The tick-borne encephalitis virus (TBEV) is one of the most common members of the Orthoflavivirus genus, which comprises the causative agents of severe diseases in humans and animals. Due to the expanding areas of orthoflavivirus infection, its differential diagnosis is highly demanded. Commercial test kits based on inactivated TBEV may not provide reliable differentiation between flaviviruses because of serological crossover in this genus. Application of recombinant domains (sE and dIII) of the TBEV Sukhar-strain protein E as antigens in an ELISA test system allowed us to identify a wide range of antibodies specific to different TBEV strains. We tested 53 sera from human patients with confirmed TBE diagnosis (the efficacy of our test system based on sE protein was 98%) and 56 sera from patients with other orthoflavivirus infections in which no positive ones were detected using our ELISA test system, thus being indicative of its 100% specificity. We also tested mouse and rabbit sera containing antibodies specific to 17 TBEV strains belonging to different subtypes; this assay exhibited high efficacy and differentiation ability in detecting antibodies against TBEV from other orthoflaviviruses such as Omsk hemorrhagic fever, Powassan, yellow fever, dengue, West Nile, Zika, and Japanese encephalitis viruses.

In the last few years, there has been a dramatic increase in the number of discovered viruses that are transmitted by arthropods. Some of them are pathogenic for humans and mammals, and the pathogenic potential of others is unknown. The genus Orthoflavivirus belongs to the family Flaviviridae and includes arboviruses that cause severe human diseases with damage to the central nervous system and hemorrhagic fevers, as well as viruses with unknown vectors and viruses specific only to insects. The latter group includes Lammi virus, first isolated from a mosquito pool in Finland. It is known that Lammi virus successfully replicates in mosquito cell lines but not in mammalian cell cultures or mice. Lammi virus reduces the reproduction of West Nile virus during superinfection and thus has the potential to reduce the spread of West Nile virus in areas where Lammi virus is already circulating. In this work, we isolated Lammi virus from a pool of adult Aedes cinereus mosquitoes that hatched from larvae/pupae collected in Saint Petersburg, Russia. This fact may indicate transovarial transmission and trans-stadial survival of the virus.

Non-human primates, due to their similarities in immune response to humans, are the preferred model for studying infectious processes and any associated cognitive impairments. Behavioral tests are indispensable for investigating pathogenesis in neuroinfections, especially those that do not manifest with noticeable clinical symptoms, as well as in the transition to a chronic form of the disease. Modeling viral infection requires specialized experimental conditions. Our work describes techniques for investigating mnemonic functions, tiredness, attentional focus, quick-wittedness, and basic behavioral responses in primates under the assumed conditions for infections with viruses that do not have an airborne route of transmission. It also outlines approaches to the training and selection of primates for virological research, as well as analyzing gender differences in learning abilities, the impact of housing conditions on the results, and the correlation between training success and behavioral test scores. These methods will allow a more detailed study of non-human primates as a model for researching cognitive and behavioral impairments under infectious and immune stress, as well as the design of less energy-intensive experiments for evaluating the efficacy and safety of therapeutic and prophylactic strategies at early stages of infection.

Non-invasive methods for monitoring the condition of laboratory animals play a key role in ensuring animal welfare and improving the reliability of scientific data. This study evaluates the effectiveness of two non-invasive approaches - daily body weight measurement and urine analysis by qPCR - for monitoring the health of BALB/c mice infected with tick-borne encephalitis virus (TBEV). Calculation of the first derivative of body weight change allowed precise determination of disease onset, which correlated with clinical symptoms and detection of viral RNA in urine. Mathematical analysis of body weight change dynamics (first derivative with type 2 cubic spline smoothing, rh = 1) showed that a derivative threshold value of ≤ −0.6 reliably distinguishes infected BALB/c mice from healthy ones (AUC = 1 in ROC analysis). Urine analysis by qPCR allowed for the detection of viral RNA as early as the second day after infection, with a peak on the seventh day. The mathematical model was further tested on C57BL/6, CBA, and BALB/c mice of different ages and proved to be suitable. The threshold values of the derivative were found to be dependent on the mouse strain. The proposed non-invasive methods offer a humane and accurate alternative to invasive procedures, contributing to higher ethical standards and quality of research in virology.

The recombinant domains (sE and dIII) of the E protein of the Suhar strain of tick-borne encephalitis virus (TBEV) were used as antigens in the ELISA test system, which allowed to identify a wide range of antibodies to various TBEV strains. We tested 53 sera from confirmed TBE patients and our sE protein-based assay was 98 % effective. 56 sera from patients with other orthoflavivirus infections were used, among which not a single positive result was detected, which indicates its 100 % specificity. We also tested mouse and rabbit sera containing antibodies to 17 TBEV strains referring to different subtypes and showed high efficiency and ability to differentiate antibodies to TBEV from antibodies to other orthoflaviviruses: Omsk hemorrhagic fever, Powassan, yellow fever, dengue, West Nile, Zika and Japanese encephalitis.