Fungal contaminated buildings and related adverse human health implications have long been a topical issue throughout the world and concern is mounting with regards to the presence of more toxigenic fungi found in buildings and the associated health risks. These risks are compounded when homes are affected by water damage as a result of water intrusion problems, in particular flooding. With the ever changing climate and unpredictable weather conditions the frequency of flash flooding has increased in recent years and is set to increase and subsequently more homes will inevitably be effected by mould contamination. The present study initially aimed to determine the types of fungi commonly detected in buildings in the United Kingdom with varied levels of water intrusion problems via a small survey using various sampling techniques and particularly aimed to determine the conditions by which growth of the toxigenic fungi Stachybotrys chartarum could occur. Penicillium, Cladosporium and Aspergillus species were the most commonly detected fungi in buildings with relatively moderate levels of water intrusion problems; Stacybotrys chartarum was only detected in building with more severe water intrusion problems. Cont/d.

Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2014-05-20T17:14:19Z No. of bitstreams: 1 Adenizar Delgado das Chagas Jr. Avanços... 2014..pdf: 11153892 bytes, checksum: 48154dea65a7a4a32a54408e694c5d30 (MD5) / Made available in DSpace on 2014-05-20T17:14:20Z (GMT). No. of bitstreams: 1 Adenizar Delgado das Chagas Jr. Avanços... 2014..pdf: 11153892 bytes, checksum: 48154dea65a7a4a32a54408e694c5d30 (MD5) Previous issue date: 2014 / Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, Brasil / A leptospirose é uma zoonose causada por espiroquetas patogênicas pertencentes ao gênero Leptospira. O modelo da doença em camundongos tem vantagens devido à ampla gama de ferramentas genéticas e imunológicas disponíveis para pesquisas básicas. A maior limitação na conduta clínica e na pesquisa experimental da leptospirose é o fraco desempenho dos métodos disponíveis para detecção direta e para quantificação de leptospiras. Foi incluído nesta tese um conjunto de três manuscritos que visam investigar o desfecho da infecção pela cepa virulenta de Leptospira interrogans nas linhagens de camundongos selvagens (A, CBA, BALB/c e C57BL/6), em camundongos óxido nítrico sintase induzível (iNOS) Knockout (KO), camundongos gene ativador de recombinação 1 (RAG1) KO , camundongos CB17 com imunodeficiência combinada grave (SCID), e os seus respectivos controles selvagens C57BL/6 e BALB/c. Investigar a confiabilidade do método de quantificação do imprint (IM), comparando os resultados obtidos com esta técnica aos obtidos com a utilização do PCR em tempo real (qPCR) para detectar e quantificar leptospiras em amostras de rim de ratos e hamsters experimentalmente infectados. Como esperado, nenhuma das linhagens de camundongos selvagens foram suscetíveis à leptospirose letal. A linhagem A e C57BL/6 exibiram altas cargas de Leptospira nas amostras de rim e as linhagens CBA e C57BL/6 desenvolveram lesões inflamatórias graves, enquanto a linhagem BALB/c provou ser a mais resistente apresentando leptospirose subclínica. Os camundongos iNOS KO e selvagem sobreviveram sem sintomas clínicos de leptospirose. A frequência e gravidade das nefrites foram significantemente menores nos camundongos iNOS KO. Todos os animais RAG1 KO e SCID morreram de leptospirose aguda, enquanto que todos os camundongos selvagens sobreviveram. A hemorragia pulmonar foi observada em 57 e 94% dos camundongos RAG 1 KO e em 83 e 100% dos camundongos SCID, usando doses de inóculos de 107 e 106 leptospiras, respectivamente. Não houve evidências de hemorragia pulmonar nos controles selvagens. Nos modelos de infecção agudo e crônico, houve correlação positiva estatisticamente significante (P < 0,05) na quantificação de leptospiras pelos métodos do qPCR e do IM. Como conclusão geral, a linhagem de camundongos A pode ser a linhagem de escolha em estudos na qual se pretende recuperar um grande número de leptospiras de rins colonizados. As linhagens CBA e C57BL/6 desenvolveram, com maior frequência, lesões inflamatórias e podem ser as mais adequadas para estudos de leptospirose associados com nefrite intersticial. A linhagem BALB/c é a mais indicada para estudar mecanismos que envolvam a imunidade inata e/ou a rápida resposta imune adaptativa. A ausência do gene do iNOS no modelo murino resultou em uma diminuição significativa da suscetibilidade para o desenvolvimento da nefrite intersticial. Além disso, a ausência de linfócitos B e T funcionais não impediu a ocorrência de hemorragia pulmonar. Estes dados fornecem fortes evidências de que a hemorragia pulmonar na leptospirose não está relacionada apenas a mecanismos autoimunes. Para a detecção e quantificação de leptospiras o método do imprint foi equivalente ao qPCR. / Leptospirosis is a zoonosis caused by pathogenic spirochaetes belonging to the genus Leptospira. The mouse disease model is advantagous due to the broad array of immunological and genetic tools available for basic research. A major limitation in the clinical management and experimental research of leptospirosis is the poor performance of the available methods in the direct detection and quantification of leptospires. This thesis includes three manuscripts that investigate the outcome of infection by a virulent strain of Leptospira interrogans in wildtype mice strains: A, CBA, BALB/c and C57BL/6; in iNOS knockout (KO) mice, recombination activating gene 1 (RAG1) KO mice and CB17 severe combined immunodeficiency (SCID) mice. To investigate whether the imprint method (IM) of quantification was reliable we compared it with against real time PCR (qPCR) for the detection and quantification of leptospires in kidney samples from rats and hamsters. As expected, none of the wildtype mice were susceptible to lethal leptospirosis. The A and C57BL/6 strains exhibited high leptospiral loads in the kidney samples and the CBA and C57BL/6 strains developed severe inflammatory lesions, whilst the BALB/c strain proved to be the most resistant to subclinical leptospirosis. The iNOS KO mice survived with no clinical symptoms of leptospirosis. The frequency and severity of nephritis was significantly lower in the iNOS KO mice. All of the RAG 1 KO and SCID animals died of acute leptospirosis, whereas all of the wildtype mice survived. Pulmonary haemorrhage was observed in 57 and 94% of Rag1 KO mice and in 83 and 100% of SCID mice, using inoculum doses of 107 and 106 leptospires, respectively. There was no evidence of pulmonary haemorrhage in the wildtype controls. In both the acute and chronic infection models, the correlation between quantification by qPCR and the IM was found to be positive and statistically significant (P < 0.05). In conclusion, the mouse A strain would be the strain of choice in studies requiring the recovery of a large number of leptospires from colonized kidneys. The CBA and C57BL/6 strains developed more inflammatory lesions and would be the most suitable for studies of leptospirosis associated with interstitial nephritis. The BALB/c strain appeared to be the most suitable for studying mechanisms involving innate immunity and/or rapid adaptive immune response. The absence of the iNOS gene in the murine model resulted in a significant decrease in susceptibility to the development of interstitial nephritis. Furthermore, the absence of functional B and T lymphocytes did not prevent the occurrence of pulmonary hemorrhage. These data provide strong evidence that pulmonary hemorrhage in leptospirosis is not only related to autoimmune mechanisms. For the detection and quantification of leptospires the imprint method was quivalent to that of qPCR. Keywords:

This study was conducted to investigate two important causes of reduced performance in sport horses. In the first study, clinical records of 201 Criollo horses, 5.59±2.11 years of age, that underwent physical examination due to lameness or reduced performance in order to document the frequency of lameness problems in these horses. A total of 223 problems were diagnosed. In the forelimbs (47.08%; n=105) and in the hindlimbs (52.9%; n=118). In the forelimbs the pain was located above the fetlock joint in 17.14% (18/105), 14.30% (15/105) in the fetlock and 68.60% (72/105) distal to the fetlock. In the hindlimbs the source of lameness was frequently located in the hock (78.80%; 93/118). Lameness above the tarsus occurred in 17.80% (21/118) and below the tarsus in 3.4% (4/118) of the cases. This study showed that Criollo horses present similar lameness problems as seen in other breeds competing in rodeo disciplines. In the second study, 38 Criollo horses, 6.63±1.53 years of age, underwent endoscopic examination of the upper and lower airways, during competitions, aiming to document the frequency of respiratory problems and evaluate its impact on the Criollo horse s performance by their competition scores. Lymphoid hyperplasia was found in 57.89% (22/38) and left laryngeal hemiplegia (grade II) in 7.89% (3/38). Exercise induced pulmonary haemorrhage was diagnosed in 21.05% (8/38). Lower respiratory inflammation indicated by the presence of mucous or purulent secretion in the trachea was seen in 78.94% (30/38). There was no difference in the performance scores between horses diagnosed with EIPH and the ones considered normal (p>0.05). However, horses with mucous/purulent secretion in the trachea showed lower performance scores compared to the EIPH group (p=0.01). The high frequency of Criollo horses in this study showing inflammation in the lower respiratory tract indicates that endoscopic examination would help to identify horses with problems before the competitions. / No primeiro estudo, foram avaliados os registros clínicos de 201cavalos Crioulos em treinamento com idade média de 5,59±2,11 anos, com o objetivo de identificar as principais causas de claudicação entre os anos de 2002 e 2009. Nestes cavalos, foram diagnosticadas 223 alterações no aparelho locomotor sendo que a claudicação foi localizada nos membros anteriores e posteriores em 47,08% (n=105) e 52,90% (n=118) respectivamente. Nos membros anteriores, 17,14% (18/105) das alterações foram diagnosticadas acima do boleto, 14,30% (15/105) no boleto e 68,60% (72/105) estavam localizadas distais ao boleto. No membro posterior 78,80% (93/118) apresentaram a origem da dor na região do tarso, 17,80% (21/118) acima do tarso e 3,40% (4/118) abaixo do tarso. Muitos dos problemas relacionados à parte distal dos membros anteriores provavelmente sejam em decorrência de descuidos com o ferrageamento e treinamento sobre superfícies duras. As articulações interfalangeanas distais são importantes fontes de dor e inflamação e mais frequentemente envolvidas em claudicações de membro anterior. Claudicações em membro posterior foram mais frequentes do que em membro anterior sendo que o tarso foi a região que mais apresentou problemas. Os cavalos Crioulos apresentam problemas semelhantes aos citados em outras raças que participam de provas de rodeio. No segundo estudo, com o objetivo de determinar a frequência de patologias respiratórias e avaliar seu possível impacto no desempenho atlético de cavalos da raça Crioula, 38 cavalos com idade média de 6,63±1,53 anos foram submetidos ao exame endoscópico das vias aéreas superiores e inferiores em etapas classificatórias ao Freio de Ouro. As notas atribuídas aos cavalos nas provas foram utilizadas como referência na avaliação do desempenho. Hiperplasia linfóide foi observada em 57,89% (22/38) e hemiplegia laríngea esquerda grau II em 7,89% (3/38). Hemorragia pulmonar induzida pelo exercício (HPIE) foi observada em 21,05% (8/38). A presença de secreção em diversos graus na traquéia, indicativo de inflamação, foi observada em 78,94% (30/38). Não se observou diferença significativa (p>0,05) no desempenho de animais que sofreram HPIE em relação aos animais sadios (sem HPIE e sem muco/catarro). Entretanto, animais com muco/catarro na traquéia apresentaram desempenho inferior (p=0,0132) aos animais que sofreram HPIE. A frequência de cavalos com presença de muco/catarro na traquéia sugere que os cavalos Crioulos podem se beneficiar do uso da endoscopia antes das competições para o diagnóstico de sinais de doença inflamatória das vias aéreas.

Much of the impetus for this research can be attributed to Kenneth W. Hinchcliff, who has studied exercise-induced pulmonary haemorrhage (EIPH) extensively. This thesis focused on EIPH in Thoroughbred racehorses competing in South Africa. Using tracheobronchoscopy, the prevalence and severity of EIPH and the association with racing performance was determined. Thereafter, the prevalence of other respiratory tract disorders and their association with racing performance is reported. This is followed by a study assessing interobserver variability using grading systems in the detection of respiratory tract disorders. Finally, there is a report on the immunopathogenesis of EIPH. Using tracheobronchoscopy after racing, the prevalence and severity of EIPH was assessed in 1,005 racehorses competing at high altitude (> 1,400 meters above sea level) and at sea level in a racing jurisdiction that does not allow the use of furosemide and nasal dilator strips. The prevalence and severity of EIPH was affected by altitude as racing at sea level was associated with a higher prevalence and greater severity of EIPH. Results also suggested that EIPH was associated with superior performance in South African Thoroughbred racehorses. Upper and lower respiratory tract disorders identified following tracheobronchoscopic examination included left arytenoid asymmetry, left laryngeal hemiplegia, epiglottic deformity, epiglottic entrapment, subepiglottic cysts, dorsal displacement of the soft palate, pharyngeal lymphoid hyperplasia (PLH), laryngeal and tracheal dirt, tracheal mucous (TM), tracheal stenosis and tracheal cartilage ring spikes in Thoroughbred racehorses after racing. Overall, there was a low prevalence of grade 2 and 3 arytenoid cartilage asymmetry, left laryngeal hemiplegia, epiglottic entrapment, subepiglottic cysts and epiglottic deformity, while more severe grades of PLH, laryngeal debris, tracheal debris, TM and tracheal cartilage ring spikes had a higher prevalence. An association with sex was identified as tracheal cartilage ring spikes occurred more often in male racehorses. Superior racing performance was identified in racehorses with grade 3 tracheal mucous and tracheal cartilage ring spikes. Endoscopic grading of EIPH, PLH, arytenoid cartilage movement (ACM), and TM was performed by 3 observers that were blinded to each racehorse’s identity and race day performance using previously established grading criteria. Excellent interobserver reliability was seen using the EIPH grading system, while the weighted kappa for PLH, ACM and TM was lower. The study demonstrated sufficient reliability for the use of the EIPH, PLH, ACM and TM grading systems in racehorses competing in South Africa. The study concluded that tracheobronchoscopy seemed to be a practical screening technique that may have prognosticative validity and clinical dependability and that would allow safe and quick assessment of the respiratory tract of a large number of racehorses in field conditions. Venous blood was collected from 10 horses in each EIPH grade classification (grade 0 to 4) following tracheobronchoscopic examinations for the determination of the presence and severity of EIPH. Following RNA isolation and cDNA synthesis, real-time PCR was used to detect equine cytokine-specific mRNA for interleukin (IL) -1, -6, -10, interferon (INF) -ã, and tumor necrosis factor (TNF) -á. Results of this study indicated that increased IL-6, and -10 mRNA production was associated with more severe forms of EIPH. Also, there was greater expression of IL-6 mRNA at sea level and TNF-á mRNA at high altitude. This study concluded that although it was unclear whether the inflammatory response observed in the study was due to pre-existing pulmonary inflammation or as a direct consequence of pulmonary bleeding, the study demonstrated a systemic correlation to pulmonary inflammation. The research reported in this thesis has contributed substantially to the determination of the prevalence, severity and affect on racing performance of respiratory tract disorders in Thoroughbred racehorses competing in South Africa. Also, determination of an association between EIPH and inflammation at a molecular level may assist future researchers in anti-cytokine therapies which may help reduce the prevalence and severity of EIPH. / Thesis (PHD)--University of Pretoria, 2007. / Companion Animal Clinical Studies / unrestricted