The Risk of Reinfection or Primary Hepatitis E Virus Infection at a Liver Transplant Center in Brazil: An Observational Cohort Study.

The hepatitis E virus is a major etiological agent of chronic hepatitis in immunosuppressed individuals. Seroprevalence in the liver transplantation setting varies according to the seroprevalence of the general population in different countries. This was a prospective cohort study of liver transplant recipients in southeastern Brazil. Recipients were systematically followed for one year, with the objective of determining the prevalence, incidence, and natural history of HEV infection in this population. We included 107 liver transplant recipients and 83 deceased donors. Positivity for anti-HEV IgG was detected in 10.2% of the recipients and in 9.7% of the donors. None of the patients tested positive for HEV RNA at baseline or during follow-up. There were no episodes of reactivation or seroconversion, even in cases of serological donor-recipient mismatch or in recipients with acute hepatitis. Acute and chronic HEV infections seem to be rare events in the region studied. That could be attributable to social, economic, and environmental factors. Our data indicate that, among liver transplant recipients, hepatitis E should be investigated only when there are elevated levels of transaminases with no defined cause, as part of the differential diagnosis of seronegative hepatitis after transplantation.

Reduction of SARS-CoV-2 viral load in saliva after rinsing with mouthwashes containing cetylpyridinium chloride: a randomized clinical study.

Background: Symptomatic patients with COVID-19 typically have a high SARS-CoV-2 viral load in their saliva. Procedures to reduce the viral load in their oral cavity are important for mitigating the viral transmission. Methods: This randomized clinical trial investigated the impact of two mouthwashes (0.075% cetylpyridinium chloride plus 0.28% zinc lactate (CPC+Zn) (n = 32), and 0.075% cetylpyridinium chloride (CPC) (n = 31)) on the viral load of SARS-CoV-2 in saliva when compared to the distilled water negative control (n = 32). Saliva was collected before (T0) and after (5 min, T1; 30 min, T2; and 60 min, T3) the intervention. Viral load in saliva was measured by qRT-PCR assays. The data in both groups was normalized for T0 and Negative Control, resulting in fold change values. Results: CPC+Zn oral solution reduced the viral load in saliva by 6.34-fold at T1, 3.6-fold at T2 and 1.9-fold at T3. Rinsing with the CPC mouthwash reduced the viral load in saliva by 2.5-fold at T1, 1.9-fold at T2 and 2.0-fold at T3. Conclusion: CPC+Zn mouthwash or with the CPC mouthwash reduced the viral load in saliva of COVID-19 patients immediately after rinsing. These reductions extended up to 60 min.

Detection of Hepatitis E Virus Genotype 3 in Feces of Capybaras (Hydrochoeris hydrochaeris) in Brazil.

Hepatitis E virus (HEV) is an emerging zoonotic pathogen associated with relevant public health issues. The aim of this study was to investigate HEV presence in free-living capybaras inhabiting urban parks in São Paulo state, Brazil. Molecular characterization of HEV positive samples was undertaken to elucidate the genetic diversity of the virus in these animals. A total of 337 fecal samples were screened for HEV using RT-qPCR and further confirmed by conventional nested RT-PCR. HEV genotype and subtype were determined using Sanger and next-generation sequencing. HEV was detected in one specimen (0.3%) and assigned as HEV-3f. The IAL-HEV_921 HEV-3f strain showed a close relationship to European swine, wild boar and human strains (90.7-93.2% nt), suggesting an interspecies transmission. Molecular epidemiology of HEV is poorly investigated in Brazil; subtype 3f has been reported in swine. This is the first report of HEV detected in capybara stool samples worldwide.

Testicular pathology in fatal COVID-19: A descriptive autopsy study.

BACKGROUND: Multi-organ damage is a common feature of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, going beyond the initially observed severe pneumonia. Evidence that the testis is also compromised is growing. OBJECTIVE: To describe the pathological findings in testes from fatal cases of COVID-19, including the detection of viral particles and antigens, and inflammatory cell subsets. MATERIALS AND METHODS: Postmortem testicular samples were obtained by percutaneous puncture from 11 deceased men and examined by reverse-transcription polymerase chain reaction (RT-PCR) for RNA detection and by light and electron microscopy (EM) for SARS-CoV-2. Immunohistochemistry (IHC) for the SARS-CoV-2 N-protein and lymphocytic and histiocytic markers was also performed. RESULTS: Eight patients had mild interstitial orchitis, composed mainly of CD68+ and TCD8+ cells. Fibrin thrombi were detected in five cases. All cases presented congestion, interstitial edema, thickening of the tubular basal membrane, decreased Leydig and Sertoli cells with reduced spermatogenesis, and strong expression of vascular cell adhesion molecule (VCAM) in vessels. IHC detected SARS-Cov-2 antigen in Leydig cells, Sertoli cells, spermatogonia, and fibroblasts in all cases. EM detected viral particles in the cytoplasm of fibroblasts, endothelium, Sertoli and Leydig cells, spermatids, and epithelial cells of the rete testis in four cases, while RT-PCR detected SARS-CoV-2 RNA in three cases. DISCUSSION AND CONCLUSION: The COVID-19-associated testicular lesion revealed a combination of orchitis, vascular changes, basal membrane thickening, Leydig and Sertoli cell scarcity, and reduced spermatogenesis associated with SARS-CoV-2 local infection that may impair hormonal function and fertility in men.

COVID-19 convalescent plasma cohort study: Evaluation of the association between both donor and recipient neutralizing antibody titers and patient outcomes.

BACKGROUND: Current evidence regarding COVID-19 convalescent plasma (CCP) transfusion practices is limited and heterogeneous. We aimed to determine the impact of the use of CCP transfusion in patients with previous circulating neutralizing antibodies (nAbs) in COVID-19. METHODS: Prospective cohort including 102 patients with COVID-19 transfused with ABO compatible CCP on days 0-2 after enrollment. Clinical status of patients was assessed using the adapted World Health Organization (WHO) ordinal scale on days 0, 5, and 14. The nAbs titration was performed using the cytopathic effect-based virus neutralization test with SARS-CoV-2 (GenBank MT126808.1). The primary outcome was clinical improvement on day 14, defined as a reduction of at least two points on the adapted WHO ordinal scale. Secondary outcomes were the number of intensive care unit (ICU)-free days and the number of invasive mechanical ventilation-free days. RESULTS: Both nAbs of CCP units transfused (p < 0.001) and nAbs of patients before CCP transfusions (p = 0.028) were associated with clinical improvements by day 14. No significant associations between nAbs of patients or CCP units transfused were observed in the number of ICU or mechanical ventilation-free days. Administration of CCP units after 10 days of symptom onset resulted in a decrease in ICU-free days (p < 0.001) and mechanical ventilation-free days (p < 0.001). CONCLUSION: Transfusion of high titer nAbs CCP units may be a determinant in clinical strategies against COVID-19. We consider these data as useful parameters to guide future CCP transfusion practices.

Anti-A and SARS-CoV-2: an intriguing association.

BACKGROUND: Blood groups and anti-A isohemagglutinin may be involved in susceptibility to SARS-CoV-2 infection. MATERIALS AND METHODS: We retrospectively studied 268 COVID-19 convalescent plasma donors and 162 COVID-19 inpatients (total 430 subjects, confirmed by RT-PCR) and 2,212 healthy volunteer first-time blood donors as a control group. These were further divided into two groups: those with anti-A (blood types O and B) and those without it (types A and AB). Titres of nucleoproteins, and neutralizing SARS-CoV-2 antibody were measured in the convalescent plasma donors and inpatients. Multivariate logistic regression and non-parametric tests were applied. RESULTS: Persons having types O or B showed less infection prevalence than those of types A or AB (OR = 0·62, 95% CI 0·50-0·78; P < 0·001), but there was no difference when COVID-19 inpatients were analysed. Immunoglobulins M, G and A were lower in COVID-19 subjects of types O or B group than those of A or AB (0·16 vs. 0·19; P = 0·03, 2·11 vs. 2·55; P = 0·02, 0·23 vs. 0·32; P = 0·03, respectively). CONCLUSION: In this retrospective cohort, COVID-19 individuals were less likely to belong to blood types O and B, and also had lower SARS-CoV-2 antibody titres than A and AB individuals. COVID-19 severity did not associate with the blood groups.

Human T-cell leukemia virus type 1 infection among Japanese immigrants and their descendants living in Southeast Brazil: A call for preventive and control responses.

Human T-cell leukemia virus type 1 (HTLV-1) has worldwide distribution and is considered endemic in southwestern Japan. HTLV-1 infection has been associated with adult T-cell leukemia/lymphoma (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) besides other diseases. This cross-sectional study aimed to investigate the prevalence, risk factors and molecular characterization of HTLV-1, among the world's largest population of Japanese immigrants and their descendants outside of Japan, in São Paulo, Southeast Brazil, as well as to analyze the phylogenetic relationship among isolates of HTLV-1. From July to December 2017, 2,139 individuals from five Japanese associations were interviewed and submitted to blood collection. All serum samples were first tested for the presence of anti-HTLV-1/2 antibodies by ELISA and then peripheral blood from individuals with positive serological results were analyzed for the presence of HTLV-1 5'LTR proviral DNA. Partial sequencing of the 5'LTR region of HTLV-1 proviral DNA was performed by Sanger. The prevalence of HTLV-1 infection was 5.1% (CI 95%: 4.2-6.0). In the multiple logistic regression model, HTLV-1 infection was associated with age ≥ 45 years, female sex, being first and second-generation Japanese immigrants, and having sexual partners with history of blood transfusion. The phylogenetic analysis revealed that all HTLV-1 were classified as Cosmopolitan (1a) subtype. Of them, 47.8% were classified as Transcontinental (A) subgroup and 52.2% as belonging to the Japanese (B) subgroup. Although most HTLV-1-infected patients were asymptomatic (97.3%), blurred vision was associated with HTLV-1 infection. The high prevalence of HTLV-1 infection found in this studied population and especially the intra- and interfamily HTLV-1 transmission presents an urgent call for preventive and control responses of this infection in Brazil.

The Molecular Characterization of Hepatitis A Virus Strains Circulating during Hepatitis A Outbreaks in São Paulo, Brazil, from September 2017 to May 2019.

Outbreaks of hepatitis A may occur in countries of medium and high socioeconomic levels in which the population generally exhibits an increased susceptibility in young adults to this infection if they are not vaccinated against the hepatitis A virus (HAV). In Europe, an outbreak involved approximately 22 European countries with 4475 cases reported from 2016 to 2018; most of them were men who have sex with men (MSM). This outbreak expanded to North and South America, including Brazil, particularly in São Paulo city with 1547 reported cases from 2016 to 2019. In the present study, we characterized the HAV strains involved in the acute hepatitis A cases identified in the reference centers of São Paulo city during this outbreak. A total of 51 cases with positive anti-HAV IgM were included, 80.4% male, 68.6% of them between 20 and 40 years old and 41.7% MSM. HAV RNA was detected in 92% (47/51) of the cases. Subgenotype IA of HAV was identified and most of the strains were closely related to that isolated in outbreaks that occurred in different European countries in 2016. These results showed the epidemiological relation between these outbreaks and reinforce the need to implement vaccination against hepatitis A for the adult population, particularly for a population with a high-risk behavior.

Pulmonary and systemic involvement in COVID-19 patients assessed with ultrasound-guided minimally invasive autopsy.

AIMS: Brazil ranks high in the number of coronavirus disease 19 (COVID-19) cases and the COVID-19 mortality rate. In this context, autopsies are important to confirm the disease, determine associated conditions, and study the pathophysiology of this novel disease. The aim of this study was to assess the systemic involvement of COVID-19. In order to follow biosafety recommendations, we used ultrasound-guided minimally invasive autopsy (MIA-US), and we present the results of 10 initial autopsies. METHODS AND RESULTS: We used MIA-US for tissue sampling of the lungs, liver, heart, kidneys, spleen, brain, skin, skeletal muscle and testis for histology, and reverse transcription polymerase chain reaction to detect severe acute respiratory syndrome coronavirus 2 RNA. All patients showed exudative/proliferative diffuse alveolar damage. There were intense pleomorphic cytopathic effects on the respiratory epithelium, including airway and alveolar cells. Fibrinous thrombi in alveolar arterioles were present in eight patients, and all patients showed a high density of alveolar megakaryocytes. Small thrombi were less frequently observed in the glomeruli, spleen, heart, dermis, testis, and liver sinusoids. The main systemic findings were associated with comorbidities, age, and sepsis, in addition to possible tissue damage due to the viral infection, such as myositis, dermatitis, myocarditis, and orchitis. CONCLUSIONS: MIA-US is safe and effective for the study of severe COVID-19. Our findings show that COVID-19 is a systemic disease causing major events in the lungs and with involvement of various organs and tissues. Pulmonary changes result from severe epithelial injury and microthrombotic vascular phenomena. These findings indicate that both epithelial and vascular injury should be addressed in therapeutic approaches.

Heterogeneous phenotype of Hereditary Xerocytosis in association with PIEZO1 variants.

Hereditary Xerocytosis (HX) is an autosomal dominantly inherited congenital hemolytic anemia associated with erythrocyte dehydration due to decreased intracellular potassium content resulting in increased mean corpuscular hemoglobin concentration. The affected members of HX families show compensated anemia with splenomegaly, hemosiderosis, and perinatal edema but are in large part transfusion independent. Functional studies show a link between mutations in mechanosensitive ion channel, encoded by PIEZO1 gene and the HX. We identified new PIEZO1 variants that are likely pathogenic in three phenotypically characterized multi-generational HX Brazilian families. Interestingly, one missense variant of the PIEZO1 gene identified, p.E2494V was associated in trans with the previously reported most frequent pathogenic duplication p.E2496ELE. The three-dimensional structure of the human protein modeled using structural coordinates of the mouse Piezo1 solved by cryo-electron microscopy (Cryo-ME) showed that the two identified variants, p.M2007L and p.T2014I, are localized to an important mechanosensitive transmembrane domain suggesting a conformational mechanism for altered channel's gating. The p.E2496ELE variant identified alters the extension of helix α1 bringing it much closer to the beam affecting the position of it structure at the end of the pore.

Clinical Applications of Point-of-Care Testing in Different Conditions.

BACKGROUND: The use of point-of-care testing (POCT) in different clinical applications is justified by the fact that the time to release the result is shortened, allowing the physician to define the diagnosis and most appropriate therapy in a shorter time. However, the negative aspects must also be highlighted and studied so that we can move forward with the use of these devices. These negative aspects include greater analytical imprecision compared to laboratory automation, the variability between different equipment from different manufacturers, the risk of inappropriate use, a low level of global regulation, higher costs compared with laboratory testing and cost ineffectiveness in terms of health care. Methods and. RESULTS: This review presents some clinical applications of POCT in different scenarios, such as for diabetes mellitus, infectious diseases, pediatrics, and chronic kidney disease, among others. CONCLUSIONS: We hope to see a global consensus on an acceptable quality standard for performing POCT that is adaptable, practical, and cost effective in primary care settings, ensuring patient safety, and minimizing the risk of harm.

Molecular diagnosis of Strongyloides stercoralis among transplant candidates.

Strongyloidiasis can occur without any symptoms or as a potentially fatal hyperinfection or disseminated infection, principally in immunosuppressed patients. Our study aimed to evaluate the application of conventional polymerase chain reaction (cPCR) and real-time PCR (qPCR). Polymerase chain reaction (PCR) and real-time PCR (qPCR) targeting the 18S rRNA gene for detection of Strongyloides stercoralis infection among transplant candidates were applied in stool samples obtained from 150 transplant candidates, preliminarily analyzed by parasitological methods. S. stercoralis larvae were visualized in 15/150 (10.0%) transplant candidates by parasitological methods. DNA from S. stercoralis was amplified in 26/150 (17.3%) and 49/150 (32.7%) stool samples of transplant candidates, using cPCR and qPCR, respectively. The results suggest that molecular methods, especially qPCR, should be used as an additional tool for diagnostic of S. stercoralis infection among transplant candidates.

Point-of-Care Testing: General Aspects.

Point-of-Care Testing (POCT) has been highlighted in the health care sector in recent decades. On the other hand, due to its low demand, POCT is at a disadvantage compared to conventional equipment, since its cost is inversely proportional to the volume of use. In addition, for the implementation of POCT to succeed, it is essential to rely on the work of a multidisciplinary team. The awareness of health professionals of the importance of each step is perhaps the critical success factor. The trend towards the continuous advancement of the use of POCT and the great potential of its contributions reinforce the need to implement quality management tools, including performance indicators, to ensure their results. This review presents some advantages and disadvantages concerning POCT and the real need to use it. A worldwide call for the availability of easy-to-use health technologies that are increasingly closer to the final user is one of the main reasons for this focus.

Hallmarks of liver lesions in pigs naturally infected by hepatitis E virus genotype 3 / Perfil das lesões histopatológicas hepáticas em suínos infectados naturalmente pelo genótipo 3 do vírus da hepatite E

Histopathological evaluation of liver from 33 pigs slaughtered for human consumption in Amazon region, previously tested by serology and molecular techniques for hepatitis E virus infection (HEV), was analysed in three groups: Group 1, negative for both HEV-RNA and anti-HEV IgG (n=10); Group 2, positive for HEV-RNA (n=13); Group 3, positive for anti-HEV IgG (n=10). Group 2 showed a significant difference among the groups for liver lesions such as lobular activity (P=0.007), periportal interface hepatitis (P=0.004), portal inflammation (P=0.028) hepatitis with lobular, portal and periportal interface activity (P=0.001). HEV detection by immunohistochemistry was performed and 3 of 6 samples of group 2 were positive. Pigs naturally infected by HEV genotype 3 present microscopic necroinflammatory liver lesions similar to HEV in humans. Liver histopathology showed be important in the diagnosis of active asymptomatic HEV infection in pigs slaughtered for human consumption because hepatic liver lesions may present distinct profiles according to molecular and serological diagnosis and in this sense, histopathology and immunohistochemistry may be an important complementary diagnostic tool.(AU)

A avaliação histopatológica hepática de 33 suínos abatidos para consumo humano na região amazônica, previamente testados para infecção pelo vírus da hepatite E (HEV) por sorologia e técnicas moleculares, foi realizada em três grupos: Grupo 1, animais negativos para HEV-RNA e anti-HEV IgG (n=10); Grupo 2, positivos para HEV-RNA (n=13); e Grupo 3, positivos para anti-HEV IgG (n=10). O grupo 2 apresentou diferenças estatísticas significantes entre os grupos em relação à presença de atividade lobular (P=0,007), hepatite periportal de interface (P=0,004), inflamação portal (P= 0.028) e atividade lobular acompanhada por inflamação portal e periportal de interface (P=0,001). A detecção imunohistoquímica do HEV foi realizada e três de seis amostras do Grupo 2 foram positivas. Suínos naturalmente infectados pelo genótipo 3 do HEV apresentam lesões necroinflamatórias no fígado similares a lesão em humanos. A histopatologia hepática demonstrou ser importante no diagnóstico de infecção ativa e assintomática por HEV em suínos abatidos para consumo humano, pois as lesões no fígado apresentaram perfis diferenciados de acordo com o diagnóstico sorológico e molecular da infecção e, neste sentido, a histopatologia e imunohistoquímica podem representar importantes ferramentas complementares de diagnóstico.(AU)

Persistence of Zika Virus in Breast Milk after Infection in Late Stage of Pregnancy.

We detected Zika virus in breast milk of a woman in Brazil infected with the virus during the 36th week of pregnancy. Virus was detected 33 days after onset of signs and symptoms and 9 days after delivery. No abnormalities were found during fetal assessment or after birth of the infant.

Prevalence of naturally occurring protease inhibitor resistance-associated variants in hemodialysis and renal transplant patients with hepatitis C virus infection.

Background NS3 protease inhibitors (PIs) were the first direct antiviral agents used for the treatment of hepatitis C virus. The combination of second-wave PIs with other direct antiviral agents enabled the use of interferon-free regimens for chronic kidney disease patients on dialysis and renal transplant (RTx) recipients, populations in which the use of interferon and ribavirin is limited. However, the occurrence of PI resistance-associated variants (RAVs), both baseline and induced by therapy, has resulted in the failure of many treatment strategies. Methods The aim of this study was to estimate the prevalence of PI RAVs and of the Q80K polymorphism in chronic kidney disease patients on hemodialysis and RTx recipients. Direct sequencing of the NS3 protease was performed in 67 patients (32 hemodialysis and 35 RTx).Results RAVs to PIs were detected in 18% of the patients: V55A (9%), V36L (1.5%), T54S (1.5%), S122N (1.5%), I170L (1.5%), and M175L (1.5%). Only 1.5% of the patients carried the Q80K polymorphism. The frequency of these mutations was more than two times higher in patients infected with GT1a (25%) than GT1b (9.7%) (P=0.1). The mutations were detected in 20% of treatment-naive patients and in 15.6% of peginterferon/ribavirin-experienced patients (P=0.64). Furthermore, no mutation that would confer high resistance to PIs was detected.Conclusion The Q80K polymorphism was rare in the population studied. The occurrence of RAVs was common, with predominance in GT1a. However, the variants observed were those associated with a low level of resistance to PIs, facilitating the use of these drugs in this special group of patients.

Characterization of hepatitis B virus infection in illicit drug users in the Marajó Archipelago, northern Brazil.

South America is considered to have a low prevalence of hepatitis B virus (HBV) infection, although areas with a relatively high prevalence have been identified in northern Brazil. Few epidemiological studies of populations at risk of HBV infection are available for this region. Given this, in the present study, we investigated the prevalence of HBV and the factors associated with infection among illicit drug users (DUs) in the Marajó Archipelago, northern Brazil. In this cross-sectional study, we collected samples and epidemiological information from DUs in 11 municipalities of the Marajó Archipelago. The diagnosis was established by ELISA and real-time PCR; and genotyping was done by multiplex real-time PCR. Statistical modeling was based on simple and multiple logistical regressions with the Hosmer-Lemeshow test. The mean age of the 466 DUs was 28.4 years, and most were male. The most-consumed illicit drugs were crack cocaine and marijuana. In all, 171 DUs were exposed to HBV, with genotypes A, D and F being identified. The factors associated with higher frequencies of HBV infection were (i) male gender, (ii) age above 35 years, (iii) anti-HIV positivity, (iv) tattoos, (v) the use of injected drugs, (vi) the use of illicit drugs for more than 3 years, (vii) sexual relations without protection, (viii) sexual relations with another DU, and (ix) more than 10 sexual partners in the past 24 months. In summary, this study provides important insights into the dynamics of HBV infection among DUs in the Marajó Archipelago. We hope that these findings will contribute to the development of strategies, actions and public health policies aimed at preventing and controlling this viral infection more effectively.

High prevalence of HBV/A1 subgenotype in native south Americans may be explained by recent economic developments in the Amazon.

Native American populations present the highest prevalence of Hepatitis B Virus (HBV) infection in the Americas, which may be associated to severe disease outcomes. Ten HBV genotypes (A­J) have been described, displaying a remarkable geographic structure, which most likely reflects historic patterns of human migrations. In this study, we characterize the HBV strains circulating in a historical sample of Native South Americans to characterize the historical viral dynamics in this population. The sample consisted of 1070 individuals belonging to 38 populations collected between 1965 and 1997. Presence of HBV DNA was checked by quantitative real-time PCR, and determination of HBV genotypes and subgenotypes was performed through sequencing and phylogenetic analysis of a fragment including part of HBsAg and Pol coding regions (S/Pol). A Bayesian Skyline Plot analysis was performed to compare the viral population dynamics of HBV/A1 strains found in Native Americans and in the general Brazilian population. A total of 109 individuals were positive for HBV DNA (~ 10%), and 70 samples were successfully sequenced and genotyped. Subgenotype A1 (HBV/A1), related to African populations and the African slave trade, was the most prevalent (66­94%). The Skyline Plot analysis showed a marked population expansion of HBV/A1 in Native Americans occurring more recently (1945­1965) than in the general Brazilian population. Our results suggest that historic processes that contributed to formation of HBV/A1 circulating in Native American are related with more recent migratory waves towards the Amazon basin, which generated a different viral dynamics in this region.