Guidelines of the Brazilian Society of Surgical Oncology for anatomopathological, immunohistochemical, and molecular testing in female tumors.

INTRODUCTION: Precision medicine has revolutionized oncology, providing more personalized diagnosis, treatment, and monitoring for patients with cancer. In the context of female-specific tumors, such as breast, ovarian, endometrial, and cervical cancer, proper tissue collection and handling are essential for obtaining tissue, immunohistochemical (IHC), and molecular data to guide therapeutic decisions. OBJECTIVES: To establish guidelines for the collection and handling of tumor tissue, to enhance the quality of samples for histopathological, IHC, genomic, and molecular analyses. These guidelines are fundamental in informing therapeutic decisions in cancer treatment. METHOD: The guidelines were developed by a multidisciplinary panel of renowned specialists between June 12, 2013 and February 12, 2024. Initially, the panel deliberated on critical and controversial topics related to conducting precision medicine studies focusing on female tumors. Subsequently, 22 pivotal topics were identified within the framework and assigned to groups. These groups reviewed relevant literature and drafted preliminary recommendations. Following this, the recommendations were reviewed by the coordinators and received unanimous approval. Finally, the groups made the final adjustments, classified the level of evidence, and ranked the recommendations. CONCLUSION: The collection of surgical samples requires minimum quality standards to enable histopathological, IHC, genomic, and molecular analyses. These analyses provide crucial data for informing therapeutic decisions, significantly impacting potential survival gains for patients with female tumors.

Evaluation of Multiple RNA Extraction Protocols for Chikungunya Virus Screening in Aedes aegypti Mosquitoes.

Chikungunya virus (Togaviridae, Alphavirus; CHIKV) is a mosquito-borne global health threat. The main urban vector of CHIKV is the Aedes aegypti mosquito, which is found throughout Brazil. Therefore, it is important to carry out laboratory tests to assist in the virus's diagnosis and surveillance. Most molecular biology methodologies use nucleic acid extraction as the first step and require quality RNA for their execution. In this context, four RNA extraction protocols were evaluated in Ae. aegypti experimentally infected with CHIKV. Six pools were tested in triplicates (n = 18), each containing 1, 5, 10, 20, 30, or 40 mosquitoes per pool (72 tests). Four commercial kits were compared: QIAamp®, Maxwell®, PureLink®, and PureLink® with TRIzol®. The QIAamp® and PureLink® with TRIzol® kits had greater sensitivity. Two negative correlations were observed: as the number of mosquitoes per pool increases, the Ct value decreases, with a higher viral load. Significant differences were found when comparing the purity and concentration of RNA. The QIAamp® protocol performed better when it came to lower Ct values and higher RNA purity and concentration. These results may provide help in CHIKV entomovirological surveillance planning.

Self-sampling for HPV testing in cervical cancer screening: A scoping review.

Cervical cancer is the third most common gynecological cancer worldwide. Its origin is linked to intraepithelial lesions caused by high-risk Human Papillomavirus (HPV) types, detected in 99.7% of cases. Early screening is essential to prevent cancer development from these lesions. Molecular methods are more specific and offer the possibility of being performed through a self-collected sample by the patient, thus contributing to increasing screening coverage for this pathology. This study aim was to map the medical-scientific literature on existing protocols for self-sampling for HPV testing in cervical cancer screening. A search strategy was developed using the following keywords and their synonyms: "self-sampling," "professional sampling," and "HPV", on the databases: MEDLINE, Cochrane Library, Virtual Health Library - BVS, Scopus, National Institute for Health Research NHS EED, Web of Science, and EMBASE. The search strategy was formulated to identify relevant studies and describe their main characteristics, such as patient acceptance of self-sampling, cost differences between the tests used, and the accuracy of self-sampling compared to the gold standard test. A total of 876 studies were found, and 33 of those studies were included in this review. Out of these, 10 studies were domized clinical trials involving 46,751 patients, and 23 observational studies included 142,795 patients. Regarding acceptance, most studies reported a preference for self-sampling. Sensitivity analyses from various studies also showed that the low cost of self-sampling kits generally increased cost-effectiveness. The study concluded that using HPV testing on self-collected samples is a viable strategy for monitoring women with HPV.

Epidemiological and Genomic Analysis of Asymptomatic SARS-CoV-2 Infections during the Delta and Omicron Epidemic Waves in São Paulo City, Brazil.

We examined the asymptomatic rates of SARS-CoV-2 infection during the Delta and Omicron waves in the city of São Paulo. Nasopharyngeal swabs were collected at strategic points of the city (open-air markets, bus terminals, airports) for SARS-CoV-2 RNA testing. Applying the questionnaire, the symptomatic individuals were excluded, and only asymptomatic cases were analyzed. During the Delta wave, a total of 4315 samples were collected, whereas 2372 samples were collected during the first Omicron wave. The incidence of the asymptomatic SARS-CoV-2 infection was 0.6% during the Delta wave and 0.8% during the Omicron wave. No statistical differences were found in the threshold amplification cycle. However, there was a statistical difference observed in the sublineage distribution between asymptomatic and symptomatic individuals. Our study determined the incidence of asymptomatic infection by monitoring individuals who remained symptom-free, thereby providing a reliable evaluation of asymptomatic SARS-CoV-2 carriage. Our findings reveal a relatively low proportion of asymptomatic cases, which could be attributed to our rigorous monitoring protocol for the presence of clinical symptoms. Investigating asymptomatic infection rates is crucial to develop and implement effective disease control strategies.

Comparison of 2 sampling methods for molecular detection of bacteria or fungi from feline hair and scale specimens.

Skin diseases of cats are among the most frequent client motivations for a veterinary consultation. Both carpet and toothbrush sampling are commonly used to obtain hair and scale samples for microbiologic testing. Although molecular tests have become more accessible and more widely used by clinicians, the ideal collection method for clinical specimens is unclear. To assess their performance in retrieving microbial DNA from clinical samples, we compared the bacterial and fungal DNA load in hair and skin scale samples collected using carpet or toothbrush methods. We evaluated sample DNA yield using fluorometry, spectrophotometry, and quantitative PCR. Despite no measurable differences in sample weight, toothbrush samples yielded significantly higher bacterial (p = 0.028) and fungal (p = 0.005) DNA loads compared to carpet samples, regardless of disease status. The toothbrush method was more effective in harvesting microbial DNA from hair and skin scale samples.

Epidemiological and genomic analysis of asymptomatic SARS-CoV-2 infections during the delta and omicron epidemic waves in São Paulo City, Brazil

We examined the asymptomatic rates of SARS-CoV-2 infection during the Delta and Omicron waves in the city of São Paulo. Nasopharyngeal swabs were collected at strategic points of the city (open-air markets, bus terminals, airports) for SARS-CoV-2 RNA testing. Applying the questionnaire, the symptomatic individuals were excluded, and only asymptomatic cases were analyzed. During the Delta wave, a total of 4315 samples were collected, whereas 2372 samples were collected during the first Omicron wave. The incidence of the asymptomatic SARS-CoV-2 infection was 0.6% during the Delta wave and 0.8% during the Omicron wave. No statistical differences were found in the threshold amplification cycle. However, there was a statistical difference observed in the sublineage distribution between asymptomatic and symptomatic individuals. Our study determined the incidence of asymptomatic infection by monitoring individuals who remained symptom-free, thereby providing a reliable evaluation of asymptomatic SARS-CoV-2 carriage. Our findings reveal a relatively low proportion of asymptomatic cases, which could be attributed to our rigorous monitoring protocol for the presence of clinical symptoms. Investigating asymptomatic infection rates is crucial to develop and implement effective disease control strategies

Reverse-Transcription Loop-Mediated Isothermal Amplification and alternative protocols for lower cost, large-scale COVID-19 testing: lessons from an emerging economy / Amplificación isotérmica mediada por bucle de transcripción inversa y protocolos alternativos para pruebas COVID-19 a gran escala y de bajo costo: lecciones de una economía emergente

Abstract Introduction: Most successful cases of COVID-19 pandemic mitigation and handling have relied on extensive reverse-transcription quantitative polymerase chain reaction (RT-qPCR). However, many emerging economies have struggled with current molecular testing demands due to economic, technical and technological constraints. Objective: To define a potential diagnostic protocol to increase testing capacity in current and post-pandemic conditions. Methods: We reviewed the literature, patents and commercial applications, for alternatives. Results: We found a good potential in saliva samples, viral inactivation and quick RNA extraction by heating; the use of an isothermal technology such as loop mediated isothermal amplification (LAMP) and naked eye test-result visualization by in-tube colorimetry or turbidity. Conclusions: Saliva samples with quick RNA extraction by heating and colorimetric LAMP are promising options for countries with economic and infrastructure limitations.

Resumen Introducción: La mayoría de los casos exitosos de mitigación y manejo de la pandemia de COVID-19 se han dado mediante pruebas basadas en la reacción en cadena de la polimerasa cuantitativa (RT-qPCR por sus siglas en inglés). Sin embargo, muchas economías emergentes han tenido problemas con las demandas actuales de pruebas moleculares debido a limitaciones económicas, técnicas y tecnológicas. Objetivo: Definir un protocolo de diagnóstico potencial para aumentar la capacidad de prueba en las condiciones actuales y posteriores a la pandemia. Métodos: Revisamos la literatura, las patentes y las aplicaciones comerciales, en busca de alternativas. Resultados: Encontramos un buen potencial en muestras de saliva, inactivación viral y extracción rápida de ARN por calentamiento; el uso de una tecnología isotérmica como la amplificación isotérmica mediada por horquillas (LAMP, por sus siglas en inglés) y la visualización del resultado de la prueba a simple vista mediante colorimetría o turbidez en el tubo. Conclusiones: Las muestras de saliva con extracción rápida de ARN por calentamiento y LAMP colorimétrico son opciones prometedoras para países con limitaciones económicas y de infraestructura.

EGFR-RAD51 fusion in lung adenocarcinoma with systemic and intracranial response to osimertinib: A case report and review of the literature.

Comprehensive next-generation sequencing panels are leading to detection of rare gene fusion events. EFGR-RAD51 fusion is a rare oncogenic finding and clinical data for management of this condition is scarce. We report a widely metastatic non-small cell lung cancer in a never-smoker young male patient with sustained near-complete systemic and intracranial response to osimertinib, a third-generation EGFR tyrosine-kinase inhibitor (TKI). We also review the available data of other TKIs in this scenario and underscore the role of comprehensive molecular testing for NSCLC.

Unlocking SARS-CoV-2 detection in low- and middle-income countries.

Low- and middle-income countries (LMICs) are significantly affected by SARS-CoV-2, partially due to their limited capacity for local production and implementation of molecular testing. Here, we provide detailed methods and validation of a molecular toolkit that can be readily produced and deployed using laboratory equipment available in LMICs. Our results show that lab-scale production of enzymes and nucleic acids can supply over 50,000 tests per production batch. The optimized one-step RT-PCR coupled to CRISPR-Cas12a-mediated detection showed a limit of detection of 102 ge/µL in a turnaround time of 2 h. The clinical validation indicated an overall sensitivity of 80%-88%, while for middle and high viral load samples (Cq ≤ 31) the sensitivity was 92%-100%. The specificity was 96%-100% regardless of viral load. Furthermore, we show that the toolkit can be used with the mobile laboratory Bento Lab, potentially enabling LMICs to implement detection services in unattended remote regions.

Optimized broad-range real-time PCR-based method for bacterial screening of platelet concentrates / Otimização de um método baseado em PCR em tempo real para rastreamento de bactérias em concentrados de plaquetas

Bacterial contamination of blood components remains a major challenge in transfusion medicine, particularly, platelet concentrates (PCs) due to the storage conditions that support bacterial proliferation. In this study, we develop a rapid, sensitive and specific real-time PCR protocol for bacterial screening of PCs. An internally controlled real-time PCR-based method was optimized and validated with our proprietary 16S Universal PCR Master Mix (IBMP/Fiocruz), which targets a conserved region of the bacterial 16S rRNA gene. Nonspecific background DNA was completely eliminated by treating the PCR Master Mix with ethidium monoazide (EMA). A lower limit of detection was observed for 10 genome equivalents with an observed Ct value of 34±1.07 in calibration curve generated with 10-fold serial dilutions of E. coli DNA. The turnaround time for processing, including microbial DNA purification, was approximately 4 hours. The developed method showed a high sensitivity with no non-specific amplification and a lower time-to-detection than traditional microbiological methods, demonstrating it to be an efficient means of screening pre-transfusion PCs.(AU)

A contaminação bacteriana dos componentes sanguíneos é um grande desafio na medicina transfusional, principalmente nos concentrados de plaquetas (PCs) devido às condições de armazenamento que favorecem a proliferação bacteriana. Neste estudo, desenvolvemos um protocolo de PCR em tempo real rápido, sensível e específico para a triagem bacteriana de PCs. Um método baseado em PCR em tempo real, controlado internamente, foi otimizado e validado com um Master Mix Universal PCR 16S (IBMP / Fiocruz), que detecta uma região conservada do gene 16S rRNA bacteriano. O background de DNA não específico foi completamente eliminado tratando a PCR Master Mix com monoazida de etídio (EMA). O limite de detecção inferior observado foi de 10 cópias equivalentes do genoma com um valor de Ct 34 ± 1,07, a curva de calibração foi gerada com diluições seriada de 10 vezes do DNA de E. coli. O tempo de processamento, incluindo a purificação microbiana do DNA, foi de aproximadamente 4 horas. O método desenvolvido mostrou alta sensibilidade sem amplificação inespecífica e menor tempo de detecção do que os métodos microbiológicos tradicionais, demonstrando ser um meio eficiente de triagem de PCs pré-transfusionais.(AU)

Optimized broad-range real-time PCR-based method for bacterial screening of platelet concentrates / Otimização de um método baseado em PCR em tempo real para rastreamento de bactérias em concentrados de plaquetas

Abstract Bacterial contamination of blood components remains a major challenge in transfusion medicine, particularly, platelet concentrates (PCs) due to the storage conditions that support bacterial proliferation. In this study, we develop a rapid, sensitive and specific real-time PCR protocol for bacterial screening of PCs. An internally controlled real-time PCR-based method was optimized and validated with our proprietary 16S Universal PCR Master Mix (IBMP/Fiocruz), which targets a conserved region of the bacterial 16S rRNA gene. Nonspecific background DNA was completely eliminated by treating the PCR Master Mix with ethidium monoazide (EMA). A lower limit of detection was observed for 10 genome equivalents with an observed Ct value of 34±1.07 in calibration curve generated with 10-fold serial dilutions of E. coli DNA. The turnaround time for processing, including microbial DNA purification, was approximately 4 hours. The developed method showed a high sensitivity with no non-specific amplification and a lower time-to-detection than traditional microbiological methods, demonstrating it to be an efficient means of screening pre-transfusion PCs.

Resumo A contaminação bacteriana dos componentes sanguíneos é um grande desafio na medicina transfusional, principalmente nos concentrados de plaquetas (PCs) devido às condições de armazenamento que favorecem a proliferação bacteriana. Neste estudo, desenvolvemos um protocolo de PCR em tempo real rápido, sensível e específico para a triagem bacteriana de PCs. Um método baseado em PCR em tempo real, controlado internamente, foi otimizado e validado com um Master Mix Universal PCR 16S (IBMP / Fiocruz), que detecta uma região conservada do gene 16S rRNA bacteriano. O background de DNA não específico foi completamente eliminado tratando a PCR Master Mix com monoazida de etídio (EMA). O limite de detecção inferior observado foi de 10 cópias equivalentes do genoma com um valor de Ct 34 ± 1,07, a curva de calibração foi gerada com diluições seriada de 10 vezes do DNA de E. coli. O tempo de processamento, incluindo a purificação microbiana do DNA, foi de aproximadamente 4 horas. O método desenvolvido mostrou alta sensibilidade sem amplificação inespecífica e menor tempo de detecção do que os métodos microbiológicos tradicionais, demonstrando ser um meio eficiente de triagem de PCs pré-transfusionais.

Métodos diagnósticos para la infección por SARS-CoV-2 / Diagnostic methods for SARS-CoV-2 infection

En diciembre de 2019 se identificó el virus SARS-CoV-2, cuya rápida propagación global puso en estado de emergencia al mundo entero, llevando al ser humano a una situación sin antecedente cercano. El objetivo de esta revisión es describir los métodos diagnósticos utilizados actualmente para identificar la infección por SARS-CoV-2. Las manifestaciones clínicas y el espectro imagenológico de la enfermedad son muy inespecíficos y no permiten realizar un diagnóstico certero. Por esta razón, es esencial una apropiada toma de muestra respiratoria en el momento y sitio anatómico adecuado para un diagnóstico preciso de COVID-19. La técnica de muestreo más utilizada es el hisopado nasofaríngeo y la prueba diagnóstica más fiable se basa en la retrotranscripción seguida por reacción en cadena de la polimerasa en tiempo real (RT-PCR). No obstante, existen otras técnicas moleculares, como también tests serológicos para detectar anticuerpos o fragmentos antigénicos del SARS-CoV-2. Más allá de la precisión diagnóstica, es importante tener en cuenta la probabilidad basal (pretest) para interpretar correctamente el resultado obtenido y aislar aquellos posibles falsos negativos. Con el objetivo de evitar la saturación del sistema de salud es imprescindible contar con información y métodos diagnósticos precisos para detectar tempranamente los focos de infección y reducir la transmisión comunitaria, utilizando eficazmente los diferentes recursos diagnósticos. (AU)

In December 2019, the SARS-CoV-2 virus was identified for the first time, whose rapid global spread put the entire world in a state of emergency, leading humans to an unprecedented situation with no immediate history. The main purpose of this review is to describe the diagnostic methods currently used to identify SARS-CoV-2 infection. The clinical manifestations and the imaging spectrum of the disease are nonspecific and do not allow an accurate diagnosis to be made. For this reason, an appropriate respiratory sampling at the right time and anatomical site is essential for an accurate diagnosis of COVID-19. The most widely used sampling technique is nasopharyngeal swab, and the most reliable diagnostic test is by reverse transcription followed by real-time polymerase chain reaction (RT-PCR). However, there are other molecular techniques, as well as serological tests to detect antibodies or antigenic fragments of SARS-CoV-2. Beyond the diagnostic precision, it is important to take into account the baseline probability (pre-test) to correctly interpret the result obtained and isolate those possible false negatives. In order to avoid saturation of the health system, it is essential to have accurate information and diagnostic methods to detect outbreaks of infection in early stages and to reduce communitary transmission, making effective use of the various diagnostic resources. Coronavirus infections/diagnosis, viral/diagnosis, pandemics, clinical laboratory techniques, real-time polymerase chain reaction, antigens, viral/analysis. (AU)

Mutational Portrait of Lung Adenocarcinoma in Brazilian Patients: Past, Present, and Future of Molecular Profiling in the Clinic.

Objectives: Approximately 60% of lung adenocarcinomas (LAs) carry mutations that can guide treatment with tyrosine-kinase inhibitors (TKI) and other targeted therapies. Data on activating mutations in EGFR and other tyrosine-kinase receptor (TKR) genes in highly admixed populations, such as that of Brazil, are scarce. In this study, we comprehensively analyzed the actionable alteration profile of LA in Brazilian patients. Materials and Methods: EGFR driver mutation data were collected from a large Brazilian LA cohort covering an 8-year period of molecular testing in a single institution. Tests were performed using three distinct methods, and demographic and histopathological data were analyzed. For a subset of patients, driver mutations in KRAS, NRAS, and BRAF and gene fusions involving TKR genes (before TKI treatment) and EGFR T790M (after TKI treatment) were assessed. Results: EGFR mutations were detected in 25% of 1,316 LAs evaluated, with exon 19 deletions and exon 21 L858R TKI sensitizing mutations representing 72.5% of all mutations. Mutation rates were higher in women and non-smokers (p < 0.001). Next-generation sequencing was very sensitive, with a lower rate of inconclusive results compared with Sanger sequencing and pyrosequencing. EGFR/RAS/BRAF hotspot gene panels were applied in 495 LA cases and detected oncogenic mutations in 51.3% of samples, most frequently in EGFR (22.4%) and KRAS (26.9%). In subgroups of 36 and 35 patients, gene fusions were detected in 11.1% of tumors and EGFR T790M resistance mutations were detected in 59% of plasma samples from patients previously treated with TKI, respectively. Conclusion: This report provides the first comprehensive actionable alteration portrait of LA in Brazil. The high rate of actionable alterations in EGFR and other driver genes in LA reinforces the need to incorporate TKI guided by molecular diagnostics into clinical routines for patients in both public and private healthcare systems.

The Main Molecular and Serological Methods for Diagnosing COVID-19: An Overview Based on the Literature.

Diagnostic tests have been considered as the main alternative for the control of coronavirus disease (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), as a correct diagnosis allows for decision making when facing the disease, particularly as there is a lack of effective therapeutic protocols and vaccines. Thus, in this review, we summarized the main diagnostic approaches currently available for the diagnosis of SARS-CoV-2 infection in humans based on studies available in article databases. The tests can be organized into two main categories: nucleic acid-based tests, recommended for the initial detection of the virus, and serological tests, recommended for assessing the disease progression. The studies have shown that the performance of diagnostic methods depends on different factors, such as the type of samples and the characteristics of each assay. It was identified that the positivity of the tests is mainly related to the onset of symptoms. We also observed that point-of-care diagnoses are considered as one of the main trends in this area, due to the low-cost and simplicity of the assay; however, the analytical performance must be critically analyzed. Thus, the COVID-19 pandemic has highlighted the critical role of diagnostic technologies in the control of infectious diseases.

Optimized broad-range real-time PCR-based method for bacterial screening of platelet concentrates

Abstract Bacterial contamination of blood components remains a major challenge in transfusion medicine, particularly, platelet concentrates (PCs) due to the storage conditions that support bacterial proliferation. In this study, we develop a rapid, sensitive and specific real-time PCR protocol for bacterial screening of PCs. An internally controlled real-time PCR-based method was optimized and validated with our proprietary 16S Universal PCR Master Mix (IBMP/Fiocruz), which targets a conserved region of the bacterial 16S rRNA gene. Nonspecific background DNA was completely eliminated by treating the PCR Master Mix with ethidium monoazide (EMA). A lower limit of detection was observed for 10 genome equivalents with an observed Ct value of 34±1.07 in calibration curve generated with 10-fold serial dilutions of E. coli DNA. The turnaround time for processing, including microbial DNA purification, was approximately 4 hours. The developed method showed a high sensitivity with no non-specific amplification and a lower time-to-detection than traditional microbiological methods, demonstrating it to be an efficient means of screening pre-transfusion PCs.

Resumo A contaminação bacteriana dos componentes sanguíneos é um grande desafio na medicina transfusional, principalmente nos concentrados de plaquetas (PCs) devido às condições de armazenamento que favorecem a proliferação bacteriana. Neste estudo, desenvolvemos um protocolo de PCR em tempo real rápido, sensível e específico para a triagem bacteriana de PCs. Um método baseado em PCR em tempo real, controlado internamente, foi otimizado e validado com um Master Mix Universal PCR 16S (IBMP / Fiocruz), que detecta uma região conservada do gene 16S rRNA bacteriano. O background de DNA não específico foi completamente eliminado tratando a PCR Master Mix com monoazida de etídio (EMA). O limite de detecção inferior observado foi de 10 cópias equivalentes do genoma com um valor de Ct 34 ± 1,07, a curva de calibração foi gerada com diluições seriada de 10 vezes do DNA de E. coli. O tempo de processamento, incluindo a purificação microbiana do DNA, foi de aproximadamente 4 horas. O método desenvolvido mostrou alta sensibilidade sem amplificação inespecífica e menor tempo de detecção do que os métodos microbiológicos tradicionais, demonstrando ser um meio eficiente de triagem de PCs pré-transfusionais.

Characterization of RHD alleles present in serologically RHD-negative donors determined by a sensitive microplate technique.

BACKGROUND AND OBJECTIVES: Weak D phenotypes with very low antigen densities and DEL phenotype may not be detected in RhD typing routine and could be typed as D-negative, leading to D alloimmunization of D-negative recipients. The present study aimed to investigate the presence of RHD-positive genotypes in blood donors typed as D-negative by an automated system using the solid-phase methodology as a confirmatory test. METHODS: Two screenings were performed in different selected donor populations. For the first screening, we selected 1403 blood donor samples typed as D-negative regardless of the CE status, and in the second screening, we selected 517 donor samples typed as D-negative C+ and/or E+. RhD typing was performed by microplate in an automated equipment (Neo-Immucor®), and the confirmatory test was performed by solid-phase technique using Capture R® technology. A multiplex PCR specific to RHD and RHDψ was performed in a pool of 6 DNA samples. Sequencing of RHD exons was performed in all RHD-positive samples, and a specific PCR was used to identify the D-CE(4-7)-D hybrid gene. RESULTS AND CONCLUSION: No weak D type was found in either screening populations. Additionally, 353 (18·4%) D-negative samples presented previously reported non-functional RHD genes, 2 samples had a DEL allele, and 6 samples demonstrated new alleles, including one novel DEL allele. Our study identified six new RHD alleles and showed that the inclusion of a confirmatory test using serological methodology with high sensitivity can reduce the frequency of weak D samples typed as D-negative.

Self-Collection and Molecular Diagnosis for Detection of Human Papillomavirus: Why Incorporate It?

PURPOSE OF REVIEW: Cervical cancer, the third cause of death by cancer among Brazil's women, is associated with human papillomavirus (HPV) infection. In some countries of South America, North America, Europe, and Oceania, initial screening for HPV DNA and subsequent follow-up with HPV-positive patients using colposcopy and cytological testing are used as preventative measures. RECENT FINDINGS: For HPV DNA detection, it is necessary to obtain cervical cells by conventional clinical collection method or self-collection of the cells that flake off from the uterine cervix and vaginal canal. Self-collection has been shown to be a viable option for obtaining samples and is a less invasive method that is more accepted by women. Thus, it can potentially decrease the limitations of the conventional clinical collection methods. The efficiency of the self-collection method aligned with the implementation of HPV molecular testing, if adopted by public and private health care systems, may extend the reach of current cervical cancer prevention efforts. In addition, considering all phases from triage to treatment, this method may reduce health care costs and the time spent by patients and health care teams to conduct examinations and collect samples.

Histophilus somni-associated syndromes in sheep from Southern Brazil

Histophilus somni is a Gram-negative bacterium that is associated with a disease complex (termed histophilosis) that can produce several clinical syndromes predominantly in cattle, but also in sheep. Histophilosis is well described in North America, Canada, and in some European countries. In Brazil, histophilosis has been described in cattle with respiratory, reproductive, and systemic disease, with only one case described in sheep. This report describes the occurrence of Histophilus somni-associated disease in sheep from Southern Brazil. Eight sheep with different clinical manifestations from five farms were investigated by a combination of pathological and molecular diagnostic methods to identify additional cases of histophilosis in sheep from Brazil. The principal pathological lesions were thrombotic meningoencephalitis, fibrinous bronchopneumonia, pulmonary abscesses, and necrotizing myocarditis. The main clinical syndromes associated with H. somni were thrombotic meningoencephalitis (n = 4), septicemia (n = 4), bronchopneumonia (n = 4), and myocarditis (n = 3). H. somni DNA was amplified from multiple tissues of all sheep with clinical syndromes of histophilosis; sequencing confirmed the PCR results. Further, PCR assays to detect Pasteurella multocida and Mannheimia haemolytica were negative. These findings confirmed the participation of H. somni in the clinical syndromes investigated during this study, and adds to the previous report of histophilosis in sheep from Brazil.(AU)

Histophilus somni-associated syndromes in sheep from Southern Brazil

Abstract Histophilus somni is a Gram-negative bacterium that is associated with a disease complex (termed histophilosis) that can produce several clinical syndromes predominantly in cattle, but also in sheep. Histophilosis is well described in North America, Canada, and in some European countries. In Brazil, histophilosis has been described in cattle with respiratory, reproductive, and systemic disease, with only one case described in sheep. This report describes the occurrence of Histophilus somni-associated disease in sheep from Southern Brazil. Eight sheep with different clinical manifestations from five farms were investigated by a combination of pathological and molecular diagnostic methods to identify additional cases of histophilosis in sheep from Brazil. The principal pathological lesions were thrombotic meningoencephalitis, fibrinous bronchopneumonia, pulmonary abscesses, and necrotizing myocarditis. The main clinical syndromes associated with H. somni were thrombotic meningoencephalitis (n = 4), septicemia (n = 4), bronchopneumonia (n = 4), and myocarditis (n = 3). H. somni DNA was amplified from multiple tissues of all sheep with clinical syndromes of histophilosis; sequencing confirmed the PCR results. Further, PCR assays to detect Pasteurella multocida and Mannheimia haemolytica were negative. These findings confirmed the participation of H. somni in the clinical syndromes investigated during this study, and adds to the previous report of histophilosis in sheep from Brazil.