The value of West Nile virus RNA detection by real-time RT-PCR in urine samples from patients with neuroinvasive forms.

INTRODUCTION: Routine laboratory screening is based on the detection of WNV specific IgM and IgG in blood and cerebrospinal fluid. Confirmation is then classically applied by real time RT-PCR (rRT-PCR) in Cerebrospinal fluid (CSF), which often gives negative results due to too short virorachia and late sampling. rRT-PCR was applied-for the first time for routine diagnosis purpose-on urine samples. METHODS: During 2018 outbreak in Tunisia, 107 patients presented WNV neurologic symptoms and were positive for WNV serology. Of them, 95 patients were sampled for urine and 35 were sampled for CSF. Qualitative rRT-PCR was performed on both type of samples. RESULTS: WNV RNA was detected in 50.5% of urine samples (48/95) and in 2.8% of CSF samples (1/35). WNV RNA was detectable from day 1 to day 41 from symptom onset, however, positive urine rate was 53.1% during the first 10 days from symptom onset. The proportions of urine-positive and urine-negative samples, based on day of collection, showed no statistical difference (p > 0.005). Cycle threshold (Ct) values ranged from 12 to 39, with no correlation with the day of collection. The lowest Ct value was detected for urine sampled on day 5 after symptom onset. A statistically significant difference was found between age groups of confirmed and non confirmed cases (p < 0.001). DISCUSSION/CONCLUSION: Our study reported the use of rRT-PCR on urine samples as a confirmatory diagnostic tool for WNV "probable cases" during an outbreak. Our findings underlined the reliability and the rapidity of this confirmatory tool, even late, and showed its superiority on CSF investigation.

The role of quantitative HBsAg in the natural history of e antigen-negative chronic hepatitis B: A Tunisian prospective study.

BACKGROUND/AIMS: During the natural course of Chronic Hepatitis B (CHB) infection, differentiation between inactive carrier (IC) and HBeAg negative CHB is a subject of ongoing debate. We studied the role of hepatitis B surface antigen (HBsAg) level as a means of differentiating between CHB and IC in a group of untreated chronic HBeAg-negative HBV-infected patients. STUDY: A total of 115 HBeAg negative carriers were enrolled and followed up for>12 months; 78 as inactive carriers (IC), and 37 as active carriers (AC). Among ACs, patients were categorized according to the highest rate of viral load: AC1 (n=23), active carriers with persistent HBV-DNA<20,000 IU/mL; AC2 (n=14), active carriers with HBV-DNA>20,000 IU/mL. RESULTS: HBsAg levels were higher in AC compared to IC patients (1607 IU/ml vs. 225 IU/ml respectively, P=0.001). Among the AC group, the 23 AC1 cases had HBsAg levels significantly lower than the 14 AC2 patients (1756 IU/mL vs. 3327 IU/mL respectively; P<10-3). HBsAg showed weak correlation with HBV-DNA in the whole cohort (r=0.44, P<0.01). The suggested cutoff value of HBsAg titer to differentiate between the two groups was 938 IU/mL. Combined single-point quantification of HBsAg (938 IU/mL) and HBV DNA (2000 IU/mL) identified IC with 87.2% specificity and 91.7% positive predictive value. CONCLUSION: This study confirms the predictability of a one-time combined HBsAg and HBV DNA measurement for true inactive carriers requiring neither strict follow-up nor antiviral treatment.

Distribution of rotavirus VP7 and VP4 genotypes circulating in Tunisia from 2009 to 2014: Emergence of the genotype G12.

Group A rotavirus (RVA) represents the most important aetiological agent of diarrhoea in children worldwide. From January 2009 to December 2014, a multi-centre study realized through 11 Tunisian cities was undertaken among children aged <5 years consulting or hospitalized for acute gastroenteritis. A total of 1127 faecal samples were collected. All samples were screened by ELISA for the presence of RVA antigen. RVA-positive samples were further analyzed by PAGE and used for G/P-genotyping by semi-nested multiplex RT-PCR. Globally, 270 specimens (24 %) were RVA-positive, with peaks observed annually between November and March. Nine different electropherotypes could be visualized by PAGE, six with a long profile (173 cases) and two with a short one (seven cases). Mixed profiles were detected in two cases. Among the 267 VP7 genotyped strains, the predominant G- genotype was G1 (39.6 %) followed by G3 (22.2 %), G4 (13 %), G9 (11.5 %), G2 (5.2 %) and G12 (5.2 %). Among the 260 VP4 genotyped strains, P[8] genotype was the predominant (74.5 %) followed by P[6] (10.4 %) and P[4] (5.5 %). A total of 257 strains (95.2 %) could be successfully G- and P-genotyped. G1P[8] was the most prevalent combination (34.4 %), followed by G3P[8] (16.3 %), G9P[8] (10.3 %), G4P[8] (8.9 %), G2P[4] (4 %), G12P[6] (2.6 %) and G12P[8] (1.9 %). Uncommon G/Pgenotype combinations, mixed infections and untypeable strains were also detected. This is the first report, in Tunisia, of multiple detection of an emerging human RVA strain, G12 genotype. This study highlighted the need for maintaining active surveillance of emerging strains in Northern Africa.