Pathogenic Yersinia enterocolitica strains increase the outer membrane permeability in response to environmental stimuli by modulating lipopolysaccharide fluidity and lipid A structure

Pathogenic biotypes of Yersinia enterocolitica (serotypes O:3, O:8, O:9, and O:13), but not environmental biotypes (serotypes O:5, O:6, O:7,8, and O:7,8,13,19), increased their permeability to hydrophobic probes when they were grown at pH 5.5 or in EGTA-supplemented (Ca(2+)-restricted) media at 37 degrees C. A similar observation was also made when representative strains of serotypes O:8 and O:5 were tested after brief contact with human monocytes. The increase in permeability was independent of the virulence plasmid. The role of lipopolysaccharide (LPS) in this phenomenon was examined by using Y. enterocolitica serotype O:8. LPS aggregates of bacteria grown in acidic or EGTA-supplemented broth took up more N-phenylnaphthylamine than LPS aggregates of bacteria grown in standard broth and also showed a marked increase in acyl chain fluidity which correlated with permeability, as determined by measurements obtained in the presence of hydrophobic dyes. No significant changes in O-antigen polymerization were observed, but lipid A acylation changed depending on the growth conditions. In standard medium at 37 degrees C, there were hexa-, penta-, and tetraacyl lipid A forms, and the pentaacyl form was dominant. The amount of tetraacyl lipid A increased in EGTA-supplemented and acidic media, and hexaacyl lipid A almost disappeared under the latter conditions. Our results suggest that pathogenic Y. enterocolitica strains modulate lipid A acylation coordinately with expression of virulence proteins, thus reducing LPS packing and increasing outer membrane permeability. The changes in permeability, LPS acyl chain fluidity, and lipid A acylation in pathogenic Y. enterocolitica strains approximate the characteristics in Yersinia pseudotuberculosis and Yersinia pestis and suggest that there is a common outer membrane pattern associated with pathogenicity.

The significance of volatile antifungal metabolites produced by trichomerma harzianum biotype Th4, in green-mould disease of commercial mushroom crops /

The aggressive mushroom competitor, Trichoderma harzianum biotype Th4, produces volatile antifungal secondary metabolites both in culture and during the disease cycle in compost. Th4 cultures produced one such compound only when cultured in the presence of Agaricus bisporus mycelium or liquid medium made from compost colonised with A. bisporus. This compound has TLC and UVabsorption and characteristics indicating that it belongs to a class of pyrone antibiotics characterised from other T. harzianum biotypes. UV absorption spectra indicated this compound was not 6-pentyl-2H-pyran-one (6PAP), the volatile antifungal metabolite widely described in Th1. Furthermore, this compound was not produced by Th1 under any culture conditions. Mycelial growth of A. bisporus, Botrytis cinerea and Sclerotium cepivorum was inhibited in the presence of this compound through volatility , diffusion and direct application. This indicates that Th4 produces novel, volatile, antifungal metabolites in the presence of A. bisporus that are likely involved in green mould disease of mushroom crops.

Examination of faecal Bifidobacterium populations in breast- and formula-fed infants during the first 18 months of life.

Bifidobacteria in the infant faecal microbiota have been the focus of much interest, especially during the exclusive milk-feeding period and in relation to the fortification of infant formulae to better mimic breast milk. However, longitudinal studies examining the diversity and dynamics of the Bifidobacterium population of infants are lacking, particularly in relation to the effects of weaning. Using a polyphasic strategy, the Bifidobacterium populations of breast- and formula-fed infants were examined during the first 18 months of life. Bifidobacterium-specific denaturing gradient gel electrophoresis demonstrated that breast-fed infants harboured greater diversity than formula-fed infants and the diversity of the infants' Bifidobacterium populations increased with weaning. Twenty-seven distinctive banding profiles were observed from ∼1100 infant isolates using ribosomal intergenic spacer analysis, 14 biotypes of which were confirmed to be members of the genus Bifidobacterium. Two profiles (H, Bifidobacterium longum subsp. infantis; and I, Bifidobacterium bifidum) were common culturable biotypes, seen in 9/10 infants, while profile E (Bifidobacterium breve) was common among breast-fed infants. Overall, inter- and intra-individual differences were observed in the Bifidobacterium populations of infants between 1 and 18 months of age, although weaning was associated with increased diversity of the infant Bifidobacterium populations. Breast-fed infants generally harboured a more complex Bifidobacterium microbiota than formula-fed infants.

Análise da variabilidade genética de Paspalum Notatum Flugge (Poaceae, Panicoideae) com o uso de marcadores moleculares, morfológicos e citometria de fluxo

O gênero Paspalum L. compreende aproximadamente 400 espécies no mundo e cerca de 220 no Brasil. Paspalum é ecologicamente e economicamente importante e tem sido utilizado como pastagem. Paspalum notatum Flügge (grama-forquilha) é uma valorosa gramínea forrageira nos subtrópicos. Esta espécie consiste de vários biótipos sexuais (diplóides) e apomíticos (tetraplóides, ocasionalmente tri e pentaplóides). Neste trabalho, os Inter Simple Sequence repeat (ISSR) foram utilizados para acessar a diversidade genética da grama-forquilha (Paspalum notatum). Os tecidos vegetativos de 95 acessos de grama-forquilha foram obtidos de vários locais da América do Sul (Brasil, Argentina e Uruguai). Um total de 91 de fragmentos reproduzível ISSR foi observado. Oitenta e nove fragmentos (97,5% do total observado) foram polimórficos. A análise de agrupamento (UPGMA) foi realizada para o conjunto de dados ISSR. Os resultados ilustram as relações genéticas entre 95 acessos de Paspalum notatum. A comparação entre dados moleculares, morfológicos e nível de ploidia foi realizada. Em resumo, os marcadores moleculares ISSR mostraram-se eficientes para distinção dos genótipos analisados e observou-se uma variabilidade ampla para a espécie. Estes resultados adicionam novas informações sobre a diversidade genética em Paspalum notatum, conseqüentemente contribuindo para o conhecimento biológico desta espécie e fornecendo subsídios para futuros programas de melhoramento genético e para programas de conservação.

Pool of Resistance Mechanisms to Glyphosate in Digitaria insularis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Controle químico de biótipos de buva (Conyza canadensis e Conyza bonariensis) resistentes ao glyphosate

Studies were conducted at Estacao Experimental de Citricultura de Bebedouro, SP to evaluate the response of glyphosate-resistant horseweed and hairy fleabane biotypes to herbicides glyphosate, bromacil + diuron, diuron e paraquat isolated and in mixture and effect of a sequential application of glyphosate. The experimental design was of complete randomized blocks with four replication and seven treatments. The herbicides were applied with costal sprayer, constant pressure with three nozzles TT110015, the equivalent spray volume was 150 L ha(-1). The control was visually evaluated, trough percentile note scale. The best results were obtained to general control of weed with diuron isolated and glyphosate in mixture with bromacil + diuron while to glyphosate-resistant horseweed and hairy fleabane there was no difference between the treatments. After sequential application to Conyza sp control, the best treatment was obtained associated with diuron and bromacil+diuron.

Only the B biotype of Bemisia tabaci is present on vegetables in São Paulo State, Brazil

Bemisia tabaci (Genn.) é considerada uma das mais importantes pragas em cultivos de hortaliças e ornamentais em todo o mundo. Baseado na análise da seqüência mitocondrial (citocromo oxidase I - mtCOI) foi proposto recentemente que B. tabaci deva ser considerado um complexo críptico de espécies, contendo 11 grupos e 24 espécies. Dois destes grupos: Middle East-Asia Minor e Mediterranean englobam os biótipos B e Q, respectivamente. Avaliou-se a sequência mtCOI de espécimes de B. tabaci coletados em regiões do estado de São Paulo, Brasil. Por PCR-RFLP utilizando-se a enzima Taq I, pôde-se observar somente o padrão típico de clivagem para o biótipo B. Comparando-se com sequências consenso, todas as moscas brancas foram classificadas no grupo Middle East-Asia Minor e puderam ser separadas em quatro haplótipos, indicando prevalência do biótipo B em áreas de pimentão (Capsicum annuum L.), tomate (Solanum lycopersicum L.), cucurbitáceas e berinjela (Solanum melongena L.) do Estado de São Paulo.

Caracterização genética de populações de Egeria najas presentes no reservatório de Jupiá e rios afluentes

A Egeria najas é uma espécie aquática submersa nativa da bacia hidrográfica do rio Paraná. Com o represamento das águas do rio para geração de energia elétrica a espécie tem mudado seu comportamento de colonização dos leitos dos rios e ocorrido em grandes maciços dentro da represa de Jupiá e rios afluentes. Essa planta tem causado problemas por obstruir a passagem de água para as turbinas de geração de energia elétrica. Coletas de material vegetativo foram realizadas na represa e afluentes do rio Paraná para análise da variabilidade isoenzimática e de DNA. A análise isoenzimática mostrou haver somente quatro classes de diferentes biotipos. No entanto, utilizando-se os procedimentos de RAPD, observou-se que a espécie possui grande variabilidade genética. O represamento das águas também está permitindo o acumulo de variabilidade no local e promovendo um aumento de variabilidade por meio de possíveis cruzamentos entre genótipos diferentes. Os resultados também possibilitaram inferir sobre as possíveis rotas de migração de material genético para colonização da represa e rios afluentes.

Sensibilidade de diferentes acessos de Egeria najas e Egeria densa aos herbicidas diquat e fluridone

O surgimento de novos genótipos de plantas daninhas aquáticas pertencentes ao gênero Egeria motivou a realização deste estudo, o qual teve como objetivo avaliar o comportamento de diferentes acessos das espécies de E. densa e E. najas em relação ao tratamento com os herbicidas diquat e fluridone. Foram avaliados acessos de diferentes biótipos de E. densa provindos dos reservatórios de Jupiá, Nova Avanhandava, Promissão e Salto Grande e de E. najas oriundos de Jupiá, Três Irmãos, Ilha Solteira e Ibitinga. O delineamento experimental adotado foi o inteiramente casualizado com cinco repetições. Os tratamentos e concentrações utilizados no estudo com E. densa e E. najas foram: diquat (Reward 240 g L-1) a 0,5 e 1,0 mg L-1 com reaplicação aos 12 dias após a primeira aplicação, e a concentração de fluridone mantida entre 3 e 20 mg L-1, além de uma testemunha sem aplicação de herbicidas. As duas espécies foram controladas de forma eficiente pelo herbicida diquat com uma ou duas aplicações, sendo ambas as concentrações eficientes, porém observaram-se respostas diferenciadas dos acessos estudados às concentrações do herbicida. Para o herbicida fluridone, as espécies e acessos estudados comportaram-se também de forma diferenciada em relação à ação do herbicida, ocorrendo acessos desde mediana a altamente suscetíveis em ambas as espécies.

Virulence characteristics and epidemiology of Yersinia enterocolitica and Yersiniae other than Y-pseudotuberculosis and Y-pestis isolated from water and sewage

Aims: To determine the species, bio-sero-phagetypes, antimicrobial drug resistance and also the pathogenic potential of 144 strains of Yersinia spp. isolated from water sources and sewage in Brazil.Methods and Results: the 144 Yersinia strains were characterized biochemically, serologically and had their antibiotic resistance and phenotypic virulence markers determined by microbiological and serological standard techniques. The Y. enterocolitica strains related to human diseases were also tested for the presence of virulence genes, by the PCR technique. The isolates were classified as Y. enterocolitica, Y. intermedia, Y. frederiksenii, Y. kristensenii and Yersinia biochemically atypical. The 144 isolates belonged to various bio-serogroups. Half of the strains showed resistance to three or more drugs. The Y. enterocolitica strains related to human diseases exhibited phenotypic virulence characteristics and virulence genes.Conclusions: Water from various sources and sewage are contaminated with Yersinia spp. in Brasil. Among these bacteria, virulent strains of Y. enterocolitica were found, with biotypes and serogroups related to human diseases.Significance and Impact of the Study: This is the first documented description of the occurrence of pathogenic Y. enterocolitica in water sources and sewage in Brazil. The occurrence of virulence strains of Y. enterocolitica shows that the environment is a potential source of human infection by this species in this country.

Differential nutritional, endocrine, and cardiovascular effects in obesity-prone and obesity-resistant rats fed standard and hypercaloric diets

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Formando lotes uniformes de reprodutores múltiplos e usando-os em acasalamentos dirigidos, em populações Nelore

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Yeasts associated with nests of the leaf-cutting ant Atta sexdens rubropilosa Forel, 1908

A total of 137 yeasts associated with the leaf-cutting ant Atta sexdens rubropilosa Forel, 1908 were characterized, being selected 93 for analysis. Twenty four species belonging to seven genera(Candida, Cryptococcus, Rhodotorula, Sporobolomyces, Tremella, Trichosporon, Pichia) were isolated from the different analysed material. The genus Candida was widely distributed, with C. homilentoma, C. colliculosa-like, C. famata and C. colliculosa being the most prevalent. A few isolates did not fit the standard descriptions and probably some of them could be new biotypes or even new species. Three strains of black yeasts were also isolated, and four others were identified as being Candida spp. The effective number of yeast species was higher in newer sponge. The origin, distribution and relative importance of these microorganisms for the ants are discussed.

EVALUATION OF DIFFERENT TECHNIQUES FOR THE DIFFERENTIATION OF PATHOGENIC AND NON PATHOGENIC STRAINS OF YERSINIA-ENTEROCOLITICA

Different methods and tests have been used to evaluate the pathogenic potential of distinct Y. enterocolitica serotypes and biotypes. We tested a total of 60 Y. enterocolitica strains, being 25 of human origin (serotype O3 biotype 4 and serotype O5 biotype 1); 6 of animal origin (serotype O3 biotype 4); 19 isolated from the environment (serotype O5.27 biotypes 1 and 2); and 8 isolated from food (serotype O5 biotype 1 and serotype 05.27 biotype 1). The methods used were based on plasmid gene expression (autoagglutination, calcium-dependence at 37 degrees C and Congo Red absorption tests), chromosomal gene expression (assays for pyrazinamidase activity, salicin fermentation and esculin hydrolysis), and invasion of HEp-2 cells. All but one of the Y. enterocolitica O3 strains, were found to be potentially pathogenic when submitted to the pyrazinamidase-salicin-esculin tests. In contrast, the results obtained with the assays related to plasmidial gene expression were not so uniform, probably due to plasmid loss. The least homogeneous results were obtained with the HEp-2 cell invasion test. Y. enterocolitica O5 behaved in a uniform manner when tested with the first two groups of tests (based on chromosomal and plasmidial gene expression), but not when tested with the HEp-2 invasion assay. The strains of serotype O5.27 biotype 1 presented a uniform behavior hen submitted to the chromosomic-related tests, showing no pathogenicity. However, they did not provide conclusive results with the tests related to plasmidial gene expression or HEp-2 cell invasion. We conclude that the tests related to chromosomal gene expression (esculin-salicin-pyrazinamidase) are simple and highly effective for the detection of potentially pathogenic Y. enterocolitica isolated from clinical cases.

Identification and characterization of different accessions of itchgrass (Rottboellia cochinchinensis)

Studies were conducted to identify and characterize different accessions of itchgrass. Seeds were collected in the counties of Aramina, Campinas, Dumont, Igarapava, Jaboticabal, and Ribeirao Preto, all in the state of São Paulo, Brazil. Accessions were characterized based on dimensions of their stomata, stomatal index (SI), and length and width of their seed (caryopses and husk). Chromosome number and length also were determined, and accessions were further differentiated using molecular markers (polymerase chain reaction [PCR]). Itchgrass from Ribeirao Preto had much longer and narrower seeds than those from the other locations, and their husks were longer as well. Accessions had similar SIs, both on the abaxial and adaxial leaf surfaces. Stomata from Campinas and Igarapava accessions were longer and wider, whereas those from Dumont and Ribeirao Preto were similar and smaller than all others. The accession from Ribeirao Preto is diploid (2n = 20); the rest are polyploid, with the total length of chromosomes smaller than all others. These differences were confirmed by molecular differentiation (PCR).