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INTRODUCTION: BK virus (BKV) infection in renal transplant patients may cause kidney allograft dysfunction and graft loss. Accurate determination of BKV viral load is critical to prevent BKV-associated nephropathy (BKVAN) but the cut-off that best predicts BKVAN remains controversial. OBJECTIVE: To evaluate the performance of a commercial and an in-house qPCR test for quantitative detection of BK virus in kidney transplant recipients. METHODS: This was a prospective study with kidney transplant recipients from two large university hospitals in Brazil. Patients were screened for BKV infection every 3 months in the first year post-transplant with a commercial and an in-house real time polymerase chain reaction (qPCR) test. BKVAN was confirmed based on histopathology. The area under the curve for plasma qPCR was determined from receiver operating characteristic analysis. RESULTS: A total of 200 patients were enrolled. Fifty-eight percent were male, 19.5% had diabetes mellitus, and 82% had the kidney transplanted from a deceased donor. BKV viremia was detected in 32.5% and BKVAN was diagnosed in 8 patients (4%). BKVAN was associated with viremia of 4.1 log copies/mL, using a commercial kit. The cut-off for the in-house assay was 6.1 log copies/mL. The linearity between the commercial kit and the in-house assay was R2=0.83. CONCLUSION: Our study shows that marked variability occurs in BKV viral load when different qPCR methodologies are used. The in-house qPCR assay proved clinically useful, a cheaper option in comparison to commercial qPCR kits. There is an urgent need to make BKV standards available to the international community.

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Abstract Introduction: BK virus (BKV) infection in renal transplant patients may cause kidney allograft dysfunction and graft loss. Accurate determination of BKV viral load is critical to prevent BKV-associated nephropathy (BKVAN) but the cut-off that best predicts BKVAN remains controversial. Objective: To evaluate the performance of a commercial and an in-house qPCR test for quantitative detection of BK virus in kidney transplant recipients. Methods: This was a prospective study with kidney transplant recipients from two large university hospitals in Brazil. Patients were screened for BKV infection every 3 months in the first year post-transplant with a commercial and an in-house real time polymerase chain reaction (qPCR) test. BKVAN was confirmed based on histopathology. The area under the curve for plasma qPCR was determined from receiver operating characteristic analysis. Results: A total of 200 patients were enrolled. Fifty-eight percent were male, 19.5% had diabetes mellitus, and 82% had the kidney transplanted from a deceased donor. BKV viremia was detected in 32.5% and BKVAN was diagnosed in 8 patients (4%). BKVAN was associated with viremia of 4.1 log copies/mL, using a commercial kit. The cut-off for the in-house assay was 6.1 log copies/mL. The linearity between the commercial kit and the in-house assay was R2=0.83. Conclusion: Our study shows that marked variability occurs in BKV viral load when different qPCR methodologies are used. The in-house qPCR assay proved clinically useful, a cheaper option in comparison to commercial qPCR kits. There is an urgent need to make BKV standards available to the international community.

Resumo Introdução: A infecção pelo vírus BK (BKV) em pacientes de transplante renal pode levar a disfunção do aloenxerto renal e perda do enxerto. A determinação precisa da carga viral do BKV é fundamental para prevenir a nefropatia associada ao BKV (BKVAN), mas o ponto de corte de melhor valor preditivo para BKVAN ainda é foco de debates. Objetivo: Avaliar o desempenho de um teste de qPCR comercial e outro desenvolvido internamente para detecção quantitativa de vírus BK em receptores de transplante renal. Métodos: O presente estudo prospectivo incluiu receptores de transplante renal de dois grandes hospitais universitários no Brasil. Os pacientes foram testados para infecção por BKV a cada três meses no primeiro ano pós-transplante com um teste comercial de reação em cadeia de polimerase quantitativa em tempo real (qPCR) e outro desenvolvido internamente. A presença de BKVAN foi confirmada com base na histopatologia. A área sob a curva para o qPCR plasmático foi determinada a partir da análise da característica de operação do receptor. Resultados: Um total de 200 pacientes foram incluídos. Cinquenta e oito por cento eram do sexo masculino, 19,5% tinham diabetes mellitus e 82% tiveram seus rins transplantados de doadores falecidos. Viremia de BKV foi detectada em 32,5% dos pacientes e oito (4%) foram diagnosticados com BKVAN. BKVAN foi associada a viremia de 4,1 log cópias/mL usando o kit comercial. O corte para o ensaio interno foi de 6,1 log cópias/mL. A linearidade entre o kit comercial e o ensaio interno foi R2 = 0,83. Conclusão: Nosso estudo demonstrou uma acentuada variabilidade na carga viral de BKV quando diferentes metodologias de qPCR foram utilizadas. O ensaio interno de qPCR mostrou-se clinicamente útil, além de ser uma opção menos onerosa em relação aos kits comerciais de qPCR. Há uma necessidade urgente de se definir padrões de BKV para a comunidade internacional.

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Abstract The diagnosis of progressive disseminated histoplasmosis is often a challenge to clinicians, especially due to the low sensitivity and long turnaround time of the classic diagnostic methods. In recent years, studies involving a variety of non-culture-based diagnostic tests have been published in the literature. We performed a systematic review by selecting studies evaluating non-culture-based diagnostic methods for progressive disseminated histoplasmosis. We searched for articles evaluating detection of antibody, antigens, as well as DNA-based diagnostic methods. A comprehensive PUBMED, Web of Science, and Cochrane Library search was performed between the years 1956 and 2016. Case reports, review articles, non-human models and series involving less than 10 patients were excluded. We found 278 articles and after initial review 18 articles were included: (12) involved antigen detection methods, (4) molecular methods, and (2) antibody detection methods. Here we demonstrate that the pursuit of new technologies is ultimately required for the early and accurate diagnosis of disseminated histoplasmosis. In particular, urinary antigen detection was the most accurate tool when compared with other diagnostic techniques.

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The diagnosis of progressive disseminated histoplasmosis is often a challenge to clinicians, especially due to the low sensitivity and long turnaround time of the classic diagnostic methods. In recent years, studies involving a variety of non-culture-based diagnostic tests have been published in the literature. We performed a systematic review by selecting studies evaluating non-culture-based diagnostic methods for progressive disseminated histoplasmosis. We searched for articles evaluating detection of antibody, antigens, as well as DNA-based diagnostic methods. A comprehensive PUBMED, Web of Science, and Cochrane Library search was performed between the years 1956 and 2016. Case reports, review articles, non-human models and series involving less than 10 patients were excluded. We found 278 articles and after initial review 18 articles were included: (12) involved antigen detection methods, (4) molecular methods, and (2) antibody detection methods. Here we demonstrate that the pursuit of new technologies is ultimately required for the early and accurate diagnosis of disseminated histoplasmosis. In particular, urinary antigen detection was the most accurate tool when compared with other diagnostic techniques.

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OBJECTIVE: Recent studies have investigated the role of the p53 gene family in reproductive processes. Each member of the gene family acts through different mechanisms: p53 is involved in genomic stability and regulation of blastocyst implantation; p63 acts as a regulator of the quality and maturation of oocytes; and p73 controls the meiotic spindle. Polymorphisms in the genes of the p53 family have been associated with female infertility. One polymorphism in MDM2, the main regulator of the p53 family, has also been associated with this condition. Although polymorphisms in the TP53 gene have been related to recurrent pregnancy loss (RPL), there have been no studies associating polymorphisms in p63 and p73 with RPL. Therefore, the aim of this study was to evaluate the role of polymorphisms in the TP63 (rs17506395), TP73 (rs2273953, rs1801173), and MDM2 (SNP309, rs2279744) genes as risk factors for RPL. STUDY DESIGN: A case-control study was conducted in 153 women with RPL and 143 fertile women with at least two living children and no history of pregnancy loss. Molecular analysis was performed by TaqMan Allelic Discrimination assay. The statistical analysis was performed using SPSS software version 20.0 and the chi-square test, Student's t-test, Mann-Whitney test and logistic regression to compare the evaluated characteristics between both groups and RPL outcome. RESULTS: The allelic and genotypic frequencies did not differ between the groups when analyzed separately, however, the interaction between the TP63 TT and MDM2 TT genotypes was shown to increase the risk of RPL (OR=2.19, CI 95%: 1.28-3.75, p=0.004), even when adjusted for alcohol consumption, smoking, number of pregnancies and ethnicity (OR=1.97, CI 95%: 1.27-3.58, p=0.025). CONCLUSIONS: Our results suggest that genes from the p53 family proteins, evaluated here, have an influence on the risk of RPL.

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The TP53 gene, first described in 1979, was identified as a tumor suppressor gene in 1989, when it became clear that its product, the p53 nuclear phosphoprotein, was frequently inactivated in many different forms of cancers. Nicknamed "guardian of the genome", TP53 occupies a central node in stress response networks. The p53 protein has a key role as transcription factor in limiting oncogenesis through several growth suppressive functions, such as initiating apoptosis, senescence, or cell cycle arrest. The p53 protein is directly inactivated in about 50% of all tumors as a result of somatic gene mutations or deletions, and over 80% of tumors demonstrate dysfunctional p53 signaling. Beyond the undeniable importance of p53 as a tumor suppressor, an increasing number of new functions for p53 have been reported, including its ability to regulate energy metabolism, to control autophagy, and to participate in various aspects of differentiation and development. Recently, studies on genetic variations in TP53 among different populations have led to the notion that the p53 protein might play an important role in regulating fertility. This review summarizes current knowledge on the basic functions of different genes of the TP53 family and TP53 pathway with respect to fertility. We also provide original analyses based on genomic and genotype databases, providing further insights into the possible roles of the TP53 pathway in human reproduction.

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BACKGROUND: The susceptibility to adverse outcome from critical illness (occurrence of sepsis, septic shock, organ dysfunction/failure, and mortality) varies dramatically due to different degrees of inflammatory response. An over expression of tumor necrosis factor alpha (TNF-α) can lead to the progression of the inflammatory condition. OBJECTIVE: We assessed the relationship of the genotype distribution of -308G >A TNF-α polymorphism with regard to the development of sepsis, septic shock, higher organ dysfunction or mortality in critically ill patients. METHODS: Observational, hospital-based cohort study of 520 critically ill Caucasian patients from southern Brazil admitted to the general ICU of São Lucas Hospital, Porto Alegre, Brazil. Patients were monitored daily from the ICU admission day to hospital discharge or death, measuring SOFA score, sepsis, and septic shock occurrences. The -308G >A TNF-α SNP effect was analyzed in the entire patient group, in patients with sepsis (349/520), and in those who developed septic shock (248/520). RESULTS: The genotypic and allelic frequencies were -308GG = 0.72; -308GA = 0.27; -308AA = 0.01; -308G = 0.85; -308A = 0.15. No associations were found with sepsis, septic shock, organ dysfunction, and/or mortality rates among the TNF-α genotypes. Our results reveal that the -308G >A TNF-α SNP alone was not predictive of severe outcomes in critically ill patients. CONCLUSION: The principal novel input of this study was the larger sample size in an investigation with -308G > A TNF-α SNP. The presence of -308A allele is not associated with sepsis, septic shock, higher organ dysfunction or mortality in critically ill patients.

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BACKGROUND: The susceptibility to adverse outcome from critical illness (occurrence of sepsis, septic shock, organ dysfunction/failure, and mortality) varies dramatically due to different degrees of inflammatory response. An over expression of tumor necrosis factor alpha (TNF-α) can lead to the progression of the inflammatory condition. OBJECTIVE: We assessed the relationship of the genotype distribution of -308G >A TNF-α polymorphism with regard to the development of sepsis, septic shock, higher organ dysfunction or mortality in critically ill patients. METHODS: Observational, hospital-based cohort study of 520 critically ill Caucasian patients from southern Brazil admitted to the general ICU of São Lucas Hospital, Porto Alegre, Brazil. Patients were monitored daily from the ICU admission day to hospital discharge or death, measuring SOFA score, sepsis, and septic shock occurrences. The -308G >A TNF-α SNP effect was analyzed in the entire patient group, in patients with sepsis (349/520), and in those who developed septic shock (248/520). RESULTS: The genotypic and allelic frequencies were -308GG = 0.72; -308GA = 0.27; -308AA = 0.01; -308G = 0.85; -308A = 0.15. No associations were found with sepsis, septic shock, organ dysfunction, and/or mortality rates among the TNF-α genotypes. Our results reveal that the -308G >A TNF-α SNP alone was not predictive of severe outcomes in critically ill patients. CONCLUSION: The principal novel input of this study was the larger sample size in an investigation with -308G > A TNF-α SNP. The presence of -308A allele is not associated with sepsis, septic shock, higher organ dysfunction or mortality in critically ill patients.

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Multiple organ failure syndrome and acute renal dysfunction share many of physiologic factors involved in their development. Recent studies correlate the susceptibility to organ dysfunction in critically ill patients with genetic inheritance. Many of them consider ACE gene could be a possible candidate to elucidate a genetic predisposition or a genetic risk factor. We aimed to examine the effects of I/D and -262A > T ACE polymorphisms in the renal function in severely ill southern Brazilians patients. A multi-organic worldwide known failure score, the SOFA (sequential organ failure assessment), was used to determine the basal health state at first day (ICU admission). Considering admission SOFA score and trend of renal function (measured by daily renal SOFA scores, with daily measure of serum creatinine and diuresis), we hypothesize that ACE polymorphisms could influence in the trend of renal function in ICU patients. A total of 153 critically ill adult patients (79 men) were included in this study. We monitored the patients daily during their entire ICU and post-ICU (hospital) stay (measured from the ICU admission day to a maximum of 224 days). We observed progression to renal failure (SOFA scores 3 and 4) in first seven days of ICU stay and need for dialysis. The general genotypic frequencies in our sample were II = 0.17; ID = 0.46; DD = 0.37 and AA = 0.30; AT = 0.55; TT = 0.15, and the allelic frequencies were I = 0.40; D = 0.60 and A = 0.56; T = 0.44. This is the first study to verify the influence of I/D and -262A > T ACE polymorphisms in acute renal dysfunction among critically ill patients. No significant association was found between genotypes or allele frequencies and the trend of the renal function. The I/D and -262A > T ACE polymorphisms have no significant impact on the trend of renal function during the first week of ICU stay, neither any influence in mortality in critically ill patients.

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A síndrome de disfunção de múltiplos órgãos e sistemas (DMOS) e a disfunção renal aguda compartilham muitos dos fatores fisiológicos envolvidos em seu desenvolvimento. Estudos recentes correlacionam suscetibilidades individuais, determinadas geneticamente, à disfunção de órgãos em pacientes criticamente enfermos, situação em que o gene da enzima conversora da angiotensina (ECA) poderia ser um candidato para elucidar predisposição ou risco genético. Nosso objetivo foi examinar os efeitos da presença de dois polimorfismos, I/D e -262A > T, na disfunção renal em pacientes agudamente graves do Sul do Brasil. O escore SOFA (sequential organ failure assessment) à admissão e a tendência da função renal (medida pelo escore renal diário do SOFA) foram determinados em pacientes de unidade de terapia intensiva (UTI). Um total de 153 pacientes adultos (79 homens) foi incluído no estudo. Houve monitoração diária da função renal durante toda a permanência na UTI e também pós-UTI. Observou-se a progressão para insuficiência renal (SOFA 3 e 4) nos primeiros sete dias de internação em UTI, bem como necessidade de diálise. As frequências genotípicas gerais em nossa amostra foram II = 0,17; ID = 0,46; DD = 0,37; e AA = 0,30; AT = 0,55; TT = 0,15; e as frequências alélicas foram I = 0,40, D = 0,60; e A = 0,56; T = 0,44. Este é o primeiro estudo para verificar a influência de polimorfismos I/D e -262A > T da ECA em disfunção renal aguda em pacientes críticos. Nenhuma associação significativa foi encontrada entre os genótipos ou as frequências alélicas e a evolução da função renal. Os polimorfismos I/D e -262A > T da ECA não têm impacto significativo sobre a evolução da função renal durante a primeira semana de internação na UTI nem exercem qualquer influência sobre a mortalidade em pacientes graves.

Multiple organ failure syndrome and acute renal dysfunction share many of physiologic factors involved in their development. Recent studies correlate the susceptibility to organ dysfunction in critically ill patients with genetic inheritance. Many of them consider ACE gene could be a possible candidate to elucidate a genetic predisposition or a genetic risk factor. We aimed to examine the effects of I/D and -262A > T ACE polymorphisms in the renal function in severely ill southern Brazilians patients. A multi-organic worldwide known failure score, the SOFA (sequential organ failure assessment), was used to determine the basal health state at first day (ICU admission). Considering admission SOFA score and trend of renal function (measured by daily renal SOFA scores, with daily measure of serum creatinine and diuresis), we hypothesize that ACE polymorphisms could influence in the trend of renal function in ICU patients. A total of 153 critically ill adult patients (79 men) were included in this study. We monitored the patients daily during their entire ICU and post-ICU (hospital) stay (measured from the ICU admission day to a maximum of 224 days). We observed progression to renal failure (SOFA scores 3 and 4) in first seven days of ICU stay and need for dialysis. The general genotypic frequencies in our sample were II = 0.17; ID = 0.46; DD = 0.37 and AA = 0.30; AT = 0.55; TT = 0.15, and the allelic frequencies were I = 0.40; D = 0.60 and A = 0.56; T = 0.44. This is the first study to verify the influence of I/D and -262A > T ACE polymorphisms in acute renal dysfunction among critically ill patients. No significant association was found between genotypes or allele frequencies and the trend of the renal function. The I/D and -262A > T ACE polymorphisms have no significant impact on the trend of renal function during the first week of ICU stay, neither any influence in mortality in critically ill patients.

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