Thesis. (M. Tech. (Dept. of Biosciences, Faculty of Applied and Computer Sciences))--Vaal University of Technology, 2010 / Methods for detecting the presence of Cryptosporidium oocysts and Giardia cysts have been developed and continuous improvement is being done to improve the recovery rate of the target protozoa. Rand Water has adopted their method for isolation and detection of Cryptosporidium oocysts and Giardia cysts in water from United State Environmental Protection Agency (US-EPA) Method 1623, 1999. In 2005 changes were made by US-EPA to the Method 1623. A study was done to improve the performance of the Rand Water Method 06 (2007) used for isolation and detection of Cryptosporidium oocysts and Giardia cysts. Three methods namely: Rand Water Method 06 (2007), US-EPA Method 1623 (2005) and Drinking Water Inspectorate standard operating procedures (2003) were compared and key different steps in the methods were identified (wrist action speed, centrifuge speed, immunomagnetic separation procedures and addition of pre-treatment steps). Different experiments were conducted to verify and evaluate the difference between two wrist action shaker speeds, three different centrifuge speeds, two slightly different immunomagnetic separation procedures and when a pre-treatment step was included in the method. Three different types of water matrices (reagent grade water, drinking water and raw water) were used for the experiments and secondary validation. Data obtained from the experiments and secondary validation was statistically analyzed to determine whether there was a significant difference in the recovery of Cryptosporidium oocysts and Giardia cysts. Secondary validation of the Rand Water Method 06 (2007) was performed by implementing the study experiments‟ findings into the method. The results indicated an increase in the recovery rate of Cryptosporidium oocysts and Giardia cysts when data was compared with the previous secondary validation report. The mean recovery of Cryptosporidium oocysts in reagent grade water samples increased from 31% to 55%, drinking water samples increased from 28% to 44% and raw water decreased from 42% to 29%. The mean recovery of Giardia cysts in reagent grade water samples increased from 31% to 41%, drinking water samples increased from 28% to 46% and raw water decreased from 42% to 32%. Furthermore, even though the recovery rate of raw water decreased the use of pre-treatment buffer reduced the number of IMS performed per sample by reducing the pellet size. Enumeration of microscope slides was also easier as there was less background interference. The optimization of the Rand Water Method 06 (2007) was successful as the recovery rate of Cryptosporidium oocysts and Giardia cysts from water increased. All the changes that were verified and that increased the recovery rate were incorporated into the improved Rand Water Method 06.

Foram estudados prospectivamente 33 pacientes cujos cistos tinham volume médio de 298,4 + 346,9 (30 - 1700) ml. A punção foi realizada sob anestesia local com agulha 18 gauge, de 20 cm. Após seu esvaziamento, injetou-se etanol absoluto no volume equivalente a um terço do volume aspirado, até o máximo de 100 ml. Os volume médios observados após um, três e seis meses foram 47,9 + 59,4 (0 - 286) ml, 25,2 + 42,8 (0 - 208) ml e 12,7 + 30 (0 - 120) ml, respectivamente. Após esse período, 30 (91%) apresentaram remissão total dos sintomas, 2 (6%) mantiveram os sintomas, sendo um com cisto residual e outro sem cisto e um (3%) apresentou melhora parcial dos sintomas / A prospective series of 33 patients with symptomatic simple renal cysts varying from 30 to 1700 ml, were considered eligible for the study. The cysts were punctured under local anesthesia with an 18 gauge, 20 centimeters needle. A volume of ethanol equivalent to one third of the aspirated volume, up to a maximum of 100 ml, was injected into the cyst and left there. After one, three and six months, the average volume of the cyst was 47.9 + 59.4 (0 - 286)ml, 25.2 + 42.8 (0 - 208)ml and 12.7 + 30 (0 - 120)ml, respectively. After this period, 30 patients (91%) showed total remission of the symptoms, two patients (6%) maintained them and one patient (3%) had partial remission of the symptoms

No description available.

No description available.

Foram estudados prospectivamente 33 pacientes cujos cistos tinham volume médio de 298,4 + 346,9 (30 - 1700) ml. A punção foi realizada sob anestesia local com agulha 18 gauge, de 20 cm. Após seu esvaziamento, injetou-se etanol absoluto no volume equivalente a um terço do volume aspirado, até o máximo de 100 ml. Os volume médios observados após um, três e seis meses foram 47,9 + 59,4 (0 - 286) ml, 25,2 + 42,8 (0 - 208) ml e 12,7 + 30 (0 - 120) ml, respectivamente. Após esse período, 30 (91%) apresentaram remissão total dos sintomas, 2 (6%) mantiveram os sintomas, sendo um com cisto residual e outro sem cisto e um (3%) apresentou melhora parcial dos sintomas / A prospective series of 33 patients with symptomatic simple renal cysts varying from 30 to 1700 ml, were considered eligible for the study. The cysts were punctured under local anesthesia with an 18 gauge, 20 centimeters needle. A volume of ethanol equivalent to one third of the aspirated volume, up to a maximum of 100 ml, was injected into the cyst and left there. After one, three and six months, the average volume of the cyst was 47.9 + 59.4 (0 - 286)ml, 25.2 + 42.8 (0 - 208)ml and 12.7 + 30 (0 - 120)ml, respectively. After this period, 30 patients (91%) showed total remission of the symptoms, two patients (6%) maintained them and one patient (3%) had partial remission of the symptoms

A hiperplasia adrenal congênita (HAC) por deficiência no P450c17 é uma doença de herança autossômica recessiva raramente relatada em pacientes 46, XX. Nosso objetivo foi o de caracterizar o fenótipo e genótipo desta doença rara revendo os dados clínicos, laboratoriais, genéticos, e da função ovariana de pacientes 46,XX de uma coorte brasileira avaliada no HCFMUSP e dos casos já publicados na literatura. Foram avaliados retrospectivamente os dados de 18 pacientes brasileiras pertencentes a 12 famílias avaliadas no HCFMUSP, e revisados os dados de literatura de pacientes de 10 coortes com deficiência da atividade P450c17 (14 pacientes, 6 delas com defeitos no CYP17A1 e 8 com defeitos no POR). Fenótipo: A maioria das pacientes apresentou-se com amenorreia primária (74 %) e 90 % das pacientes não desenvolveram pubarca; 69,5 % das pacientes apresentavam hipertensão arterial no momento do diagnóstico. Observamos uma alta incidência de distúrbios psiquiátricos (76%), como depressão, ansiedade e síndrome do pânico, em nossa coorte, mas não foram encontrados relatos na literatura. O ultrassom mostrou um aumento de pelo menos um dos ovários em 87 % dos pacientes antes do tratamento e macrocistos ovarianos em 65%, 6 pacientes (26%) referiram terem sido submetidas a cirurgia anterior para tratamento de torção ou ruptura de ovário. O tratamento com dexametasona, estrogênio e progesterona resultou em redução efetiva do volume ovariano. Todos os pacientes apresentaram níveis basais elevados de progesterona e LH e redução dos níveis de androgênios. Não observamos correlação entre os níveis de LH, da relação LH/FSH e de progesterona com o volume ovariano nos dois defeitos. Genótipo: O estudo molecular revelou que 17 pacientes de nossa coorte apresentavam mutações inativadoras no gene CYP17A1 e 1 paciente no gene POR. Duas novas mutações foram identificadas no gene CYP17A1, a p.R362H no éxon 6 e a p.G478S no éxon 8. A mutação mais prevalente no CYP17A1 foi a p.W406R identificada em 41 % das nossas famílias. Algumas das mutações no CYP17A1 foram encontradas apenas na coorte brasileira, mas a mutação p.A287P encontrada no gene POR em uma das nossas pacientes é a mais prevalente na literatura. Em relação à etnia, houve um predomínio dos defeitos moleculares no CYP17A1 em pacientes chinesas e brasileiras, enquanto que os defeitos no gene POR foram mais relatados nas pacientes europeias e norte - americanas. Em conclusão, a análise retrospectiva do fenótipo, genótipo e morfologia ovariana de trinta e duas pacientes 46,XX portadoras de deficiência da atividade P450c17, nos permitiu destacar os seguintes aspectos: a importância da dosagem da progesterona basal para este diagnóstico, a alta prevalência de aumento dos ovários com a presença de macrocistos ovarianos com risco de torção, e de transtornos psiquiátricos identificados em nossa coorte. Acreditamos que esta revisão possa contribuir para o diagnóstico mais precoce desta rara doença congênita / Congenital adrenal hyperplasia due to P450c17 deficiency is rarely reported on 46,XX patients. Our aim was to characterize the phenotype and genotype of this rare disorder reviewing the clinical, laboratory, genetic and ovarian imaging of 46,XX patients of our cohort and of other already reported cases. We retrospectively reviewed 32 patients with deficiency of P450c17 activity: 18 Brazilian patients belonging to 12 families and 10 cohorts already published (fourteen 46,XX patients, 6 of them with CYP17A1 defects and 8 with POR defects). Phenotype: most patients had primary amenorrhea (74%) and 90% of the patients did not develop pubarche; 69.5% of the patients had blood hypertension at diagnosis. We observed a high incidence of psychiatric disorders such as depression, anxiety and panic (76%) in our cohort but no reports were found in the literature. Ultrasound showed an increase of at least one of the ovaries in 87% of the patients before treatment and ovarian macrocysts in 65% of them; 6 patients (26%) had had previous surgery for twisting or ovarian rupture. Treatment with dexamethasone, estrogen and progesterone resulted in ovarian volume reduction. All patients showed elevated basal progesterone and LH and levels, and decreased androgen levels. There was no correlation between the levels of LH and progesterone and of LH/FSH ratio and the ovarian volume in both defects. Genotype: the molecular study showed that 17 patients from our cohort had inactivating mutation in the CYP17A1 gene and 1 in POR gene. Two novel mutations were identified in the CYP17A1 gene, the p.R362H in exon 6 and p.G478S in exon 8. The most prevalent mutation in the CYP17A1 was the p.W406R, identified in 41% of our families. Some of the CYP17A1 mutations were found only in the Brazilian cohort, but the mutation p.A287P found in the POR gene is the most prevalent in the literature. Regarding the ethnicity of the defects, there was a predominance of Chinese and Brazilian patients with defects in the CYP17A1 whereas defects in POR were most reported in European and North-American subjects.Conclusion: In this data review of thirty-two 46,XX patients with dysfunction of P450c17 activity we characterized the phenotype and genotype of this rare disorder and emphasize: the importance of basal progesterone measurement for this diagnosis, the high prevalence of ovarian macrocysts with risk of twisting, and the psychiatric disorders. We believe that this review may contribute to the early diagnosis of this disorder

A hiperplasia adrenal congênita (HAC) por deficiência no P450c17 é uma doença de herança autossômica recessiva raramente relatada em pacientes 46, XX. Nosso objetivo foi o de caracterizar o fenótipo e genótipo desta doença rara revendo os dados clínicos, laboratoriais, genéticos, e da função ovariana de pacientes 46,XX de uma coorte brasileira avaliada no HCFMUSP e dos casos já publicados na literatura. Foram avaliados retrospectivamente os dados de 18 pacientes brasileiras pertencentes a 12 famílias avaliadas no HCFMUSP, e revisados os dados de literatura de pacientes de 10 coortes com deficiência da atividade P450c17 (14 pacientes, 6 delas com defeitos no CYP17A1 e 8 com defeitos no POR). Fenótipo: A maioria das pacientes apresentou-se com amenorreia primária (74 %) e 90 % das pacientes não desenvolveram pubarca; 69,5 % das pacientes apresentavam hipertensão arterial no momento do diagnóstico. Observamos uma alta incidência de distúrbios psiquiátricos (76%), como depressão, ansiedade e síndrome do pânico, em nossa coorte, mas não foram encontrados relatos na literatura. O ultrassom mostrou um aumento de pelo menos um dos ovários em 87 % dos pacientes antes do tratamento e macrocistos ovarianos em 65%, 6 pacientes (26%) referiram terem sido submetidas a cirurgia anterior para tratamento de torção ou ruptura de ovário. O tratamento com dexametasona, estrogênio e progesterona resultou em redução efetiva do volume ovariano. Todos os pacientes apresentaram níveis basais elevados de progesterona e LH e redução dos níveis de androgênios. Não observamos correlação entre os níveis de LH, da relação LH/FSH e de progesterona com o volume ovariano nos dois defeitos. Genótipo: O estudo molecular revelou que 17 pacientes de nossa coorte apresentavam mutações inativadoras no gene CYP17A1 e 1 paciente no gene POR. Duas novas mutações foram identificadas no gene CYP17A1, a p.R362H no éxon 6 e a p.G478S no éxon 8. A mutação mais prevalente no CYP17A1 foi a p.W406R identificada em 41 % das nossas famílias. Algumas das mutações no CYP17A1 foram encontradas apenas na coorte brasileira, mas a mutação p.A287P encontrada no gene POR em uma das nossas pacientes é a mais prevalente na literatura. Em relação à etnia, houve um predomínio dos defeitos moleculares no CYP17A1 em pacientes chinesas e brasileiras, enquanto que os defeitos no gene POR foram mais relatados nas pacientes europeias e norte - americanas. Em conclusão, a análise retrospectiva do fenótipo, genótipo e morfologia ovariana de trinta e duas pacientes 46,XX portadoras de deficiência da atividade P450c17, nos permitiu destacar os seguintes aspectos: a importância da dosagem da progesterona basal para este diagnóstico, a alta prevalência de aumento dos ovários com a presença de macrocistos ovarianos com risco de torção, e de transtornos psiquiátricos identificados em nossa coorte. Acreditamos que esta revisão possa contribuir para o diagnóstico mais precoce desta rara doença congênita / Congenital adrenal hyperplasia due to P450c17 deficiency is rarely reported on 46,XX patients. Our aim was to characterize the phenotype and genotype of this rare disorder reviewing the clinical, laboratory, genetic and ovarian imaging of 46,XX patients of our cohort and of other already reported cases. We retrospectively reviewed 32 patients with deficiency of P450c17 activity: 18 Brazilian patients belonging to 12 families and 10 cohorts already published (fourteen 46,XX patients, 6 of them with CYP17A1 defects and 8 with POR defects). Phenotype: most patients had primary amenorrhea (74%) and 90% of the patients did not develop pubarche; 69.5% of the patients had blood hypertension at diagnosis. We observed a high incidence of psychiatric disorders such as depression, anxiety and panic (76%) in our cohort but no reports were found in the literature. Ultrasound showed an increase of at least one of the ovaries in 87% of the patients before treatment and ovarian macrocysts in 65% of them; 6 patients (26%) had had previous surgery for twisting or ovarian rupture. Treatment with dexamethasone, estrogen and progesterone resulted in ovarian volume reduction. All patients showed elevated basal progesterone and LH and levels, and decreased androgen levels. There was no correlation between the levels of LH and progesterone and of LH/FSH ratio and the ovarian volume in both defects. Genotype: the molecular study showed that 17 patients from our cohort had inactivating mutation in the CYP17A1 gene and 1 in POR gene. Two novel mutations were identified in the CYP17A1 gene, the p.R362H in exon 6 and p.G478S in exon 8. The most prevalent mutation in the CYP17A1 was the p.W406R, identified in 41% of our families. Some of the CYP17A1 mutations were found only in the Brazilian cohort, but the mutation p.A287P found in the POR gene is the most prevalent in the literature. Regarding the ethnicity of the defects, there was a predominance of Chinese and Brazilian patients with defects in the CYP17A1 whereas defects in POR were most reported in European and North-American subjects.Conclusion: In this data review of thirty-two 46,XX patients with dysfunction of P450c17 activity we characterized the phenotype and genotype of this rare disorder and emphasize: the importance of basal progesterone measurement for this diagnosis, the high prevalence of ovarian macrocysts with risk of twisting, and the psychiatric disorders. We believe that this review may contribute to the early diagnosis of this disorder

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