Detection of antibodies to the hepatitis E virus in domestic reindeer (Rangifer tarandus) in the Republic of Sakha (Yakutia).

INTRODUCTION: Although domestic pigs and wild boars are the main reservoir of zoonotic hepatitis E virus (HEV) genotypes in temperate countries, the presence of antibodies to HEV (anti-HEV) in the indigenous population of circumpolar territories, i.e. outside the habitat of wild and domestic pigs, indicates the presence of an alternative reservoir of the virus. Reindeer (Rangifer tarandus) may be a potential reservoir for HEV in the polar regions. The purpose of the study was to determine the prevalence of anti-HEV among domestic reindeer in the Republic of Sakha (Yakutia). MATERIALS AND METHODS: Sera from 497 domestic reindeer from the Oymyakon (n = 425) and Ust-Yansky districts (n = 72) of the Republic of Sakha (Yakutia) were tested for anti-HEV. A commercial ELISA kit DS-ELISA-ANTI-HEV-G (Diagnostic Systems-Stolitsa LLC, Russia) was used for detection of anti-HEV IgG, but a rabbit polyclonal antibody against deer IgG labeled with horseradish peroxidase (KPL, USA) at a dilution of 1 : 100 in phosphate-buffered saline were used instead of the human specific conjugate from the kit. RESULTS: The average detection rate of anti-HEV in reindeer sera was 15.5% (95% CI: 12.6-19.0%). The detection rate of anti-HEV significantly increased with age, from 3.5% (95% CI: 1.1-9.0%) in calves aged 3-6 months to 25.0% (95% CI: 1.6 -36.5%) in deer aged 2-4 years (p < 0.0001). From this age group, anti-HEV detection rates reached a plateau, not differing significantly between older age groups (p > 0.05). The average anti-HEV detection rate among reindeer 2 years of age and older was 19.0% (95% CI: 15.3-23.4%). There were no statistically significant differences in the frequency of anti-HEV detection between female and male reindeer, both among adult animals and among calves. CONCLUSION: The observed anti-HEV detection rates among domestic reindeer in the Republic of Sakha (Yakutia) indicate that infection caused by HEV or an antigenically similar virus is common in these animals. The dynamics of antibody accumulation in the reindeer population indicates that infection apparently occurs during the first two years of life.

[Detection of markers of hepatitis B and D virus infection in biological media and dried blood spots.]

The aim of this study was to assess the rates of detection of the major markers of infection with hepatitis B and Delta (D) viruses in serum, saliva and dry blood dots (DBS) as a possible option for serological studies among the population of the endemic region in conditions of limited laboratory resources. For this purpose, paired samples of blood serum and DBS, blood serum and saliva from patients with chronic hepatitis B with Delta agent living in the Republic of Tyva, which is endemic for this disease. HBsAg was detected in 289 (100%) serum samples, in 88/92 (95.7%) saliva samples, in 60/80 (75%) DBS samples, stored three years at room temperature, and in 111/117 (94.9%) DBS stored one year at the same conditions. Anti-HBcore was detected in 209 (100%) serum samples, while in saliva and DBS samples this marker was detected in only 13.04% (12/92) and 19.7% (23/117), respectively. Anti-HDV antibodies in serum were detected in 209 (100%) samples collected from patients in 2017-2018. In saliva and DBS anti-HDV were not detected in any sample. This difference in the detection rates of anti-HBcore and anti-HDV might be accounted for the fact that the HBV core protein is a very strong immunogen, indusing the production of anti-HBcore in high concentrations. Probably, the concentration of anti-HDV is much lower, which explains its absence in saliva and DBS in patients with hepatitis B+D. Samples of biological media (saliva), as well as DBS can serve as an alternative material for the detection of HBsAg in screening and research prevalence studies. Meanwhile, the definition of anti-HDV in such media is not possible due to the false negative results. Due to the high probability of superinfection with HDV in patients with HBV in endemic areas, the detection of HBsAg in alternative media (saliva or DBS) should be followed by testing for anti-HDV in serum samples.

Natural infection of captive cynomolgus monkeys (Macaca fascicularis) with hepatitis E virus genotype 4.

Data on natural HEV infection of infection in monkeys are limited. We report a case of hepatitis E virus genotype 4 infection in captive non-human primates (cynomolgus monkeys) imported from Vietnam. Phylogenetic analysis demonstrated that HEV infection was not the result of spillover from single source of infection, but rather the persistent circulation of HEV-4 among cynomolgus monkeys or multiple infections by related strains from a human or swine reservoir.

[Belgorod region - the territory endemic for hepatitis E.]

INTRODUCTION: Belgorod region is the territory with the highest incidence of hepatitis E in the Russian Federation. OBJECTIVES: The aim of the study was to comprehensively characterize the circulation of hepatitis E virus (HEV) in the Belgorod region, including the study of population immunity to the virus, determining the prevalence of infection among the pig population and analysis of the genetic diversity of HEV from patients and animals. MATERIAL AND METHODS: Serum samples of a conditionally healthy population (n = 2027) of all age groups were tested for anti-HEV IgG and IgM by ELISA with commercial assays. HEV RNA was determined in fecal samples from pigs aged 2-4 months (n = 526), in sewage samples from pig farms (n = 10), as well as in stool samples from patients with hepatitis E (n = 6) using reverse transcription polymerase chain reaction (RT-PCR). Phylogenetic analysis was performed for an amplified 300 nt fragment corresponding to HEV open reading frame 2. RESULTS AND DISCUSSION: The prevalence of anti-HEV IgG in general population averaged 16.4% (95% CI: 14.8-18.1; 332/2027). The proportion of individuals who had both anti-HEV IgM and IgG averaged 2.8% (95% CI: 2.2-3.6; 57/2027). The incidence rate of anti-HEV IgG increased with age, from 2.8% (95% CI: 1.3-5.8) in children aged 1-14 years to 40.1% (95% CI: 34.9-45.6) in people 70 years or older. The detection rate of HEV RNA in pigs was 20% (95% CI: 16.8-23.6; 105/526). HEV RNA was detected in 2 out of 10 sewage samples. The HEV sequences isolated from patients with hepatitis E, pigs, and sewage samples in Belgorod region belonged to the HEV genotype 3, had a 95-100% homology, and formed common clusters on a phylogenetic tree. CONCLUSIONS: The high prevalence of HEV in pigs population has led to the formation of an endemic territory in the Belgorod region, which is the center of pig breeding. Measures aimed at reducing the circulation of HEV among pig population and decontamination of sewage from pig farms are necessary to control HEV infection.

[The duration of preservation of anamnestic antibodies to hepatitis E virus.]

The purpose of the study was to determine the duration of antibody response against hepatitis E virus (anti-HEV). Veterans of the war in Afghanistan who were in this endemic region in the late 1970s and early 1980s were tested for anti-HEV. On average, 20 years after the end of military service in Afghanistan, the rate of seropositivity was 30.0% (95/317), which was significantly higher compared to positivity rates in males who were at military service in the territory of Russia during the same period (3.9%, 8/208). At an average of 29.5 years after the visit to Afghanistan anti-HEV prevalence in veterans dropped to 20.0% (21/105), but still significantly exceeded the seroprevalence in general population (3.8%). Serum samples from elderly individuals (>60 years) without known risk factors of infection were also tested for anti-HEV IgG and IgM (n = 896). Anti-HEV IgG detection rates in the elderly from two regions of Russia exceeded those observed of primary blood donors (18.0-27.8% vs. 4.5-10.0%, p<0.01). The detection of anti-HEV IgM in individuals above 60 years (2.7-6.9%) indicates a current or recent infection. Thus, anti-HEV IgG can persist several decades after the infection. This might account for the wide anti-HEV prevalence among the elderly. At the same time, a significant proportion of individuals exposed to HEV lose detectable anti-HEV IgG within 20-30 years. The detection of anti-HEV IgM among the elderly indicates the viral circulation in older age groups, suggesting the need for testing for hepatitis E markers in elderly patients with liver diseases.

[The standardization of diagnostic of hepatitis E.]

The study was carried out to develop approaches to standardization of laboratory diagnostic of hepatitis E. The three stages of standardization are establishment of analytical sensitivity of molecular test for detection of RNA of virus of hepatitis E; establishment of analytical sensitivity in International Units of enzyme-linked immunosorbent assay testing widely applied in Russia for detection of anti-virus of hepatitis E; And development of national reference material - standard anti-virus of hepatitis E IgG validated relatively to International standard. The results of study permitted to develop tools for standardizing of laboratory diagnostic of hepatitis E and epidemiological control of the given function - molecular test for detecting RNA of virus of hepatitis E with sensitivity within range of 1250250 IU/ml, data concerning analytical sensitivity of commercial enzyme-linked immunosorbent assay testing for detecting anti-virus of hepatitis E (0.25 IU/ml) and national standard of anti-virus of hepatitis E with concentration of 5 IU/ml.

[Prevalence of hepatitis E markers in children].

AIM: Frequency of detection determination for past and current hepatitis E virus (HEV) infection markers in children with immune suppression, as well as children with normal immune status. MATERIALS AND METHODS: The presence of HEV markers (anti-HEV IgG and IgM, HEV RNA) was studied in 609 sera samples of children with neurologic pathologies, 87 samples--from children with immune deficiencies, as well as 3122 samples from conditionally healthy children of 6 regions of Russia. The children were divided into 5 age groups. Anti-HEV IgG and IgM determination was carried out in EIA, HEV RNA--by RT-PCR. RESULTS: The frequency of detection of anamnestic anti-HEV IgG turned out to be significantly higher among immune-compromised. children compared with healthy children (5.7% against 1.4%, p < 0.05). Anti-HEV IgM, that testify to current or recent infection, were also detected significantly more frequently among children with immune-suppression (1.1-1.6%) compared with healthy children (0.25%, p < 0.05). HEV RNA was detected in 1 child with the absence of anti-HEV IgM and IgG. Nucleotide sequence analysis of HEV confirmed membership of this isolate in genotype 3, that is prevalent in non-endemic territories. CONCLUSION: The data obtained have demonstrated, that HEV-infection is prevalent among children in Russia and its course is, probably, asymptomatic in most cases. Immune suppression is a factor of increased risk of infection of children with HEV.