Identification of genes expressed by Cryptococcus gattii during iron deprivation

Cryptococcus neoformans and C. gattii are pathogenic yeasts that cause life-threatening diseases in humans and animals. Iron is an essential nutrient for virtually every organism as it functions as a cofactor in numerous essential enzymatic reactions. In the literature, the competition for iron between microbes and mammalian hosts during infection is well documented. In this study, we used representational difference analysis (RDA) in order to gain a better understanding of how C. gattii responds to iron starvation. A total of 15 and 29 genes were identified as having altered expression levels due to iron depletion after 3 h and 12 h, respectively. Of these, eight genes were identified in both libraries. The transcripts were related to many biological processes, such as cell cycle, ergosterol metabolism, cell wall organization, transportation, translation, cell respiration and the stress response. These data suggest a remodeling of C. gattii metabolism during conditions of iron deprivation.

Distinct chitinases are expressed during various growth phases of the human pathogen Paracoccidioides brasiliensis

The aim of this work was the partial purification and subsequent evaluation of chitinase expression during the various growth phases of Paracoccidioides brasiliensis. Initially, PbCTS1r was expressed as a recombinant protein and displayed enzymatic activity against 4-MU-[N-acetylglucosamine (GlcNAc)]3 and 4-MU-(GlcNAc)2. Two proteins, 45 kDa and 39 kDa in size, were partially purified from P. brasiliensis yeast crude extract using cation-exchange chromatography coupled with HPLC and were characterised as PbCTS1 and PbCTS2, respectively. Anti-PbCTS1r antibody recognised two proteins in the crude extracts of yeast and the transitional stage between mycelial and yeast phases. In crude extracts of mycelium, only the 45 kDa protein was detected. However, quantitative real-time polymerase chain reaction led to the detection of small quantities of Pbcts2 transcript in the mycelial phase. In the yeast cell wall extract, only the 39 kDa protein was detected. Moreover, both proteins were secreted by the yeast parasitic phase, suggesting that these proteins participate in the modulation of the fungal environment. Phylogenetic analysis of the predicted PbCTS1 and PbCTS2 proteins indicated that they code for distinct chitinases in P. brasiliensis. During evolution, P. brasiliensis could have acquired the paralogues Pbcts1 and Pbcts2 for growth and survival in diverse environments in both saprophytic and parasitic phases.

Virulence factors and antimicrobial resistance of Escherichia coli isolated from urinary tract of swine in southern of Brazil / Fatores de virulência e resistência aos antimicrobianos de Escherichia coli isoladas do trato urinário de suínos do Sul do Brasil

The present study determined the molecular and resistance patterns of E. coli isolates from urinary tract of swine in Southern of Brazil. Molecular characterization of urinary vesicle samples was performed by PCR detection of virulence factors from ETEC, STEC and UPEC. From a total of 82 E. coli isolates, 34 (38.63 percent) harbored one or more virulence factors. The frequency of virulence factors genes detected by PCR were: pap (10.97 percent), hlyA (10.97 percent), iha (9.75 percent), lt (8.53 percent), sta (7.31 percent) sfa (6.09 percent), f4 (4.87 percent), f5 (4.87 percent), stb (4.87 percent), f6 (1.21 percent) and f41 (1.21 percent). Isolates were resistant to penicillin (95.12 percent), lincomycin (93.9 percent), erythromycin (92.68 percent), tetracycline (90.24 percent), amoxicillin (82.92 percent), ampicillin (74.39 percent), josamycin (79.26 percent), norfloxacin (58.53 percent), enrofloxacin (57.31 percent), gentamicin (39.02 percent), neomycin (37.8 percent), apramycin (30.48 percent), colistine (30.48 percent) and cefalexin (6.09 percent). A number of 32 (39.02 percent) E. coli isolates harbored plasmids.

O presente estudo teve por objetivo determinar os padrões moleculares e de resistência aos antimicrobianos de isolados de E. coli provenientes do trato urinário de suínos no Sul do Brasil. Os fatores estudados dividiram os patotipos ETEC, STEC e UPEC. Trinta e quatro (38,63 por cento) isolados avaliados apresentavam um ou mais dos fatores de virulência pesquisados. A freqüência dos genes de virulência detectados foram: pap (10,97 por cento), hlyA (10,97 por cento), iha (9,75 por cento), lt (8,53 por cento), sta (7,31 por cento) sfa (6,09 por cento), f4 (4,87 por cento), f5 (4,87 por cento), stb (4,87 por cento), f6 (1,21 por cento) e f41 (1,21 por cento). Os isolados foram resistentes à penicilina (95,12 por cento), lincomicina (93,9 por cento), eritromicina (92,68 por cento), tetraciclina (90,24 por cento), amoxacilina (82,92 por cento), ampicilina (74,39 por cento), josamicina (79,26 por cento), norfloxacina (58,53 por cento), enrofloxacina (57,31 por cento), gentamicina (39,02 por cento), neomicina (37,8 por cento), apramicina (30,48 por cento), colistina (30,48 por cento) e cefalexina (6,09 por cento). Trinta e dois (39,02 por cento) isolados de E. coli continham plasmídeos.

Comparative genome analysis of proteases, oligopeptide uptake and secretion systems in Mycoplasma spp

Mycoplasmas are very fastidious in their nutritional requirements for in vitro growth and have limited biosynthetic capacity, a reflection of their reduced genomes. As a result, these bacteria depend upon external metabolites for nutrition and growth and have developed dependence on their hosts for survival and maintenance. Protein degradation and peptide importation play an important role in Mycoplasma spp. nutrition, and proteases can play a role in host adaptation and pathogenicity. Here, we present a general survey on the genes involved in protein degradation, secretion and importation, comparing all available Mollicute genomes.

Carbohydrate assimilation profiles of Brazilian Candida dubliniensis isolates based on ID 32C system

Dezenove culturas de C. dubliniensis isoladas no Brasil, previamente identificadas através de métodos genotípicos, foram avaliadas pelo kit comercial ID 32C (bioMerieux). Treze culturas foram identificadas como C. dubliniensis, mas as demais (seis) evidenciaram perfil duvidoso, embora o software do sistema sugerisse 83,6% de chances das mesmas pertencerem à espécie C. dubliniensis. A literatura tem registrado grande variabilidade fenotípica com esta espécie e, por isto, as identificações obtidas com este sistema deverão ser consideradas como presuntivas.

Susceptibility to heat and antifungal agents of Cryptococcus neoformans var. neoformans (serotype D) isolated from Eucalyptus spp in Rio Grande do Sul, Brazil

Este estudo foi realizado com os primeiros isolados ambientais de C. neoformans sorotipo D, obtidos no Rio Grande do Sul. Objetivando-se avaliar a susceptibilidade a agentes antifúngicos de forma mais rigorosa, utilizou-se a técnica de referência proposta pelo NCCLS, Caldo Yeast Nitrogen Base (YNB) proposto por Ghannoum et al., Antibiotic medium 3, caldo YNB adequado à metodologia do NCCLS e o E-test. Os resultados indicaram que todos os isolados foram sensíveis à anfotericina B (0,0625-0,5 µg/mL), fluconazol (0,125-4,0 µg/mL), itraconazol (0,031-0,25 µg/ml) e fluorocitosina (0,125-4,0 µg/mL) através das técnicas empregadas. Nos testes de termotolerância (47ºC/30 min), observou-se que as culturas de C. neoformans sorotipo D são mais sensíveis do que as de C. neoformans sorotipo A.

In vitro susceptibility to antifungal agents of clinical and environmental Cryptococcus neoformans isolated in Southern of Brazil

The purpose of the present study was to compare the susceptibility to four antifungal agents of 69 Cryptococcus neoformans strains isolated from AIDS patients with that of 13 C. neoformans strains isolated from the environment. Based on the NCCLS M27-A methodology the Minimal Inhibitory Concentrations (MICs) obtained for amphotericin B, itraconazole and ketoconazole were very similar for clinical and environmental isolates. Clinical isolates were less susceptible to fluconazole than environmental isolates. The significance of these findings and aspects concerning the importance, role and difficulties of C. neoformans susceptibility testing are also discussed

Isolation of streptomycetes strain inhibitors of toxigenic fungi: partial characterization of their chitinolytic system

En la búsqueda de biocontroladores potenciales de hongos toxicogénicos, se aislaron desde el suelo 27 cepas de streptomycetes spp. y se desarrollaron pruebas de confrontación contra cepas toxicogénicas de: aspergillus parasiticus (productor de aflatoxinas t-2 y ht-2) y fusarium graminearum (productor de deoxinivalenol y nivalenol). El desarrollo de al menos uno de los hongos toxicogénicos fue inhibido por el 63 porciento de los streptomyces aislados, 41 porciento de las cepas de streptomyces inhibieron el crecimiento de al menos dos de los hongos probados, y el 36 porciento de los streptomyces fue efectivo contra los tres hongos. Además, fue analizada la capacidad de los aislamiento de degradar quitina coloidal y posteriormente fueron caracterizados los complejos quitinolíticos de dos de las cepas de streptomyces. El 66 porciento de los streptomyces spp. degradaron quitina coloidal en las pruebas en placa. La secreción de quitinasas de dos aislamientos fue ensayada empleando métodos colorimétricos y geles de actividad, y los productos de degradación a partir de diferentes sustratos fueron analizados por cromatografía en capa delgada. Los resultados indicaron que uno de los streptomyces (c112), posee actividad endoquitinasa pero no n-acetilglucosaminidasa y que en el complejo de enzimas quitinolíticas de la cepa (c103) una actividad de n-acetil-glucosaminidasa