Erythroid differentiation and protoporphyrin IX down-regulate frataxin expression in Friend cells: characterization of frataxin expression compared to molecules involved in iron metabolism and hemoglobinization

Friedreich ataxia (FA) Is caused by decreased frataxin expression that results in mitochondrial iron (Fe) overload. However, the role of frataxin in mammalian Fe metabolism remains unclear. In this investigation we examined the function of frataxin in Fe metabolism by implementing a well-characterized model of erythroid differentiation, namely, Friend cells induced using dimethyl sulfoxide (DMSO). We have characterized the changes in frataxin expression compared to molecules that play key roles in Fe metabolism (the transferrin receptor [TfR] and the Fe transporter Nramp2) and hemoglobinization (beta-globin). DMSO induction of hemoglobinization results in a marked decrease in frataxin gene (Frda) expression and protein levels. To a lesser extent, Nramp2 messenger RNA (mRNA) levels were also decreased on erythroid differentiation, whereas TfR and beta-globin mRNA levels increased. Intracellular Fe depletion using desferrioxamine or pyridoxal isonicotinoyl hydrazone, which chelate cytoplasmic or cytoplasmic and mitochondrial Fe pools, respectively, have no effect on frataxin expression. Furthermore, cytoplasmic or mitochondrial Fe loading of induced Friend cells with ferric ammonium citrate, or the heme synthesis inhibitor, succinylacetone, respectively, also had no effect on frataxin expression. Although frataxin has been suggested by others to be a mitochondrial ferritin, the lack of effect of intracellular Fe levels on frataxin expression is not consistent with an Fe storage role. Significantly, protoporphyrin IX down-regulates frataxin protein levels, suggesting a regulatory role of frataxin in Fe or heme metabolism. Because decreased frataxin expression leads to mitochondrial Fe loading in FA, our data suggest that reduced frataxin expression during erythroid differentiation results in mitochondrial Fe sequestration for heme biosynthesis. (C) 2002 by The American Society of Hematology.

Detection of West Nile virus antigen in mosquitoes and avian tissues by a monoclonal antibody-based capture enzyme immunoassay

An antigen capture immunoassay to detect West Nile (WN) virus antigen in infected mosquitoes and avian tissues has been developed. With this assay purified WN virus was detected at a concentration of 32 pg/0.1 ml, and antigen in infected suckling mouse brain and laboratory-infected mosquito pools could be detected when the WN virus titer was 10(2.1) to 10(3.7) PFU/0.1 ml. In a blindly coded set of field-collected mosquito pools (n = 100), this assay detected WN virus antigen in 12 of 18 (66.7%) TaqMan-positive pools, whereas traditional reverse transcriptase PCR detected 10 of 18 (55.5%) positive pools. A sample set of 73 organ homogenates from naturally infected American crows was also examined by WN virus antigen capture immunoassay and TaqMan for the presence of WN virus. The antigen capture assay detected antigen in 30 of 34 (88.2%) TaqMan-positive tissues. Based upon a TaqMan-generated standard curve of infectious WN virus, the limit of detection in the antigen capture assay for avian tissue homogenates was approximately 10(3) PFU/0.1 ml. The recommended WN virus antigen capture protocol, which includes a capture assay followed by a confirmatory inhibition assay used to retest presumptive positive samples, could distinguish between the closely related WN and St. Louis encephalitis viruses in virus-infected mosquito pools and avian tissues. Therefore, this immunoassay demonstrates adequate sensitivity and specificity for surveillance of WN virus activity in mosquito vectors and avian hosts, and, in addition, it is easy to perform and relatively inexpensive compared with the TaqMan assay.

Balancing acts: Respiratory sensations, motor control and human posture

1. The present brief review covers some novel aspects of integration between respiration and movement of the body. 2. There are potent viscerosomatic reflexes in animals involving small-diameter pulmonary afferents that, when excited, would limit exercise. However, recent studies using lobeline injections to excite pulmonary afferents in awake humans suggest that there is no evoked reflex motoneuronal inhibition. Instead, the noxious respiratory sensations generated by the vagal afferents may be crucial in the decision to stop exercise. 3. While respiratory movements may affect limb movements, the control of the trunk and limbs can involve interaction (and even interference) with key respiratory muscles, such as the diaphragm. Recent studies have revealed that not only does the diaphragm receive feed-forward drive prior to some limb movements, but that it also contracts both phasically and tonically during repetitive limb movements. 4. Thus, challenges to posture can indirectly challenge ventilation, while coordinated diaphragm contraction may contribute to control of the trunk.

Characterisation of multiple Caribbean ciguatoxins and congeners in individual specimens of horse-eye jack (Caranx latus) by high-performance liquid chromatography/mass spectrometry

We studied the variation in toxin profiles of purified extracts of 10 individual specimens and two pools of ciguateric Caranx latus. High-performance liquid chromatography/mass spectrometry (HPLC/MS) identified in all individual samples at least seven Caribbean ciguatoxins (C-CTXs) comprising C-CTX-1 and its epimer C-CTX-2 ([M + H](+) m/z 1141.58), and five new C-CTX congeners with pseudo-molecular ions at m/z 1141.58, 1143.60, 1157.57, 1159.58, and 1127.57. In some samples, additional C-CTX isomers were detected with [M + H](+) ions at m/z 1141.58 (two), 1143.60 (one) and 1157.57 (two). The two low-toxic pools contained only four to six ciguatoxins. The comparison in relative proportions of four different mass classes ([M + H](+) at m/z 1141, 1143, 1157 and 1127) showed that the group at m/z 1157 increased (2-20%) with flesh toxicity. More than 80% of group m/z 1141 comprised C-CTX-1, C-CTX-2 and their isomer C-CTX-1 a whose level in this group correlated with fish toxicity. Contrary to low-toxic fishes, high-risk specimens had C-CTX-1 levels

Microhabitat use by the rainbowfish Melanotaenia duboulayi in a subtropical Australian stream

Microhabitat use and feeding behavior of the rainbowfish Melanotaenia duboulayi (Castelnau) were investigated in a slow-flowing stream adjacent to riparian forest in south-eastern Queensland, Australia. Fish were more abundant in vegetated areas, but did not enter dense Vallisneria beds, where predators were observed. In sunny conditions shoals of juveniles occurred near the water surface feeding floating material on the surface, but larger fish tended to occur at the bottom near submerged vegetation, often utilizing the overhanging aquatic plant community as a refuge and food source. In the middle of the day, juveniles and small fish seemed to show behavioral thermoregulation at the surface in the warmest site. Under cloudy conditions, however, fish of all sizes preferred deeper water. The present study suggests that in still and sunny pools thermal change caused by sunlight influences the microhabitat choice of small fish. A field experiment using a kingfisher model implies that fish swimming at the surface could escape from aerial predators in sunlit conditions by responding to moving shadows, but could not do so under cloudy conditions.

Polymerase chain reaction tests for the identification of Ross River, Kunjin and Murray Valley encephalitis virus infections in horses

Objective To develop and validate specific, sensitive and rapid diagnostic tests using RT-PCR for the detection of Ross River virus (RRV), Kunjin virus (KV) and Murray Valley encephalitis virus (MVEV) infections in horses. Methods Primer sets based on nucleotide sequence encoding the envelope glycoprotein E2 of RRV and on the nonstructural protein 5 (NS5) of KV and MVEV were designed and used in single round PCRs to test for the respective viruses in infected cell cultures and, in the case of RRV, in samples of horse blood and synovial fluid. Results The primer pairs designed for each of the three viruses amplified a product of expected size from prototype viruses that were grown in cell culture. The identity of each of the products was confirmed by nucleotide sequencing indicating that in the context used the RT-PCRs were specific. RRV was detected in serums from 8 horses for which there were clinical signs consistent with RRV infection such that an acute-phase serum sample was taken and submitted for RRV serology testing. The RRV RT-PCR was analytically sensitive in that it was estimated to detect as little as 50 TCID50 of RRV per mL of serum and was specific in that the primer pairs did not amplify other products from the 8 serum samples. The RRV primers also detected virus in three independent mosquito pools known to contain RRV by virus isolation in cell culture. Samples from horses suspected to be infected with KV and MVEV were not available. Conclusion Despite much anecdotal and serological evidence for infection of horses with RRV actual infection and associated clinical disease are infrequently confirmed. The availability of a specific and analytically sensitive RT-PCR for the detection of RRV provides additional opportunities to confirm the presence of this virus in clinical samples. The RTPCR primers for the diagnosis of KV and MVEV infections were shown to be specific for cell culture grown viruses but the further validation of these tests requires the availability of appropriate clinical samples from infected horses.

Field evaluation of a sentinel mosquito (Diptera : culicidae) trap system to detect Japanese encephalitis in remote Australia

Incursions of Japanese encephalitis (JE) virus into northern Queensland are currently monitored using sentinel pigs. However, the maintenance of these pigs is expensive, and because pigs are the major amplifying hosts of the virus, they may contribute to JE transmission. Therefore, we evaluated a mosquito-based detection system to potentially replace the sentinel pigs. Single, inactivated JE-infected Culex annulirostris Skuse and C. sitiens Wiedemann were placed into pools of uninfected mosquitoes that were housed in a Mosquito Magnet Pro (MM) trap set under wet season field conditions in Cairns, Queensland for 0, 7, or 14 d. JE viral RNA was detected (cycling threshold [CT] = 40) in 11/ 12, 10/14, and 2/5 pools containing 200, 1,000, and 5,000 mosquitoes, respectively, using a TaqMan real-time reverse transcription-polymerase chain reaction (RT-PCR). The ability to detect virus was not affected by the length of time pools were maintained under field conditions, although the CT score tended to increase with field exposure time. Furthermore, JE viral RNA was detected in three pools of 1,000 mosquitoes collected from Badu Island using a MM trap. These results indicated that a mosquito trap system employing self-powered traps, such as the MosquitoMagnet, and a real-time PCR system, could be used to monitor for JE in remote areas.

Nitrogen cycling in degraded Leucaena leucocephala-Brachiaria decumbens pastures on an acid infertile soil in south-east Queensland, Australia

A grazing trial was conducted to quantify N cycling in degraded Leucaena leucocephala (leucaena)-Brachiaria decumbens (signal grass) pastures grown on an acid, infertile, podzolic soil in south-east Queensland. Nitrogen accumulation and cycling in leucaena-signal grass pastures were evaluated for 9 weeks until all of the leucaena on offer (mean 600 kg edible dry matter (EDM)/ha, 28% of total pasture EDM) was consumed. Nitrogen pools in the grass, leucaena, soil, cattle liveweight, faeces and urine were estimated. The podzolic soil (pH 4.8-5.9) was found to be deficient in P, Ca and K. Leucaena leaf tissues contained deficient levels of N, P and Ca. Grass tissues were deficient in N and P. Grazing was found to cycle 65% of N on offer in pasture herbage. However, due to the effect of the plant nutrient imbalances described above, biological N fixation by leucaena contributed only 15 kg/ha N to the pasture system over the 9-month regrowth period, of which 13 kg/ha N was cycled. Cattle retained 1.8 kg/ha N (8% of total N consumed) in body tissue and the remainder was excreted in dung and urine in approximately equal proportions. Mineral soil N concentrations did not change significantly (-3.5 kg/ha N) over the trial period. The ramifications of grazing and fertiliser management strategies, and implications for pasture rundown and sustainability are discussed.

Seringas hipodérmicas descartáveis versus reutilizáveis: estudo de possíveis efeitos sobre o vírus da vacina viva, atenuada contra o sarampo

Objetivou-se verificar entre seringas hipodérmicas descartáveis e reutilizáveis qual interfere mais com o vírus vivo presente na vacina contra o sarampo. Vacinas pertencentes a dois lotes foram reconstituídas com os dois tipos de seringas, de modo a formarem dois pools distintos, mantidos à temperatura de +2 a+8°C e protegidos da luz. De cada lote foram realizadas, no mínimo, seis titulações em paralelo, com amostragem a cada hora, de zero a seis horas após reconstituição. A análise estatística dos resultados obtidos nas titulações, feita pelo sistema de retas de regressão demonstrou que embora as vacinas manipuladas com ambos os tipos de seringas apresentassem um decréscimo de título estatisticamente significativo com o decorrer das horas, ele foi bem mais acentuado para as vacinas reconstituídas com seringas reutilizáveis. A menor interferência das descartáveis no título da vacina viva, atenuada contra o sarampo, demonstrou que a preconização e uso desse tipo de seringas pela Secretaria de Estado da Saúde de São Paulo é o ideal e recomendável, por preservar mais a vacina desde a reconstituição até sua administração e, conseqüentemente, a sua eficácia na prevenção dessa infecção.

Populations of larvae of Anopheles spp. in natural breeding sites in Western Venezuela, an area of refractory malaria

Studies have been un dertaken into on the diversity and relative abundance of larvae of Anopheles (Nyssorhynchus) spp. in 22 permanent or temporary pools in an area of 70 km² in the eastern piedmont of the Venezuela Andes, between the mountains and the plains, an area in which malaria is refractory and A. nuñeztovari is present. Twelve species were identified, the most frequent, abundant and sympatric being A. triannulatus, A. albitarsis, A. nuñeztovari, A. oswaldoi and A. strodei. The samples from the permanent pools showed greater diversity of species and greater numbers of larvae than the samples from the temporary pools. The existence of the same larval associations in pools of other localities in the eastern piedmont of the Venezuelan Andes suggests the possibility of the making an ecological map of the breeding sites of A. nuñeztovari and for these anophelines in a region extending for 430 km.

Immune cell changes in response to a swimming training session during a 24-h recovery period

Understanding the impact of training sessions on the immune response is crucial for the adequate periodization of training, to prevent both a negative influence on health and a performance impairment of the athlete. This study evaluated acute systemic immune cell changes in response to an actual swimming session, during a 24-h recovery period, controlling for sex, menstrual cycle phases, maturity, and age group. Competitive swimmers (30 females, 15 ± 1.3 years old; and 35 males, 16.5 ± 2.1 years old) performed a high-intensity training session. Blood samples were collected before, immediately after, 2 h after, and 24 h after exercise. Standard procedures for the assessment of leukogram by automated counting (Coulter LH 750, Beckman) and lymphocytes subsets by flow cytometry (FACS Calibur BD, Biosciences) were used. Subjects were grouped according to competitive age groups and pubertal Tanner stages. Menstrual cycle phase was monitored. The training session induced neutrophilia, lymphopenia, and a low eosinophil count, lasting for at least 2 h, independent of sex and maturity. At 24 h postexercise, the acquired immunity of juniors (15-17 years old), expressed by total lymphocytes and total T lymphocytes (CD3+), was not fully recovered. This should be accounted for when planning a weekly training program. The observed lymphopenia suggests a lower immune surveillance at the end of the session that may depress the immunity of athletes, highlighting the need for extra care when athletes are exposed to aggressive environmental agents such as swimming pools

Estudo de caso. A implementação do projeto de internalização do RX do Centro de Diagnóstico Pneumológico do Porto

Portugal atravessa uma crise económico-financeira. A austeridade económica é prioritária, e a área de saúde não é exceção. Nesta conjuntura justifica-se de todo a realização de análises económicas na saúde. Este estudo é uma análise custo - mínimo ao Projeto de Internalização do RX do Centro Diagnóstico Pneumológico (CDP) do Porto com o objetivo de rentabilizar o equipamento radiológico instalado no CDP do Porto, pretende-se usar a capacidade máxima instalada e permitida pelo equipamento para produção de radiografias a utentes provenientes dos Agrupamentos de Centros de Saúde (ACES) do Porto. O Projeto de Internalização do RX do CDP do Porto é um projeto da Administração Regional de Saúde (ARS) do Norte, que pretende encaminhar os utentes dos ACES do Porto para realização de exames radiográficos no RX do CDP do Porto. Para proceder à análise económica foram identificados os custos e os benefícios relevantes do RX do CDP do Porto e elaboradas 3 estimativas: 1. Realização dos exames radiográficos do CDP do Porto a utentes do CDP do Porto e realização de exames radiográficos a utentes dos ACES do Porto nos gabinetes de radiologia convencionados pela ARSNorte; 2. Projeto de Internalização do RX do CDP do Porto com realização dos exames radiográficos até esgotar a capacidade máxima instalada do equipamento radiológico com recurso a utentes dos ACES do Porto; 3. Realização dos exames radiográficos nos ACES do Porto e dos exames do CDP do Porto nos gabinetes de radiologia convencionados. Os resultados da análise levaram à conclusão que a implementação do Projeto de Internalização do RX do Porto é rentável, ou seja, o projeto permitirá ganhos financeiros para a ARSNorte se for implementado na sua plenitude.

Avaliação do ambiente térmico em piscinas cobertas

Este estudo teve como objectivo avaliar as condições do ambiente térmico em piscinas cobertas através de duas abordagens distintas. A abordagem objectiva consistiu na análise da temperatura da água e dos parâmetros de ambiente térmico. A abordagem subjectiva teve por base a aplicação de um questionário. Os resultados dos parâmetros físicos apresentaram-se elevados na maioria das situações avaliadas, nomeadamente no que respeita à temperatura da água e à humidade relativa. A análise subjectiva evidenciou percentagens de insatisfeitos inferiores à obtida na análise objectiva. Verificou-se ainda que os utilizadores das piscinas preferem ambientes considerados ligeiramente quentes ou quentes.

Primeiro registro de Aedes (Stegomyia) albopictus no Estado do Ceará, Brasil

Pela primeira vez é registrada a ocorrência de Aedes (Stegomyia) albopictus em área urbana da cidade de Fortaleza, no Estado do Ceará, Brasil. De janeiro a julho de 2005 foram utilizadas ovitrampas para a coleta de ovos de Aedes spp., os quais foram mantidos em laboratório para desenvolvimento até a fase adulta. Os mosquitos resultantes foram identificados e submetidos a testes para o isolamento dos vírus da dengue. Foram identificados 13 espécimes de Aedes albopictus, todos fêmeas. Não foi isolado vírus da dengue em nenhum dos pools de mosquitos. Apesar de o Aedes albopictus não ter sido incriminado por surtos de dengue no Brasil, não se pode descartar a possibilidade da transmissão dos vírus da dengue por tais mosquitos.

Estudo da transferência das cloraminas da água para o ar duma piscina coberta

Mestrado em Engenharia Química