Large spontaneous HBV DNA fluctuations and potential usefulness of a single-point measurement of combined HBV DNA and quantitative HBsAg for the exclusion of HBeAg-negative chronic hepatitis B: A prospective Tunisian cohort study.

BACKGROUND AND STUDY AIM: During the natural course of HBeAg-negative chronic hepatitis B (CHB), fluctuations in hepatitis B virus (HBV) DNA and alanine aminotransferase (ALT) levels are often observed, making the classification of patients difficult. We aimed to describe spontaneous short-term HBV DNA level fluctuations and to assess the usefulness of qHBsAg in Tunisian patients with HBeAg-negative chronic HBV infection. PATIENTS AND METHODS: We included 174 treatment-naive Tunisian patients with HBeAg-negative chronic HBeAg-negative HBV infection. A prospective 1-year follow-up was conducted with serial determinations of HBV DNA, ALT levels, and qHBsAg. The patients were classified into three groups: inactive carriers (G1), patients with negative HBeAg CHB (G2), and patients with an "indeterminate state" (G3). For the latter group, a liver biopsy was indicated. RESULTS: Only genotype D was detected. During follow-up, 21.6% and 19.5% of patients with a low initial (<2,000 IU/ml) and intermediate viral load (2,000-20,000 IU/ml) experienced a subsequent increase in their HBV DNA levels above 2,000 and 20,000 IU/ml, respectively. Significant variations in viral load were observed in 61.1% of patients at 6-month intervals. Among the 174 patients, 89 (51.1%) belonged to G1, 33 (19%) to G2, and 52 (29.9%) to G3. Fourteen patients have undergone a liver biopsy, of whom seven showed moderate to severe liver disease. Combination of HBV DNA < 2,000 IU/ml and qHBsAg < 832 IU/ml excluded CHB in 98.4% of cases. A cutoff point for qHBsAg < 100 IU/ml associated with an annual decline of > 0.5 log 10 IU/ml is a good predictor marker of functional cure for hepatitis B. CONCLUSIONS: This study highlights the large short-term fluctuations in HBV DNA in patients with HBeAg-negative chronic HBeAg-negative HBV infection with genotype D. Thus, using the cutoff value of 832 for qHBsAg combined with that of 2,000 for HBV DNA makes it possible to exclude CHB for most patients.

Significant impact of COVID-19 pandemic on the circulation of respiratory viruses in Tunisia, 2020-2021.

Background: The COVID-19 pandemic changed the typical patterns of respiratory infections globally. While SARS-CoV-2 illness exhibited explosive growth since 2020, the activity of other respiratory viruses fell below historical seasonal norms. The objective of this study was to assess the prevalence of seasonal respiratory viruses during the COVID-19 pandemic in Tunisia. Methods: This is a retrospective cross-sectional study including 284 nasopharyngeal samples tested negative for SARS-CoV-2 during the period October 2020-May 2021. All samples were screened for fifteen common respiratory viruses. Either a fast syndromic approach using Biofire FILM ARRAY respiratory 2.1 (RP2.1) Panel, or end-point multiplex RT-PCRs detecting RNA viruses and Real-Time PCR detecting Adenoviruses were used. Results: Overall, 30.6% (87/284) of samples were positive for at least one virus. Mixed infections were detected in 3.4% of positive cases. Enterovirus/Rhinovirus (HEV/HRV) was the most detected virus throughout the study period, especially during December 2020 (33.3% of all HEV/HRV being detected). During the 2020-2021 winter season, neither Respiratory Syncytial Virus nor Influenza Viruses circulation was observed. Metapneumovirus and Parainfluenza Viruses infections were detected during the spring season. The highest rate of respiratory viruses detection was observed in children and adults aged [0-10] years (50%) and [31-40] years (40%). HEV/HRV was the most detected virus regardless of age group. Conclusions: Public health measures used to prevent SARS-CoV-2 spread in Tunisia were also effective to reduce transmission of the other respiratory viruses, especially Influenza. The higher resistance of HEV/HRV in the environment could explain their predominance and continuous circulation during this period.

Development of a simple genotyping method based on indel mutations to rapidly screen SARS-CoV-2 circulating variants: Delta, Omicron BA.1 and BA.2.

The high need of rapid and flexible tools that facilitate the identification of circulating SARS-CoV-2 Variants of Concern (VOCs) remains crucial for public health system monitoring. Here, we develop allele-specific (AS)-qPCR assays targeting three recurrent indel mutations, ΔEF156-157, Ins214EPE and ΔLPP24-26, in spike (S) gene to identify the Delta VOC and the Omicron sublineages BA.1 and BA.2, respectively. After verification of the analytical specificity of each primer set, two duplex qPCR assays with melting curve analysis were performed to screen 129 COVID-19 cases confirmed between December 31, 2021 and February 01, 2022 in Sfax, Tunisia. The first duplex assay targeting ΔEF156-157 and Ins214EPE mutations successfully detected the Delta VOC in 39 cases and Omicron BA.1 in 83 cases. All the remaining cases (n = 7) were identified as Omicron BA.2, by the second duplex assay targeting Ins214EPE and ΔLPP24-26 mutations. The results of the screening method were in perfect concordance with those of S gene partial sequencing. In conclusion, our findings provide a simple and flexible screening method for more rapid and reliable monitoring of circulating VOCs. We highly recommend its implementation to guide public health policies.

Evidence of SARS-CoV-2 symptomatic reinfection in four healthcare professionals from the same hospital despite the presence of antibodies.

OBJECTIVES: Since the onset of the COVID-19 pandemic, cases of reinfection with SARS-CoV-2 have been reported, raising additional public health concerns. SARS-CoV-2 reinfection was assessed in healthcare workers (HCWs) in Tunisia because they are at the greatest exposure to infection by different variants. METHODS: We conducted whole-genome sequencing of the viral RNA from clinical specimens collected during the initial infection and the suspected reinfection from 4 HCWs, who were working at the Habib Bourguiba University Hospital (Sfax, Tunisia) and retested positive for SARS-CoV-2 through reverse transcriptase-polymerase chain reaction (RT-PCR) after recovery from a first infection. A total of 8 viral RNAs from the patients' respiratory specimens were obtained, which allowed us to characterize the differences between viral genomes from initial infection and positive retest. The serology status for total Ig, IgG, and IgM against SARS-CoV-2 was also determined and followed after the first infection. RESULTS: We confirmed through whole-genome sequencing of the viral samples that all 4 cases experienced a reinfection event. The interval between the 2 infection events ranged between 45 and 141 days, and symptoms were milder in the second infection for 2 patients and more severe for the remaining 2 patients. Reinfection occurred in all 4 patients despite the presence of antibodies in 3 of them. CONCLUSION: This study adds to the rapidly growing evidence of COVID-19 reinfection, where viral sequences were used to confirm infection by distinct isolates of SARS-CoV-2 in HCWs. These findings suggest that individuals who are exposed to different SARS-CoV-2 variants might not acquire sufficiently protective immunity through natural infection and emphasize the necessity of their vaccination and the regular follow-up of their immune status both in quantitative and qualitative terms.

The Delta variant wave in Tunisia: Genetic diversity, spatio-temporal distribution and evidence of the spread of a divergent AY.122 sub-lineage.

Introduction: The Delta variant posed an increased risk to global public health and rapidly replaced the pre-existent variants worldwide. In this study, the genetic diversity and the spatio-temporal dynamics of 662 SARS-CoV2 genomes obtained during the Delta wave across Tunisia were investigated. Methods: Viral whole genome and partial S-segment sequencing was performed using Illumina and Sanger platforms, respectively and lineage assignemnt was assessed using Pangolin version 1.2.4 and scorpio version 3.4.X. Phylogenetic and phylogeographic analyses were achieved using IQ-Tree and Beast programs. Results: The age distribution of the infected cases showed a large peak between 25 to 50 years. Twelve Delta sub-lineages were detected nation-wide with AY.122 being the predominant variant representing 94.6% of sequences. AY.122 sequences were highly related and shared the amino-acid change ORF1a:A498V, the synonymous mutations 2746T>C, 3037C>T, 8986C>T, 11332A>G in ORF1a and 23683C>T in the S gene with respect to the Wuhan reference genome (NC_045512.2). Spatio-temporal analysis indicates that the larger cities of Nabeul, Tunis and Kairouan constituted epicenters for the AY.122 sub-lineage and subsequent dispersion to the rest of the country. Discussion: This study adds more knowledge about the Delta variant and sub-variants distribution worldwide by documenting genomic and epidemiological data from Tunisia, a North African region. Such results may be helpful to the understanding of future COVID-19 waves and variants.

Treatment of chronic hepatitis C with sofosbuvir in a hemodialysis patient: a case report.

The treatment of chronic hepatitis C virus (HCV) infection in chronic hemodialysis patients remains an issue of great concern for nephrologists. In 2008 the kidney disease improving global outcomes working group suggested the use of pegylated interferon in end stage kidney disease patients treated by dialysis. Since then, series and some clinical trials on different direct-acting antiviral agents have shown better efficacy and tolerance than interferon-based regimens. Data on the efficacy, tolerance and the right dose of sofosbuvir in this population are still unclear. We report a case of chronic HCV genotype 1b infection in a 47-year-old patient on maintenance hemodialysis successfully treated by a combination of sofosbuvir and ledipasvir for 12 weeks. Evolution was marked by the complete regression of the hepatic cytolysis, a complete and sustained virologic response with HCV viral load undetectable for a 24 months follow-up period. No adverse reaction was found. The treatment of HCV genotype 1 or 4 infection in patients on maintenance hemodialysis is possible with sofosbuvir based regimens with a good efficacy/safety ratio in the absence of current recommended drugs for patients with eGFR<30ml/min/1.73m2. The prescription of sofosbuvir should be encouraged amongst this population in this setting.

Focus on hepatitis C virus genotype distribution in Tunisia prior to elimination: a 16-year retrospective study.

With the introduction of direct-acting antiviral treatment (DAA), Tunisia has committed to achieving the international goal of eliminating viral hepatitis. Because the specific DAA prescribed depends on viral genotype, viral genotyping remains of great importance. The aim of the present study was to outline the trends in the distribution of HCV genotypes from 2002 to 2017 in the Tunisian general population in order to guide authorities towards the most appropriate therapeutic strategies for preventing HCV infection. A total of 2532 blood samples were collected over a 16-year period and from all regions of Tunisia. Genotyping showed that genotype 1 (subtype 1b) was the most prevalent genotype in the country (n = 2012; 79.5%), followed by genotype 2 (n = 339; 13.3%). Genotypes 3, 4 and 5 were detected in 4.8%, 2.2% and 0.1% of the country's population, respectively. Mixed infections with different HCV genotypes were detected in 0.1% of the population (one case each of genotypes 1b + 4, 1b + 2 and 2 + 4). Interestingly, a significant increase in genotypes 2, 3 and 4 was observed over time (p = 0.03). Sixteen different subtypes were detected over the study period, most of which were subtypes of genotype 2, and some of these subtypes appeared to be new. Patients infected with genotypes 1a, 3 and 4 were significantly younger than those infected with genotypes 1b and 2 (p < 0.01). Furthermore, genotypes 1b and 2 were detected more often in women than men, while genotypes 1a and 3 were detected mostly in men (P < 0.01). Our study confirms a large predominance of genotype1/subtype1b in Tunisia and shows a significant increase in the prevalence of other genotypes over time. These findings reinforce the need for an additional HCV genotype survey to improve the design of treatment strategies in Tunisia.

The intrathecal polyspecific antiviral immune response (MRZ reaction): A potential cerebrospinal fluid marker for multiple sclerosis diagnosis.

We tested the performance of MRZ-reaction, an intrathecal humoral immune response against-Measles (M), Rubella (R) and Varicella Zoster (Z) viruses, in multiple sclerosis (MS) diagnosis. The MRZ-reaction was significantly more positive in MS than in non-MS group with a specificity of 91.9%. In MS group, the RZ-profile was the most prevalent and the R-specific antibody-index was correlated to the number of oligoclonal bands (OCB) in CSF. Interestingly, the MRZ-reaction was detected in 53% of OCB-negative-MS patients. The MRZ-reaction seems to be a relevant CSF diagnostic marker of MS disease. The likely relation between its positivity and the vaccination status deserves to be investigated.

Clinical epidemiological and molecular aspects of rubella outbreak with high number of neurological cases, Tunisia 2011-2012.

BACKGROUND: A large and unusually prolonged rubella outbreak occurred in Tunisia from April 2011 to July 2012 and was characterized by a high number of neurological cases. OBJECTIVES: To describe the outbreak and to perform virus genotyping of isolated virus strains. STUDY DESIGN: From January 2011 to December 2012, 5000 sera for serological diagnosis of acute rubella and 31 cerebrospinal fluid from patients with neurological symptoms were tested for the presence of rubella immunoglobulins G and M. Real-time PCR was performed on 49 throat swabs, 21 cerebrospinal fluid and 27 serum samples. Positive samples were assessed for virus genotyping by sequencing and the obtained sequences were compared to those previously isolated in the country. RESULTS: Acute rubella was confirmed in 280 patients including 15 neonates, 217 children and adults with mild rash and 48 patients with severe rubella (mainly encephalitis, n = 39). Most of acquired rubella cases (60.7%) were aged over 12 years with a male predominance observed in the age group 12-25 years (79%). Females belonged essentially to the unvaccinated age groups under 12 and over 25 years. Among the 23 samples tested positive by real-time PCR, six could be genotyped and clustered with either the 1E genotype, previously detected in Tunisia, or the 2B genotype which has never been isolated in Tunisia before. CONCLUSIONS: Gender and age distributions of the patients reflect the impact of the selective rubella vaccination program adopted in Tunisia since 2005. Genotype 1E continues to circulate and genotype 2B was probably recently introduced in Tunisia.