Identification of Staphylococcus spp. isolated during the ripening process of a traditional minas cheese / Identificação de Staphylococcus spp. isolados durante o processo de maturação de um queijo-de-minas tradicional

The population dynamics of Staphylococcus spp. was studied during the ripening of Canastra Minas cheese at three farms located in the State of Minas Gerais, Brazil. The presence of coagulase (coa), thermonuclease (nuc), and enterotoxin (sea, seb, sec, and sed) genes was investigated in Staphylococcus strains isolated during the 60-day cheese-ripening period. The presence of the staphylococcal enterotoxins A, C, and D was also investigated in the cheese samples. Cheese samples that were matured for 0, 7, 15, 30, and 45 days presented staphylococci counts from 10³ to 10(8)cfu/g. All isolates considered coagulase-positive by physiological tests had the coa gene. However, no association was observed between the results obtained with biochemical tests and those obtained by PCR using gene-specific primers for coagulase-negative strains. Coagulase and thermonuclease genes occurred simultaneously in 41.3 percent of Staphylococcus spp. tested. None of the investigated Staphylococcus strains expressed enterotoxins SEA, SEB, SEC, and SED. Enterotoxins A, C, and D were not detected in any of the cheese samples.

Estudou-se a dinâmica das populações de Staphylococcus spp. durante a maturação do queijo Canastra, em três fazendas localizadas no estado de Minas Gerais. A presença dos genes que codificam para a produção das enzimas coagulase (coa), termonuclease (nuc) e produção de enterotoxinas (sea, seb, sec e sed), em linhagens de Staphylococcus isoladas durante os 60 dias de maturação do queijo foi analisada. Também foi investigada a presença de enterotoxina estafilocócica A, C e D nas amostras de queijo. As amostras de queijo com 0, 7, 15, 30 e 45 dias de maturação apresentaram contagens de Staphylococcus spp. entre 10³ e 10(8)ufc / g. Todos os isolados coagulase positivo nos testes fisiológicos apresentaram o gene coa. Não foi observada associação entre os resultados obtidos com os testes bioquímicos e aqueles obtidos com a PCR usando iniciadores gene-específicos para linhagens coagulase negativa. Os genes da coagulase e termonuclease ocorreram simultaneamente em 41,3 por cento dos Staphylococcus spp. testados. Nenhum dos isolados de Staphylococcus apresentou os genes que codificam para a produção das enterotoxinas SEA, SEB, SEC ou SED. As enterotoxinas A, C ou D não foram detectadas em nenhuma das amostras de queijo analisadas.

Diversity and frequency of yeasts from the dorsum of the tongue and necrotic root canals associated with primary apical periodontitis.

AIMS: To determine the frequency and diversity of yeasts from the dorsum of the tongue and necrotic root canals with teeth associated with primary apical periodontitis. METHODOLOGY: Detailed medical and dental histories of 168 patients were recorded. The samples were collected from the dorsum of tongue and from 168 teeth with root canals contained necrotic pulps. Yeasts were isolated on Sabouraud agar with 100 mg L(-1) chloramphenicol, purified and characterized by standard methods. Identification was confirmed by EI1 PCR fingerprint technique. Yeast isolates of uncertain identity or with a different genetic fingerprint profile from the reference strains were identified by sequencing the D1/D2 variable domains of the large subunit rDNA. RESULTS: Yeasts were isolated from 22.6% of teeth sampled and from 45.8% of tongue samples. Candida albicans was the most frequently isolated species at both investigated sites but other species were also found. Saccharomyces cerevisiae and Kluyveromyces lactis were recovered from the tongue. CONCLUSIONS: Although the detection of yeasts in the root canal does not imply an involvement in the disease process, the study suggests a frequency of Candida spp. in primary endodontic infections that deserves further clarification.

Thermogenesis in birds.

The article discusses the importance of avian skeletal muscle as a source for heat generation by means of both shivering and non-shivering. Non-shivering thermogenesis in birds is still a polemic issue. Recent evidence at the molecular/cellular level indicates, however, that this type of heat generation may also exist among birds. The involvement of the sarcoplasmic reticulum calcium ATPase in non-shivering thermogenesis is discussed in-depth.

Cloning and functional characterization of an uncoupling protein homolog in hummingbirds.

The cDNA of an uncoupling protein (UCP) homolog has been cloned from the swallow-tailed hummingbird, Eupetomena macroura. The hummingbird uncoupling protein (HmUCP) cDNA was amplified from pectoral muscle (flight muscle) using RT-PCR and primers for conserved domains of various known UCP homologs. The rapid amplification of cDNA ends (RACE) method was used to complete the cloning of the 5' and 3' ends of the open reading frame. The HmUCP coding region contains 915 nucleotides, and the deduced protein sequence consists of 304 amino acids, being approximately 72, 70, and 55% identical to human UCP3, UCP2, and UCP1, respectively. The uncoupling activity of this novel protein was characterized in yeast. In this expression system, the 12CA5-tagged HmUCP fusion protein was detected by Western blot in the enriched mitochondrial fraction. Similarly to rat UCP1, HmUCP decreased the mitochondrial membrane potential as measured in whole yeast by uptake of the fluorescent potential-sensitive dye 3',3-dihexyloxacarbocyanine iodide. The HmUCP mRNA is primarily expressed in skeletal muscle, but high levels can also be detected in heart and liver, as assessed by Northern blot analysis. Lowering the room's temperature to 12-14 degrees C triggered the cycle torpor/rewarming, typical of hummingbirds. Both in the pectoral muscle and heart, HmUCP mRNA levels were 1.5- to 3.4-fold higher during torpor. In conclusion, this is the first report of an UCP homolog in birds. The data indicate that HmUCP has the potential to function as an UCP and could play a thermogenic role during rewarming.

Uncoupling proteins 1 and 3 are regulated differently.

Using a heterologous yeast expression system, we have previously found a marked discordance between the effects of uncoupling protein (UCP) 1 and UCP3L on basal O(2) consumption in whole yeast versus isolated mitochondria. In whole yeast, UCP3L produces a greater stimulation of basal O(2) consumption, while in isolated mitochondria, UCP1 produces a much greater effect. As shown previously and in this report, UCP3L, in contrast to UCP1, is not inhibited by purine nucleotides. In the present study, we addressed two hypothetical mechanisms that could account for the observed discordance: (i) in whole yeast, purine nucleotides inhibit UCP1 but not UCP3L and (ii) preparations of isolated mitochondria lack an activator of UCP3L that is normally present in vivo. By use of a mutant of UCP1 that lacks purine nucleotide inhibition, it is demonstrated that cytosolic concentrations of purine nucleotides present in yeast effectively inhibit UCP1 activity. This suggests that the lower activity of UCP1 compared to UCP3L in whole yeast is due to purine nucleotide inhibition of UCP1 but not UCP3L. As potential activators of UCP3L we tested free fatty acids in whole yeast and isolated mitochondria. While UCP1 was strongly activated by free fatty acids, no stimulatory effect on UCP3L was observed. In summary, this study indicates that UCP1 and UCP3L differ in their regulation by purine nucleotides and free fatty acids. This different regulation may be related to different physiological functions of the two proteins.