Viral Protein VP1 Virus-like Particles (VLP) of CVB4 Induces Protective Immunity against Lethal Challenges with Diabetogenic E2 and Wild Type JBV Strains in Mice Model.

Several epidemiological studies demonstrated that coxsackievirus B4 (CVB4) causes viral pancreatitis and can ultimately result in type 1 diabetes mellitus (T1D). Prevention of CVB4 infection is therefore highly desirable. There is currently no vaccine or antiviral therapeutic reagent in clinical use. VLP are structurally similar to native virus particles and therefore are far better immunogens than any other subunit vaccines. Many studies have shown the potential of capsid protein VP1 on providing protective effects from different viral strains. In this study, we contributed towards the development of a CVB4 VLP-based vaccine from the total protein VP1 of the diabetogenic CVB4E2 strain and assessed whether it could induce a protective immunity against both the wild-type CVB4JBV and the diabetogenic CVB4E2 strains in mice model. Serum samples, taken from mice immunized with VLP, were assayed in vitro for their anti-CVB4 neutralizing activity and in vivo for protective activity. We show that VLP vaccine generates robust immune responses that protect mice from lethal challenges. Results demonstrate that CVB4 VP1 capsid proteins expressed in insect cells have the intrinsic capacity to assemble into non-infectious VLP, which afforded protection from CVB4 infection to mice when used as a vaccine.

Molecular epidemiology and clinical characterization of group A rotavirus infections in Tunisian children with acute gastroenteritis.

Rotaviruses are the most common cause of severe viral gastroenteritis in early childhood worldwide. Thus, the objectives of our study were to determine the molecular epidemiology and the clinical features of rotavirus gastroenteritis in Tunisia. Between January 2003 and April 2007, a prospective study was conducted on 788 stool samples collected from children under 12 years of age who were suffering from acute gastroenteritis. Rotavirus was detected by multiplex RT-PCR in 27% (n = 213) of samples, among them 79.3% (n = 169) cases were monoinfections. The frequency of rotavirus infections was significantly higher among inpatients (29%) than among outpatients (13%) (P < 0.001). The seasonal distribution of rotavirus diarrhea showed a winter peak, with an unusual peak from June to September. The mean duration of hospitalization was 6.5 ± 8.1 days and the mean age was 15.8 ± 22.8 months for rotavirus monoinfections. Fever, vomiting, abdominal pain, and dehydration were observed in 88, 98, 13, and 80 cases, respectively, in children with rotavirus monoinfections. G3P[8] (45.6%) and G1P[8] (23.9%) were the most common genotypes found in our study. The determination of rotavirus infection prevalence and the characterization of the rotavirus strains circulating will help us to better understand the molecular biology and epidemiology of the disease in our country.

Bacteriological and physico-chemical assessment of wastewater in different region of Tunisia: impact on human health.

BACKGROUND: In many parts of the world, health problems and diseases have often been caused by discharging untreated or inadequately treated wastewater. In this study, we aimed to control physico-chemical parameters in wastewater samples. Also, microbiological analyses were done to reveal Salmonella strains and each Escherichia coli (E.coli) pathotype. FINDINGS: Sixty wastewater samples were collected from fifteen different regions of Tunisia. All physico-chemical parameters (pH, residual free chlorine, total suspended solids, biological oxygen demand, and chemical oxygen demand) were evaluated.For microbiological analyses, samples were filtered to concentrate bacteria. DNA was extracted by boiling and subjected to polymerase chain reaction (PCR) using different pairs of primers.The mean pH values recorded for the sampling point were above the WHO pH tolerance limit. The total suspended solids (TSS) concentrations varied between 240 mg/L and 733 mg/L in entrance points and between 13 mg/L and 76 mg/L in exit points. In entrance points, the studied wastewater has an average COD concentration that varied between 795 mg/mL to 1420 mg/mL. Whereas, BOD concentration of the wastewater ranged between 270 mg/L to 610 mg/L. In exit points, COD concentration varied between 59 mg/L and 141 mg/L, whereas BOD concentration ranged from 15 mg/L to 87 mg/L.The bacteriological control of wastewaters showed that, in entrance points, Escherichia coli (E.coli) was detected at the rate of 76.6%. Three E.coli pathotypes were found: ETEC (53.3%), EAEC (16.6%) and EIEC (6.6%).Concerning the ETEC isolated strains, 8 of 16 (50%) have only the heat-labile toxin gene, 5 of 16 (31.2%) present only the heat-stable toxin gene and 3 of 16 (18.7%) of strains possess both heat-labile toxin gene and heat-stable toxin gene. In exist point, the same pathotypes were found but all detected ETEC strains present only the "est" gene.Concerning Salmonella isolated strains; percentages of 66.6% and 20% were found in entrance and exit points respectively. CONCLUSIONS: Wastewaters contain a large amount of pathogenic bacteria that present a real impact on human health. Assessment wastewater treatment stations have to consider in account enterobacterial pathogens as potential pathogens that should be correctly controlled.

Molecular detection of genogroup I sapovirus in Tunisian children suffering from acute gastroenteritis.

This study investigated the prevalence of sapovirus infections in children with acute gastroenteritis in Monastir region, Tunisia, from January 2003 to April 2007. Sapovirus was characterized by sequence and phylogenetic analyses of the partial polymerase gene. From 788 fecal specimens tested, 6 (0.8%) were positive for sapovirus, of these, 4 (66.7%) were monoinfections. All sapovirus positive samples were detected in outpatient, contrary to norovirus which was significantly more frequent in hospitalized children than in outpatients (14.5 vs. 9.5%, P = 0.03). The mean age of children with sapovirus infections was 11 ± 5.56 months (range 6-19 months). Sapovirus isolates were detected in March and between September and December 2003. Fever, vomiting, abdominal pain, and dehydration were not observed in patients with sapovirus infections. Analysis of nucleotide and amino acid sequences revealed that all 6 Tunisian sapovirus strains clustered in the GGI/1 genotype and strains were identical in the region sequenced, sharing 90.2% nucleotide identity with the reference strain Sapporo/82/JP (U65427). This represents the first finding of sapovirus infections in North Africa and especially in Tunisia. The data indicate that, contrary to norovirus which can cause severe diarrhea and is an important etiologic agent in hospitalized cases, sapovirus causes mild gastroenteritis in Tunisian children.

Aichi virus IgG seroprevalence in Tunisia parallels genomic detection and clinical presentation in children with gastroenteritis.

Aichi virus has been described as a novel causative agent of gastroenteritis in humans. In this study, we report the seroprevalence distribution of Aichi virus in Tunisia. A panel of 1,000 sera was screened by applying an enzyme-linked immunosorbent assay for immunoglobulin G specific for Aichi virus. A considerable prevalence (92%) of antibody to Aichi virus was found across all age groups. The specific anti-Aichi virus antibodies increased with age, from a high rate (68.8%) in children under 10 years old to about 100% in persons more than 60 years old. We found a statistically significant increase in levels of antibody to Aichi virus according to the age of patients. Immunoglobulin M antibodies were detected among five children. A high frequency of Aichi virus monoinfections in hospitalized children with severe gastroenteritis was previously observed in Tunisia. Aichi virus causes diarrhea with dehydration, fever, and vomiting. This work is the first to establish a correlation between the high seroprevalence of specific Aichi virus antibodies, clinical presentation, and a high frequency of isolation of Aichi virus by genomic characterization in stools of children suffering from gastroenteritis. Our data show the importance and emerging character of Aichi virus in the viral etiology of pediatric gastroenteritis.

[Detection of enteric viruses in wastewater of Monastir region by RT-PCR method]. / Détection des virus entériques dans les eaux usées de la région de monastir par RT-PCR.

BACKGROUND: Wastewater frequently has been contaminated by enteric viruses, which can cause gastroenteritis, hepatitis, and other viral diseases. Since current water treatments do not ensure their complete removal and they become contaminants of the water environment. AIM: In order to monitor and assess the prevalence and the seasonal incidence of enteric viruses contaminating wastewater, a total of 93 samples were collected from the stations of Sayada-Lamta-Bouhjar and El Frina. METHODS: The purpose of this study is to establish a rapid, sensitive and specific RT-PCR method using highly conserved primers to detect enteric viruses (caliciviruses, enteroviruses and hepatitis A viruses) circulating in the area of Monastir, during the period from October 2005 to July 2006. RESULTS: Our study shows that the caliciviruses were detected in 67.56% of the wastewater samples and present an essentially winter distribution with a specific peak in spring. The enteroviruses were detected in 35.13% of the samples tested and present a regular circulation during the year with a predominance in the summer and early fall. Finally, the presence of hepatitis A viruses was noted especially in winter in 10.81% of the wastewater samples. CONCLUSION: We notice that current wastewater treatment methods are sometimes not efficient to eliminate enteric viruses.

Molecular epidemiology of human astrovirus and adenovirus serotypes 40/41 strains related to acute diarrhea in Tunisian children.

Human astrovirus (AstV) and adenovirus types 40 and 41 (AdV 40/41) are responsible for epidemic and endemic acute gastroenteritis in children and adults. The present study was designed to evaluate the prevalence and genetic diversity of enteric viruses in children in Tunisia. A total of 788 fecal samples were collected during a 4-year period in the region of Monastir, from children under 12 years old, hospitalized or presenting in dispensaries with symptoms of acute gastroenteritis. AstV and AdV40/41 were detected by immunoenzymatic methods and confirmed by PCR/RT-PCR and sequence analysis. Phylogenetic analyses were performed for nucleotide homology with reference strains. AstV and AdV40/41 were characterized as a causative agent in 28 (3.6%) and 18 (2.3%) of the fecal samples, respectively. Phylogenetic analysis showed that the AstVs belonged to the serotypes 3 (n = 4; 14.3%) and 1 (n = 24; 85.7%), and the enteric AdVs to the serotypes 40 (n = 1; 5.6%) and 41 (n = 17; 94.4%). This is the first report that describes the molecular epidemiology of AstV and AdV40/41 in Tunisian children. Their respective detection rate was very low, far below that of rotavirus and norovirus. The genetic diversity among these two viruses is relatively limited and varies depending on the area.

Detection and genomic characterization of Aichi viruses in stool samples from children in Monastir, Tunisia.

Aichi virus has been associated with acute gastroenteritis in adults and children. Stool samples were collected from 788 Tunisian children suffering from diarrhea. Aichi virus was found in 4.1% of the cases. The high proportion of monoinfections and the high frequency of hospitalizations support the role of Aichi virus in pediatric gastroenteritis.

Molecular epidemiology of norovirus gastroenteritis investigated using samples collected from children in Tunisia during a four-year period: detection of the norovirus variant GGII.4 Hunter as early as January 2003.

Human noroviruses (NoVs) cause epidemic and endemic acute gastroenteritis in children and adults. To study the prevalence and genetic diversity of NoV in children in Tunisia, a total of 788 fecal samples were collected during a 4-year period in the region of Monastir, from children 12 years of age or younger, hospitalized or presenting in dispensaries with symptoms of acute gastroenteritis. NoV was detected by reverse transcription-PCR and confirmed by sequence analysis. This is the first report that describes the molecular epidemiology of NoV in Tunisian children: NoVs were characterized as the causative agent in 128 (16.2%) of the samples. Fourteen samples contained a mixture of two NoVs, and 33 samples were coinfected with additional enteric viruses. Eight distinct NoV genotypes were detected (GGI.2, GGI.4, GGII.1, GGII.4, GGII.8, GGII.14, GGIIb/GGII.2, and GGIIb/GGII.3). GGII.4 was the most prevalent genotype, accounting for 83 (64.8%) cases. Interestingly the GGII.4 variant Hunter, described as spreading all over the world in 2004, was found in Tunisia as early as January 2003. The delay of 1 year between the isolation in Tunisia and the worldwide emergence is somewhat surprising, considering the importance of the contacts between North Africa and Europe particularly. Nevertheless, this illustrates the idea that sporadic gastroenteritis cases may be a reservoir for emerging epidemic NoV strains.

Acute infantile gastroenteritis associated with human enteric viruses in Tunisia.

This prospective study, conducted from January 2003 to June 2005, investigated the incidence and the clinical role of various enteric viruses responsible for infantile gastroenteritis in 632 Tunisian children presenting in dispensaries (380 children) or hospitalized (252 children) for acute diarrhea. At least one enteric virus was found in each of 276 samples (43.7%). A single pathogen was observed in 234 samples, and mixed infections were found in 42 samples. In terms of frequency, rotavirus and norovirus were detected in 22.5 and 17.4% of the samples, respectively, followed by astrovirus (4.1%), Aichi virus (3.5%), adenovirus types 40 and 41 (2.7%), and sapovirus (1.0%). The seasonal distribution of viral gastroenteritis showed a winter peak but also an unusual peak from May to September. The severity of the diarrhea was evaluated for hospitalized infants. No significant differences were observed between rotavirus and norovirus infections with regard to the incidence and the clinical severity of the disease, especially in dehydration.