The Neogene of the Lower Tagus Basin (Portugal)

Revista Española de Paleontologia 19 (2), 229-242

Areias de Vale de Santarém. Estratigrafia e Palinologia

The palynologic study of several boreholes for lignite prospection at the "Vale de Santarém sands" is presented. Height spores and 18 pollen forms have been identified. The quantitative and qualitative results are shown in table 2. Four palynologic associations (A - D) corresponding to climatic changes have been characterized. Correlations are established with the plant associations of Rio Maior Basin. Correlation between the boreholes allow a better understanding of the small, Vale de Santarém basin infilling.

Intestinal microsporidiosis in HIV-positive patients with chronic unexplained diarrhea in Rio de Janeiro, Brazil: diagnosis, clinical presentation and follow-up

After the diagnosis of two cases of microsporidial intestinal infection in 1992, in Rio de Janeiro, we have started looking for this parasite in HIV-infected patients with chronic unexplained diarrhea. We have studied 13 patients from Hospital Evandro Chagas, IOC-FIOCRUZ. Fecal specimens from these patients were examined for the presence of Cryptosporidia and Microsporidia, in addition to routine examination. Spores of Microsporidia were found in the stools of 6 (46.1%) of the 13 patients studied, with 2 histological jejunal confirmations. The Microsporidia-infected patients presented chronic diarrhea with about 6 loose to watery bowel movements a day. Five infected patients were treated with Metronidazole (1.5 g/day). They initially showed a good clinical response, but they never stopped eliminating spores. After about the 4th week of therapy, their diarrhea returned. Two patients utilized Albendazole (400 mg/day-4 weeks) with a similar initial improvement and recurrence of the diarrhea. Intestinal Microsporidiosis seems to be a marker of advanced stages of AIDS, since 5 of our 6 infected patients were dead after a 6 month period of follow-up. The present study indicates that intestinal microsporidiosis may be a burgeoning problem in HIV-infected patients with chronic diarrhea in Brazil, which deserves further investigation.

Evaluation of the allergenicity of spore and mycelia extracts of Pisolithus tinctorius

The antigenic and allergenic chemical analysis of spore and mycelia extracts of Pisolithus tinctorius was carried out. The spores were collected from basidiocarps in plantations of Eucalyptus spp and the mycelia from culture in MNM medium. With basis on the fungus growth curve, the mycelia masses were obtained after 10, 20, 30, and 40 days of incubation, which correspond, respectively, to the beginning, middle and end of the log phase, and beginning of the decline phase. The mycelia masses, together with the spores, were submitted to the action of three extractors (Coca, Tris-HCl, and ammonium bicarbonate). The contents of carbohydrates and proteins were determined. The SDS-PAGE electrophoretical analysis revealed separate fractions in these extracts, besides common fractions, in function of cultivation time and extraction methods. The selected extracts for the allergic tests were the ones with the highest number of fractions. The prick-tests were conducted in 374 patients – rural workers, eucalyptus plantation works and college students. The positivity to the "prick test" with the antigenic extract of P. tinctorius was, respectively, 3.78%, 28.20% and 6.40%. Most prick-test positive patients (82.75%) also presented symptoms of respiratory allergy (asthma and rhinitis). There was no reactivity difference when the spore and mycelia extracts were employed. The analysis of the positive patients’ sera revealed the presence of IgE specific to the P. tinctorius antigens. Since Pisolithus tinctorius is found as mycorrhyza of Eucalyptus spp, and this plant is used in reforestation in most countries, the importance of that fungus should be regarded as a possible cause of respiratory allergies, especially in occupationally exposed workers

A 3-YEAR FOLLOW-UP OF A BRAZILIAN AIDS PATIENT WITH PROTRACTED DIARRHEA CAUSED BY Enterocytozoon bieneusi

Enterocytozoon bieneusi is the most prevalent microsporidian parasite that causes gastrointestinal infection in persons with AIDS. Microsporidia are increasingly recognized as important opportunistic pathogens all over the world but in Brazil only few cases have been reported due either to the non awareness of the clinical presentation of the disease or to difficulties in the laboratory diagnosis. We report a 3-year follow-up of a Brazilian HIV-positive patient in whom microsporidial spores were detected in stools and were identified as E. bieneusi using electron microscopy and PCR. The patient presented with chronic diarrhea, CD4 T-lymphocytes count below 100/mm3 and microsporidial spores were consistently detected in stools. Albendazole was given to the patient in several occasions with transient relief of the diarrhea, which reappeared as soon as the drug was discontinued. Nevertheless, a diarrhea-free period with weight gain up to 18 Kg occurred when a combination of nucleoside and protease inhibitors was initiated as part of the antiviral treatment.

Clinical and diagnostic aspects of intestinal microsporidiosis in HIV-infected patients with chronic diarrhea in Rio de Janeiro, Brazil

The objectives of this study were to determine both the prevalence of microsporidial intestinal infection and the clinical outcome of the disease in a cohort of 40 HIV-infected patients presenting with chronic diarrhea in Rio de Janeiro, Brazil. Each patient, after clinical evaluation, had stools and intestinal fragments examined for viral, bacterial and parasitic pathogens. Microsporidia were found in 11 patients (27.5%) either in stools or in duodenal or ileal biopsies. Microsporidial spores were found more frequently in stools than in biopsy fragments. Samples examined using transmission electron microscopy (n=3) or polymerase chain reaction (n=6) confirmed Enterocytozoon bieneusi as the causative agent. Microsporidia were the only potential enteric pathogens found in 5 of the 11 patients. Other pathogens were also detected in the intestinal tract of 21 patients, but diarrhea remained unexplained in 8. We concluded that microsporidial infection is frequently found in HIV infected persons in Rio de Janeiro, and it seems to be a marker of advanced stage of AIDS.

Airborne fungi in the city of Porto Alegre, Rio Grande do Sul, Brazil

Knowledge of anemophilous fungi in a given city or region is important for the ecological diagnosis and specific treatment of allergic manifestations induced by inhaled allergens. In order to diagnose the presence of anemophilous fungi, several qualitative and quantitative techniques are used depending on the study place. This study of fungal air spores was performed with a Rotorod Sampler®, an equipment which samples the air through a plastic rod attached to an electric engine that makes it spin fast enough to collect the particles in the air. The samples were collected once a week during 24 hours using the standard cycle of the manufacturers. A total of 52 samples were obtained from April 2000 through March 2001. The results revealed prevalence of ascosporos (50.49%), Cladosporium (17.86%), Aspergillus/Penicillium (15.03%), basidiosporos (3.84%), rusts (3.82%), and Helminthosporium (2.49%), and a lesser frequency of Botrytis (1.22%), Alternaria (1.19%), smuts (0.90%), Curvularia (0.87%), Nigrospora (0.61%), and Fusarium (0.08%). Also, 1.59% of the spores detected here could not be identified by the systematic key used. More fungal spores were observed during the summer than during the autumn.

Airborne fungi isolated from Fortaleza city, State of Ceará, Brazil

Airbone fungi are considered important causes of allergic rhinitis and allergic asthma. The knowledge of these fungi in a city or region is important for the ecological diagnosis and specific treatment of allergic manifestations induced by inhalation of fungal allergens. The airborne fungi of Fortaleza, State of Ceará, Brazil, were studied during a one year period. Five hundred and twenty Petri dishes with Sabouraud dextrose agar medium were exposed at ten different locations in the city. The dishes exposed yielded one thousand and five hundred and twenty one colonies of twenty four genera. The most predominants were: Aspergillus (44.7%), Penicillium (13.3%), Curvularia (9.8%), Cladosporium (6.8%), Mycelia sterilia (6.0%), Fusarium (3.5%), Rhizopus (3.1%), Drechslera (2.6%), Alternaria (2.4%) and Absidia (2.2%). The results shown that Aspergillus, Penicillium, Mycelia sterilia, Fusarium and Alternaria were found during all months in the year. Absidia was more frequent during the dry season. Anemophilous fungi and the high concentration of spores in the air are important because may result in an increased number of people with allergic respiratory disease.

Assembly and function of a protein cross-linking enzyme during bacterial spore morphogenesis

Sporulation in Bacillus subtilis culminates with the formation of a dormant endospore. The endospore (or spore) is one of the most resilient cell types known and can remain viable in the environment for extended periods of time. Contributing to the spore’s resistance and its ability to interact with and monitor its immediate environment is the coat, the outermost layer of B. subtilis spores. The coat is composed by over 70 different proteins, which are produced at different stages in sporulation and orderly assembled around the developing spore.(...)

Description of microsporidia in simulids: molecular and morphological characterization of microsporidia in the larvae of Simulium pertinax Kollar (Diptera: Simuliidae)

IntroductionMicrosporidia constitute the most common black fly pathogens, although the species' diversity, seasonal occurrence and transmission mechanisms remain poorly understood. Infections by this agent are often chronic and non-lethal, but they can cause reduced fecundity and decreased longevity. The objective of this study was to identify microsporidia infecting Simulium (Chirostilbia) pertinax (Kollar, 1832) larvae from Caraguatatuba, State of São Paulo, Brazil, by molecular and morphological characterization.MethodsLarvae were collected at a single point in a stream in a rural area of the city and were kept under artificial aeration until analysis. Polydispyrenia spp. infection was characterized by the presence of at least 32 mononuclear spores measuring 6.9 ± 1.0 × 5.0 ± 0.7µm in persistent sporophorous vesicles. Similarly, Amblyospora spp. were characterized by the presence of eight uninucleate spores measuring 4.5 × 3.5µm in sporophorous vesicles.ResultsThe molecular analysis confirmed the presence of microsporidian DNA in the 8 samples (prevalence of 0.51%). Six samples (Brazilian larvae) were related to Polydispyrenia simulii and Caudospora palustris reference sequences but in separate clusters. One sample was clustered with Amblyospora spp. Edhazardia aedis was the positive control taxon.ConclusionsSamples identified as Polydispyrenia spp. and Amblyospora spp. were grouped with P. simulii and Amblyospora spp., respectively, corroborating previous results. However, the 16S gene tree showed a considerable distance between the black fly-infecting Amblyospora spp. and the mosquito-infecting spp. This distance suggests that these two groups are not congeneric. Additional genomic region evaluation is necessary to obtain a coherent phylogeny for this group.

Exploring the relationship between toxin and spore production in the human enteric pathogen Clostridium difficile

RESUMO: Clostridium difficile é presentemente a principal causa de doença gastrointestinal associada à utilização de antibióticos em adultos. C. difficile é uma bactéria Gram-positiva, obrigatoriamente anaeróbica, capaz de formar endósporos. Tem-se verificado um aumento dos casos de doença associada a C. difficile com sintomas mais severos, elevadas taxas de morbilidade, mortalidade e recorrência, em parte, devido à emergência de estirpes mais virulentas, mas também devido à má gestão do uso de antibióticos. C. difficile produz duas toxinas, TcdA e TcdB, que são os principais fatores de virulência e responsáveis pelos sintomas da doença. Estas são codificadas a partir do Locus de Patogenicidade (PaLoc) que codifica ainda para um regulador positivo, TcdR, uma holina, TcdE, e um regulador negativo, TcdC. Os esporos resistentes ao oxigénio são essenciais para a transmissão do organismo e recorrência da doença. A expressão dos genes do PaLoc ocorre em células vegetativas, no final da fase de crescimento exponencial, e em células em esporulação. Neste trabalho construímos dois mutantes de eliminação em fase dos genes tcdR e tcdE. Mostrámos que a auto-regulação do gene tcdR não é significativa. No entanto, tcdR é sempre necessário para a expressão dos genes presentes no PaLoc. Trabalho anterior mostrou que, com a exceção de tcdC, os demais genes do PaLoc são expressos no pré-esporo. Mostrámos aqui que TcdA é detectada à superfície do esporo maduro e que a eliminação do tcdE não influencia a acumulação de TcdA no meio de cultura ou em associação às células ou ao esporo. Estas observações têm consequências para o nosso entendimento do processo infecioso: sugeremque o esporo possa ser também um veículo para a entrega da toxina nos estágios iniciais da infecção, que TcdA possa ser libertada durante a germinação do esporo, e que o esporo possa utilizar o mesmo receptor reconhecido por TcdA para a ligação à mucosa do cólon.---------------------------ABSTRACT: Clostridium difficile is currently the major cause of antibiotic-associated gastrointestinal diseases in adults. This is a Gram-positive bacterium, endospore-forming and an obligate anaerobe that colonizes the gastrointestinal tract. Recent years have seen a rise in C. difficile associated disease (CDAD) cases, associated with more severe disease symptoms, higher rates of morbidity, mortality and recurrence, which were mostly caused due to the emergence of “hypervirulent” strains but also due to changing patterns of antibiotics use. C. difficile produces two potent toxins, TcdA and TcdB, which are the main virulence factors and the responsible for the disease symptoms. These are codified from a Pathogenicity Locus (PaLoc), composed also by the positive regulator, TcdR, the holin-like protein, TcdE, and a negative regulator, TcdC. Besides the toxins, the oxygen-resistant spores are also essential for transmission of the organism through diarrhea; moreover, spores can accumulate in the environment or in the host, which will cause disease recurrence. The expression of the PaLoc genes occurs in vegetative cells, at the end of the exponential growth phase, and in sporulating cells. In this work, we constructed two in-frame deletion mutants of tcdR and tcdE. We showed that the positive auto regulation of tcdR is not significant. However, tcdR is always necessary for the expression of the PaLoc genes. A previous work showed that, except tcdC, all the PaLoc genes are expressed in the forespore. Here, we detected TcdA at the spore surface. Furthermore, we showed that the in-frame deletion of tcdE does not affect the accumulation of TcdA in the culture medium or in association with cells or spores. This data was important for us to conclude about the infeccious process: it suggests that the spore may be the vehicle for the delivery of TcdA in early stages of infection, that TcdA may be released during spores germination and that this spore may use the same receptor recognized by TcdA to bind to the colonic mucosa.

Determinants for the subcellular localization of the inner and outer spore coat hubs in Bacillus subtilis

Endospores, or spores for simplicity, are a highly resistant cell type produced by some bacterial species under adverse conditions. Two main protective layers contribute to the resilience of spores: the cortex, composed of peptidoglycan, and the outermost proteinaceous coat. In Bacillus subtilis, the coat comprises up to 80 different proteins, organized into four sublayers: the basement layer, the inner coat, the outer coat and the crust. These proteins are synthesized at different times during sporulation and deposited at the spore surface in multiple coordinated waves. Central to coat formation is a group of morphogenetic proteins that guide the assembly of the coat components. Targeting of the coat proteins to the surface of the developing spore is mainly controlled by the SpoIVA morphogenetic ATPase. In a second stage, the coat proteins fully encircle the spore, a process termed encasement that requires the morphogenetic protein SpoVID. Assembly of the inner coat requires SafA, whereas formation of the outer coat and the crust requires CotE. SafA interacts directly with the N terminus of SpoVID. (...)

Microscopy- or DNA-based analyses: Which methodology gives a truer picture of stream-dwelling decomposer fungal diversity?

We assessed aquatic hyphomycete diversity in autumn and spring on oak leaves decomposing in five streams along a gradient of eutrophication in the Northwest of Portugal. Diversity was assessed through microscopy-based (identification by spore morphology) and DNA-based techniques (Denaturing Gradient Gel Electrophoresis and 454 pyrosequencing). Pyrosequencing revealed five times greater diversity than DGGE. About 21% of all aquatic hyphomycete species were exclusively detected by pyrosequencing and 26% exclusively by spore identification. In some streams, more than half of the recorded species would have remained undetected if we had relied only on spore identification. Nevertheless, in spring aquatic hyphomycete diversity was higher based on spore identification, probably because many species occurring in this season are not yet connected to ITS barcodes in genetic databases. Pyrosequencing was a powerful tool for revealing aquatic hyphomycete diversity on decomposing plant litter in streams and we strongly encourage researchers to continue the effort in barcoding fungal species.

Nitrificação e aproveitamento de alguns adubos nitrogenados

The rate of nitrification of several nitrogenous fertilizers (ammonium sulfate, nitre-chalk, ureia, and cottonseed meal) was studied in three soils, namely, "terra roxa legítima", a red soil derived from basalt, "terra roxa misturada", a soil also derived from basalt but with a higher proportion of sand, and "areito Corumbataí", a sandy soil. The effects of the following treatments on nitrification were considered: addition of limestone of micronutrients (Fe, Cu, Zn, Mn, and Mo), and inoculation with a suspension of spores of Aspergillus wentii, a heterotrophic nitrifier. The results showed that: in "terra roxa legítima" limestone had no influence on the nitrification rate, whereas the micronutrients estimulated the oxidation of nitre-chalk, cottonseed meal and urea; inoculation with A. wentii helped only the nitrification of ammonium sulfate and of the cottonseed meal; the latter, in all the treatments employed gave use to a smaller amount of nitrates; in "terra roxa misturada", all the fertilizers depending upon the treatments they were subjected to, presented maximum values for nitrification; limestone estimulated the oxidation of ammonium sulfate as well as the mineralization of the cottonseed meal; the addition of micronutrients helped the nitrification of all the fertilizers, except that of urea; inoculation showed a benefical influence on the nitrification of ammonium sulfate and cottonseed meal; in "arenito de Corumbatai", the amounts of nitrates produced was roughly the same for all the fertilizers investigated; limestone estimulated the nitrification of nitro-chalk, ammonium sulfate and cottonseed meal whilst the addition of micronutrients benefited only the latter two; the inoculation with A. wentii helped the oxidation of all the fertilizers. In order to study the availability of the various fertilizers above discussed, two plant growing experiments were carried cut, one in pots, using the three soil types and another one in the field, with "terra roxa misturada". In "arenito de Corumbatai" there was no significant difference in the yield both of straw and rice grains for none of the fertilizers: Chilean nitrate of soda was used as a control; ho marked agreement could be detected between the data concerning nitrification and the yield results. In "terra roxa legítima", ammonium sulfate won the competition and there was a good parallelism between nitrification and yield. In "terra roxa misturada", there was no statistical difference among the various fertilizers; the agreement between nitrification and yields was reasonable. In the field (corn), Chilean nitrate, ammonium sulfate and nitro-chalk were clearly beter than urea and cottonseed meal which did not differ from the minus nitrogen plots.

Propagação da samambaia-de-metro

The author studied the possibility of propagating "Samambaia de Metro" Polypodium Subauriculatum, by means of spores, since vegetative multiplication is hard to obtain for large numbers of plants. Five treatments were tried, with 4 replications. I - Coarse "Flores Floor" and fine "Flores Floor". II - Coarse Flores Floor and moss. III - Coarse sand, Flores Floor and brick powder VI - Coarse sand, Flores Floor and brick powder V - Coarse sand and Flores Floor. The best germination was obtained with treatment V (Coarse sand et the boltom), topesed by Flores Floor. The pots were permanently kept on a tray pul of water, in a Greenhouse. This method provided over 4.000 offspring, which at 2 ½ years of age were of a size mitable for ornamental use.