Hepatitis C Virus as a Possible Helper Virus in Human Hepatitis Delta Virus Infection.

Previous studies reported that the hepatitis C virus (HCV) could help disseminate the hepatitis D virus (HDV) in vivo through the unrelated hepatitis B virus (HBV), but with essentially inconclusive results. To try to shed light on this still-debated topic, 146 anti-HCV-positive subjects (of whom 91 HCV/HIV co-infected, and 43 with prior HCV eradication) were screened for anti-HDV antibodies (anti-HD), after careful selection for negativity to any serologic or virologic marker of current or past HBV infection. One single HCV/HIV co-infected patient (0.7%) tested highly positive for anti-HD, but with no positive HDV-RNA. Her husband, in turn, was a HCV/HIV co-infected subject with a previous contact with HBV. While conducting a thorough review of the relevant literature, the authors attempted to exhaustively describe the medical history of both the anti-HD-positive patient and her partner, believing it to be the key to dissecting the possible complex mechanisms of HDV transmission from one subject to another, and speculating that in the present case, it may have been HCV itself that behaved as an HDV helper virus. In conclusion, this preliminary research, while needing further validation in large prospective studies, provided some further evidence of a role of HCV in HDV dissemination in humans.

Complete coding sequence of an Aedes flavivirus strain isolated from Aedes albopictus collected in Northern Italy.

Complete genome data for the globally distributed Aedes flavivirus (AEFV) is scarce. We analyzed a new Italian AEFV strain isolated from Aedes albopictus. The results demonstrated genetic diversity among Italian AEFVs. The high similarity between AEFV genomes across geographically distant regions suggests long distance spreading via invasive host mosquito species.

Longitudinal monitoring of SARS-CoV-2 viral load in self-collected saliva from health care workers during breakthrough infections to spare working days.

IMPORTANCE: Real-time quantitative PCR (RT-qPCR) on nasopharyngeal swabs (NPS) has been used as the standard method for detecting and monitoring SARS-CoV-2 infection during the pandemic. However, NPS collection often causes discomfort and poses a higher risk of transmission to health care workers (HCW). Furthermore, RT-qPCR only provides relative quantification and does not allow distinguishing those samples with residual, no longer active infection, whereas droplet digital PCR (ddPCR) allows for precise quantification of viral load, offering greater sensitivity and reproducibility. This study highlights the effectiveness of using self-collected saliva as a convenient and reliable sampling method. By utilizing ddPCR to measure the SARS-CoV-2 viral load in saliva samples, individuals with low or undetectable viral loads can be quickly identified. This approach is particularly advantageous for surveillance programs targeting HCW, as it enables the early identification and release of uninfected personnel, minimizing lost workdays. Additionally, analyzing viral load in saliva samples by ddPCR is valuable in determining virus shedding duration across different SARS-CoV-2 variants, informing transmission and disease control. Finally, testing saliva could overcome the detection of historic cases due to prolonged RNA swabbing past-infection and the unnecessary exclusion of those individuals from the workplace.

Baseline Plasma Osteopontin Protein Elevation Predicts Adverse Outcomes in Hospitalized COVID-19 Patients.

More than three years have passed since the first case, and COVID-19 is still a health concern, with several open issues such as the lack of reliable predictors of a patient's outcome. Osteopontin (OPN) is involved in inflammatory response to infection and in thrombosis driven by chronic inflammation, thus being a potential biomarker for COVID-19. The aim of the study was to evaluate OPN for predicting negative (death or need of ICU admission) or positive (discharge and/or clinical resolution within the first 14 days of hospitalization) outcome. We enrolled 133 hospitalized, moderate-to-severe COVID-19 patients in a prospective observational study between January and May 2021. Circulating OPN levels were measured by ELISA at admission and at day 7. The results showed a significant correlation between higher plasma concentrations of OPN at hospital admission and a worsening clinical condition. At multivariate analysis, after correction for demographic (age and gender) and variables of disease severity (NEWS2 and PiO2/FiO2), OPN measured at baseline predicted an adverse prognosis with an odds ratio of 1.01 (C.I. 1.0-1.01). At ROC curve analysis, baseline OPN levels higher than 437 ng/mL predicted a severe disease evolution with 53% sensitivity and 83% specificity (area under the curve 0.649, p = 0.011, likelihood ratio of 1.76, (95% confidence interval (CI): 1.35-2.28)). Our data show that OPN levels determined at the admission to hospital wards might represent a promising biomarker for early stratification of patients' COVID-19 severity. Taken together, these results highlight the involvement of OPN in COVID-19 evolution, especially in dysregulated immune response conditions, and the possible use of OPN measurements as a prognostic tool in COVID-19.

Baseline plasma SARS-CoV-2 RNA detection predicts an adverse COVID-19 evolution in moderate to severe hospitalized patients.

BACKGROUND: SARS-CoV-2 is a single-stranded RNA virus, known to be the causative agent of COVID-19. As the resulting disease shows a very heterogeneous range of clinical manifestations, the identification of early biomarkers allowing patients stratification according to the expected disease severity is still an unmet clinical need. METHODS: In this observational prospective cohort study, 137 consecutive patients, testing positive for SARS-CoV-2 infection by nasopharyngeal swab RT-PCR or antigenic test, were enrolled to evaluate their plasma viral load at the time of hospitalization. RESULTS: Even if all of them had a molecular diagnosis of COVID-19, only 29 patients showed a detectable plasma SARS-CoV-2 RNAemia. Such viremic patients also showed other clinical and laboratory finding alterations (increased troponin I, IL-6, RDW-CV, and creatinine levels along with decreased platelet count and glomerular filtration rate). A plasma detectable RNA viral load predicted in hospital death or ICU admission with an odds ratio of 3.53 (CI: 1.44-8.64, P=0.0058), while the lack of a detectable viral load was associated with a faster recovery, with an odds ratio of 4.06 (CI: 1.72-9.59, P=0.0014). These findings were confirmed in multivariate models including age, sex and baseline National Early Warning Score 2 and arterial oxygen tension over inspired oxygen fraction ratio. CONCLUSIONS: Our data thus suggest that plasma viral RNA load at the time of hospital admission could represent a useful independent biomarker allowing early patients' stratification according to the expected disease evolution, and driving clinical decisions tailored on the specific needs of the individual patient.

Unusual presentation of fatal disseminated varicella zoster virus infection in a patient with lupus nephritis: a case report.

BACKGROUND: The risk of life-threatening complications, such as visceral disseminated varicella zoster virus (VZV) infection, is greater in immunosuppressed individuals, such as systemic lupus erythematosus (SLE) patients. CASE PRESENTATION: Here, a case is reported of a Caucasian woman diagnosed with lupus nephritis and anti-phospholipid syndrome, who was subjected to mycophenolate mofetil and high-dose steroid remission-induction therapy. Two months later she developed abdominal pain followed by a fatal rapid multi-organ failure. As no typical skin rashes were evident, death was initially attributed to catastrophic anti-phospholipid syndrome. However, autopsy and virological examinations on archival material revealed a disseminated VZV infection. CONCLUSIONS: Overall, this case highlights the importance of having a high clinical suspicion of fatal VZV infections in heavily immunosuppressed SLE patients even when typical signs and symptoms are lacking.

Strigolactone Analogs Are Promising Antiviral Agents for the Treatment of Human Cytomegalovirus Infection.

The human cytomegalovirus (HCMV) is a widespread pathogen and is associated with severe diseases in immunocompromised individuals. Moreover, HCMV infection is the most frequent cause of congenital malformation in developed countries. Although nucleoside analogs have been successfully employed against HCMV, their use is hampered by the occurrence of serious side effects. There is thus an urgent clinical need for less toxic, but highly effective, antiviral drugs. Strigolactones (SLs) are a novel class of plant hormones with a multifaceted activity. While their role in plant-related fields has been extensively explored, their effects on human cells and their potential applications in medicine are far from being fully exploited. In particular, their antiviral activity has never been investigated. In the present study, a panel of SL analogs has been assessed for antiviral activity against HCMV. We demonstrate that TH-EGO and EDOT-EGO significantly inhibit HCMV replication in vitro, impairing late protein expression. Moreover, we show that the SL-dependent induction of apoptosis in HCMV-infected cells is a contributing mechanism to SL antiviral properties. Overall, our results indicate that SLs may be a promising alternative to nucleoside analogs for the treatment of HCMV infections.

Interplay of PNPLA3 and HSD17B13 Variants in Modulating the Risk of Hepatocellular Carcinoma among Hepatitis C Patients.

A single-nucleotide polymorphism causing a C to G change in the PNPLA3 gene (rs738409) is associated with disease severity and development of hepatocellular carcinoma (HCC) in nonalcoholic fatty liver disease; the insertion variant rs72613567:TA of the 17ß-hydroxysteroid dehydrogenase type 13 (HSD17B13) mitigates this detrimental effect. Our aim was to evaluate if the same holds true in chronic hepatitis C virus infection (HCV). With a case control retrospective study design, we selected 110 patients who developed HCC on a background of HCV infection, matching each patient for sex and age (±30 months) to three HCV-infected, non-HCC patients. All participants underwent genotyping for PNPLA3 and HSD17B13 gene variants. Both univariate and multivariate analyses of risk factors for advanced disease and HCC were performed. Carriage of PNPLA3 G∗ allele was associated with a trend of progressively more severe liver disease, from mild fibrosis to significant fibrosis, cirrhosis, and HCC (p = 0.007). When the HSD17B13:TA status of these patients was taken into account, the abovementioned trend was strengthened among HSD17B13 major allele homozygotes and completely blunted among carriers of the minor allele (p = 0.0003 and 0.953, respectively). In a conditional logistic regression model including diabetes and AST to platelet ratio index among predictor variables, the unfavourable genetic profile characterized by the coexistence of the PNPLA3 minor allele and HSD17B13 major allele (vs. all other possible combinations) was an independent risk factor for HCC (OR = 2.00, 95% CI: 1.23-3.26) together with a history of alcohol abuse. In conclusion, carriage of the combination PNPLA3 minor allele and HSD17B13 major allele may represent a risk factor for HCC among HCV-infected patients. The interplay between the two genes may explain some of the controversy on this topic and may be exploited to stratify HCC risk in hepatitis C.

Interferon signature in immunosuppressed patients with lower respiratory tract infections: dosage on bronchoalveolar lavage.

BACKGROUND: Interferon signature (IS) is the measure of transcripts belonging to pathways of interferon activation. Viral infections can interfere with the interferon pathway, in particular herpesvirus present in immunocompromised hosts. The aim of our study was to evaluate if herpesvirus infections in immunocompromised patients with lower respiratory tract infections (LRTI) could lead to IS alterations. METHODS: We measured IS transcription of six genes on bronchoalveolar lavage of immunocompromised patients with LRTI (IFI27, IFI44, IFIT1, ISG15, RSAD2, SIGLEC1). Patients were divided in three groups based on Epstein-Barr virus (EBV) and other herpesviruses coinfections. RESULTS: We included 56 patients, 10 without and 17 with only EBV reactivation (respectively N and E groups) and 29 with EBV and other herpesviruses (group C). IS was higher in group C (P=0.01) compared to other ones, but single gene expressions were different among groups: IFI27 was higher whereas IFIT1 and ISG15 were lower in group C (P<0.05). CONCLUSIONS: The continuous stimulation of interferon cascade by herpesviruses enhances IS. The analysis of IS in immunocompromised population is possible by limiting the use of IFI27, IFIT1, ISG15 genes. Our preliminary results seem to indicate that IS is a useful biomarker of cellular response to herpesvirus infection in immunocompromised patients.

Use of an innovative and non-invasive device for virologic sampling of cough aerosols in patients with community and hospital acquired pneumonia: a pilot study.

BACKGROUND: The aetiology of lower respiratory tract infections is challenging to investigate. Despite the wide array of diagnostic tools, invasive techniques, such as bronchoalveolar lavage (BAL), are often required to obtain adequate specimens. PneumoniaCheckTM is a new device that collects aerosol particles from cough, allowing microbiological analyses. Up to now it has been tested only for bacteria detection, but no study has investigated its usefulness for virus identification. METHODS: In this pilot study we included 12 consecutive patients with pneumonia. After testing cough adequacy via a peak flow meter, a sampling with PneumoniaCheckTM was collected and a BAL was performed in each patient. Microbiological analyses for virus identification were performed on each sample and concordance between the two techniques was tested (sensitivity, specificity and positive/negative predictive values), taking BAL results as reference. RESULTS: BAL was considered adequate in 10 patients. Among them, a viral pathogen was identified by PneumoniaCheckTM 6 times, each on different samples, whereas BAL allowed to detect the presence of a virus on 7 patients (14 positivities). Overall, the specificity for PneumoniaCheckTM to detect a virus was 100%, whereas the sensitivity was 66%. When considering only herpes viruses, PneumoniaCheckTM showed a lower sensitivity, detecting a virus in 1/4 of infected patients (25%). CONCLUSIONS: In this pilot study PneumoniaCheckTM showed a good correlation with BAL for non-herpes virologic identification in pneumonia patients, providing excellent specificity. Further studies on larger population are needed to confirm these results and define its place in the panorama of rapid diagnostic tests for lower respiratory tract infections.

Is HERV-K and HERV-W Expression Regulated by miR-155 in Kidney Transplant Patients with Human Cytomegalovirus Infection?

According to the latest update, 2,578 unique mature micro-RNAs (miRNAs) are currently annotated in the human genome and participate in the regulation of multiple events, such as cellular proliferation or apoptosis. A previous study analyzing global miRNA expression patterns in GH cells (high human endogenous retrovirus, HERV, K vs. low) showed that 2 miRNAs (miR-663 and miR-638) are differentially regulated and exhibit expression parallel to that of HERV-K. The aim of this study was to evaluate HERV-K and -W pol gene and miR-155 expression in kidney transplant recipients and the possible relationship between them. The comparison between kidney transplant patients negative for human cytomegalovirus (HCMV) infection and positive patients showed a significant difference in terms of miR-155 expression (p = 0.0111). We demonstrated that HERV-K and -W pol gene expression was significantly higher in CMV-infected kidney transplant recipients versus those not infected as previously reported by our groups. Our correlation data suggest that miR-155 are not directly involved in regulating the HERV notwithstanding that we together observed increased expression of HERV-K and -W and diminished expression of miR-155 in HCMV-infected human kidney transplant recipients.

Gas6 as a predictor of esophageal varices in patients affected by hepatitis C virus related-chronic liver disease.

AIM: Plasma Gas6 was tested as an alternative to Baveno VI criteria (liver stiffness <20 kPa and platelet count >150 × 109/l) in an endoscopy-sparing strategy. METHODS:  A total of 160 patients with chronic hepatitis C and advanced fibrosis/cirrhosis underwent, on the same occasion, liver elastography, upper endoscopy, a platelet count and serum Gas6 measurement. RESULTS:  A total of 74/160 (46%) patients had esophageal varices, that were small (diameter <5 mm) in 57/160 (34%) and large in 17/160 (11%) cases. A total of 34/160 (21%) patients satisfied Baveno VI criteria, according to which screening for esophageal varices could have been omitted; 1/34 had large varices (sensitivity 94%). A plasma Gas6 value <45 ng/ml, detected in 34/160 (21%) patients, was also 94% sensitive. CONCLUSION: Plasma Gas6 might represent a feasible alternative to Baveno VI criteria when transient elastography is unavailable/unsuccessful.

mir-155 expression is downregulated in kidney transplant patients with human cytomegalovirus infection.

MicroRNAs are small, noncoding RNAs that control expression of target genes through inhibiting protein translation or inducing degradation of mRNA transcripts of target genes. According to the latest update, 2578 unique mature miRNAs are currently annotated in the human genome and participate in the regulation of multiple events, such as cellular proliferation or apoptosis. Herpesvirus family comprehends many viruses able to control and modulate host-cell processes permitting the survival by a latency phase after primary infection. Recently has been attested that Human Cytomegalovirus, which belongs to Herpesvirus family, can alter human miRNAs expression in vitro, and, in particular, downregulate mir-155 expression. In this study 20 kidney transplant patients positive to Human Cytomegalovirus infection and 11 negative were enrolled. The patients' positive to Human Cytomegalovirus infections have been subdivided into two groups: one group including patients with a low viral load and one including patients with a high viral load. The mir-155 expression profile has been evaluated by a stem-loop Real Time PCR in all these conditions to observe differences among the groups and compare the results obtained with the literature. The comparison between kidney transplant patients negative to Human Cytomegalovirus infection and patients with a high viral load showed a not significant difference in terms of mir-155 expression. However, considering low viral load group or the group including both high and low viral load patients, mir-155 expression levels decreased significantly. Considering this data together, it is possible confirm data published before and assert that Human Cytomegalovirus is responsible of mir-155 downregulation.

A novel TaqMAMA assay for allelic discrimination of TLR9 rs352140 polymorphism.

TaqMAMA is an allele-specific PCR-based (ASPCR) method that may be suitable for broad and cost-effective genotyping applications in all types of laboratories. There is evidence that interactions between some toll like receptors (TLRs) with viruses influence both the immune response and outcome of HCMV infection. We developed a TaqMAMA genotyping assay for the detection of rs352140 TLR9 polymorphism in transplant recipients with and without HCMV infections. Performance parameters to ensure a solid pre-validation protocol have been here argued. We analysed a population of 74 kidney transplants recipients subdivided in 58 HCMV PCR positive and 16 HCMV PCR negative in the post-transplant routine control. All 74 samples were tested with 31/74 (41.9%) homozygotes (11 CC and 20 TT) and 43/74 (58.1%) heterozygotes (CT). Our preliminary data suggest that there is no correlation between TLR9 rs352140 polymorphism and frequency of HCMV infection.

Nation-wide measure of variability in HCMV, EBV and BKV DNA quantification among centers involved in monitoring transplanted patients.

BACKGROUND: Inter-laboratory variability in quantifying pathogens involved in viral disease following transplantation may have a great impact on patient care, especially when pre-emptive strategies are used for prevention. OBJECTIVES: The aim of this study was to analyze the variability in quantifying CMV, EBV and BKV DNA from 15 virology laboratories of the Italian Infections in Transplant Working Group (GLaIT) involved in monitoring transplanted patients. STUDY DESIGN: Panels from international Quality Control programs for Molecular Diagnostics (QCMD, year 2012), specific for the detection of CMV in plasma, CMV in whole blood (WB), EBV and BKV were used. Intra- and inter-laboratory variability, as well as, deviations from QCMD consensus values were measured. RESULTS: 100% specificity was obtained with all panels. A sensitivity of 100% was achieved for EBV and BKV evaluations. Three CMV samples, with concentrations below 3 log10 copies/ml, were not detected by a few centers. Mean intra-laboratory variability (% CV) was 1.6 for CMV plasma and 3.0 for CMV WB. Mean inter-laboratory variability (% CV) was below 15% for all of the tested panels. Inter-laboratory variability was higher for CMV in WB with respect to the CMV plasma panel (3.0 vs 1.6% CV). The percentiles 87.7%, 58.6%, 89.6% and 74.7% fell within±0.5 log10 difference of the consensus values for CMV plasma, CMV WB, EBV and BKV panels, respectively. CONCLUSIONS: An acceptable intra- and inter-laboratory variability, in comparison with international standards was observed in this study. However, further harmonization in viral genome quantification is a reasonable goal for the future.

Human Cytomegalovirus Impact on mir-146a Expression Levels in Kidney Transplant Patients.

It is known that several viruses can alter or modulate the transcriptional and translational activity of host cells to obtain a rapid and efficient replication. In particular, Human Cytomegalovirus (HCMV) can interact with host cell at multiple levels, even modulating the expression of small signaling molecules called microRNAs. Especially, human miRNA mir-146a expression seems to be downregulated by HCMV infection in vitro. The aim of this study was to evaluate mir-146a expression in kidney transplant patients during HCMV infection. Sixty-four serum samples from 22 kidney transplant patients were analyzed and subdivided in three groups (high viral load, low viral load, and absent viral load). Mir-146a expression for each sample has been evaluated by a specific stem-loop Real Time polymerase chain reaction, and a statistical analysis was performed. Expression levels of mir-146a were similar among the three groups tested showing no statistical significant difference. Results obtained did not confirm data previously reported in literature, but the change of mir-146a expression levels has to be more clearly defined as it could not be directly caused by virus replication.