Optimización de unidades formadoras de colonias y detección del virus de papiloma humano en sangre de cordón umbilical / Colony-forming units optimization and human papillomavirus detection in umbilical cord blood

Resumen Introducción: Se requiere analizar diversos parámetros para el control de calidad adecuado de las unidades de sangre de cordón umbilical (USCU) cuando se utilizan con fines terapéuticos. Objetivo: Optimizar las unidades formadoras de colonias (UFC) de cultivos clonogénicos y detectar el genoma del virus del papiloma humano (VPH) en USCU. Métodos: Se incluyeron 141 muestras de sangre de cordón umbilical (SCU), de segmento y de UFC de cultivos clonogénicos de USCU. Se realizó extracción de ADN, cuantificación y amplificación por PCR del gen endógeno GAPDH. Se detectó el gen L1 del VPH con los oligonucleótidos MY09/MY11 y GP5/GP6+; los productos de PCR se migraron en electroforesis de agarosa. El ADN purificado de las UFC se analizó mediante electroforesis de agarosa y algunos ADN, con la técnica sequence specific priming. Resultados: La concentración de ADN extraído de UFC fue superior comparada con la de SCU (p = 0.0041) y la de segmento (p < 0.0001); así como la de SCU comparada con la de segmento (p < 0.0001). Todas las muestras fueron positivas para la amplificación de GAPDH y negativas para MY09/MY11 y GP5/GP6+. Conclusiones: Las USCU criopreservadas fueron VPH netativas; además, es factible obtener ADN en altas concentraciones y con alta pureza a partir de UFC de los cultivos clonogénicos.

Abstract Introduction: Analysis of several markers is required for adequate quality control in umbilical cord blood units (UCBU) when are used for therapeutic purposes. Objective: To optimize colony-forming units (CFU) from clonogenic cultures and to detect the human papillomavirus (HPV) genome in UCBU. Methods: One hundred and forty-one umbilical cord blood (UCB), segment or CFU samples from UCBU clonogenic cultures were included. DNA extraction, quantification and endogenous GAPDH gene PCR amplification were carried out. Subsequently, HPV L1 gene was detected using the MY09/MY11 and GP5/GP6+ oligonucleotides. PCR products were analyzed with electrophoresis in agarose gel. CFU-extracted purified DNA was analyzed by electrophoresis in agarose gel, as well as some DNAs, using the SSP technique. Results: CFU-extracted DNA concentration was higher in comparison with that of UCB (p = 0.0041) and that of the segment (p < 0.0001), as well as that of UCB in comparison with that of the segment (p < 0.0001). All samples were positive for GAPDH amplification and negative for MY09/MY11 and GP5/GP6+. Conclusions: Cryopreserved UCBUs were HPV-negative. Obtaining CFU DNA from clonogenic cultures with high concentrations and purity is feasible.

Prevalencia de los genotipos de HPV en lesiones pre invasoras de alto grado de malignidad y cáncer de cuello uterino en la población del Hospital de Clínicas: Montevideo-Uruguay / Prevalence of HPV genotypes in cervical high risk intraepithelial lesions and cervical cancer in the population of Hospital de Clínicas: Montevideo-Uruguay / Prevalência de genótipos de HPV em lesões pré-invasivas com alto grau de malignidade e câncer de colo do útero na população do Hospital de Clínicas: Montevidéu, Uruguai

El Cáncer de Cuello Uterino (CCU) es un problema de Salud pública a nivel mundial. Su indiscutible asociación con el Virus del papiloma humano (HPV) hace necesario su estudio. El objetivo de este trabajo, es conocer la prevalencia de los diferentes genotipos de HPV, en lesiones pre invasoras de alto grado de malignidad (HSIL) y/o cáncer de cuello uterino. Material y Métodos: Todas las Mujeres que fueron derivadas a pol de TGI del H Clínicas entre enero del 2011 y diciembre de 2012, por un PAP sospechoso de lesión y en las que se confirmó luego un HSIL o cáncer cervical fueron tipificadas. Se recabaron datos de edad, tipo de lesión y genotipificación. La extracción de ADN viral se realizó a partir de muestras cervico vaginales conservadas en medio de transporte comercial (Sacace) mediante el kit QIAamp DNA Mini Kit (QIAGEN). Se buscaron 14 genotipos de alto riesgo. Resultados: Se tipificaron 75 pacientes, 19 con CCU y 56 con HSIL. El HPV 16 fue el más prevalente en un 61,5 % para las infecciones únicas y en un casi 100 % para las múltiples y un 60% para el total de las lesiones, seguido en prevalencia por los HPV 31,33 y 45. El HPV 18 fue muy poco prevalente. Conclusiones: En esta muestra, la prevalencia del HPV 16 está acorde con las publicaciones nacionales siendo el más frecuente. El HPV 18 tiene muy baja prevalencia siendo 2 casos en 75, siempre en infecciones múltiples.

Cervical Cancer (CC) is a public health problem worldwide. Its indisputable association with the Human Papilloma Virus (HPV) makes its study necessary. The objective of this work is to know the prevalence of the different HPV genotypes, in pre-invasive high-grade malignant lesions (HSIL) and / or cervical cancer. Material and Methods: All women who were referred to Low genital tract service in the Hospital de Clinicas between January 2011 and December 2012, for a PAP suspected of injury and in which HSIL or cervical cancer was later confirmed were typified. Data on age, type of lesion and genotyping were collected. The viral DNA extraction was carried out from cervico-vaginal samples preserved in commercial transport medium (Sacace) using the QIAamp DNA Mini Kit (QIAGEN). 14 high-risk genotypes were searched. Results: 75 patients were typified, 19 with CCU and 56 with HSIL. HPV 16 was the most prevalent in 61.5% for single infections and almost 100% for multiple infections and 60% for all lesions, followed in prevalence by HPV 31,33 and 45. The HPV 18 was very rare. Conclusions: In this sample, the prevalence of HPV 16 is in accordance with national publications, being the most frequent. HPV 18 has a very low prevalence, being 2 cases in 75, always in multiple infections.

O câncer cervical é um problema de saúde pública em todo o mundo. E um câncer com uma ligação comprovada com o vírus do papiloma humano. O objetivo é conhecer a prevalência dos diferentes genótipos do HPV em mulheres que apresentam neoplasias pré-invasivas de alto grau e câncer de colo do útero, que foram tratadas e diagnosticadas no Hospital de Clínicas entre janeiro de 2011 e dezembro de 2012. Material e Métodos: o estudo foi realizado em 75 pacientes do Hospital de Clinicas, com diagnóstico histológico de Câncer Cervical e lesões intraepiteliais de alto grau para as quais foi realizado o tipageme do HPV alto risco. Resultados: 75 pacientes foram tipificados, 19 com CCU e 56 com HSIL. O HPV 16 foi o mais prevalente em 61,5% para infecções únicas e quase 100% para infecções múltiplas e 60% para todas as lesões, seguido em prevalência pelo HPV 31,33 e 45. O HPV 18 era muito raro. Conclusões: Nesta amostra, a prevalência do HPV 16 está de acordo com as publicações nacionais, sendo a mais frequente. O HPV 18 tem prevalência muito baixa, sendo 2 casos em 75, sempre em infecções múltiplas.

Interpretación de resultados de laboratorio para diagnóstico de COVID-19, 6 de mayo del 2020 / Interpretation of laboratory results for COVID-19 diagnosis, 6 May 2020 / Interpretação dos resultados laboratoriais para o diagnóstico de COVID-19, 6 de maio de 2020

La confirmación etiológica de la infección por el virus que causa la COVID-19 (SARS-CoV-2) solo puede hacerse mediante ensayos de laboratorio. La interpretación adecuada de los resultados obtenidos en cualquier tipo de ensayo debe ser realizada cuidadosamente y teniendo en cuenta la dinámica de la infección (momento en que se toma una muestra y la calidad de dicha muestra) y el objetivo por el cual se toma una muestra (diagnóstico, seroprevalencia, etc.).

Etiological confirmation of COVID-19 virus (SARS-CoV-2) infection can only be made by laboratory tests. In general, the currently available assays for COVID-19 can be classified into two groups: • The first group (virological tests) includes tests that can detect the presence of the components of the virus (genetic material or antigens).

A interpretação adequada dos resultados obtidos em qualquer tipo de ensaio deve ser realizada com cuidado e levar em consideração a dinâmica da infecção (quando a amostra é coletada) e o objetivo para o qual a amostra é coletada (diagnóstico, soroprevalência, etc.).

Polyomaviruses BK and JC DNA infection in peripheral blood cells from blood donors

ABSTRACT Objectives: To investigate the prevalence of human polyomavirus (BK and JC viruses) infection in peripheral blood mononuclear cells of healthy blood donors. Methods: The study included 250 healthy blood donors. Five-milliliter blood was drawn into sterile EDTA tubes and PBMCs were isolated from whole blood. The isolated PBMCs were counted and stored at −70 °C for future investigation. DNA was extracted and subjected to simple, sensitive and specific semi-nested PCR as well as QPCR using both general and specific primers for different assays. Results: Of 250 blood samples, 66 (26.4%) were positive for BKV DNA (146-34,514 copies/106 cells). JC DNA was found in 45 (18%) blood samples (65-21,250 copies/106 cells). Co-infection with these viruses were found in 11 (4.4%) out of 250 blood samples. Discussion: Our study provides important data on polyomavirus infection in peripheral blood mononuclear leukocytes in immunocompetent individuals. These data indicate significant differences between the prevalence of BKV and JCV infection in healthy blood donors. The prevalence of BK and JC virus infection is higher in the age range 30-39 years compared to other age ranges.

Molecular identification of viral agents associated with acute conjunctivitis: a prospective controlled study

Abstract Background: Viral conjunctivitis are the most frequent infections in ophthalmology clinics. The diagnosis is usually relying on clinical findings and medical history. However, topical antibiotics are often used unnecessarily addition to symptomatic treatment because of unsure agents. We aimed to detect the Adenovirus, Coxsackievirus and Enterovirus from conjunctiva and pharyngeal samples of patients. Methods: The conjunctiva and pharyngeal samples of the patients with conjunctivitis were taken by Virocult transport media and kept at -80 ºC up to study day. Adenovirus spp, Enterovirus 70 and Enterovirus 71, Coxsackie A24 and Coxsackie A16 were detected by real-time PCR. Samples from healthy health care workers of ophthalmology clinic were used for control group. Results: A total of 176 samples (conjunctival and pharyngeal samples of 62 patient and 26 healthy subjects) were included. The mean age of 34 (55.7%) male and 27 (44.3%) female patients was 34 ± 17. Twenty five (40.3%) of the patients were receiving antibiotic drops at first visit. The main etiologic agent in conjunctival samples was found to be Adenovirus (46/62, 74.2%) followed by Enterovirus 70 (4/62, 6.4%) and Enterovirus 71 (4/62, 6.4%). Coxsackievirus 16 and 24 were also found in 2 patients (1/62 each, 1.6%). Pharyngeal samples were also positive for Adenovirus (20/62, 32.3%), Enterovirus 70 and 71 (7/62, 11.3% and 5/62, 8.1% respectively), Coxsackievirus 16 and 24 (2/62, 3.2% and 1/61, 1.6%). Conclusions: It is very difficult in viral conjunctivitis to make clinical differentiation caused by different agents because of common clinical signs and symptoms. In routine clinical work, the viral conjunctivitis usually related with Adenovirus. But almost one fourth of the patients' conjunctivitis were not related to Adenovirus, which shows the importance of the laboratory diagnostics. True diagnosis plays an important role on prevention of contamination and unnecessary use of antibiotics in viral conjunctivitis.

Detection and molecular characterisation of a diagnosis escape variant associated with occult hepatitis B virus in Brazil

BACKGROUND Many studies have identified mutations in the hepatitis B surface antigen (HBsAg) as important factors limiting the ability of commercial serological assays to detect this viral antigen. However, an association between mutations in the HBsAg gene and the occurrence of occult HBV infection (OBI) in patients has not been established. OBJECTIVES To detect hepatitis B virus (HBV) DNA in patients with anti-HBc as a unique serological marker, a previously published, cost-effective TaqMan-based real-time polymerase chain reaction (PCR) test with minor groove binding probes was adapted for use in this study. The current study also aimed to investigate HBsAg mutations and genotypes of HBV in OBI at the Viral Hepatitis Ambulatory Clinic in Rio de Janeiro to determine any possible association. METHODS Intra-assay and inter-assay reproducibility were determined, and the mean coefficient of variation values obtained were 2.07 and 3.5, respectively. Probit analysis indicated that the 95% detection level was 25 IU/mL. The prevalence of OBI was investigated in 35 serum samples with an ‘anti-HBc alone’ profile from individuals who attended our clinic between 2011 and 2013. FINDINGS HBV DNA was detected in only one sample, resulting in an OBI rate of 2.9%. Nucleotide sequencing of the pre-S/S region was performed to genotype and analyse mutations within the HBsAg gene of this HBV DNA. The HBV in the OBI case was classified as sub-genotype A1, and a sequence analysis of the small S gene revealed 12 mutations in the major hydrophilic region compared to the consensus A1 sequence. Most of these mutations occurred in amino acid residues that have been reported as clinically relevant because they have been implicated in vaccine escape and/or inability to detect HBsAg by commercial serological assays. MAIN CONCLUSIONS Our study suggests the importance of specific HBsAg mutations, different from those in D, B, and C genotypes, in sub-genotype A1 HBV associated with OBI.

Prevalence of occult hepatitis b infection in iranian cancer patients before chemotherapy treatment / Prevalência de infecção oculta da hepatite B em pacientes iranianos com câncer antes do tratamento de quimioterapia

ABSTRACT Background Occult hepatitis B infection is characterized by negative hepatitis B surface antigen (HBsAg) and also detectable hepatitis B virus (HBV) -DNA, with or without hepatitis B core antibody (anti-HBc). HBV reactivation in individuals under immunosuppressive therapy is critical, occurring in occult HBV. Objective In this study, we aimed to determine the prevalence of occult HBV infection among hepatitis B surface antigen negative in cancer patients before receiving chemotherapy. Methods Sera from 204 cancer patients who were negative for HBsAg, were tested for anti-HBc antibodies. The samples that were negative for HBsAg but positive for anti-HBc also examined for HBV-DNA by polymerase chain reaction (PCR). Results Of the 204 HBsAg negative blood samples, 11 (5.4%) samples were positive for anti-HBc antibodies. HBV-DNA was detected in 9/11 (81%) of anti-HBc positive samples. Occult HBV infection in hematological cancers was more than solid cancers, 4.8% and 4.3% respectively. There was no significant difference in HBc antibody positivity based on vaccination, previous blood transfusions, history of familial hepatitis or biochemical parameters (ALT, AST, total and direct bilirubin levels) (P>0.05). Conclusion Screening of occult HBV infection by HBsAg, HBV DNA and anti HB core antibody should be suggested as a routine investigation in cancer patients before receiving chemotherapy.

RESUMO Contexto A infecção oculta da hepatite B caracteriza-se por antígeno de superfície da hepatite B (AgHBs) negativo com vírus detectável da hepatite B (HBV) -DNA, com ou sem anticorpo de núcleo da hepatite B (anti-HBc). A reativação do HBV em indivíduos sob terapia imunossupressora é crítica, originando a infecção oculta pelo VHB. Objetivo Este estudo teve como objetivo determinar a prevalência de infecção oculta pelo VHB entre em pacientes com câncer e com antígeno de superfície da hepatite B negativo antes de receber quimioterapia. Métodos Soro de 204 pacientes com câncer que foram negativos para AgHBs, foram testados para anticorpos anti-HBc. As amostras que foram negativos para AgHBs, mas positivo para anti-HBc foram também examinadas para HBV-DNA, por reação em cadeia da polimerase. Resultados Entre 204 amostras de sangue AgHBs negativas, 11 (5,4%) foram positivos para anticorpos anti-HBc. HBV-DNA foi detectado em 9/11 (81%) das amostras positivas de anti-HBc. Infecção oculta de VHB em câncer hematológico foi maior que em cânceres sólidos, 4,8% e 4,3% respectivamente. Não houve diferença significativa na positividade anti-HBc, com base na vacinação, transfusões de sangue anteriores, história de hepatite familiar ou parâmetros bioquímicos (ALT, AST, total e níveis de bilirrubina total) (P & gt; 0,05). Conclusão A triagem de infecção oculta por AgHBs, HBV-DNA e anti-anticorpo de núcleo HB deve ser sugerida como uma investigação de rotina em pacientes com câncer antes de receber a quimioterapia.

The role of human adenoviruses type 41 in acute diarrheal disease in Minas Gerais after rotavirus vaccination

Abstract Human adenovirus species F (HAdV-F) type 40 and 41 are commonly associated with acute diarrheal disease (ADD) across the world. Despite being the largest state in southeastern Brazil and having the second largest number of inhabitants, there is no information in the State of Minas Gerais regarding the role of HAdV-F in the etiology of ADD. This study was performed to determine the prevalence, to verify the epidemiological aspects of infection, and to characterize the strains of human adenoviruses (HAdV) detected. A total of 377 diarrheal fecal samples were obtained between January 2007 and August 2011 from inpatient and outpatient children of age ranging from 0 to 12 years. All samples were previously tested for rotavirus, norovirus, and astrovirus, and 314 of 377 were negative. The viral DNA was extracted, amplified using the polymerase chain reaction and the HAdV-positive samples were sequenced and phylogenetically analyzed. Statistical analyses were performed using the Chi-square test (p < 0.05), considering two conditions: the total of samples tested (377) and the total of negative samples for the remaining viruses tested (314). The overall prevalence of HAdV was 12.47% (47/377); and in 76.60% (36/47) of the positive samples, this virus was the only infectious agent detected. The phylogenetic analysis of partial sequences of 32 positive samples revealed that they all clustered with the HAdV-F type 41. The statistical analysis showed that there was no correlation between the onset of the HAdV infection and the origin of the samples (inpatients or outpatients) in the two conditions tested: the total of samples tested (p = 0.598) and the total of negative samples for the remaining viruses tested (p = 0.614). There was a significant association in the occurrence of infection in children aged 0–12 months for the condition 1 (p = 0.030) as well as condition 2 (p = 0.019). The occurrence of infections due to HAdV did not coincide with a pattern of seasonal distribution. These data indicate the significant involvement of HAdV-F type 41 in the etiology of ADD in Minas Gerais, which demonstrates the importance of other viral agents in the development of the disease after the introduction of rotavirus vaccine immunization.

Assessment of Epstein-Barr virus nucleic acids in gastric but not in breast cancer by next-generation sequencing of pooled Mexican samples

Gastric (GC) and breast (BrC) cancer are two of the most common and deadly tumours. Different lines of evidence suggest a possible causative role of viral infections for both GC and BrC. Wide genome sequencing (WGS) technologies allow searching for viral agents in tissues of patients with cancer. These technologies have already contributed to establish virus-cancer associations as well as to discovery new tumour viruses. The objective of this study was to document possible associations of viral infection with GC and BrC in Mexican patients. In order to gain idea about cost effective conditions of experimental sequencing, we first carried out an in silico simulation of WGS. The next-generation-platform IlluminaGallx was then used to sequence GC and BrC tumour samples. While we did not find viral sequences in tissues from BrC patients, multiple reads matching Epstein-Barr virus (EBV) sequences were found in GC tissues. An end-point polymerase chain reaction confirmed an enrichment of EBV sequences in one of the GC samples sequenced, validating the next-generation sequencing-bioinformatics pipeline.

An in-house real-time polymerase chain reaction: standardisation and comparison with the Cobas Amplicor HBV monitor and Cobas AmpliPrep/Cobas TaqMan HBV tests for the quantification of hepatitis B virus DNA

This study aimed to standardise an in-house real-time polymerase chain reaction (rtPCR) to allow quantification of hepatitis B virus (HBV) DNA in serum or plasma samples, and to compare this method with two commercial assays, the Cobas Amplicor HBV monitor and the Cobas AmpliPrep/Cobas TaqMan HBV test. Samples from 397 patients from the state of São Paulo were analysed by all three methods. Fifty-two samples were from patients who were human immunodeficiency virus and hepatitis C virus positive, but HBV negative. Genotypes were characterised, and the viral load was measure in each sample. The in-house rtPCR showed an excellent success rate compared with commercial tests; inter-assay and intra-assay coefficients correlated with commercial tests (r = 0.96 and r = 0.913, p < 0.001) and the in-house test showed no genotype-dependent differences in detection and quantification rates. The in-house assay tested in this study could be used for screening and quantifying HBV DNA in order to monitor patients during therapy.

Association of human papillomavirus and Chlamydia trachomatis with intraepithelial alterations in cervix samples

The influence of different infectious agents and their association with human papillomavirus (HPV) in cervical carcinogenesis have not been completely elucidated. This study describes the association between cytological changes in cervical epithelium and the detection of the most relevant aetiological agents of sexually transmitted diseases. Samples collected from 169 patients were evaluated by conventional cytology followed by molecular analysis to detect HPV DNA, Chlamydia trachomatis, herpes simplex virus 1 and 2,Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis, andTreponema pallidum, besides genotyping for most common high-risk HPV. An association between cytological lesions and different behavioural habits such as smoking and sedentariness was observed. Intraepithelial lesions were also associated with HPV and C. trachomatis detection. An association was also found between both simple and multiple genotype infection and cytological changes. The investigation of HPV and C. trachomatisproved its importance and may be considered in the future for including in screening programs, since these factors are linked to the early diagnosis of patients with precursor lesions of cervical cancer.

Bonafide, type-specific human papillomavirus persistence among HIV-positive pregnant women: predictive value for cytological abnormalities, a longitudinal cohort study

This study investigated the rate of human papillomavirus (HPV) persistence, associated risk factors, and predictors of cytological alteration outcomes in a cohort of human immunodeficiency virus-infected pregnant women over an 18-month period. HPV was typed through L1 gene sequencing in cervical smears collected during gestation and at 12 months after delivery. Outcomes were defined as nonpersistence (clearance of the HPV in the 2nd sample), re-infection (detection of different types of HPV in the 2 samples), and type-specific HPV persistence (the same HPV type found in both samples). An unfavourable cytological outcome was considered when the second exam showed progression to squamous intraepithelial lesion or high squamous intraepithelial lesion. Ninety patients were studied. HPV DNA persistence occurred in 50% of the cases composed of type-specific persistence (30%) or re-infection (20%). A low CD4+T-cell count at entry was a risk factor for type-specific, re-infection, or HPV DNA persistence. The odds ratio (OR) was almost three times higher in the type-specific group when compared with the re-infection group (OR = 2.8; 95% confidence interval: 0.43-22.79). Our findings show that bonafide (type-specific) HPV persistence is a stronger predictor for the development of cytological abnormalities, highlighting the need for HPV typing as opposed to HPV DNA testing in the clinical setting.

DENGUE OUTBREAK IN MATO GROSSO STATE, MIDWESTERN BRAZIL / Surto de dengue em Mato Grosso, centro-oeste do Brasil

Dengue virus (DENV) is the most frequent arbovirus worldwide. In this study, we report a large outbreak in Mato Grosso State (MT). Serum samples from 604 patients with acute febrile illness for less than five days were inoculated in C6/36 cells, then infected cells were subjected to an indirect immunofluorescence test for DENV serotypes and yellow fever virus. Serum samples were submitted to a multiplex-semi-nested-RT-PCR for 11 flaviviruses. DENV-4 was isolated in 150/604 (24.8%) and DENV-1 in 19/604 (3.1%) specimens. By RT-PCR, 331 (54.8%) samples tested positive for DENV; 321 had single infections (DENV-4 n = 305; DENV-1 n = 15; DENV-3 n = 1), nine had co-infections of DENV-1/DENV-4, and one of DENV-2/DENV-4. DENV-4 was detected in 315/331 (95.2%) positive patients from 17 municipalities, and DENV-1 in 24/331 (7.2%) patients from five cities in north-central MT and the city of Cuiaba. The incidence of infection was higher in patients aged 20-39 (142/331; 42.9%). The NS5 partial nucleotide sequence of DENV-1 was most similar to that of genotype V, DENV-2 to Southeast Asian/American, DENV-3 to genotype III, and DENV-4 to genotype II strains, considered the most frequent strains in Brazil. This outbreak coincided with the introduction of DENV-4 in the state. Cuiaba was hyperendemic for the four DENV serotypes, highlighting the necessity for arbovirus surveillance in MT.

O vírus da dengue (DENV) é o arbovirus mais frequente no mundo. Neste estudo, é relatada uma epidemia de grandes proporções no estado de Mato Grosso (MT). Amostras de soro de 604 pacientes com doença febril aguda a menos de 5 dias foram inoculadas em células C6/36 seguida de Imunofluorescência indireta para os sorotipos do DENV e vírus da febre amarela e submetidas a multiplex-semi-nested-RT-PCR para 11 flavivírus. O DENV-4 foi isolado em 150/604 (24,8%) e DENV-1 em 19/604 (3,1%) amostras. Por RT-PCR, 331 (54,8%) pacientes foram positivos para DENV; 321 com infecções únicas (DENV-4 n=305; DENV-1 n=15; DENV-3 n=1), nove co-infecções entre DENV-1/DENV-4 e uma com DENV-2/DENV-4. O DENV-4 foi detectado em 315/331 (95,2%) pacientes de 17 municípios e o DENV-1 em 24/331 (7,2%) pacientes de 5 cidades da região centro-norte de MT e em Cuiabá. A incidência de infecção foi maior em pacientes de 20-39 anos (142/331; 42,9%). As sequências de nucleotídeos de região do gene NS5 do DENV-1 apresentaram maior similaridade com o genótipo V, do DENV-2 com Sudeste Asiático/Americano, DENV-3 com genótipo III e DENV-4 com genótipo II, considerados os mais frequentes no Brasil. Esta epidemia coincidiu com a introdução do DENV-4 no estado. Cuiabá foi considerada hiperendêmica para os quatro sorotipos do DENV, ressaltando a necessidade de vigilância para arbovírus em MT.

Herpes viruses and human papilloma virus in nasal polyposis and controls / Herpesvírus e vírus do papiloma humano na polipose nasal e controles

ABSTRACT INTRODUCTION: Chronic rhinosinusitis with nasal polyps is a multifactorial disease entity with an unclear pathogenesis. Contradictory data exist in the literature on the potential implication of viral elements in adult patients with chronic rhinosinusitis. OBJECTIVE: To compare the prevalence of human herpes viruses (1-6) and Human Papilloma Virus in adult patients with chronic rhinosinusitis with nasal polyps and healthy controls. METHODS: Viral DNA presence was evaluated by real-time polymerase chain reaction application to nasal polyps specimens from 91 chronic rhinosinusitis with nasal polyps patients and nasal turbinate mucosa from 38 healthy controls. RESULTS: Epstein-Barr virus positivity was higher in nasal polyps (24/91; 26.4%) versus controls (4/38; 10.5%), but the difference did not reach significance (p = 0.06). Human herpes virus-6 positivity was lower in nasal polyps (13/91; 14.29%) versus controls (10/38; 26.32%,p = 0.13). In chronic rhinosinusitis with nasal polyps group, 1 sample was herpes simplex virus-1-positive (1/91; 1.1%), and another was cytomegalovirus-positive (1/91; 1.1%), versus none in controls. No sample was positive for herpes simplex virus-2, varicella-zoster virus, high-risk-human papilloma viruses (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59) and low-risk-human papilloma viruses (6, 11). CONCLUSION: Differences in Epstein-Barr virus and human herpes virus-6 positivity among patients with chronic rhinosinusitis with nasal polyps and healthy controls are not statistically significant, weakening the likelihood of their implication in chronic rhinosinusitis with nasal polyps pathogenesis.

RESUMO INTRODUÇÃO: A rinossinusite crônica com pólipos é uma doença multifatorial de etiopatogênese ainda não definida. Existem dados contraditórios na literatura sobre a implicação potencial de elementos virais na etiologia de pólipos nasossinusais. OBJETIVO: Comparar a prevalência de herpes vírus humanos (1-6) e papiloma vírus humano em pacientes adultos com rinossinusite crônica com pólipos nasais (CRwNP) e controles saudáveis. MÉTODO: A presença de DNA viral foi avaliada por PCR em tempo real, em amostras de pólipos nasais de 91 pacientes com CRwNP e na mucosa das conchas nasais de 38 controles saudáveis. RESULTADOS: A positividade do EBV foi maior nos pólipos nasais (24/91; 26,4%) do que nos controles (4/38; 10,5%), mas a diferença não foi significante (p = 0,06). O HHV-6 apresentou positividade menor nos pólipos nasais (13/91; 14,29%) do que os controles (10/38; 26,32%), (p = 0,13). No grupo CRwNP, uma amostra foi positiva para o vírus herpes simples (HSV-1) (1/91; 1,1%), e uma para citomegalovírus (CMV) (1/91; 1,1%); e nenhuma amostra foi positiva no grupo controle. Não houve amostra positiva para HSV-2, VZV, HR-HPV (16,18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59) e LR-HPV (6,11). CONCLUSÃO: Diferenças de positividade do EBV e HHV-6 entre pacientes com CRwNP e controles saudáveis não são estatisticamente significantes, enfraquecendo a probabilidade de sua implicação na patogênese da CRwNP.

Occult hepatitis B virus infection in liver transplant patients in a Brazilian referral center

Estimates of occult hepatitis B virus (HBV) infection prevalence varies among different studies depending on the prevalence of HBV infection in the study population and on the sensitivity of the assay used to detect HBV DNA. We investigated the prevalence of occult HBV infection in cirrhotic patients undergoing liver transplantation in a Brazilian referral center. Frozen liver samples from 68 adults were analyzed using a nested polymerase chain reaction assay for HBV DNA. The specificity of the amplified HBV sequences was confirmed by direct sequencing of the amplicons. The patient population comprised 49 (72.1%) males and 19 (27.9%) females with a median age of 53 years (range=18-67 years). Occult HBV infection was diagnosed in three (4.4%) patients. The etiologies of the underlying chronic liver disease in these cases were alcohol abuse, HBV infection, and cryptogenic cirrhosis. Two of the patients with cryptic HBV infection also presented hepatocellular carcinoma. Markers of previous HBV infection were available in two patients with occult HBV infection and were negative in both. In conclusion, using a sensitive nested polymerase chain reaction assay to detect HBV DNA in frozen liver tissue, we found a low prevalence of occult HBV infection in cirrhotic patients undergoing liver transplant, probably due to the low prevalence of HBV infection in our population.

Prevalence of high-risk human papillomavirus by cobas 4800 HPV test in urban Peru

Background: Molecular tests allow the detection of high-risk human papillomavirus in cervical samples, playing an important role in the prevention of cervical cancer. Objectives: We performed a study to determine the prevalence of HPV 16, HPV 18 and other high-risk human papillomavirus (pool 12 genotypes) in Peruvian females from diverse urban areas using the cobas 4800 HPV test. Methods: Routine cervical samples collected in our laboratory were analyzed by cobas 4800 HPV test. Results: A total of 2247 samples from female patients aged 17–79 years were tested. high-risk human papillomavirus was positive in 775 (34.49%) samples. Of these, 641 (82.71%) were single infections and 134 (17.29%) were multiple infections. The positivity rates for HPV 16, HPV 18, and other high-risk human papillomavirus were 10.77%, 2.0%, and 28.08%, respectively. In multiple high-risk human papillomavirus infections, the concomitance of HPV 16 and other high-risk human papillomavirus was more prevalent (13.42%). Conclusion: Our study showed high prevalence of high-risk human papillomavirus in urban Peru, mainly among young women. In both single and multiple infections other high-risk human papillomavirus were more prevalent than HPV 16 and HPV 18, which might influence vaccine impact in our country. Furthermore, the cobas 4800 HPV test may be considered a useful tool for HPV molecular diagnosis. .

High prevalence of occult hepatitis B virus genotype H infection among children with clinical hepatitis in west Mexico

Studies on the prevalence of infection with hepatitis B virus (HBV) among children are scarce in Latin American countries, especially in Mexico. This study was aimed to investigate the prevalence of HBV infection, occult hepatitis B infection (OBI) and HBV genotypes among children with clinical hepatitis. In total, 215 children with clinical hepatitis were evaluated for HBV infection. HBV serological markers and HBV DNA were analysed. OBI diagnosis and HBV genotyping was performed. HBV infection was found in 11.2% of children with clinical hepatitis. Among these HBV DNA positive-infected children, OBI was identified in 87.5% (n = 21/24) of the cases and 12.5% (n = 3/24) were positive for both HBV DNA and hepatitis B surface antigen. OBI was more frequent among children who had not been vaccinated against hepatitis B (p < 0.05) than in those who had been vaccinated. HBV genotype H was prevalent in 71% of the children followed by genotype G (8%) and genotype A (4%). In conclusion, OBI is common among Mexican children with clinical hepatitis and is associated with HBV genotype H. The results show the importance of the molecular diagnosis of HBV infection in Mexican paediatric patients with clinical hepatitis and emphasise the necessity of reinforcing hepatitis B vaccination in children.

Assessment of immunological changes in Epstein-Barr virus co-infection in Egyptian chronic HCV patients

Epstein-Barr virus (EBV) plays a major role in liver pathology. Similar to other members of the herpesvirus family, EBV establishes a persistent infection in more than 90% of adults. The aim of this study was to evaluate the impact of EBV and chronic hepatitis C co-infection (HCV) on biochemical and immunological responses in patients. The study was conducted in 62 patients and 33 apparently healthy controls. Patients were divided into three groups: group I, consisting of 31 patients with chronic hepatitis C infection (CHC), group II, consisting of eight patients with EBV infection and without HCV infection and group III, consisting of 23 patients with EBV and chronic HCV. The percentage of CD3+ cells, helper CD4+ cells and CD19+ B-cells was measured by flow cytometry. Human interferon-γ (IFN-γ) and interleukin (IL)-15 levels were measured by an ELISA. The levels of liver alanine aminotransferase and aspartate aminotransferase enzymes were higher in EBV/HCV patients compared to that in EBV and HCV mono-infected patients. EBV/HCV patients had significantly reduced percentages of CD3+ and CD4+ cells compared to EBV patients. Serum IFN-γ levels were significantly reduced in EBV/HCV patients (3.86 pg/mL) compared to CHC patients (6.76 pg/mL) and normal controls (4.69 pg/mL). A significant increase in serum IL-15 levels was observed in EBV/HCV patients (67.7 pg/mL) compared to EBV patients (29.3 pg/mL). Taken together, these observations suggest that HCV and EBV co-infection can potentiate immune response dampening in patients.

High incidence of oncogenic HPV genotypes found in women from Southern Brazil

Oncogenic HPV genotypes are strongly associated with premalignant and malignant cervical lesion. The purpose was to determine human papillomavirus (HPV) prevalence and genotypes, and to estimate cervical cancer risk factor associations. Cervical samples were obtained from 251 women seeking gynecological care at the Pelotas School of Medicine Clinic. This is a cross-sectional study. HPV-DNA was amplified by nested-PCR using MY09/11 and GP5/6 primers, and the sequencing was used for genotyping. Sociodemographic and behavioral risk factors were obtained by closed questionnaire, and its relationship to HPV infection prevalence were analyzed. Statistical analyses were performed using SPSS 16.0 software, and differences were considered significant at p < 0.05. As results, the prevalence of HPV infection was 29.9%. The most frequent genotype was HPV-16 (41.3%), followed by HPV-18 (17.3%), and HPV-33 (9.3%). Others nine HPV genotypes were also found. On this population, prevalence of oncogenic HPV genotypes was high, but does not seem to confer relationship with the risk factors investigated. Future investigations in larger populations are necessary, for the proposition of more appropriated monitoring strategies and treatment according to the Brazilian health service reality, as well as patients.

Human papillomavirus risk factors for infection and genotype distribution in aboriginal women from Northern Argentina / Virus papiloma humano (hpv) en mujeres de la etnia Pilagá del nordeste argentino: Factores de riesgo de infección y distribución de tipos virales

The mortality rate for cervical cancer (CC) in Northern Argentina is three times higher than the average for the country (7.8 deaths/100 000 women). We determined the prevalence and genotype distribution of human papillomavirus (HPV) in 227 sexually active women of the native Pilagá community in Formosa, Argentina. We also conducted an HPV-16 variant analysis and studied several community factors that might play a role in viral entry and infection. Endo- and exocervical samples were tested for HPV DNA with MY09/11-PCR or with GP5+/6+-PCR. HPV was detected in 46.7% of the samples and 21 different types were found; the most frequent being HPV-16 (19.4%), -6 and -18 (5.3%), -58 (3.5%) and -31 and -33 (3.1%). In relation to HPV-16 variants, 68.2% were European and 31.8% Asian-American. Among the cofactors analyzed only disposal of human excreta to the open air (P=0.01) was significantly associated with HPV infection. Our prevalence estimates clearly show that Pilagá women are highly exposed to or infected with high risk HPV types and therefore are at a high risk of developing precancerous lesions and eventually CC at the population level.

La tasa de mortalidad por cáncer cervical (CC) en la región norte de la Argentina es tres veces más alta que la media del país (7.8 muertes/100 000 mujeres). En el presente trabajo se determinó la prevalencia de infección por virus papiloma humano (VPH) y la distribución y frecuencia de los genotipos en 227 mujeres sexualmente activas de la etnia aborigen Pilagá (Formosa, Argentina). También se realizó un análisis de las variantes intratípicas de VPH-16 presentes en la comunidad y se analizaron diversos factores socioculturales que podrían tener algún rol destacado en la transmisión de la infección viral. Se estudiaron muestras de células endo-exocervicales mediante PCR basadas en los cebadores MY09/11 y GP5+/6+ con posterior restricción enzimática y/o hibridación dot-blot. La infección por VPH fue detectada en el 46.7% de las mujeres analizadas. Fueron identificados 21 genotipos, de los cuales los más frecuentes fueron HPV-16 (19.4%), -6 y -18 (5.3%), -58 (3.5%) y -31 y -33 (3.1%). Respecto al HPV-16, se encontraron 68.2% de variantes europeas y 31.8% de asiático-americanas. Entre los cofactores analizados, solo la disposición de excretas al aire libre estuvo significativamente asociada con la infección por VPH (P = 0.01). Los datos obtenidos reflejan que la comunidad Pilagá está altamente expuesta a las infecciones por genotipos de alto riesgo de VPH, lo cual puede estar asociado a una alta incidencia de lesiones cervicales preneoplásicas y neoplásicas.