Antibody-dependent enhancement representing in vitro infective progeny virus titer correlates with the viremia level in dengue patients

Dengue virus (DENV) causes dengue fever (DF) and dengue hemorrhagic fever in humans. Some DF patients suddenly develop severe symptoms around the defervescent period. Although the pathogenic mechanism of the severe symptoms has not been fully elucidated, the viremia level in the early phase has been shown to correlate with the disease severity. One of the hypotheses is that a phenomenon called antibody-dependent enhancement (ADE) of infection leads to high level of viremia. To examine the plausibility of this hypothesis, we examined the relationship between in vitro ADE activity and in vivo viral load quantity in six patients with dengue diseases. Blood samples were collected at multiple time points between the acute and defervescent phases, and the balance between neutralizing and enhancing activities against the autologous and prototype viruses was examined. As the antibody levels against DENV were rapidly increased, ADE activity was decreased over time or partially maintained against some viruses at low serum dilution. In addition, positive correlations were observed between ADE activity representing in vitro progeny virus production and viremia levels in patient plasma samples. The measurement of ADE activity in dengue-seropositive samples may help to predict the level of viral load in the subsequent DENV infection.

Detection of genotype-1 of dengue virus serotype 3 for the first time and complete genome analysis of dengue viruses during the 2018 epidemic in Mandalay, Upper Myanmar

Background Dengue (DEN) is a neglected tropical disease, and surveillance of dengue virus (DENV) serotypes and genotypes is critical for the early detection of outbreaks. Risk factors for outbreaks include the emergence of new genotypes and serotype shifting. Methodology and principal findings To understand the genomic and viral characteristics of DENV-infected patients, we conducted a cross-sectional descriptive study among pediatric patients admitted at the 550-bedded Mandalay Children Hospital during the 2018 DEN endemic season. We conducted virus isolation, serological tests, viremia level measurement, and whole-genome sequencing. Among the 202 serum samples, we detected 85 samples with DENV (46 DENV-1, 10 DENV-3, 26 DENV-4 and three multiple serotype co-infections) via reverse transcription quantitative/real-time PCR (RT-qPCR), and we obtained 49 DENV isolates (31 DENV-1, 10 DENV-3 and 8 DEN-4). We did not detect DENV-2 in this study. The viral genome levels in serum did not differ significantly among virus serotypes, infection status (primary versus secondary) and disease severity. Based on the phylogenetic analysis, we identified DENV-1 genotype-1, DENV-4 genotype-1 and DENV-3 genotype-3 and genotype-1 which was detected for the first time. Next-generation sequencing analysis revealed greater frequencies of nonsynonymous and synonymous mutations per gene in the nonstructural genes. Moreover, mutation rates were also higher among DENV-1. Conclusion/Significance In conclusion, there was an increasing trend of DENV-3 cases during DENV endemic season in 2018 with the first detection of the genotype 1. However, DENV-1 has remained the predominant serotype in this study area since 2013, and we identified stop codon mutations in the DENV-1 genome. This report is the first to feature a complete genome analysis of the strains of DENV-3 and DENV-4 circulating among pediatric patients in Myanmar. This study highlighted the importance of annual surveillance for a better understanding of the molecular epidemiology of DENVs.

Antibody-dependent enhancement representing in vitro infective progeny virus titer correlates with the viremia level in dengue patients

ABSTRACTDengue virus (DENV) distributes throughout tropical and subtropical countries and causes dengue fever (DF) and dengue hemorrhagic fever in humans. Some DF patients suddenly develop severe symptoms after the defervescent period. Although the pathogenic mechanism of the severe symptoms has not been fully elucidated, the viremia level in the early phase has been shown to correlate with the disease severity. One of the hypotheses is that a phenomenon called antibody-dependent enhancement (ADE) of infection leads to a high level of viremia. To examine the plausibility of this hypothesis, we examined the relationship between in vitro ADE activity and in vivo viral load quantity in six patients with dengue diseases. An autologous DENV strain was isolated from each of the six patients. Blood samples were then collected at multiple time points between the acute and defervescent phases, and the balance between neutralizing and enhancing activities against the autologous and prototype viruses was examined. As the antibody levels against DENV were rapidly increased, ADE activity was decreased over time or partially maintained against some viruses at low serum dilution. In addition, positive correlations were observed between ADE activity representing in vitro progeny virus production and viremia levels in patient plasma samples. Therefore, the measurement of ADE activity in dengue-seropositive samples may help to predict the impact of viral load in the subsequent DENV infection.IMPORTANCEIt has not been fully elucidated how the phenomenon of antibody-dependent enhancement (ADE) affects the pathogenesis of severe dengue diseases, although high viremia levels have been epidemiologically demonstrated to be associated with the disease severity. Here, we show that ADE in the acute-phase patient sera exhibited significantly different activities against autologous and lab strains than ADE in the defervescent-phase sera. Further, the enhancement of progeny virus production activity, which is one of the factors to evaluate ADE in vitro, was significantly correlated with the levels of viral load in the patient blood circulation. This suggests that measurement of the in vitro enhancing progeny virus titers might be used to predict the impact of in vivo DENV viremia level. Our present findings could contribute to a method to forecast disease severity for seropositive populations who would be at risk of developing severe disease in the event of heterotypic DENV infection.

A prospective study in hepatitis C virus treatment-naïve patients showing rheumatologic extra-hepatic manifestations of hepatitis C with associated risk factors: efficacy and safety using sofosbuvir-based direct antiviral therapy

Background To study the most common rheumatologic manifestations of hepatitis C viral (HCV) infection in Egyptian patients and associated risk factors with assessment the effect of current therapies on these manifestations. A prospective study was carried out to HCV patients attending the tropical medicine department referred to rheumatology department, over a year. A total of 204 hepatitis C virus treatment-naïve patients aged 21–71 years old suffering from rheumatologic manifestations were recruited, and history taking, general and musculoskeletal examination, laboratory and serological investigations, imaging, and liver fibrosis findings were assessed: baseline, end of treatment, and 12 weeks later, either sofosbuvir/ribavirin or sofosbuvir/simeprevir regimens and through three consecutive visits: joint activity and functional scores were taken. Results Common observed rheumatologic manifestations were fibromyalgia (74.5%), arthralgia (73.5%), Raynaud’s phenomenon (54.9%), peripheral neuropathy (29.4%),chronic fatigue syndrome and purpura (24.5%), arthritis (16.7%), Sicca symptoms and skin ulcers (9.8%), and vasculitic CNS involvement (5.9%), mostly seen in females. VAS and FAS scales have improved across visits (p value < 0.001) with lowered number and percentage of arthralgia (tender joint counts), arthritis (swollen joint counts), improvement of fibromyalgia, purpura, peripheral neuropathy, anemia, and thrombocytopenia (p < 0.001). Common reported risk factors were barber shaving (52 %), dental procedures (44.1%), and surgical interventions (36.3%). Non-reactive cases shown by HCV-PCR response increased at the end of study reaching 62%. The percent of improvement was significantly higher in patients receiving sofosbuvir/simeprevir regimen (100%) versus sofosbuvir/ribavirin (58.2%). Conclusion Direct antiviral drugs seem to improve the rheumatic extra-hepatic manifestations of HCV patients and lowering viremia level especially sofosbuvir/simeprevir regimen in hepatitis C treatment-naïve patients.

Diagnosis of west Nile neuroinvasive disease in humans

West Nile virus (WNV) is arbovirus distributed all around the world. In humans, 80% of infection cases are asymptomatic. In 20% of infected people, a febrile self-limiting illness is reported. WNV has the potential for fatal neuroinvasive disease. In 1% of cases, the infection may result in neuroinvasive disease with permanent neurological consequences or death outcome. Neurological forms may vary presenting with encephalitis, meningitis, meningoencephalitis or acute flaccid paralysis. Outbreaks with neurological forms of WNV infection were recorded in different areas of Greece, Italy, Romania, Hungary and Serbia. During the period from 2013 to 2016, 114 samples of cerebrospinal fluid and 107 serum samples were taken from 114 patients suspected of WNV neuroinvasive disease (WNND). The presence of specific anti-WNV IgM and IgG antibodies in cerebrospinal fluid (CSF) and sera samples were tested by WNV IgM and IgG ELISA (Euroimmun, Germany). In addition, 48 samples of CSF or/and serum of people with suspected WNV infection were examined by commercial molecular tests - real time RT-PCR (WNV Real-TM, Sacace biotechnologies, Italy). The IgM antibodies against WNV were present in 25.4% (29/114) of CSF samples, and in 31.8% (34/107) of serum samples tested from 114 patients suspected of WNND. The IgG antibodies against WNV were detected in 3.5% (4/114) of CSF samples, and in 11.2% (12/107) of serum samples. The WNV RNA was detected by real time RT-PCR test in 7 out of 48 (14.6%) CSF or/and serum samples. In this study, detection of IgM antibodies in CSF is more frequent than detection of WNV RNA in CSF or serum samples. WNV RNA detection in CSF is confirmatory diagnostic test but has limited utility in the diagnosis of WNV neuroinvasive disease due to low viremia level at the time of clinical presentation of the disease. The limitations in the use of ELISA IgM test are linked to cross - reactivity among flaviviruses and long persistence of IgM antibodies in the serum and CSF.

Circulating CD34+ hematopoietic stem/progenitor cells paralleled with level of viremia in patients chronically infected with hepatitis B virus

Introduction: Liver regeneration is a heterogeneous process involving proliferation of different cell types in response to injury. Bone marrow derived stem cells may be involved in this process, by making contribution to parenchymal restoration and cellular replacement. We aimed to investigate the correlation between level of circulating mobilized CD34+ hematopoietic stem progenitor cells (HSPCs) and viremia level in patients chronically infected with hepatitis B virus (HBV). Methods: Blood samples were prospectively collected for assessing percentage and absolute counts of circulating CD34+ HSPCs and viral load level using flow cytometry and RT-PCR respectively. Patients with chronic hepatitis B (CHB) (n = 30), Entecavir (ETV) treated subjects (n = 30) and 20 age and gender matched healthy controls were enrolled in this study. Results were expressed as mean ± SD. Results and discussion: A significant increase in circulating CD34+ HSPCs level was observed in CHB patients (5 ± 3.1, 324 ± 195 × 103/ml) as compared to ETV treated subjects (0.57 ± 0.27,1022 ± 325) and healthy controls (0.53 ± 0.37, 694 ± 254, P < 0.001) in regards to percentage and absolute counts respectively. Levels of CD34+ HSPCs strongly and positively correlated with HBV DNA viral load levels in CHB patients (r2 = 0.8417, 0.649, P < 0.001).Thus, in chronic liver disorders (CHB), when reduced regenerative capacity of hepatocytes is reached, BMSCs mobilization occurs and their level increases in peripheral blood. The level of circulating CD34+ cells in peripheral blood of CHB patients paralleled with the hepatitis B viral load.

The occurrence of liver steatosis in patients with chronic hepatitis C - the experience of the Clinical Center of Vojvodina

Background/Aim. Hepatic steatosis in patients with chronic hepatitis C occurs in about half of the cases. Its occurrence is influenced by factors of the host and viral factors and its importance lies in the fact that it reduces the success of antiviral therapy based on interferon in the treatment of chronic hepatitis C and that, associated with other factors, exacerbates liver disease. The aim of this study was to determine the prevalence and severity of steatosis in patients with chronic hepatitis C and to determine the factors that affect its occurrence. Methods. The study included 123 patients with chronic hepatitis C with diagnosis of liver steatosis made by liver biopsy and histopathological examination according to which ? 5% of hepatocytes was affected by fatty change. Based on the presence of steatosis, the patients were divided into two groups: 43 patients with steatosis and 80 patients without steatosis. The influence of certain factors on the occurrence of steatosis was examined using standard statistical methods. Results. Liver steatosis was found in 34.96% of patients with chronic hepatitis C, and a majority of patients (76.74%) had mild steatosis. Of the examined predictive factors for the occurrence of steatosis, statistical significance in its occurrence was connected to elevated body mass index (BMI), genotype 3 hepatitis C virus (HCV) and HCV viremia. Conclusion. Hepatic steatosis often occurs in people with chronic hepatitis C, and most often it is mild. The occurrence of hepatic steatosis in our sample was most often affected by genotype 3 HCV and HCV viremia level. Hepatic steatosis can reduce the success of antiviral therapy based on interferon and negatively affect chronic liver disease course. Therefore, we need to recognize it, treat it and make it withdraw.