Design of a University Course for the Training of Biology Teachers in a Virtual Environment (Analysis of Results Taking into Account Students' Attitudes to the E-learning)

Over the last decade, the concept of "designing e-learning" in the field of higher education has been in the focus of various research communities. Given the increasing number of e-learning publications, the issue of a university course model is becoming a widely discussed topic. The aim of the article is to present a specific solution related to the design of a traditionally organized course in electronic, for the preparation of future teachers of biology. The research process of designing a theoretically based model of pedagogical activity within the framework of a university e-learning course, including stepwise steps in its creation, following the phases of the ADDIE model, is briefly described. Using statistical tests to verify hypotheses (relative percentages), a comparison of the results obtained, taking into account the attitudes of the students before and after the e-learning, was made

Synthesis and Inhibiting Activity of Some 4-Hydroxycoumarin Derivatives on HIV-1 Protease

Six novel 4-hydroxycoumarin derivatives were rationally synthesized, verified, and characterized by molecular docking using crystal HIV-1 protease. Molecular docking studies predicted antiprotease activity of (7) and (10). The most significant functional groups, responsible for the interaction with HIV-1 protease by hydrogen bonds formation are pyran oxygen, atom, lactone carbonyl oxygen and one of the hydroxyl groups. The newly synthesized compounds were biologically tested in MT-4 cells for inhibiting HIV-1 replication, exploring the protection of cells from the cytopathic effect of HIV measured by cell survival in MTT test. One derivative −7 showed 76–78% inhibition of virus infectivity with IC50 = 0.01 nM, much less than the maximal nontoxic concentration (1 mM). Antiprotease activity of 7 in two different concentrations was detected to be 25%. Nevertheless, the results of study of (7) encourage using it as a pharmacophore for further synthesis and evaluation of anti-HIV activity

HEALTHCARE WORKERS IN BULGARIA - ARE THEY PROTECTED FROM VACCINE-PREVENTABLE INFECTIONS?

Background: Healthcare workers (HCWs) are at increased risk of exposure to many viral infections, including vaccine preventable diseases (VPDs) such as measles, mumps and rubella (MMR) as compared to non-HCWs. Immunity of HCWs against these viruses is mandatory in a healthcare setting due to possible exposure from patients or colleagues. Aim: To provide an assessment of anti-measles, mumps and rubella IgG seropositivity among Bulgarian HCWs employed in hospitals and regional health inspectorates (RHI), as an indicator of protective immunity against MMR in this risk group.  Materials and Methods: In the current study, 181 HCWs from Infectious Units in regional hospitals in the country, and HCWs from the RHI, involved in the monitoring and surveillance of MMR cases in Bulgaria were screened. Serum specimens from all participants were tested by a commercial indirect enzyme-linked immunosorbent assay (Anti-Measles, Anti-Mumps, Anti-Rubella IgG EIA-Euroimmun®, Germany) for presence of IgG antibodies against measles, mumps and rubella, as an indicator of protective immunity.  Results: The study included 181 HCWs, 25 male and 156 female, aged 22 to 66 years. The average protective seroprevalence for measles, mumps and rubella was 82.9%, 76.2% and 92.3% percent, respectively. The highest share of negative results were obtained for mumps-specific IgG – 23.2% (42/181), followed by measles 16.6% (60/181) and rubella-specific IgG 7.7% (19/181). Regarding the age distribution, the highest number of HCWs non-immune to measles and mumps was found among the 31- 40-year olds, and against mumps – among the 41-50-year-olds. Conclusion: HCWs are at greater risk of contracting infections than the general population because of contact with sick patients or infectious material. Infected healthcare workers can spread nosocomial diseases to vulnerable patients with more severe illness, leading to complications and even death. Therefore, the vaccination status of HCWs must be strictly monitored

Serological and Molecular Detection of Coxiella Burnetii in Clinical Samples from Veterinarians and Cattle Farm Workers from Gabrovo Region, Bulgaria

Coxiella burnetii, which causes Q fever, is a highly infectious agent that is widespread around the world.  During the last decades, the number of cases reported in Bulgaria varied from year to year. The present study aimed to determine the frequency of C. burnetii infection using ELISA and conventional PCR among freelance veterinarians and cattle farm workers in Gabrovo province, Bulgaria. In the period April 2020 to June, 2021 a total of 154 blood samples of target group was tested in the National Reference Laboratory of Cell cultures, rickettsia and oncogenic viruses (NRL CCROV) at NCIPD - Sofia. Diagnosis of C. burnetii was performed by indirect enzyme-linked immunosorbent assay ELISA (anti-Coxiella burnetii ph. II IgG/IgM) and by end-point PCR technique (to detect the sodB gene region of C. burnetii). By indirect ELISA assay of the tested 154 clinical samples, anti-C. burnetii positive ph. II IgM antibodies were registered in 37% of samples. A relatively high percentage are affected in the active age between 50-60 years old. Anti-C. burnetii positive ph. II IgG antibodies were proven at 50% of tested samples. A positive PCR signal for C. burnetii DNA was obtained at 37/154 (20% of samples) and follows the above reported trend of acute infection of active age patients. Around 10% of tested samples were positive for three C. burnetii laboratory markers. We conclude that Q fever is endemic in Bulgaria. More research is necessary in different Bulgarian regions to set the human risk groups, to diagnose acute and chronic Q fever and to determine the economic impact of Q fever in the cattle industry. In the NRL CCROV was developed diagnostic scheme including complex methods to improve early laboratory diagnosis of C. burnetii, allowing taking proper treatment of suspected with Q fever patients

Mini Review: Q Fever (Coxiellosis): Epidemiology, Pathogenesis and Current Laboratory Diagnosis

Q fever is zooantroponozis with global distribution caused by the strictly intracellular bacterium Coxiella burnetii. Causative agent of Q fever is an obligate intracellular parasite, classified in the genus Coxiella, family Coxiellaceae, class Gammaproteobacteria. The importance of the disease was assessed both in terms of human health and the serious economic damage they cause on livestock. Clinical manifestation of Q fever in humans is characterized by a wide variety - from asymptomatic infection to a chronic disease that can be fatal. Several basic methods have been developed to detection of C. burnetii. PCR and C. burnetii genomic sequences in whole blood are a sensitive and safe method of detection, with >90% sensitivity. A four-fold or greater rise of (CF) antibody (phase 2) between the paired sera is also diagnostic approach. Sensitivity of a four-fold rise in titre has been estimated as 73% ÷78% and specificity has been estimated as 90%, respectively. EIA is method with highly sensitive and specific. EIA detect IgM and then IgG antibodies which develop to phase II antigens in 10 to 14 days from symptom onset. IFA tests are of particular value for confirmation of acute infection and for diagnosis of chronic infection with high sensitivity. The technique detected IgG, IgM and IgA immunoglobulin classes. Suitable specimens for C. burnetii detection are blood samples. Although scientific interest in Q fever has always existed, a number of facts concerning the unforeseen nature of the epidemic, various clinical manifestations both in humans and in animals, the opportunities for chronic and other features of infection remain unclear. For this reason, timely and highly sensitive laboratory diagnosis is crucial for the outcome of the disease and subsequent treatment and monitoring

EVALUATION OF INTERACTIONS OF SARS-COV-2 STRUCTURAL PROTEINS WITH SPECIFIC ANTIBODIES BY SPR ASSAY

Background: The World Health Organization admitted that the vaccination against Covid 19 limited the deaths, but not the spread of the disease. This requires a method allowing a specific, rapid and accurate diagnosis of the disease. We report a SPR assay that meets the requirements and can be applied no only for SARS Cov-2 diagnosis but as a tool for early diagnosis of otherinfections. Methods: Surface plasmon resonance (SPR) method was used to identify the binding of S/N protein to monoclonal antibodies. N-protein monoclonal antibody (NP mAb), S-protein monoclonal antibody (SP mAb), and receptor bind domain (RBD) antibody were used as recognition molecules. Ligands were deposited by the matrix-assisted laser evaporation (MAPLE) method, which guarantees maximum interaction specificity. Results: We registered S/N protein binding to the corresponding mAbs and S protein to RBD antibody with high sensitivity: the interactions were observed at protein concentration about 130 femtomoles (fM). A very good specificity was observed: the measured S protein binding activity to NP mAb was below the limit of detection (LOD). The same was noticed for N protein binding to SP mAb. Conclusions: The presented SPR assay possesses high sensitivity and selectivity and provides quantitative analysis. This makes it applicable for following the evolution of acute SARS-CoV-2 infection, especially at the early stages of viral replication which can be clinically useful

Dried Blood Spots as a Clinical Samples for Laboratory Diagnosis and Surveillance of Vaccine-Preventable Diseases in Bulgaria

In recent years the dried blood spots (DBS) had new and innovative applications in medicine, neonatology, virology and microbiology. This study aimed to evaluation of the frequency of detection of viral IgM/IgG markers in dried blood spots and introducing an easy-to-implement protocol for serum extraction in measles, mumps and rubella surveillance. The total 204 clinical samples (102 serum samples and 102 dried blood spots) collected from 102 patients were included. All specimens were tested for presence of specific viral markers (IgM and IgG antibodies) by a commercial indirect enzyme-linked immunosorbent assay (ELISA). Of all tested patients, three (3/102, 2.94%, 95% CI: 0 ÷ 6.22) were confirmed for acute measles infection and two (2/102, 1.96%, 95% CI: 0 ÷ 4.65) for mumps. Double positive ELISA-IgM results were found in their serum samples and DBS. No acute rubella infection and rubella IgM marker were detected in both clinical samples. By immunoassay analysis of all 102 patients, measles, mumps and rubella IgG were found in 83/102 (81%, 95% CI: 73.40 ÷ 88.60), 76/102 (75%, 95% CI: 66.60 ÷ 83.40) and 79/102 (77%, 95% CI: 68.83 ÷ 85.17) serum samples.  Comparative results were obtained in the adequately obtained DBS. Viral IgG seroprevalence in DBS were obtained in 79/102 (77%, 95% CI: 68.83 ÷ 85.17) for measles, 69/102 (68%, 95% CI: 58.67 ÷ 77.33) for mumps and 73/102 (72%, 95% CI: 63 ÷ 81) for rubella, respectively. Double negative results for each screened viral markers were proven in six tested patients.The study shown higher extinction value (Ratio and NovaTec units) in DBS compared to serum samples of same persons were calculated. Our studies show over 90% coincidence in combined ELISA assay of viral markers against measles, mumps, and rubella in serum samples and DBS. DBS clinical approach is non-aggressive and more acceptable to the public (including young children, pregnant women, etc.). It has a variety of new and innovative applications in medicine and in particular in the laboratory diagnosis of acute and past (presence of protective immunity) measles, mumps and rubella infection in the phase of elimination

A Review on Dried Blood Spots (DBS) as Alternative, Archival Material for Detection of Viral Agents (Measles, Mumps, Rubella, Hepatitis B Virus)

In recent years there appears a variety of new and innovative applications of the dried blood spots. The areas of their range of application are medicine, neonatology, virology, microbiology, toxicology and pharmacokinetics, metabolic exchange, therapeutic drug monitoring, toxicology, and control of environmental pollution. The advantages of DBS technology can be combined into four main groups: (1) compared to conventional venipuncture, requires less blood volume, which is especially important in pediatrics and neonatology; (2) the procedure for blood collection is easy, inexpensive and noninvasive; (3) the risk of bacterial contamination or hemolysis is minimal; and (4) DBS can be maintained for a long time with almost no impact on the quality of the analysis. In recent years is increasing the application of DBS as method for seroepidemiological survey with focus viral infections: measles, mumps, rubella and hepatitis B virus. The DBS technique is optimized as an alternative approach (non-invasive, inexpensive, not requiring trained staff and cold chain for transport and storage) of venipuncture collection of clinical material in virology.This method facilitates the scientific researches about the concentration of virus specific antibodies in peripheral blood taken from a finger or heel; determining the percentage susceptibility / protection of the studied group of patients againt vaccine-preventable infectious - measles, mumps, rubella and hepatitis B; social benefits - non-invasive technique for testing of small children and infants and applications in regions in the countries with not well developed logistics infrastructure

Infectious Agents and Miscarriage in Bulgaria

A number of infection agents have been linked to miscarriage and to other adverse outcomes of pregnancy, such as stillbirth and preterm delivery. The purpose of present study was to determine the implication and prevalence of viral agents (parvovirus B19, rubella, CMV and adenoviruses) and Chlamydia trachomatis in the etiology of miscarriage during the first and second trimester of pregnancy. A total of 62 serum samples from women with miscarriage (n=32) and control group of healthy women (n=30) for period January 2015 – June 2016 were tested by ELISA (detected of specific IgM/IgG antibodies) and PCR (detected of specific genomic region) assays. The possible role of B19V, Ch. trachomatis and adenoviruses for miscarriages were detected in 6/32 (18.75%) by ELISA and in 7/32 (21,87%) by PCR methods. The seroprevalence of protective IgG antibodies in the highest level was proven against rubella 25/32 (78,12%) and the lowest – against adenoviruses (1/32, 6,25%). All tested healthy women in the control group had a negative result for acute infection for the five tested infectious agents. The detailed study aimed at enriching the diagnostic palette of these infectious pathogens is necessary for understanding the exact mechanisms behind infection-induced miscarriages and could lead to effective treatment and thus prevention

Pojavnost bakterije Coxiella burnetii u mliječnim stadima u pokrajini Gabrovo, Bugarska, ustanovljena serološkim i molekularnim analizama uzoraka mlijeka iz cisterni

This study investigated the occurrence of Coxiella burnetii in dairy cattle, goat herds and sheep flocks in the Gabrovo Region, central Bulgaria, to identify the potential sources of Coxiella burnetii infection diagnosed in veterinarians and farm workers in this region. To detect infection on livestock farms, we tested bulk-tank milk (BTM) for the presence of antibodies and/or the Coxiella burnetiid genome using ELISA and PCR, respectively. A total of 81 herds were tested, including 23 dairy cattle herds, 43 sheep herds, 9 goat flocks, and 6 mixed flocks (sheep and goats). By ELISA, antibodies against Coxiella burnetii were detected in 30.4% of the BTM tested samples from cattle farms, 60.4% of the sheep farms, and 11.1% of the goat BTM samples. The results were inconclusive in 6.98% of the tested sheep milk samples and 11.1% of the goat milk samples. There was a statistically significant correlation between the herd size and the ELISA S/P % values on the dairy cattle farms. Excretion of the pathogen in milk was detected by PCR in 9 out of 67 BTM samples, including 5 out of 19 cattle BTM samples, 3 out of 39 sheep BTM samples and 1 out of 5 goat BTM ones. The results indicate that C. burnetii infection is widely prevalent in the region, which calls for adequate control and prophylactic measures to reduce the health risks from the transmission of this zoonosis to humans.U ovom je radu istraživana pojavnost bakterije Coxiella burnetii u stadima mliječnih krava, koza i ovaca u pokrajini Gabrovo, središnja Bugarska. Rezultati bi trebali pomoći pri otkrivanju potencijalnih izvora zaraze navedenom bakterijom u veterinara i poljoprivrednih radnika u tom području. Kako bi se otkrila infekcija na stočnim farmama, mlijeko iz velikih spremnika (BTM) testirano je na prisutnost protutijela i/ili genoma bakterije Coxiella burnetii primjenom testa ELISA i PCR-om. Testirano je ukupno 81 stado, uključujući 23 stada mliječnih krava, 43 stada ovaca, 9 stada koza i 6 mješovitih stada (ovce i koze). Testom ELISA otkrivena su protutijela na bakteriju Coxiella burnetii u 30,4% uzoraka iz BTM-a s farmi krava, 60,4% s farmi ovaca i 11,1% uzoraka BTM-a s farmi koza. Pouzdani zaključci nisu mogli biti doneseni u slučaju 6,98% uzoraka mlijeka ovaca i 11,1% uzoraka mlijeka koza. Uočena je statistički znakovita korelacija između veličine stada i postotne vrijednosti ELISA S/P u mliječnih krava. Izlučivanje patogena u mlijeko otkriveno je PCR-om u 9 od 67 uzoraka BTM-a, uključujući 5 od 19 uzoraka mlijeka krava, 3 od 39 uzoraka mlijeka ovaca i 1 od 5 uzoraka mlijeka koza. Rezultati su pokazali da je infekcija bakterijom C. burnetii široko rasprostranjena u pokrajini Gabrovo, što zahtijeva odgovarajuće mjere kontrole i profilakse kako bi se smanjili zdravstveni rizici od prijenosa ovog zoonotskog uzročnika na ljude