Molecular characterization of environmental Cryptococcus neoformans VNII isolates in Jos, Plateau State, Nigeria.

Cryptococcus neoformans and Cryptococcus gattii are encapsulated yeasts able to cause fatal neurological infections in both human and other mammals. Cryptococcosis is the most common fungal infection of the central nervous system and has a huge burden in sub-Saharan Africa and South East Asia. Bird excreta are considered an environmental reservoir for C. neoformans in urban areas, therefore a study aimed at isolating and characterizing this yeast is important in disease management. In this study, one hundred samples of pigeon droppings were collected in Jos, Plateau State, Nigeria. C. neoformans was isolated from three samples and initially identified using standard phenotypic and biochemical tests. Molecular analysis revealed that all three isolates belonged to C. neoformans genotype VNII, mating type α and were assigned to the sequence type ST43 by multilocus sequence typing analysis. This study reports, for the first time, the molecular characterization of C. neoformans in Nigeria, where little is still known about the environmental distribution of the genotypes, serotypes and mating types of this important human pathogen.

Multilocus microsatellite analysis of European and African Candida glabrata isolates.

This study aimed to elucidate the genetic relatedness and epidemiology of 127 clinical and environmental Candida glabrata isolates from Europe and Africa using multilocus microsatellite analysis. Each isolate was first identified using phenotypic and molecular methods and subsequently, six unlinked microsatellite loci were analyzed using automated fluorescent genotyping. Genetic relationships were estimated using the minimum-spanning tree (MStree) method. Microsatellite analyses revealed the existence of 47 different genotypes. The fungal population showed an irregular distribution owing to the over-representation of genetically different infectious haplotypes. The most common genotype was MG-9, which was frequently found in both European and African isolates. In conclusion, the data reported here emphasize the role of specific C. glabrata genotypes in human infections for at least some decades and highlight the widespread distribution of some isolates, which seem to be more able to cause disease than others.

Potential association of specific Candida parapsilosis genotypes, bloodstream infections and colonization of health workers' hands.

Fungal nosocomial infections continue to be a serious problem among hospitalized patients, decreasing quality of life and adding millions of euros to healthcare costs. The aim of this study was to describe the pattern of fungi associated with the hands of healthcare workers and to genotype Candida parapsilosis isolates in order to understand whether their high clinical prevalence stems from endemic nosocomial genotypes or from the real emergence of epidemiologically-unrelated strains. Approximately 39% (50/129) of healthcare workers were positive for yeasts and among 77 different fungal isolates recovered, C. parapsilosis was the most frequent (44/77; 57%). Twenty-seven diverse genotypes were obtained by microsatellite analysis of 42 selected blood and hand isolates. Most of the isolates from hands showed a new, unrelated, genotype, whereas a particular group of closely related genotypes prevailed in blood samples. Some of the latter genotypes were also found on the hands of healthcare workers, indicating a persistence of these clones within our hospital. C. parapsilosis genotypes from the hands were much more heterogeneous than clinical ones, thus reflecting a high genetic diversity among isolates, which is notably unusual and unexpected for this species.

Looking for Candida nivariensis and C. bracarensis among a large Italian collection of C. glabrata isolates: results of the FIMUA working group.

Two recently described pathogenic Candida species, C. nivariensis and C. bracarensis, share many phenotypic characteristics with C. glabrata and are easily misidentified as such. The aim of this study was to determine the occurrence of these cryptic species in Italy. One thousand yeast isolates collected in 14 Italian regions and identified as C. glabrata by phenotypic and biochemical methods were included in this study: 928 were screened on CHROMagar and 72 were analysed by a multiplex PCR. None of these cryptic species was identified despite the nationwide distribution and the variety of biological origin of the isolates.

Differentiation of aflatoxin-producing and non-producing strains of Aspergillus flavus group.

AIMS: Three conventional methods and a multiplex PCR procedure with a set of four primers (Quadruplex-PCR) were used to differentiate between aflatoxin-producing and non-producing strains of the Aspergillus flavus group. METHODS AND RESULTS: By combining sets of primers for aflR, nor-1, ver-1 and omt-A genes of the aflatoxin biosynthetic pathway, Quadruplex-PCR showed that aflatoxinogenic strains gave a quadruplet pattern, indicating the presence of all the genes involved in the aflatoxin biosynthetic pathway which encode for functional products. Non-aflatoxinogenic strains gave varying results with one, two, three or four banding patterns. A banding pattern in three non-aflatoxinogenic strains resulted in non-differentiation between these and aflatoxinogenic strains. CONCLUSION AND SIGNIFICANCE AND IMPACT OF THE STUDY: Because conventional methods are time-consuming, further studies are needed to develop a rapid and objective technique that permits complete differentiation between aflatoxin-producing and non-producing strains of the A. flavus group.

Intra-specific diversity of Aureobasidium pullulans strains isolated from rocks and other habitats assessed by physiological methods and by random amplified polymorphic DNA (RAPD).

Intra-specific diversity of Aureobasidium pullulans strains isolated from environmental sources and from stones was studied by assessment of morphological, biochemical and physiological characters as well as random amplified polymorphic DNA (RAPD) using microsatellite or minisatellite DNA primers (GTG)5, (GACA)4, M13. The results showed that both classical and molecular techniques evidenced a phenotypic and genetic diversity of analysed A. pullulans strains. A different behaviour was observed in reference to the growth responses with D-glucosamine, citrate, galactitol and with different salt concentrations and range of growth temperature. Molecular analysis partially confirmed the data obtained with biochemical and physiological tests, additionally showing common fragments in all strains, to be used for a possible application as 'in situ' probes and for a rapid identification of A. pullulans strains.

Killer activity at different pHs against Cryptococcus neoformans var. neoformans serotype A by environmental yeast isolates.

Yeast isolates that share the same habitats as Cryptococcus neoformans var. neoformans serotype A in a restricted Mediterranean area were assayed in order to verify their killer activity against Cr. neoformans strains isolated from clinical and environmental sources. Many of the environmental yeast isolates expressed the killer phenomenon against the assayed strains of Cr. neoformans. Two species of Candida: Candida parapsilosis and Candida famata, and Pichia carsonii, were the most active killers at pH 4.6, 5.0 and 5.6 levels encountered in pigeon and canary guanos. Killer activity by C. parapsilosis is reported for the first time. The authors hypothesized that the killer phenomenon exerted by yeast species with heavy killer activity against Cr. neoformans would lend themselves for use as biological control agents against sensitive strains of Cr. neoformans when directly inoculated into the habitats of Cr. neoformans.

Molecular subtyping of clinical and environmental strains of Cryptococcus neoformans variety neoformans serotype A isolated from southern Italy.

Analysis of ribosomal DNA (rDNA) restriction fragment-length polymorphism (RFLP) and random amplification of polymorphic DNA (RAPD) was used to investigate the genetic variability and biogeographic distribution of clinical and environmental strains of Cryptococcus neoformans isolated from a limited area of southern Italy, where the selection of a predominant cryptococcal genotype could be expected. All isolates belonged to the species Cr. neoformans variety neoformans serotype A. RFLP analysis of a specific rDNA fragment allowed the distinction of strains of Cr. neoformans from closely related fungal reference species, but neither intraspecies nor intravarieties polymorphism was detected. On the contrary, RAPD fingerprints produced by priming with four different primers [(GTG)5, (GACA)4, M13 core sequence and the 8-mer oligonucleotide (GCGGACGG)] were able to characterize the isolates up to the individual level, indicating the presence of marked heterogeneity among Cr. neoformans serotype A strains in southern Italy.

Comparison of phospholipase production in Cryptococcus neoformans isolates from AIDS patients and bird droppings.

Secreted phospholipase has been recently proposed as a virulence determinant in Cryptococcus neoformans as well as Candida albicans. This issue of cryptococcal phospholipase requires screening of phospholipase production in a larger number of isolates from clinical and environmental sources. In this study we examined phospholipase production in a total of 67 C. neoformans isolates from AIDS patients and bird droppings by using the egg-yolk plate method. Phenoloxidase activity, capsule size and growth at 37 degrees C were also measured in these strains in order to observe a possible relationship between phospholipase production of different C. neoformans strains and its virulence. Four of the 21 AIDS strains at 28 degrees C and 1 at 37 degrees C did not produce phospholipase, respectively. In contrast, 38 and 34 of the 46 bird dropping strains were negative for phospholipase production at 28, and 37 degrees C, respectively. Statistical analysis revealed a significant difference in phospholipase production, capsule size and growth ability at 37 degrees C, but not phenoloxidase activity, between the AIDS and the bird dropping strains. The highly prevalent distribution of phospholipase activity in the AIDS strains suggests a role of the enzyme in invading the host.

Serotyping of Cryptococcus neoformans isolates from environmental and clinical sources in extreme southern Italy (Calabria and Sicily, central Mediterranean area).

Ninety-seven environmental and four clinical isolates of Cryptococcus neoformans from several localities in extreme southern Italy were serotyped. All proved to be Cr. neoformans var. neoformans serotype A. The homogeneity of the serotypes suggests that geographical climatological conditions may play a role in Cr. neoformans serotype diffusion. It was furthermore revealed that, unlike the data reported for the serotypes of Cr. neoformans in mainland Italy, Cr. neoformans serotype D was not present in the examined sites in extreme southern Italy.

Genetic relatedness and diversity of Cryptococcus neoformans strains in the Maltese Islands.

The genetic relatedness of clinical and environmental Cryptococcus neoformans strains in the Maltese Islands was investigated by randomly amplified polymorphic DNA fingerprinting with four primers. The clinical strains isolate over the course of 1 year from AIDS patients showed identical fingerprints. The electrophoretic patterns of the two clinical strains were also the most common patterns among the environmental strains, but the patterns among the environmental strains showed a wide variability and no correlation with the site of isolation.

Evidence of canary droppings as an important reservoir of Cryptococcus neoformans.

After preliminary results had shown the occurrence of Cryptococcus neoformans in canary droppings in southern Italy, the increasing epidemiological interest in cryptococcosis and the habitats of the agent led to more extensive studies. 180 samples of canary droppings were collected from pet shops and private households in two towns (Messina and Reggio Calabria) of southern Italy and culturally examined for C. neoformans. The examination was carried out with the help of the brown colour effect (BCE) specific of the C. neoformans colony on Guizotia abyssinica creatinine agar (Staib agar). 48 samples (26.6%) were positive; of these, 32/108 (29.6%) were from pet shops and 16/72 (22.2%) from private households. The colony growth was not mucous, i.e. microscopically the blastospores were found to be little or not encapsulated. All C. neoformans strains tested by the auxanographic method showed a weak assimilation of creatinine. The variety status of the isolates was tested with canavanine-glycine-bromothymol-blue (CGB) agar; all strains tested (n = 420) were found to be C. neoformans var. neoformans. In the light of extensive studies on C. neoformans and the canary in the pre-AIDS era in Germany, the present observations in Italy ask for further basic research, epidemiological studies, and education of the public.

Cryptococcal meningoencephalitis in a patient with Hodgkin's lymphoma: successful treatment with fluconazole.

The authors report a case of cryptococcal meningoencephalitis in a 22-year-old male with Hodgkin's disease at third stage B. Cryptococcus neoformans was isolated from cerebral spinal fluid (CSF) and blood specimens. Latex antigen agglutination tests and India ink preparations also were positive for this pathogenic yeast. Drug treatment for a week with 20 mg i.v./day of amphotericin B in combination with 2.5 g i.v./day of 5-fluorocytosine did not lead to any improvement in the patient. The patient became asymptomatic after the first week of treatment with a daily dose of 400 mg of fluconazole, administered intravenously. After two months of this regimen, all laboratory tests on CSF and blood specimens were negative.

An outbreak of Tinea pedis and Tinea cruris in a tyre factory in Messina, Italy.

A case of an epidemic by Tinea cruris and Tinea pedis in a tyre factory in Sicily is reported. There was noticeable spread of the causative agents throughout the environment. They were mainly transmitted by wash-basins in the footbaths, by footboards of showers and in smaller numbers by towels. Trichosporon beigelii was the most common fungus isolated from the environment, followed by Trichophyton rubrum, Trichophyton mentagrophytes, Epidermophyton floccosum, Candida parapsilosis and Geotrichum candidum.

[Mycological diagnosis and study of mycetes isolated from an area along the Tyrrhenian coast in the province of Catanzaro (Calabria)]. / Indagine e diagnosi micologica di miceti isolati su un tratto del litorale Tirrenico della provincia di Catanzaro (Calabria).

10 areas along the Tyrrhenian coast were studied: 37 genera of hyphomycetes were isolated from 100 sand samples. The genera Aspergillus, Fusarium, Alternaria, Penicillium, Dendryphiopsis, were constantly present. Many of the fungi isolated occurred only rarely and were represented by Trichoderma, Sepedonium, Cladosporium and Chrysosporium. The genus Geotrichum was the most prevalent of the unpigmented yeasts which were predominant (59.6%), while Rhodotorula was the main genus in pink yeasts (40.4%). The species Chrysosporium keratinophilum and Chrysosporium tropicum were the only representatives of keratinophilic fungi; Dermatophytes were not isolated. Furthermore the hyphomycetes were osmophilic genera and for this reason they may be regarded as specific sand flora. Hence the data proves useful to us, indicating the negligible degree of human and animal influence of these sea shores.