Characterization of clinical isolates of pathogenic <Emphasis Type="Italic">Nocardia</Emphasis> strains and related actinomycetes in Thailand from 1996 to 2003

In Thailand from 1996 to 2003, 171 strains of pathogenic aerobic actinomycetes from clinical specimens were isolated. Of those strains, 134 were mycolic acid containing actinomycetes, including 96 strains of Nocardia species. Others included 10 strains of Gordonia, 14 strains of Rhodococcus, and 22 strains of Mycobacterium. One strain each of the genera Tsukamurella and Corynebacterium were also isolated. Also identified were 27 strains of non-mycolic acid containing actinomycetes. Our identification studies of 96 strains of Nocardia species showed that significant pathogens in Thailand were N. beijingensis (18 strains), N. cyriacigeorgica (13 strains), and N. farcinica (34 strains); the most prevalent species was N. farcinica (35.4%). We also isolated four strains of N. asiatica, five strains of N. asteroides sensu stricto, four strains of N. nova, seven strains of N. otitidiscaviarum, eight strains of N. transvalensis, and two strains of N. pseudobrasiliensis.     

Three new basidiomycetous yeasts,Pseudozyma alboarmeniaca sp. nov., Pseudozyma crassa sp. nov. and Pseudozyma siamensis sp. nov. isolated from Thai patients

We previously reported the first isolation of Pseudozyma species from the blood of Thai patients. In this study, three additional new Pseudozyma species were isolated from clinical specimens from Thai patients. The Pseudozyma species showed relatively low sensitivity to azole antifungal agents. The names proposed for these isolates are Pseudozyma alboarmeniaca (DMST 17135^T = JCM 12454^T = CBS 9961^T), Pseudozyma crassa (DMST 17136^T = JCM 12455^T = CBS 9959^T) and Pseudozyma siamensis (DMST 17137^T = JCM 12456^T ?CBS 9960^T), where DMST is Department of Medical Sciences Culture Collection, JCM is Japan Collection of Microorganisms and CBS is Centraalbureau voor Schimmelcultures.

     

Random amplified polymorphic DNA (RAPD) analysis ofPenicillium marneffei strains isolated from AIDS patients in Thailand

Results of random amplified polymorphic DNA (RAPD) analysis using three different primers showed that 16 strains ofPenicillium marneffei isolated from AIDS patients in Thailand belonged to a genetically homogenous group, but different slightly from an isolate from bamboo rat in China. Six PCR fragments (from about, 200 to 600 bp) that were commonly observed in the RAPD fingerprint of all strains were extracted and sequented. Usefulness of this sequence information for identification ofP. marneffei is discussed.     

Genotyping and Antifungal Drug Susceptibility of the Pathogenic Yeast <Emphasis Type="Italic">Trichosporon asahii</Emphasis> Isolated from Thai Patients

Trichosporonosis due to Trichosporon asahii is a life-threatening infection with a very poor prognosis. We analyzed the genotype of intergenic transcribed spacer (IGS) region 1 of the rRNA gene and determined the drug susceptibility of 101 T. asahii isolates obtained from Thai patients to collect basic information on trichosporonosis in Thailand. Of the five genotypes in the IGS region identified in this study, types 1 and 3 were predominant in Thailand. The distribution in Thailand differs from that in other countries, suggesting that there is a geographic substructure among T. asahii clinical isolates. Voriconazole appeared to be the most active drug.     

Real-time PCR assay to detect DNA in sera for the diagnosis of deep-seated trichosporonosis

Deep-seated trichosporonosis due to Trichosporon asahii is life-threatening and has high mortality. A real-time PCR assay to detect T. asahii DNA in sera for diagnosis of this fungal infection was developed. The assay showed a higher sensitivity than polysaccharide antigen detection method. Our new real-time PCR assay may be used for diagnosing deep-seated trichosporonosis due to T. asahii.     

The first isolation of ustilaginomycetous anamorphic yeasts,Pseudozyma species,from patients' blood and a description of two new species: P. parantarctica and P. thailandica

The genus Pseudozyma is ustilaginomycetous anamorphic yeasts, and are mainly isolated from plants. We isolated three Pseudozyma strains from the blood of patients in Thailand. While one isolate was identified as P. antarctica by rDNA sequence analysis, the other two were considered to be new species and were named P. parantarctica and P. thailandica. The three isolates proved to be resistant to 5-flucytosine, and P. thailandica was also resistant to fluconazole and itraconazole. As far as we know, this is the first isolation of Pseudozyma strains from humans. The two new species are described.     

Genotoxicity of the Kishon River,Israel: the application of an in vitro cellular assay

The alkaline comet assay, a sensitive method for DNA strand breaks and alkali labile site detection in individual cells, was employed here as an ecotoxicological monitoring tool for evaluation of genotoxicity in the Kishon River, Israel. This river, the most polluted river in Israel, has recently elicited major public concern with regard to cancer incidences in people who have dived there over many years. Five water samples were collected every odd month throughout the year 2001, from four localities. The comet assay was employed on fish hepatoma cell line RTH-149. Cells were exposed for 2h, in triplicate, to Kishon water: medium (1:1) samples that were pre-adjusted for pH and salinity percentage levels. Three DNA damage parameters (comet percentages, score of damage, and cumulative tail lengths of the comet), revealed significantly higher genotoxic values in Kishon water-treated cells as compared with the controls (up to 2.4, 3.2, and 3.6-fold, respectively). Part of the sampling sites revealed higher genotoxicity than other polluted sites. The results of this study demonstrate that the comet assay with RTH-149 cells can be successfully applied to a variety of aquatic samples revealing freshwater, marine and estuary conditions. The method found to be fast, sensitive, and suitable for monitoring programs.     

The intergenic spacer region of the ribosomal RNA gene of Penicillium marneffei shows almost no DNA sequence diversity

The fungus Penicillium marneffei causes fatal systemic infections and is endemic in many parts of South-East Asia, especially Thailand. The intergenic spacer (IGS) region, the most variable region of rRNA genes, was found to be highly conserved among 58 P. marneffei strains. IGS analysis might not be suitable for molecular epidemiological analysis of P. marneffei infections.     

Comparative evaluation of Trichosporon asahii susceptibility using ASTY colorimetric microdilution and CLSI M27‐A2 broth microdilution reference methods

The in vitro activity of AMPH‐B, 5‐FC, FLCZ, MCZ, ITCZ, and VRCZ against 50 isolates of T. asahii was determined using CLSI M27‐A2 microdilution and ASTY colorimetric methods. Observed agreement ranged from 96 to 100% according to the drug. Overall, the agreement between two methods was 97.7%. The ASTY colorimetric method was thus determined to be comparable to the CLSI reference method when testing the susceptibility of T. asahii to a variety of antifungal agents.