Air-borne fungi in the air of Barcelona (Spain). II. The genus Alternaria

The genus Alternaria Nees formed one of the more important components of the fungus population of the air at Barcelone (Spain), during a two-year study, from February 1976 through January 1978. Results were based only on studies of colonies obtained by gravity-exposed plates.The occurrence of this genus was greatly affected by climatic conditions. In general, however, Alternaria appeared to prefer warmer weather.A total of six species of Alternaria was identified, of which, Alternaria alternata (Fries) Keissler was by far the commonest, representing 71.3% of all colonies.     

Lung infection with gamma-herpesvirus induces progressive pulmonary fibrosis in Th2-biased mice

Idiopathic pulmonary fibrosis (IPF) is a progressive, fibrotic lung disease of unknown etiology. A viral pathogenesis in IPF has been suggested since >95% of IPF patients have evidence of chronic pulmonary infection with one or more herpesviruses. To determine whether pulmonary infection with herpesvirus can cause lung fibrosis, we infected mice with the murine gamma-herpesvirus 68 (MHV68). Because IPF patients have a T helper type 2 (Th2) pulmonary phenotype, we used IFN-gammaR-/-, a strain of mice biased to develop Th2 responses. Chronic MHV68 infection of IFN-gammaR-/- mice resulted in progressive deposition of interstitial collagen as shown by light and electron microscopy. A significant decrease in tidal volume paralleled the collagen deposition. Five features typically seen in IPF, increased transforming growth factor-beta expression, myofibroblast transformation, production of Th2 cytokines, hyperplasia of type II cells, and increased expression of matrix metalloproteinase-7, were also present in chronically infected IFN-gammaR-/- mice. There also was altered synthesis of surfactant proteins, which is seen in some patients with familial IPF. MHV68 viral protein was found in type II alveolar epithelial cells, especially in lung areas with extensive alveolar remodeling. In summary, chronic herpesvirus pulmonary infection in IFN-gammaR-/- mice causes progressive pulmonary fibrosis and many of the pathological features seen in IPF.     

Production of saccharogenic and dextrinogenic amylases by Rhizomucor pusillus A 13.36

A newly-isolated thermophilic strain of the zygomycete fungus Rhizomucor pusillus 13.36 produced highly active dextrinogenic and saccharogenic enzymes. Cassava pulp was a good alternative substrate for amylase production. Dextrinogenic and saccharogenic amylases exhibited optimum activities at a pH of 4.0-4.5 and 5.0 respectively and at a temperature of 75 degrees C. The enzymes were highly thermostable, with no detectable loss of saccharogenic or dextrinogenic activity after 1 h and 6 h at 60 degrees C, respectively. The saccharogenic activity was inhibited by Ca(2+) while the dextrinogenic was indifferent to this ion. Both activities were inhibited by Fe(2+) and Cu(2+) Hydrolysis of soluble starch by the crude enzyme yielded 66% glucose, 19.5% maltose, 7.7% maltotriose and 6.6% oligosaccharides.     

A rapid and efficient method for purifying high quality total RNA from peaches (Prunus persica) for functional genomics analyses

Prunus persica has been proposed as a genomic model for deciduous trees and the Rosaceae family. Optimized protocols for RNA isolation are necessary to further advance studies in this model species such that functional genomics analyses may be performed. Here we present an optimized protocol to rapidly and efficiently purify high quality total RNA from peach fruits (Prunus persica). Isolating high-quality RNA from fruit tissue is often difficult due to large quantities of polysaccharides and polyphenolic compounds that accumulate in this tissue and co-purify with the RNA. Here we demonstrate that a modified version of the method used to isolate RNA from pine trees and the woody plant Cinnamomun tenuipilum is ideal for isolating high quality RNA from the fruits of Prunus persica. This RNA may be used for many functional genomic based experiments such as RT-PCR and the construction of large-insert cDNA libraries.     

Detection of prion epitopes on PrP and PrP of transmissible spongiform encephalopathies using specific monoclonal antibodies to PrP

Amino acid residues 90-120 of the prion protein (PrP) are likely to be critical for the conversion of PrP(c) to PrP(sc) in the transmissible spongiform encephalopathies. We raised 10 monoclonal antibodies against the 90-120 amino acid region, mapped the epitope specificity of these anti-PrP antibodies, and investigated the expression of epitopes recognized by the antibodies in both PrP(c) and PrP(sc). Four out of five of the anti-PrP antibodies raised in a prion knockout mouse immunized with the linear peptide of PrP90-120 could detect PrP(sc) in 'native' and denatured forms and PrP(c) in normal cells, as well as recognize epitopes within PrP93-112 residues. In contrast, the other six anti-PrP reagents, including five raised from the two knockout mice immunized with conformationally modified PrP90-120 peptide, could detect PrP(c) and recognize epitopes within PrP93-107 residues. Four of these reagents could also detect denatured PrP(sc) on western blots but not PrP(sc) plaques in brain tissue. The results indicate that residues PrP93-102 are exposed in PrP(c) but are buried upon conversion to the PrP(sc) isoform. Furthermore, PrP103-107 residues are partially buried in PrP(sc) while only the PrP107-112 epitope remains exposed, suggesting that the region PrP93-112 undergoes conformational changes during its conversion to PrP(sc).     

Efficient preservation in a silicon oxide matrix of Escherichia coli, producer of recombinant proteins

The aim of this work was to study the use of silicon oxide matrices for the immobilization and preservation of recombinant-protein-producing bacteria. We immobilized Escherichia coli BL21 transformants containing different expression plasmids. One contained DNA coding for a T-cell receptor β chain, which was expressed as inclusion bodies in the cytoplasm. The other two encoded bacterial superantigens Staphylococcal Enterotoxin G and Streptococcal Superantigen, which were expressed as soluble proteins in the periplasm. The properties of immobilization and storage stability in inorganic matrices prepared from two precursors, silicon dioxide and tetraethoxysilane, were studied. Immobilized E. coli was stored in sealed tubes at 4 and 20°C and the number of viable cells and level of recombinant protein production were analyzed weekly. Different tests showed that the biochemical characteristics of immobilized E. coli remained intact. At both temperatures selected, we found that the number of bacteria in silicon dioxide-derived matrix was of the same order of magnitude (10⁹ cfu ml⁻¹) as before immobilization, for 2 months. After 2 weeks, cells immobilized in an alkoxide-derived matrix decreased to 10⁴ cfu ml⁻¹ at 4°C, and no viable cells were detected at 20°C. We found that immobilized bacteria could be used as a starter to produce recombinant proteins with yields comparable to those obtained from glycerol stocks: 15 mg l⁻¹ for superantigens and 2 mg l⁻¹ for T-cell receptor β chain. These results contribute to the development of methods for microbial cell preservation under field conditions. Martín F. Desimone and Mauricio C. De Marzi contributed equally to this work     

The efficacy of short tandem repeat polymorphisms versus single-nucleotide polymorphisms for resolving population structure

Accurately resolving population structure in a sample is important for both linkage and association studies. In this study we investigated the power of single-nucleotide polymorphisms (SNPs) in detecting population structure in a sample of 286 unrelated individuals. We varied the number of SNPs to determine how many are required to approach the degree of resolution obtained with the Collaborative Study on the Genetics of Alcoholism (COGA) short tandem repeat polymorphisms (STRPs). In addition, we selected SNPs with varying minor allele frequencies (MAFs) to determine whether low or high frequency SNPs are more efficient in resolving population structure. We conclude that a set of at least 100 evenly spaced SNPs with MAFs of 40-50% is required to resolve population structure in this dataset. If SNPs with lower MAFs are used, then more than 250 SNPs may be required to obtain reliable results.     

Social cues and hormone levels in male Octodon degus (Rodentia): a field test of the Challenge Hypothesis

Social interactions are important factors determining and regulating individual behaviors. Testosterone has been related to agonistic interactions, while glucocorticoids have been related to social stress, especially during interactions of dominance. We compared testosterone and cortisol concentrations in male degus (Octodon degus, Rodentia) under laboratory conditions without male social interactions, with data from wild males in nature. Under natural conditions, males should present higher levels of testosterone during the breeding season due to social interactions (Challenge Hypothesis). Alternatively, intense social instability could act as a stressing environment, raising glucocorticoids, which inhibit testosterone concentrations. Our results show a significant increase in agonistic interactions between males during the breeding season, and disappearance of non-agonistic male interactions during this period. Hormone levels in breeding season show nonsignificant differences between laboratory groups, but testosterone concentrations in field males were significantly higher than in laboratory males. Testosterone levels were similar among pre-breeding and breeding periods, but in field animals the concentration was approximately 30% higher than in laboratory degus. In field animals, we found two different mating strategies: resident males, with territorial behavior, and transient males, displayed an opportunistic approach to females. Finally, cortisol presents a similar pattern in both laboratory and field animals; pre-breeding values of cortisol are higher than during the breeding season. This suggests that social interactions in O. degus activate a rise in testosterone, supporting the Challenge Hypothesis, and could be considered as partial support of the Social Stress Hypothesis.     

The T-cell receptor in primates: identifying and sequencing new owl monkey TRBV gene sub-groups

The New World primate Aotus nancymaae (owl monkey) has been shown to be an excellent experimental model when studying malarial parasites. Characterising the T-cell receptor (TR) repertoire by means of the different variable beta (TRBV) genes displayed contributes to a better understanding of these lymphocytes role in the response against several malarial antigens. This study describes identifying and characterising eleven new TRBV gene sub-groups in cDNA from Aotus nancymaaes peripheral blood lymphocytes; these 11 gene sequences displayed homology to the previously reported human TRBV3, TRBV10, TRBV11, TRBV14, TRBV18, TRBV19, TRBV20, TRBV25, TRBV27, TRBV29 and TRBV30 sub-groups, resulting in 83% overall homology at the amino acid level. An additional Aotus sequence was found having similarity with the human TRBJ-2–7*01 gene. Evolutionary relationships amongst these sequences and the homologous genes from both New and Old World primates have shown that the TRBV repertoire has been maintained in the species being studied, displaying varying association patterns and substitution rates, depending on the sub-group being studied. The degree of identity observed when comparing human and Aotus genes suggests that these species might have a similar TRBV repertoire.