West Nile virus infections in Hungary: Epidemiological update and phylogenetic analysis of the Hungarian virus strains between 2015 and 2022.

Following the introduction of the West Nile virus (WNV) into Hungary in 2004, it has shortly become one of the most important human arbovirus infections, with a gradually increasing number of cases. The study aimed to summarize the current epidemiological situation in Hungary and sequence the WNV PCR-positive clinical specimens and virus isolates by next-generation whole genome sequencing (NGS) to obtain a detailed phylogenetic analysis of the circulating virus strains. Whole blood and urine samples from confirmed WNV-infected patients and WNV isolates were investigated by reverse transcription PCR assays. Genome sequencing was carried out by Sanger-method, followed by NGS on the Illumina MiSeq platform. Altogether 499 human infections were diagnosed between 2004 and 2022. A particularly remarkable increase in human WNV infections was observed in 2018, while the number of reported cases significantly decreased during the COVID-19 pandemic. Between 2015 and 2022, 15 WNV isolates, and 10 PCR-positive clinical specimens were investigated by NGS. Phylogenetic analysis revealed that the major European WNV lineage 2 clades, namely the Eastern European (or Russian) and the Central European (or Hungarian) clades, are presented in Hungary. Strains of the Balkan and other European clusters within the Central European clade are co-circulating in the country, following a characteristic geographical distribution. In Hungary, the presence and co-circulation of multiple lineage 2 WNV strains could be identified in the last few years. Therefore, in light of the 2018 WNV outbreak, sequence-based typing of the currently circulating strains could highly support outbreak investigations.

West Nile and Usutu virus seroprevalence in Hungary: A nationwide serosurvey among blood donors in 2019.

In Hungary, West Nile virus (WNV) has been responsible for 459 laboratory confirmed human cases between 2004 and 2019, while the first human Usutu virus (USUV) infection was confirmed only in 2018. A comprehensive serosurvey was conducted among blood donors to assess the WNV and USUV seroprevalence in 2019, one year after the largest European WNV epidemic. Altogether, 3005 plasma samples were collected and screened for WNV and USUV specific Immunoglobulin G (IgG) antibodies by Enzyme-Linked Immunosorbent Assay (ELISA). All reactive samples were further tested for tick-borne encephalitis virus IgG antibodies by ELISA. Indirect immunofluorescence test and microneutralization assay were used as confirmatory methods. Overall, the WNV seroprevalence was 4.32%, and in five blood donors USUV seropositivity was confirmed. The highest seroprevalence was measured in Central, Eastern and Southern Hungary, while the Western part of the country proved to be less affected. There was a statistically strong association between the WNV seroprevalence of 2019 and the cumulative incidence in the period of 2004 and 2019 calculated for every NUTS 3 region. The last WNV serological screening was performed in 2016 and the prevalence of anti-WNV IgG proved to be 2.19%. One year after the 2018 WNV outbreak, a significant increase in seroprevalence was observed in the Hungarian population and evidence for USUV seropositivity was also obtained. The spatial pattern of seroprevalence can support the identification of high-risk areas raising awareness of the need for increased surveillance, such as screening vector, equine, and avian populations. The communication with general practitioners and other professionals in primary health care services can support the early identification of acute human cases. Education and awareness-raising on the importance of protection against mosquito vectors amongst residents are also important parts of preventive measures.