Liver and Inflammatory Biomarkers Are Related to High Mortality in Hospitalized Patients with COVID-19 in Brazilian Amazon Region.

COVID-19 is a multisystem disease with many clinical manifestations, including liver damage and inflammation. The objective of this study is to analyze inflammation biomarkers in relation to the clinical outcome and respiratory symptoms of COVID-19. This is a retrospective cohort of patients with COVID-19 admitted to the Hospital Regional do Baixo Amazonas from 2020 to 2022. Data were collected from electronic medical records from admission to the 30th day of hospitalization and soon after hospital discharge. A total of 397 patients were included in the study. In the longitudinal follow-up of liver markers, a significant difference was found for AST on day 14, with a higher median in the death group. Among the hematological markers, lymphopenia was observed throughout the follow-up, with the death group having the most altered values. When comparing the evolution of biomarkers in the Non-Invasive Ventilation (NIV) and Invasive Mechanical Ventilation (IMV) groups, AST showed a significant difference only on day 14 and GGT on day 1, being greater in the IMV group, and indirect bilirubin on day 7 being more altered in the NIV group. In conclusion, death during hospitalization or a more severe form of COVID-19 was related to significant changes in liver and inflammatory biomarkers.

SARS-CoV-2 and dialysis: humoral response, clinical and laboratory impacts before vaccination.

BACKGROUND: Patients with kidney disease on Hemodialysis (HD) are susceptible to Coronavirus Disease (COVID-19) due to multiple risk factors. AIM: This study aims to report the prevalence of antibodies against SARS-CoV-2 among patients on hemodialysis before vaccination in Brazil and to compare with clinical, demographic, and laboratory data. METHODS: Blood samples from 398 Chronic Kidney Disease (CKD) patients treated in three different private institutions in Rio de Janeiro State, Brazil were submitted to the total anti-SARS-CoV-2 testing. Kidney, liver, and hematological markers were also determined. Respiratory samples were tested by real-time PCR for SARS-CoV-2 RNA and positive samples were subjected to high-throughput sequencing on the MinION device. RESULTS: Overall, anti-SARS-CoV-2 prevalence was 54.5 % (217/398) and two individuals had SARS-CoV-2 RNA with variant B.1.1. High anti-SARS-CoV-2 seroprevalence was found in male gender and those with hospital admission in the last 3-months before the inclusion in the study. Lower red blood cell count was observed in the anti-SARS-CoV-2 seropositive group. High levels of anti-SARS-CoV-2 were found in those who reported symptoms, had low levels of eosinophils and low hematocrit, and who practiced physical activity. CONCLUSION: High prevalence of anti-SARS-CoV-2 was found in CKD patients before the universal immunization in Brazil suggesting that dialysis patients were highly exposed to SARS-CoV-2.

SARS-CoV-2 and dialysis: humoral response, clinical and laboratory impacts before vaccination

Abstract Background Patients with kidney disease on Hemodialysis (HD) are susceptible to Coronavirus Disease (COVID-19) due to multiple risk factors. Aim This study aims to report the prevalence of antibodies against SARS-CoV-2 among patients on hemodialysis before vaccination in Brazil and to compare with clinical, demographic, and laboratory data. Methods Blood samples from 398 Chronic Kidney Disease (CKD) patients treated in three different private institutions in Rio de Janeiro State, Brazil were submitted to the total anti-SARS-CoV-2 testing. Kidney, liver, and hematological markers were also determined. Respiratory samples were tested by real-time PCR for SARS-CoV-2 RNA and positive samples were subjected to high-throughput sequencing on the MinION device. Results Overall, anti-SARS-CoV-2 prevalence was 54.5 % (217/398) and two individuals had SARS-CoV-2 RNA with variant B.1.1. High anti-SARS-CoV-2 seroprevalence was found in male gender and those with hospital admission in the last 3-months before the inclusion in the study. Lower red blood cell count was observed in the anti-SARS-CoV-2 seropositive group. High levels of anti-SARS-CoV-2 were found in those who reported symptoms, had low levels of eosinophils and low hematocrit, and who practiced physical activity. Conclusion High prevalence of anti-SARS-CoV-2 was found in CKD patients before the universal immunization in Brazil suggesting that dialysis patients were highly exposed to SARS-CoV-2.

Seroprevalence of anti-SARS-CoV-2 in patients with hepatitis B and C: a pre-vaccination study

Abstract The serological markers for the diagnosis of COVID-19 plays an important role in the epidemiological investigation of the pandemic. This study aims to assess the prevalence of anti-SARS-CoV-2 in hepatitis B and C patients in a pre-vaccination of COVID-19 period. Between March 2020 and January 2021, 199 serum samples from individuals with HBsAg/HBV DNA or anti-HCV/HCV RNA positivity were tested for antibodies (IgM and IgG) against SARS-CoV-2 using Electrochemiluminescent Immunoassay (ECLIA). Among these, 50.3 % (100/199) tested positive for hepatitis C virus infection and 49.7 % (99/199) for hepatitis B virus, confirmed through molecular and serological diagnosis. The anti-SARS-CoV-2 seroprevalence was 24.1 % (48/199) in this population, with 23.23 % (23/99) hepatitis B and 25 % (25/100) hepatitis C patients tested positive for anti-SARS-CoV-2. The higher seroprevalence of anti-SARS-CoV-2 (16.58 %, 33/199) was detected among those over-40 years of age and the month of November 2020 had the highest number of detections 9 % (18/199) with the majority living in impoverished and neglected neighborhoods in the city of Rio de Janeiro. We found a high prevalence of anti-SARS-CoV-2 in patients with viral hepatitis before COVID-19 vaccination. This demonstrates the high exposure of this population during the period of social isolation.

Seroprevalence of antibodies against herpesvirus type 2 in a female prison population in Mato Grosso / Soroprevalência de anticorpos contra herpesvírus tipo 2 em população prisional feminina mato-grossense

Descrever a soroprevalência de anticorpos contra herpes vírus simples 2 em reeducandas de uma cadeia pública feminina de Mato Grosso no ano de 2016. Trata-se de um estudo transversal com abordagem quantitativa, realizado com 50 reeducandas reclusas de uma cadeia pública feminina de Mato Grosso. A coleta de dados foi realizada por meio de entrevista com 50 mulheres. Para determinar a soroprevalência da infecção por HSV-2, foram analisadas amostras de soro pelo método ELISA em busca de anticorpos do tipo IgG no Laboratório de Imunologia Viral do Instituto Oswaldo Cruz ­ RJ. A soroprevalência de HSV-2 encontrada na população avaliada foi de 80%, valor muito superior ao relatado na população geral brasileira e em outras estudos com populações prisionais em todo o mundo. O perfil das reeducandas predominou entre mulheres jovens, pardas, com baixa escolaridade, solteiras e com renda mensal baixa. O presente estudo encontrou alta soroprevalência de anticorpos contra HSV-2 nesta população. Esses dados fornecem importantes informações que podem auxiliar na implementação de ações efetivas que melhor previnam e controlem a herpes genital, bem como as demais ISTs em populações encarceradas.

High prevalence of hepatitis A and E viruses in environmental and clinical samples from West Argentina

ABSTRACT Environmental surveillance of water sources is important to monitoring viral hepatitis transmission in clinical settings. This study investigated the circulation of hepatitis A (HAV) and E (HEV) viruses in sewage and clinical samples from Argentina. Between 2016 and 2017, 80 raw sewage samples and 86 clinical samples (stool and serum) from suspected cases of hepatitis A and hepatitis E were obtained. HAV and HEV were tested by both real-time and nested PCR. Positive samples were sequenced for genotype determination and phylogenetic analysis. Overall, HAV was recovered in 39% of sewage samples and 61.1% of clinical samples. HEV was detected in 22.5% of sewage samples and 15.9% of clinical samples. HAV was found more frequently in sewage during the winter and in clinical samples in spring; HEV was more prevalent in sewage during summer and in clinical samples in autumn. All HAV isolates belonged to genotype IA and HEV isolates belonged to genotype 3, the most prevalent genotypes in South America. High prevalence of HAV and HEV in environmental and clinical samples in Mendoza, Argentina was observed. These findings reinforce the importance of environmental surveillance and implementation of health strategies to control the spread of HAV and HEV in developing countries.

Evaluation of herpesvirus members on hospital admission in patients with systemic lupus erythematous shows higher frequency of Epstein-Barr virus and its associated renal dysfunction / Avaliação de membros da família Herpesviridae na admissão hospitalar de pacientes com lúpus eritematoso sistêmico mostra maior frequência do vírus Epstein-Barr e sua disfunção renal associada

Abstract Introduction: Members of the Herpesviridae family have been described in patients with systemic lupus erythematous (SLE), but the clinical impact on renal function is not well known. Methods: HSV1, HSV2, VZV, EBV, CMV, HHV-6, HHV-7, and HHV-8 were evaluated by molecular biology on admission in blood samples from 40 consecutive SLE patients hospitalized for lupus activity. Results: Patients were 90.0% female, 77.5% non-white, with average age of 32.7 ± 13.6 years. We found positivity for EBV (65.0%), CMV (30.0%), HSV-1 (30.0%), HHV-6 (12.5%), and HHV-7 (7.5%). For all viruses, age, SLEDAI, hematological tests, ferritin, LDH, C-reactive protein, and erythrocyte sedimentation rate (ESR) were not significant. However, EBV positivity was a significant factor for higher serum creatinine (3.0 ± 2.8 vs. 0.9 ± 0.8; P = 0.001) and urea (86 ± 51 vs. 50 ± 46; P = 0.03). Moreover, positive cases for EBV only or with combined co-infections (66.7%-CMV; 58.3%-HSV-1) or negative for EBV only were evaluated by Kruskal-Wallis test again showed statistical significance for serum creatinine and urea (both P ≤ 0.01), with posttest also showing statistical differences for renal dysfunction and EBV presence (alone or in combined co-infections). The presence of EBV viral load was also significant for nephrotic-range proteinuria, renal flare, and the need for hemodialysis. Conclusion: Members of the Herpeviridae family (mainly EBV, HSV-1 and CMV) are common on hospital admission of SLE patients, reaching 65% for EBV, which seems to be associated with renal dysfunction and could reflect a previous association or overlapping disease, which is not well understood.

Resumo Introdução: Membros da família Herpesviridae tem sido descritos em pacientes com lúpus eritematoso sistêmico (LES), mas o impacto clínico na função renal não é bem conhecido. Métodos: Avaliou-se HSV1, HSV2, VZV, EBV, CMV, HHV-6, HHV-7, HHV-8 por biologia molecular na admissão em amostras sanguíneas de 40 pacientes com LES consecutivos hospitalizados por atividade lúpica. Resultados: Pacientes 90,0% mulheres, 77,5% não brancos, idade média 32,7 ± 13,6 anos. Encontramos positividade para EBV (65,0%), CMV (30,0%), HSV-1 (30,0%), HHV-6 (12,5%), HHV-7 (7,5%). Para todos os vírus, idade, SLEDAI, exames hematológicos, ferritina, LDH, proteína C reativa, velocidade de hemossedimentação não foram significativos. Entretanto, positividade para EBV foi estatisticamente significativo para creatinina (3,0 ± 2,8 vs. 0,9 ± 0,8; P = 0,001) e ureia (86 ± 51 vs. 50 ± 46; P = 0,03) séricas mais elevadas. Ademais, casos positivos para EBV isolado ou com coinfecções combinadas (66,7%-CMV; 58,3%-HSV-1) ou negativos apenas para EBV foram avaliados pelo teste Kruskal-Wallis e novamente mostraram significância estatística para creatinina e ureia séricas (ambas P ≤ 0,01), com pós-teste mostrando também diferenças estatísticas para disfunção renal e presença de EBV (sozinho ou em coinfecções combinadas). A presença de carga viral do EBV também foi significativa para proteinúria de faixa nefrótica, inflamação aguda, necessidade de hemodiálise. Conclusão: Membros da família Herpeviridae (principalmente EBV, HSV-1, CMV) são comuns na admissão de pacientes com LES, chegando a 65% para EBV, que parece associar-se à disfunção renal podendo refletir associação prévia ou doença sobreposta, o que não é bem compreendido.

Hepatitis A and E among immigrants and refugees in Central Brazil.

OBJECTIVE: To estimate the prevalence of hepatitis A virus (HAV) and hepatitis E virus (HEV) among immigrants and refugees in Goiás, Central Brazil. METHODS: Overall, 355 individuals were interviewed, and blood samples were tested for anti-HAV and anti-HEV IgG. Anti-HEV-positive samples were similarly tested for HEV RNA. RESULTS: All participants were from Latin American countries, most of whom, young adult males. The overall anti-HAV IgG prevalence was 87.4% (95%CI: 83.5-90.4), of whom 94.9%, 75.6%, and 60% were from Haiti, Venezuela, and other Latin American countries, respectively (p < 0.001). Age above 19 years and more than 36 months residing in Brazil were associated with a higher prevalence of previous HAV and HEV infection, respectively. Of the children eligible for HAV vaccination according to the National Immunization Program, only eight (44%) had been vaccinated. The overall anti-HEV IgG prevalence was 6.5% (95%CI: 4.4-9.5). All anti-HEV IgG-positive individuals were Haitians, including a child born in Brazil. HEV RNA was detected in two of the anti-HEV IgG-positive samples. CONCLUSION: The survey detected a high prevalence of anti-HAV and anti-HEV IgG among immigrants and refugees, and active HEV infection among some Haitian participants. Prevention measures are urgently required to interrupt enteric virus transmission in this emergent and vulnerable population.

Evaluation of herpesvirus members on hospital admission in patients with systemic lupus erythematous shows higher frequency of Epstein-Barr virus and its associated renal dysfunction.

INTRODUCTION: Members of the Herpesviridae family have been described in patients with systemic lupus erythematous (SLE), but the clinical impact on renal function is not well known. METHODS: HSV1, HSV2, VZV, EBV, CMV, HHV-6, HHV-7, and HHV-8 were evaluated by molecular biology on admission in blood samples from 40 consecutive SLE patients hospitalized for lupus activity. RESULTS: Patients were 90.0% female, 77.5% non-white, with average age of 32.7 ± 13.6 years. We found positivity for EBV (65.0%), CMV (30.0%), HSV-1 (30.0%), HHV-6 (12.5%), and HHV-7 (7.5%). For all viruses, age, SLEDAI, hematological tests, ferritin, LDH, C-reactive protein, and erythrocyte sedimentation rate (ESR) were not significant. However, EBV positivity was a significant factor for higher serum creatinine (3.0 ± 2.8 vs. 0.9 ± 0.8; P = 0.001) and urea (86 ± 51 vs. 50 ± 46; P = 0.03). Moreover, positive cases for EBV only or with combined co-infections (66.7%-CMV; 58.3%-HSV-1) or negative for EBV only were evaluated by Kruskal-Wallis test again showed statistical significance for serum creatinine and urea (both P ≤ 0.01), with posttest also showing statistical differences for renal dysfunction and EBV presence (alone or in combined co-infections). The presence of EBV viral load was also significant for nephrotic-range proteinuria, renal flare, and the need for hemodialysis. CONCLUSION: Members of the Herpeviridae family (mainly EBV, HSV-1 and CMV) are common on hospital admission of SLE patients, reaching 65% for EBV, which seems to be associated with renal dysfunction and could reflect a previous association or overlapping disease, which is not well understood.

Hepatitis A and E among immigrants and refugees in Central Brazil

ABSTRACT OBJECTIVE To estimate the prevalence of hepatitis A virus (HAV) and hepatitis E virus (HEV) among immigrants and refugees in Goiás, Central Brazil. METHODS Overall, 355 individuals were interviewed, and blood samples were tested for anti-HAV and anti-HEV IgG. Anti-HEV-positive samples were similarly tested for HEV RNA. RESULTS All participants were from Latin American countries, most of whom, young adult males. The overall anti-HAV IgG prevalence was 87.4% (95%CI: 83.5-90.4), of whom 94.9%, 75.6%, and 60% were from Haiti, Venezuela, and other Latin American countries, respectively (p < 0.001). Age above 19 years and more than 36 months residing in Brazil were associated with a higher prevalence of previous HAV and HEV infection, respectively. Of the children eligible for HAV vaccination according to the National Immunization Program, only eight (44%) had been vaccinated. The overall anti-HEV IgG prevalence was 6.5% (95%CI: 4.4-9.5). All anti-HEV IgG-positive individuals were Haitians, including a child born in Brazil. HEV RNA was detected in two of the anti-HEV IgG-positive samples. CONCLUSION The survey detected a high prevalence of anti-HAV and anti-HEV IgG among immigrants and refugees, and active HEV infection among some Haitian participants. Prevention measures are urgently required to interrupt enteric virus transmission in this emergent and vulnerable population.

Introduction of human gammaherpesvirus 8 genotypes A, B, and C into Brazil from multiple geographic regions

Variations in the open reading frame (ORF) K1 gene sequence of human gammaherpesvirus 8 (HHV-8) has led to the identification of 6 major genotypic clades (A, B, C, D, E, and F) in specimens isolated from around the world. These clades exhibit clear clustering among individuals in different ethnic groups and from different geographic regions. The human population of Brazil varies greatly in ethnicity because of multiple immigration events from Africa, Europe, Asia, and indigenous communities. However, there is scant information about the HHV-8 genotypes currently circulating in Brazil. Here, we describe HHV-8 genotypic diversity in isolates from Brazilian HIV-infected patients living with Kaposi's sarcoma (KS) by analysis of the complete ORF-K1 region. We also identified the most likely geographic origins of these different Brazilian genotypes. We extracted HHV-8 DNA (24 positive samples) from individuals with HIV/KS from the states of São Paulo and Rio de Janeiro, amplified the ORF-K1 gene using nested PCR (about 870 base pairs), performed sequencing and phylogenetic analysis, and then calculated the mean genetic distances of Brazilian sequences from sequences in other regions of the world (523 sequences analyzed). Phylogenetic analysis showed that genotypes C, A, and B were present in 45.8 %, 29.2 % and 25 % of the isolates from Brazil, respectively. These isolates grouped into separate clades, rather than a single monophyletic cluster. Mean genetic distance analyses suggested that these genotypes were introduced into the Brazil multiple times from different geographical regions. HHV-8/A isolates appear to be from Ukraine, Russia, and the Tartar ethnic group; HHV-8/B isolates appear to be from Congo and Democratic Republic of the Congo; and HHV-8/C isolates appear to be from Australia, Algeria, England, and French Guiana. These results contribute to a better understanding of the genetic diversity and origins of HHV-8 strains circulating in Brazil, and will provide a foundation for further epidemiological and evolutionary studies of HHV-8

Novel variants of human herpesvirus 2 from Brazilian HIV-1 coinfected subjects.

BACKGROUND Human herpesvirus 2 (HHV-2) have DNA genome with a limited genetic variability and have been classified into two clades. OBJECTIVES To identify and characterise six HHV-2 isolates derived from Brazilian women. METHODS HHV-2 isolates were performed polymerase chain reaction (PCR) and sequencing of 2250 pb of the glycoprotein B (gB) coding regions. FINDINGS Four HHV-2 isolates were classified into clade B, while the remaining two, derived from HIV-1 co-infected women, showed a notable genetic divergence (> 1%). MAIN CONCLUSION The results reveal novel HHV-2 variants. The impact of these novel variants on HHV-2 pathogenesis and HIV/HHV-2 coinfection need to be investigated.

Diagnosis of human herpes virus 1 and 2 (HHV-1 and HHV-2): use of a synthetic standard curve for absolute quantification by real time polymerase chain reaction

The use of quantitative real time polymerase chain reaction (qPCR) for herpesvirus detection has improved the sensitivity and specificity of diagnosis, as it is able to detect shedding episodes in the absence of clinical lesions and diagnose clinical specimens that have low viral loads. With an aim to improve the detection and quantification of herpesvirus by qPCR, synthetic standard curves for human herpesvirus 1 and 2 (HHV-1 and HHV-2) targeting regions gD and gG, respectively, were designed and evaluated. The results show that synthetic curves can replace DNA standard curves in diagnostic herpes qPCR.

Poor sensitivity of rapid tests for the detection of antibodies to the hepatitis B virus: implications for field studies

Rapid tests (RTs) can be used as an alternative method for the conventional diagnosis of hepatitis B virus (HBV). This study aims to evaluate antibodies to HBsAg (anti-HBs) and antibodies to HBeAg (anti-HBe) RTs under different Brazilian settings. The following three groups were included: GI: viral hepatitis outpatient services; GII: low resource areas; and GIII: crack users and beauticians. Imuno-rápido anti-HBsAg™ and Imuno-rápido anti-HBeAg™ RTs were evaluated and showed specificities greater than 95% in all groups. The sensitivity values to anti-HBs were 50.38%, 51.05% and 46.73% and the sensitivity values to anti-HBe were 76.99%, 10.34% and 11.76% in the GI, GII and GIII groups, respectively. The assays had a low sensitivity and high specificity, which indicated their use for screening in regions endemic for HBV.

Poor sensitivity of rapid tests for the detection of antibodies to the hepatitis B virus: implications for field studies.

Rapid tests (RTs) can be used as an alternative method for the conventional diagnosis of hepatitis B virus (HBV). This study aims to evaluate antibodies to HBsAg (anti-HBs) and antibodies to HBeAg (anti-HBe) RTs under different Brazilian settings. The following three groups were included: GI: viral hepatitis outpatient services; GII: low resource areas; and GIII: crack users and beauticians. Imuno-rápido anti-HBsAg™ and Imuno-rápido anti-HBeAg™ RTs were evaluated and showed specificities greater than 95% in all groups. The sensitivity values to anti-HBs were 50.38%, 51.05% and 46.73% and the sensitivity values to anti-HBe were 76.99%, 10.34% and 11.76% in the GI, GII and GIII groups, respectively. The assays had a low sensitivity and high specificity, which indicated their use for screening in regions endemic for HBV.

Diagnosis of human herpes virus 1 and 2 (HHV-1 and HHV-2): use of a synthetic standard curve for absolute quantification by real time polymerase chain reaction.

The use of quantitative real time polymerase chain reaction (qPCR) for herpesvirus detection has improved the sensitivity and specificity of diagnosis, as it is able to detect shedding episodes in the absence of clinical lesions and diagnose clinical specimens that have low viral loads. With an aim to improve the detection and quantification of herpesvirus by qPCR, synthetic standard curves for human herpesvirus 1 and 2 (HHV-1 and HHV-2) targeting regions gD and gG, respectively, were designed and evaluated. The results show that synthetic curves can replace DNA standard curves in diagnostic herpes qPCR.

RNA interference inhibits herpes simplex virus type 1 isolated from saliva samples and mucocutaneous lesions.

The aim of this study was to evaluate the use of RNA interference to inhibit herpes simplex virus type-1 replication in vitro. For herpes simplex virus type-1 gene silencing, three different small interfering RNAs (siRNAs) targeting the herpes simplex virus type-1 UL39 gene (sequence si-UL 39-1, si-UL 39-2, and si-UL 39-3) were used, which encode the large subunit of ribonucleotide reductase, an essential enzyme for DNA synthesis. Herpes simplex virus type-1 was isolated from saliva samples and mucocutaneous lesions from infected patients. All mucocutaneous lesions' samples were positive for herpes simplex virus type-1 by real-time PCR and by virus isolation; all herpes simplex virus type-1 from saliva samples were positive by real-time PCR and 50% were positive by virus isolation. The levels of herpes simplex virus type-1 DNA remaining after siRNA treatment were assessed by real-time PCR, whose results demonstrated that the effect of siRNAs on gene expression depends on siRNA concentration. The three siRNA sequences used were able to inhibit viral replication, assessed by real-time PCR and plaque assays and among them, the sequence si-UL 39-1 was the most effective. This sequence inhibited 99% of herpes simplex virus type-1 replication. The results demonstrate that silencing herpes simplex virus type-1 UL39 expression by siRNAs effectively inhibits herpes simplex virus type-1 replication, suggesting that siRNA based antiviral strategy may be a potential therapeutic alternative.

Detection of replication-defective hepatitis A virus based on the correlation between real-time polymerase chain reaction and ELISA in situ results.

ELISA in situ can be used to titrate hepatitis A virus (HAV) particles and real-time polymerase chain reaction (RT-PCR) has been shown to be a fast method to quantify the HAV genome. Precise quantification of viral concentration is necessary to distinguish between infectious and non-infectious particles. The purpose of this study was to compare cell culture and RT-PCR quantification results and determine whether HAV genome quantification can be correlated with infectivity. For this purpose, three stocks of undiluted, five-fold diluted and 10-fold diluted HAV were prepared to inoculate cells in a 96-well plate. Monolayers were then incubated for seven, 10 and 14 days and the correlation between the ELISA in situ and RT-PCR results was evaluated. At 10 days post-incubation, the highest viral load was observed in all stocks of HAV via RT-PCR (10(5) copies/mL) (p = 0.0002), while ELISA revealed the highest quantity of particles after 14 days (optical density = 0.24, p < 0.001). At seven days post-infection, there was a significant statistical correlation between the results of the two methods, indicating equivalents titres of particles and HAV genome during this period of infection. The results reported here indicate that the duration of growth of HAV in cell culture must be taken into account to correlate genome quantification with infectivity.

Detection of replication-defective hepatitis A virus based on the correlation between real-time polymerase chain reaction and ELISA in situ results

ELISA in situ can be used to titrate hepatitis A virus (HAV) particles and real-time polymerase chain reaction (RT-PCR) has been shown to be a fast method to quantify the HAV genome. Precise quantification of viral concentration is necessary to distinguish between infectious and non-infectious particles. The purpose of this study was to compare cell culture and RT-PCR quantification results and determine whether HAV genome quantification can be correlated with infectivity. For this purpose, three stocks of undiluted, five-fold diluted and 10-fold diluted HAV were prepared to inoculate cells in a 96-well plate. Monolayers were then incubated for seven, 10 and 14 days and the correlation between the ELISA in situ and RT-PCR results was evaluated. At 10 days post-incubation, the highest viral load was observed in all stocks of HAV via RT-PCR (10(5) copies/mL) (p = 0.0002), while ELISA revealed the highest quantity of particles after 14 days (optical density = 0.24, p < 0.001). At seven days post-infection, there was a significant statistical correlation between the results of the two methods, indicating equivalents titres of particles and HAV genome during this period of infection. The results reported here indicate that the duration of growth of HAV in cell culture must be taken into account to correlate genome quantification with infectivity.

An immunoenzymatic assay for the diagnosis of hepatitis A utilising immunoglobulin Y.

The detection of anti-hepatitis A virus (HAV) antibody levels by diagnostic kits in the convalescent period of disease generally use immunoglobulin G (IgG), which is expensive. An alternative to IgG is immunoglobulin Y (IgY), an immunoglobulin antibody encountered in birds and reptiles. The aim of this study was to develop a competitive immunoenzymatic assay to measure total anti-HAV antibody levels using anti-HAV IgY as the capture and conjugated immunoglobulins. For this purpose, anti-HAV IgY was conjugated to horseradish peroxidase (HRP) and the optimal dilution of HRP-conjugated antibodies was evaluated to establish the competitive immuneenzymatic assay. The results obtained from our "in-house" assay were plotted on a receiver operator curve, which showed a sensitivity of 95% and a specificity of 98.8%, demonstrating that a competitive anti-HAV IgY immunoenzymatic assay developed "in house" could be used as an alternative to commercial assays that utilise IgG.