The danger of the emergence of new viral diseases and their rapid spread demands apparatuses for continuous rapid monitoring in real time. This requires the creation of new bioanalytical methods that overcome the shortcomings of existing ones and are applicable for point-of-care diagnostics. For this purpose, a variety of biosensors have been developed and tested in proof-of-concept studies, but none of them have been introduced for commercial use so far. Given the importance of the problem, in this study, long-period grating (LPG) and surface plasmon resonance (SPR) biosensors, based on antibody detection, were examined, and their capabilities for SARS-CoV-2 structural proteins detection were established. Supersensitive detections of structural proteins in the order of several femtomoles were achieved by the LPG method, while the SPR method demonstrated a sensitivity of about one hundred femtomoles. The studied biosensors are compatible in sensitivity with ELISA and rapid antigen tests but, in contrast, they are quantitative, which makes them applicable for acute SARS-CoV-2 infection detection, especially during the early stages of viral replication.

Given the medical and social significance of Helicobacter pylori infection, timely and reliable diagnosis of the disease is required. The traditional invasive and non-invasive conventional diagnostic techniques have several limitations. Recently, opportunities for new diagnostic methods have appeared based on the recent advance in the study of H. pylori outer membrane proteins and their identified receptors. In the present study we assess the way in which outer membrane protein–cell receptor reactions are applicable in establishing a reliable diagnosis. Herein, as well as in other previous studies of ours, we explore the reliability of the binding reaction between the best characterized H. pylori adhesin BabA and its receptor, the blood antigen Le^b. For the purpose we developed surface plasmon resonance (SPR) and double resonance long period grating (DR LPG) biosensors based on the BabA–Le^b binding reaction for diagnosing H. pylori infection. In SPR detection, the sensitivity was estimated at 3000 CFU/mL—a much higher sensitivity than that of the RUT test. The DR LPG biosensor proved to be superior in terms of accuracy and sensitivity—concentrations as low as 10² CFU/mL were detected.

Background: Q fever is a zoonosis, included in category B of particularly dangerous infectious agents and as such merits careful surveillance and regular updating of the information about its distribution. Aim: This observational retrospective study aimed to provide an overview of Q fever incidence in Bulgaria in the period 2011 to 2017. Methods: Aggregated surveillance data from Bulgaria’s mandatory surveillance system, laboratory data on individual samples received at the National Reference Laboratory Rickettsiae and Cell Cultures and outbreak reports sent by the regional health authorities to the National Centre of Infectious and Parasitic Diseases, were used in this analysis. Cases were described by year, region, age group and most commonly identified risk behaviours. Results: A total of 139 confirmed cases were reported in the study period (average annual incidence: 0.27 cases/100,000 inhabitants). No seasonality or trend in reported cases was observed. Cases were mostly sporadic, with two small outbreaks in 2017. Identified risk behaviours among cases were occupational exposure and consumption of milk and dairy products, although exposure data were incomplete. The male/female ratio was 1.4. The identification and resolution of the two rural outbreaks in 2017 with a total of 18 cases involved good practices: active case finding and collaboration between public health and veterinary authorities. Conclusion: Between 2011 and 2017, Bulgaria retained low Q fever incidence, mostly sporadic cases and two small outbreaks. Occupational exposure and consumption of milk and dairy products were the most often reported likely exposures among cases. The outbreak investigations demonstrate the application of good control practices.

One of the first clinical observations related to COVID-19 identified hematological dysfunctions. These were explained by theoretical modeling, which predicted that motifs from SARS-CoV-2 structural proteins could bind to porphyrin. At present, there is very little experimental data that could provide reliable information about possible interactions. The surface plasmon resonance (SPR) method and double resonance long period grating (DR LPG) were used to identify the binding of S/N protein and the receptor bind domain (RBD) to hemoglobin (Hb) and myoglobin (Mb). SPR transducers were functionalized with Hb and Mb, while LPG transducers, were only with Hb. Ligands were deposited by the matrix-assisted laser evaporation (MAPLE) method, which guarantees maximum interaction specificity. The experiments carried out showed S/N protein binding to Hb and Mb and RBD binding to Hb. Apart from that, they demonstrated that chemically-inactivated virus-like particles (VLPs) interact with Hb. The binding activity of S/N- and RBD proteins was assessed. It was found that protein binding fully inhibited heme functionality. The registered N protein binding to Hb/Mb is the first experimental fact that supports theoretical predictions. This fact suggests another function of this protein, not only binding RNA. The lower RBD binding activity reveals that other functional groups of S protein participate in the interaction. The high-affinity binding of these proteins to Hb provides an excellent opportunity for assessing the effectiveness of inhibitors targeting S/N proteins.

The cytotoxicity and the antivirus activity of native hemocyanin, RtH, derived from the Bulgarian marine mollusk Rapana thomasiana and its structural isoform, RtH2, against HSV replication was evaluated on three HSV strains - two wt strains, TM (HSV 1) and Bja (HSV 2), and one ACV^R mutant with tk gene mutation, DD (HSV 2). The experiments were performed on continuous RD 64 cells and three HSV 1 and HSV 2 strains were used, two mutants sensitive to acyclovir and one resistant mutant. Both compounds were found to be effective inhibitors of wt HSV replication. Both compounds did not exhibit any effect on the infectious virus yield on ACV^R mutant. The most promising, active and selective, anti-HSV agent, especially to genital herpes virus, was found to be the functional unit of native hemocyanin - RtH2. RtH2 did not induce apoptosis/ necrosis 8 h after virus infection and the target of its action, was found to be the viral but not the host cell DNA.

Cell-based assays are a valuable tool for examination of virus–host cell interactions and drug discovery processes, allowing for a more physiological setting compared to biochemical assays. Despite the fact that cell-based SPR assays are label-free and thus provide all the associated benefits, they have never been used to study viral growth kinetics and to predict drug antiviral response in cells. In this study, we prove the concept that the cell-based SPR assay can be applied in the kinetic analysis of the early stages of viral infection of cells and the antiviral drug activity in the infected cells. For this purpose, cells immobilized on the SPR slides were infected with human coronavirus HCov-229E and treated with hydroxychloroquine. The SPR response was measured at different time intervals within the early stages of infection. Methyl Thiazolyl Tetrazolium (MTT) assay was used to provide the reference data. We found that the results of the SPR and MTT assays were consistent, and SPR is a reliable tool in investigating virus–host cell interaction and the mechanism of action of viral inhibitors. SPR assay was more sensitive and accurate in the first hours of infection within the first replication cycle, whereas the MTT assay was not so effective. After the second replication cycle, noise was generated by the destruction of the cell layer and by the remnants of dead cells, and masks useful SPR signals.