Kluyveromyces aestuarii, a potential environmental quality indicator yeast for mangroves in the State of Rio de Janeiro, Brazil

Kluyveromyces aestuarii was found in sediments from 7 of 8 mangroves in Rio de Janeiro; and absent only at one site with heavy plastic bag pollution. Its presence suggests influence in other habitats from a mangrove and its absence in a mangrove suggests some non-fecal pollution or other habitat alteration.

Kluyveromyces aestuarii, a potential environmental quality indicator yeast for mangroves in the State of Rio de Janeiro, Brazil.

Kluyveromyces aestuarii was found in sediments from 7 of 8 mangroves in Rio de Janeiro; and absent only at one site with heavy plastic bag pollution. Its presence suggests influence in other habitats from a mangrove and its absence in a mangrove suggests some non- fecal pollution or other habitat alteration.

Differential recovery of Candida species from subgingival sites in human immunodeficiency virus-positive and healthy children from Rio de Janeiro, Brazil.

The prevalence of subgingival Candida species was studied in 52 human immunodeficiency virus (HIV)-positive and 42 HIV-negative children. Candida was cultured from 22 (42.3%) and 3 (7.1%) HIV-infected and control children, respectively. C. albicans was the most common Candida species isolated from HIV-infected children, followed by C. dubliniensis, C. glabrata, and C. tropicalis. In the HIV-positive group, the prevalence of Candida isolation was significantly higher in children who presented with low CD4(+)-T-lymphocyte counts, elevated viral loads, and gingivitis.

Yeast-Yeast Interactions in Guava and Tomato Fruits.

The host specificity of yeast-yeast interactions was investigated for two yeast types, represented by six pairs of Pichia membranifaciens clade yeasts (Pichia membranifaciens or Issatchenkia occidentalis) with apiculate yeasts (Kloeckera apis, Kloeckera africana, or Saccharomycodes ludwigii), commonly found in fruits. Competitive interactions between the two types were detected in both ripe tomato and guava fruit pulp. The differences in growth rates and carrying capacities depended on fruit type (host) and culture conditions (monocultures versus bicultures). These differences were probably due to nutrient composition of each fruit. Pichia membranifaciens did not show host dependent responses, but the apiculate yeasts and Issatchenkia occidentalis did. Depending on yeast strain and culture conditions (i.e., monoculture or biculture), carbon, nitrogen, and vitamins were investigated as potential limiting growth factors in guava fruit. Both singular and multiple limiting nutrients were implicated.

Contamination of expressed human breast milk with an epidemic multiresistant Staphylococcus aureus clone.

Nosocomial infections caused by methicillin-resistant Staphylococcus aureus (MRSA) are a major cause of outbreaks in intensive care units. Infants make up a sector of the population that presents a high risk for MRSA infections. Mother-to-infant transmission has been indicated as a possible cause of MRSA infections in neonates. The occurrence and characteristics of MRSA in samples of banked human milk were investigated by selective culture, antibiogram and pulsed-field gel electrophoresis. MRSA contamination was found in 11% of 500 samples of expressed, fresh-frozen milk from 500 different donors at five Brazilian milk banks. The great majority of the contaminated samples passed breast milk quality control criteria for dispensing as raw milk under Brazilian and American guidelines. Most of the MRSA isolates belonged to the Brazilian epidemic clone, which is reported to be widespread in several Brazilian states, in Argentina and in Portugal. It is concluded that expressed breast milk can be a reservoir of multiresistant S. aureus epidemic clones. Studies are necessary to assess the source of contamination and potential role of MRSA-contaminated milk in the transmission of MRSA to neonates.

Methicillin-resistant Staphylococcus aureus in human milk.

We collected and analyzed 500 samples of human milk, from five Brazilian cities (100 from each) to detect methicillin-resistant strains of Staphylococcus aureus (MRSA) producing enterotoxins. We found 57 strains of MRSA, and the mecA gene, responsible for resistance, was detected in all of them using a specific molecular probe. We examined 40 strains for the presence of four enterotoxins, after selecting a subset that included all strains from each region, except for the largest sample, from which 10 were randomly selected. Among these two presented enterotoxin B, and growth in human colostrum and trypicase soy broth. After 5 h of incubation at 37 degrees C, population sizes were already higher than 9.4 x 10(5) UFC/ml and enterotoxin was released into culture medium and colostrum. Our results stress the importance of hygiene, sanitary measures, and appropriate preservation conditions to avoid the proliferation of S. aureus in human milk.

Methicillin-resistant Staphylococcus aureus in human milk

We collected and analyzed 500 samples of human milk, from five Brazilian cities (100 from each) to detect methicillin-resistant strains of Staphylococcus aureus (MRSA) producing enterotoxins. We found 57 strains of MRSA, and the mecA gene, responsible for resistance, was detected in all of them using a specific molecular probe. We examined 40 strains for the presence of four enterotoxins, after selecting a subset that included all strains from each region, except for the largest sample, from which 10 were randomly selected. Among these two presented enterotoxin B, and growth in human colostrum and trypicase soy broth. After 5 h of incubation at 37§C, population sizes were already higher than 9.4 x 105 UFC/ml and enterotoxin was released into culture medium and colostrum. Our results stress the importance of hygiene, sanitary measures, and appropriate preservation conditions to avoid the proliferation of S. aureus in human milk.

Yeast communities associated with sugarcane in Campos, Rio de Janeiro, Brazil.

Yeast communities associated with sugarcane leaves, stems and rhizosphere during different phases of plant development were studied near Campos, in Rio de Janeiro, Brazil. Atmospheric temperature, soil granulometry and pH, and sugar cane juice degree Brix and pH were determined. Yeast communities associated with sugarcane were obtained after cellular extraction by shaking, blending and shaking plus sonication, and cultured on Yeast Nitrogen Base Agar plus glucose (0.5%) and Yeast Extract-Malt Extract Agar. No significant differences in yeast counts were found among the cellular extraction treatments and culture media. 230 yeast cultures were identified according to standard methods, and distinct yeast communities were found for each substrate studied. The prevalent species isolated from sugarcane were Cryptococcus laurentii, Cryptococcus albidus, Rhodotorula mucilaginosa and Debaryomyces hansenii.

Natural transformation and availability of transforming DNA to Acinetobacter calcoaceticus in soil microcosms.

A small microcosm, based on optimized in vitro transformation conditions, was used to study the ecological factors affecting the transformation of Acinetobacter calcoaceticus BD413 in soil. The transforming DNA used was A. calcoaceticus homologous chromosomal DNA with an inserted gene cassette containing a kanamycin resistance gene, nptII. The effects of soil type (silt loam or loamy sand), bacterial cell density, time of residence of A. calcoaceticus or of DNA in soil before transformation, transformation period, and nutrient input were investigated. There were clear inhibitory effects of the soil matrix on transformation and DNA availability. A. calcoaceticus cells reached stationary phase and lost the ability to be transformed shortly after introduction into sterile soil. The use of an initially small number of A. calcoaceticus cells and nutrients, resulting in bacterial growth, enhanced transformation frequencies within a limited period. The availability of introduced DNA for transformation of A. calcoaceticus cells disappeared within a few hours in soil. Differences in transformation frequencies between soils were found; A. calcoaceticus cells were transformed at a higher rate and for a longer period in a silt loam than in a loamy sand. Physical separation of DNA and A. calcoaceticus cells had a negative effect on transformation. Transformation was also detected in nonsterile soil microcosms, albeit only in the presence of added nutrients and at a reduced frequency. These results suggest that chromosomal DNA released into soil rapidly becomes unavailable for transformation of A. calcoaceticus. In addition, strain BD413 quickly loses the ability to receive, stabilize, and/or express exogenous DNA after introduction into soil.

The incidence of killer activity and extracellular proteases in tropical yeast communities.

The presence of killer and proteolytic yeasts was studied among 944 isolates representing 105 species from tropical yeast communities. We found 13 killer toxin producing species, with Pichia kluyveri being the most frequent. Other killer yeast isolates were Candida apis, Candida bombicola, Candida fructus, Candida krusei, Candida sorbosa, Hanseniaspora uvarum, Issatchenkia occidentalis, Kloeckera apis, Kluyveromyces marxianus, Pichia membranaefaciens, Pichia ohmeri-like, and Sporobolomyces roseus. The communities from which killer yeasts were isolated had strains sensitive to them, and there were interspecific and intraspecific differences in the spectra of their killer activities. Pichia kluyveri had the broadest spectra of activity against sensitive isolates, and it apparently produced different toxins. The coexistence of sensitive and killer yeasts using the same substrate suggests that there is spatial separation in microhabitats or temporal separation in different stages of successions. Basidiomycetous yeasts were more frequently proteolytic than ascomycetous yeasts. Extracellular proteases could be important for the yeasts to have access to more nitrogen nutrients and obtain a better balance with available carbon sources.

Dipodascus starmeri sp. nov., a new species of yeast occurring in cactus necroses.

In a previous publication describing the geographic distribution of yeasts associated with cactus necroses (W. T. Starmer, M.-A. Lachance, H. J. Phaff, and W. B. Heed, Evol. Biol. 24:253-296, 1990), 127 isolates were identified as strains of Candida ingens van der Walt et van Kerken on the basis of morphology and certain phenotypic characteristics. Here we show by using DNA hybridization and additional phenotypic properties that these strains were misidentified and that they represent a minimum of three separate species that can be differentiated from C. ingens and from each other by utilization of 2-propanol or acetone, sensitivity to digitonin, utilization of L-lysine as a sole nitrogen source, vitamin dependence, NaCl tolerance, lipolytic activity, and habitat. One of the new species is haploid and heterothallic, and its teleomorph represents the genus Dipodascus. We describe Dipodascus starmeri sp. nov. The phylogenetic relationship of D. starmeri with other members of the genus Dipodascus and its anamorph, the genus Geotrichum, was estimated from ribosomal DNA nucleotide sequence divergence. The type strain, a heterothallic haploid isolate, is UCD-FST 72-316 (= CBS 780.96 = ATCC 200546 = NRRL Y-17816). The complementary mating type is UCD-FST 81-513.3 (= CBS 781.96 = ATCC 200547 = NRRL Y-17817).

PCR amplification of the rDNA internal transcribed spacer region for differentiation of Saccharomyces cultures.

The size of the internal transcribed spacer (ITS) region as measured by gel electrophoresis of PCR products, amplified by primers ITS1 and ITS4, was over 800 bp for all Saccharomyces sensu stricto species, but yeasts belonging to other Saccharomyces species had a shorter ITS region, making this characteristic potentially useful in the identification of Saccharomyces isolates. The ITS product length was homogeneous within the species Saccharomyces cerevisiae.

Selection of rhizosphere-competent Pseudomonas strains as biocontrol agents in tropical soils.

Pseudomonas strains were isolated from the rhizosphere of maize grown in yellow-red latosol from Rio de Janeiro, Brazil, to serve as a delivery system for heterologous genes and for risk assessment studies in tropical soils. Selected strains were modified by insertion of the cryIVB gene from Bacillus thuringiensis and tested for pathogenicity gene expression against larvae of a susceptible model species, Anopheles aquasalis. Modified strains Br8 and Br12 showed similar survival performance to their parental strains, and presented a viable density of 10(7) c.f.u./g dry soil 30 days after release. A strain of P. fluorescens (Br12) that presented positive results for gene expression and the best survival performance, was selected for risk assessment studies in soil microcosms.

Yeast succession in the Amazon fruit Parahancornia amapa as resource partitioning among Drosophila spp.

The succession of yeasts colonizing the fallen ripe amapa fruit, from Parahancornia amapa, was examined. The occupation of the substrate depended on both the competitive interactions of yeast species, such as the production of killer toxins, and the selective dispersion by the drosophilid guild of the amapa fruit. The yeast community associated with this Amazon fruit differed from those isolated from other fruits in the same forest. The physiological profile of these yeasts was mostly restricted to the assimilation of a few simple carbon sources, mainly L-sorbose, D-glycerol, DL-lactate, cellobiose, and salicin. Common fruit-associated yeasts of the genera Kloeckera and Hanseniaspora, Candida guilliermondii, and Candida krusei colonized fruits during the first three days after the fruit fell. These yeasts were dispersed and served as food for the invader Drosophila malerkotliana. The resident flies of the Drosophila willistoni group fed selectively on patches of yeasts colonizing fruits 3 to 10 days after the fruit fell. The killer toxin-producing yeasts Pichia kluyveri var. kluyveri and Candida fructus were probably involved in the exclusion of some species during the intermediate stages of fruit deterioration. An increase in pH, inhibiting toxin activity and the depletion of simple sugars, may have promoted an increase in yeast diversity in the later stages of decomposition. The yeast succession provided a patchy environment for the drosophilids sharing this ephemeral substrate.

Ascomycetous yeast communities of marine invertebrates in a southeast Brazilian mangrove ecosystem.

The ascomycetous yeast communities associated with 3 bivalve mollusk, and 4 crab species were studied in the mangrove at Coroa Grande on Sepetiba Bay in Rio de Janeiro, Brazil. These were made up mostly of diverse but sparse and apparently allochtonous yeast populations. The striking exception was a prevalent population of the species Kluyveromyces aestuarii, which predominated the yeast communities of 2 detritus feeding crabs, Sesarma rectum and Uca spp., and the shipworm Neoteredo reynei. However, K. aestuarii was absent from the omnivorous crabs Aratus pisonii and Goniopsis cruentata, and the clam Anomalocardia brasiliana, and was rare in the clam Tagelus plebeius from mostly submerged more sandy sediments. Pichia membranaefaciens, Candida valida-like, Candida krusei, Candida sorbosa, Candida colliculosa-like, Candida famata-like, Kloeckera spp., Candida guilliermondii, Candida albicans, Candida silvae, Geotrichum spp., Rhodotorula spp., Cryptococcus spp., and the methylotrophic yeast Candida boidinii were frequently isolated. The 322 ascomycetous yeast cultures representing 252 isolates from crabs and mollusks were classified as 40 species that fit standard descriptions, and 44 putative new species. The ascomycetous yeast communities of the mangrove ecosystem include many new biotypes that require better taxonomic definition.

A new genetically isolated population of the Saccharomyces sensu stricto complex from Brazil.

Genetic and karyotypic studies of some Saccharomyces sensu stricto yeasts from Brazil revealed a genetically isolated population which apparently represents a new sibling species of S. cerevisiae.

Yeast communities as descriptors of habitat use by the Drosophila fasciola subgroup (repleta group) in Atlantic rain forests.

Yeast communities associated with four species of the Drosophila fasciola subgroup (repleta group) in tropical rain forests were surveyed in an abandoned orchard, and rain forest sites of Rio de Janeiro and Ilha Grande, State of Rio de Janeiro, Brazil. Adult flies of Drosophila carolinae, Drosophila coroica, Drosophila fascioloides and Drosophila onca frequently carried Candida colliculosa, Geotrichum sp, Kloeckera apiculata and a Pichia membranaefaciens-like species. The most frequent yeasts in the crop of flies included Candida collicullosa, C. krusei, Pichia kluyveri and a P. membranaefaciens-like species. The physiological abilities and species composition of these yeast communities differed from those of other forest-inhabiting Drosophila. The narrow feeding niches of the fasciola subgroup suggested the use of only part of the substrates available to the flies as food in the forest environment, as noted previously for cactophilic Drosophila serido (mulleri subgroup of the repleta group) in a sand dune ecosystem. The cactophilic yeasts that were isolated have not been previously found in forests. The fasciola subgroup probably used epiphytic cactus substrates as breeding and feeding sites in the forest. The physiological profile of yeasts associated with the fasciola flies was broader than that of yeasts associated with the cactophilic Drosophila serido, suggesting that the fasciola subgroup represents an older lineage from which the South American repleta species evolved.

Yeast communities of the cactus Pilosocereus arrabidae and associated insects in the Sandy coastal plains of southeastern Brazil.

The yeast communities from necrotic tissues, decaying flowers and fruits, and from larval feeding sites of the moth Sigelgaita sp. in the cactus Pilosocereus arrabidae were surveyed in three restinga ecosystems in Southeastern Brazil. Insects associated with these substrates were sampled to verify the vectoring of yeasts. The cactus Pilosocereus arrabidae was shown to have four different yeast communities associated with it. Necrotic stems had a diverse yeast community with the prevalent species Pichia barkeri, Candida sonorensis, Pichia cactophila, Geotrichum sp., Myxozyma mucilagina and Sporopachydermia sp. A, representing about 80% of the total isolates. Pichia sp. A and a Candida domercqii-like species represented more than 90% of the yeast isolates from decaying flowers. Fruits had a heterogeneous yeast community with typical fruit yeasts of the genus Kloeckera, basidiomicetous anamorphs of the genus Cryptococcus, the black yeast Aureobasidium pullulans, Pichia sp. A, a Candida domercqii-like species, and some cactophilic yeasts, especially Clavispora opuntiae. The feeding site of Sigelgaita sp. larvae had Clavispora opuntiae as the prevalent species. Insect vectors are suggested as one the most important factors influencing the composition of these yeast communities.

Yeast communities of the cactus Pilosocereus arrabidae as resources for larval and adult stages of Drosophila serido.

The feeding behavior of Drosophila serido on the yeast communities of necrotic stem tissue of Pilosocereus arrabidae were studied in a sand dune ecosystem of Rio de Janeiro, Brazil. The prevalence of cactophilic yeasts including Pichia barkeri, Candida sonorensis and Geotrichum sp. in the crops and external surfaces of D. serido reflected its association with the cactus habitat. The effective number of yeasts vectored on the surface of flies was higher than that in the crops. Also overlap between the yeasts from stems and from crops was partial suggesting selective feeding by the flies in the substrates visited. The females had a higher effective number of yeast species and a lower similarity than males with the yeast community of P. arrabidae. This was probably related to the search for oviposition sites by females. The presence of Pichia thermotolerans-like and Pichia amethionina var pachycereana in the flies, but not in P. arrabidae stems, indicated that D. serido was not limited to this cactus species. The larvae and adults lived in different patches with the adults feeding in patches with higher yeast species richness. The larvae had a narrower feeding niche and higher overlap with P. arrabidae, and preferred P. barkeri and Pichia cactophila as food. Adult flies fed on patches with the most frequent yeasts except for P. cactophila. Pichia caribaea was found in higher frequency in the adult crops than in the stems. Our data suggested that there was food selection and diet partitioning between adult and larval stages of D. serido.

Yeasts and coliform bacteria of water accumulated in bromeliads of mangrove and sand dune ecosystems of southeast Brazil.

Yeasts and coliform bacteria were isolated from water that accumulated in the central cups and adjacent leaf axilae of two bromeliads, Neoregelia cruenta of a coastal sand dune and Quesnelia quesneliana of a mangrove ecosystem near the city of Rio de Janeiro, Brazil. The mean total coliform counts were above 10,000 per 100 mL for waters of both plants, but the mean fecal coliform counts were only 74 per 100 mL for Q. quesneliana and mostly undetected in water from N. cruenta. Of 90 fecal coliform isolates, 51 were typical of Escherichia coli in colony morphology and indol, methyl red, Volges-Proskauer, and citrate (IMViC) tests. Seven representatives of the typical E. coli cultures were identified as this species, but the identifications of nine other coliform bacteria were mostly dubious. The yeast community of N. cruenta was typical of plant surfaces with basidiomycetous yeasts anamorphs, and the black yeast Aureobasidium pullulans was prevalent. Quesnelia quesneliana had a substantial proportion of ascomycetous yeasts and their anamorphs, including a probable new biotype of Saccharomyces unisporus. Our results suggested that the microbial communities in bromeliad waters are typically autochtonous and not contaminants.