatening disease, a survey of bamboo rats for natural infections by Penicillium marneffei was carried out in the central plains of Thailand during June-September, 1987. Thirty-one lesser bamboo rats (Cannomys badius) and eight hoary bamboo rats (Rhizomys pruinosus) were trapped. Portions of their internal organs were cultured to determine if they had been infected by P. marneffei. Six each of C. badius (19.4%) and R. pruinosus (75%) yielded cultures of this unique, dimorphic Penicillium species. All of the isolates were readily converted to their unicellular form that multiplies by the process of schizogony by incubating them at 37 degrees C on plates of brain heart infusion agar. Their identity was further confirmed by a specific immunological test. Among the internal organs of the positive rats, the lungs had the highest positivity (83.3%), next in decreased order of frequency were the liver (33.3%) and the pancreas (33.3%). The use and value of domestic and wild animals in locating and demarcating endemic areas of geophilic fungal pathogens are discussed. Penicilliosis marneffei is considered to be a zooanthroponosis--a disease that occurs in lower animals, as well as, humans.

Hygienic status of food handlers.

The study demonstrated bacterial species on hands and nails of food-handlers before and after hand-washing. Those were Staphylococcus spp., Streptococcus spp., Micrococcus spp., Bacillus spp., Diphtheroid, Aeromonas hydrophila, Klebsiella pneumoniae, Acinetobacter, Enterobacter cloacae, Escherichia coli, Pseudomonas spp., Proteus mirabilis, Serratia spp., Citrobacter freundii. Before hand washing, each food-handler harboured one to eight bacterial species. After hand-washing (eight with water from plastic boxes, 97 from pipe water, 57 out of 97 (58.8%) used soap or detergent with water), disappearance of one to four bacterial strains from hands and nails were found in 47.6 per cent of food-handlers. Cultures of water used for washing from eight plastic boxes yielded Staph. spp., Strep. spp., Aeromonas hydrophila, Kleb.pneumoniae, Acinetobacter anitratus, Enterobacter cloacae. From pipe water, Diphtheroid in 4, 4.1 per cent Micrococcus in 1, 1.03 per cent were shown. Comparing bacterial species found in food-handlers with long nails and short nails, 4-8 more species were revealed in the former than the latter for 35.7 per cent. After hand-washing, there was recontamination of bacterial species in 17 food-handlers. This was probably due to dirty napkins or dresses during hand-drying or from water in plastic boxes.

Persistent bacterial endocarditis caused by three isolates of satellite streptococci.

Three isolates of satellite streptococci were cultivated from the blood of a patient affected persistent bacterial endocarditis. They had distinguishable ultrastructural abnormalities. Their cell wall architecture changed from a fuzzy coat (first isolate) to a thick electron transparent layer covered with a rough fuzzy coat (second isolate), and to electron dense globular material which detached from the wall in small patches (third isolate). The antibiotics probably played an important role in changing their architecture. These three isolates were probably derived from the same strain, since they had common biochemical characteristics and they were isolated from the same patient during the course of his endocarditis.

Cholecystitis associated with Enterobacter agglomerans.

A case report of chronic cholecystitis due to Enterobacter agglomerans, occurred in a 54-year old female with homozygous beta-thalassemia. The patient responded successfully to cholecystectomy and sulfamethoxazole + trimetroprim therapy. The source of the infection was not known, however, cystic duct obstruction and immune deficit were thought to be the predisposing causes.

Distribution of IMVC biogroups of Aeromonas hydrophila and Aeromonas sobria isolated from human, fish and water.

Aeromonas sobria and A.hydrophila were isolated from infected fish in ratio of 3.5:1 during the outbreak of fish infections from December 1982 to February 1983, while isolates from human diarrheic stool was 1 :2. On the basis of IMVC reactions 138 isolates of motile aeromonads could be divided into 11 biogroups, with biogroup 4 showing statistically significant association with infections. Nine biogroups of aeromonads which were isolated from infected fish reflected that the outbreak was not caused by a single type of bacteria. There may have been some common factors which acted as predisposing causes. The possibility of zoonosis spreading of this epidemic infection of fish was low, because the majority of the infective agents in man and fish were different.

Free coagulase and thermostable nuclease reactions of staphylococci: a comparison.

Two-hundred-fifty-five staphylococcal isolates (clinical) were tested for both free coagulase (CGase) and thermostable nuclease (TNase) production. Two-hundred-ten of the 255 isolates were identified as Staphylococcus aureus when both strongly positive TNase and CGase reactions (human and rabbit plasma) were observed with these strains. Forty-five of the 255 isolates were identified as Staphylococcus species, other than S. aureus when either weakly positive TNase and CGase (rabbit plasma) reactions or negative TNase and CGase (rabbit and human plasma) reactions were observed. This experiment suggest that the identification of staphylococcal isolates as S. aureus can be based upon demonstration of both strongly positive TNase and CGase reactions.