TABLE OF CONTENT

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1. A RAPID AND VERSATILE ASSAY FOR SCREENING OF AAC(6')-IB-CR IN MULTIDRUG-RESISTANT ENTEROCBACTERIACEAE

I. N. Ivanov, S. Sabtcheva, T. Saga, Y. Ishii, M. Каки, T. Kantardjiev

ABSTRACT
Plasmid-mediated quinolone resistance has emerged as an important clinical problem due to the rapid spread in clinical isolates of Enterobacteriaceae. Despite conferring only low levelofresistancetociprofloxacinandnorfloxacinthe plasmid-mediated ACC(6’)-lb-cr acetyltransferase is able to promote the selection and accumulation of high-level resistant chromosomal mutants during quinolone treatment. Therefore, in order to guide therapy, rapid assays for its detection are necessary. This study investigates one such method which is based on the Bccl-Restriction Fragment Length Polymorphism (RFLP). It removes the disadvantages of previously used techniques and is considered cost-effective and versatile.


2. PUTATIVE AGGREGATION - RELATED SURFACE PROTEINS IN BIFIDOBACTERIUM LONGUM 1/2

T. Yungareva, Z. Urshev, Z. Dimitrov, M. Michaylova

ABSTRACT
Earlier observations have shown that Bifidobacterium longum 1/2 demonstrates autoaggregation properties or co-aggregates with some enteropathogenic bacteria. In this study we aimed to collect initial data that will point to the potential factors that promote the aggregation properties of this strain. An overnight culture was grown in modified TS broth, washed and treated with different solutions with or without proteases to test the effect on the aggregation of cells. Surface protein extracts obtained with 5.0M LiCI were separated by SDS-PAGE. The genome of B. longum 1/2 was used to perform in silico analyses of candidate cell surface proteins that may be involved in aggregation. Trypsin, chymotrypsin and pepsin treatments drastically reduced the extent of aggregation pointing to the proteinaceous nature of the factor involved in cell-to-cell interaction. For cells treated with LiCI a partial loss of auto-aggregation and coaggregation abilities was observed that correlated with the extraction of a specific protein with MW of approximately 13 kDa. In B. longum 1/2 four sortase genes were found, only one of which was part of a putative pilus-encoding gene cluster as it contained the typical major (FimA) and minor (FimB) pilin subunit genes. The results from this initial study suggest that the aggregation phenotype of B. longum 1/2 may be attributed either to a LiCI-extractable13 kDa surface protein or to the presence of pili-encoding gene cluster in the genome of this strain.


3. MICROBIAL DIVERSITY OF MALE GENITOURINARY TRACT MICROBIOME IN DNIPRO CITY, UKRAINE

Sklyar T.V., Voronkova O.S., Papiashvili M.H.

ABSTRACT
The most common causes of male reproductive system disorders are infectious and inflammatory diseases of the genitourinary tract. Obligate and opportunistic pathogens are involved as causative agents of different pathological conditions and can provoke complications, raising the need for a complex therapy with antibiotics. By performing real-time PCR assay using the “Androflor” test-system, the composition of genitourinary tract microbiota was determined in 67 men. Of them, 16% of cases were healthy individuals. The following obligate pathogens were detected: Chlamydia trachomatis (3.0%), Mycoplasma genitalium (6.0%), Trichomonas vaginalis (1.5%) and Neisseria gonorrhoeae (1.5%). Opportunistic microorganisms were identified in 71.6% of cases. Among them, staphylococci were the most commonly isolated (75.0%) with prevalence of Staphylococcus epidermidis (58.3%). We also identified Enterococcus faecalis and members of Enterobacteriaceae (52.1%). In 22.4% of cases there were associations of more than 2 pathogens or opportunistic microorganisms. The number of microorganisms exhibited some deviations as compared with microbiota of healthy people. The majority of microorganisms or their associations had quantity of 2.5 - 4.9 Ig CFU/ml, which is a significant level indicating microbial disorder. More than 74% of opportunistic microorganisms were susceptible to fluoroquinolones and cephalosporins. Low susceptibility was determined for penicillins, macrolides and tetracyclines. The use of molecular genetic methods significantly speeds up the identification process, which in turn enables early initiation of treatment.


4. SELECTIVE PRESSURE ANALYSIS OF HUMAN RESPIRATORY SYNCYTIAL VIRUSES CIRCULATING IN BULGARIA

I. Georgieva, S. Angelova, N. Korsun

ABSTRACT
In order to investigate the genetic and evolutionary diversity of co-circulating genotypes of human respiratory syncytial viruses (HRSV) in Bulgaria we conducted selective pressure analysis of the second hypervariable region of the attachment G protein of HRSV. The study included representative numbers of all HRSV-positive nasopharyngea samples collected from children aged under 5 years, during two consecutive winter seasons (2014/2015 and 2015/2016). The HRSVs were detected using Real-timeRT-PCR assay. The G genes were partially sequenced and tested for positive selection. For HRSV group A, selection pressure analysis revealed purifying selection in the ON1 genotype. Three codons at position 274, 297 and 298 were positively selected by ai least two different methods. For HRSV group B no positive selection site was detected in the BAS genotype. This is the first report describing the selective pressure analysis of HRSV genotypes circulating in Bulgaria.


5. RT-PCR SYSTEMS FOR DETECTION OF ARBOVIRUSES

I. Trifonova, I. Christova, E. Panayotova, T. Gladnishka

ABSTRACT
The number of vector-borne viruses (tick- and mosquito-borne) is huge. They have widespread distribution and potential to cause conditions ranging from mild self-limiting illnesses to severe diseases with fatal outcome. Viruses of particular importance that belong to this group are Tick-borne encephalitis virus (TBEV), West Nile fever virus (WNV), causative agents of dengue fever (DENV), yellow fever (YFV), Japanese encephalitis (JEV), as well as zika (ZIKV) and Toskana (TOSV) viruses. Among them, TBE and WNV are known to cause infections in Bulgaria. However,on a global scale the possibility of importing infections is an actual issue and the preparedness for fast and accurate diagnosis is very important. In order to improve diagnostic capabilities for early and reliable detection of arboviruses, genetic methods for direct detection were developed and tested with clinical samples, reference materials and samples received from external quality assessment. In recent years, a number of new PCR protocols were implemented for routine diagnostics in our laboratory, including genetic methods for WNV (Real-time PCR detection of lineage 1 and 2), ZIKV (conventional and Real-time PCR) and Real-time detection protocols for TBEV, DENV, YFV, TOSV, ZIKV and Usutu virus. Reliability of the introduced methods was confirmed by international external quality assessment controls as well as detection of the specific RNAs in two patients with WNF and two patients with imported dengue fever.


6. SEROPREVALENCE STUDY FOR WEST NILE VIRUS IN BULGARIA

I. Christova, I. Trifonova, E. Panayotova, S. Tchakarova, T. Gladnishka, E. Taseva

ABSTRACT
A nationwide seroprevalence study was conducted in order to assess local circulation and risk of human infections with West Nile virus (WNV) in Bulgaria. In total, 1451 residents of all 28 districts in Bulgaria were tested for WNV-specific IgG antibodies. The survey found overall seroprevalence of 1.5%. The highest WNV seroprevalence was found in Sofia Province and districts near the river Danube. The results showed that WNV infection seems to be more widespread in the country than it has been described.


7. CHALLENGES IN THE DIAGNOSIS AND TREATMENT OF ALLERGY TO ALTERNARIA ALTERNATE AND OTHER MOULD SPECIES

D. Hristova, MD

ABSTRACT
Moulds are involved in the pathogenesis of a number of respiratory diseases. It is estimated that 2-6% of the general population is sensitized to moulds. Terms such as sensitization, colonisation and invasive infection can be discussed in regard to mould-sensitive individuals. Clinical manifestations are predominantly determined by the immunological status. Alternaria is among the most common moulds in the world. The incidence of asthma is three times higher among mould-sensitized individuals and up to 70% of them show positive skin tests to A. alternata. However, cross-reactivity among moulds makes the accurate diagnose difficult. Alt A1 is a marker for actual sensitization to A. alternata. The additional testing ofenolase and Mn superoxide dismutase is useful in assessing the sensitization to Pleosporaceae.


8. METHODS FOR DIAGNOSIS OF SYSTEMIC CANDIDOSIS

L. Boyanova

ABSTRACT
The aim of this review is to present the common laboratory methods for diagnosis of invasive candidosis, a fungal disease caused by species of the genus Candida. We are conducting a study which is part of the surveillance system BulSTAR investigating the etiological structure of infections in Bulgaria. The extracted BulSTAR data evaluating Candida blood culture isolates from Bulgarian hospitals showed higher number of isolates than previous years. The most frequently isolated species were: Candida albicans, followed by Candida parapsilosis, C. glabrata, C. krusei, C. tropicalis and other Candida non-albicans strains (19). The current study confirmed the increase in the number of isolated C. non-albicans strains. The most common methods in the diagnosis of systemic candidosis are blood culture, antigen and antibody detection in body fluids (serum, plasma), histopathology (biopsy) and PCR-based molecular techniques.


9. TOXOPLASMOSIS AND PREGNANCY, DIAGNOSTIC AND THERAPEUTIC APPROACHES

R. Harizanov, N. Tsvetkova, I. Kaftandjiev, I. Rainova. Boyanova

ABSTRACT
Background: Healthcare for women, especially during pregnancy, is of paramount importance to public health policies. The aim of this study is to present our experience in the management of pregnancy-related toxoplasmosis.

Methods: A group of 301 pregnant women, 9 newborn and 3 stillborn children was examined for infection with Toxoplasma gondii. Laboratory diagnosis was performed with ELISA IgG, IgM, IgA, ELISA IgG avidity and PCR. Data were processed using descriptive statistics and groups were compared with the Mann-Whitney test.

Results: Examination for anti-Toxoplasma gondii IgG showed positive results in 148 cases (seroprevalence of 49.2%). Of them, IgM antibodies were found in 67 (45.3%). The difference in IgG median values among women positive for IgG but negative for IgM, and women positive for IgG and IgM, was statistically significant. All women with positive results for specific IgM showed positive results for specific IgG. They were examined for determination of IgG avidity. Low or borderline avidity was found in 24 patients (35.8%). We conducted spiramycin therapy until delivery in pregnant women with serologic data for primary toxoplasmosis and low or borderline avidity (n=24).This group was monitored regularly with ultrasonography for pathological morphological changes in the foetus and such were not found.

Conclusions: Preventive care provides time to apply measures against mother-to-foetus transmission of several diseases, including toxoplasmosis. Tests for toxoplasmosis prior pregnancy are of utmost importance to avoid the risk of a possible congenital infection and emotional distress in pregnant women.


10. APPLICATION OF SiO2/HYDROXYPROPYL CELLULOSE HYBRID MATERIALS DOPED WITH Zn IONS IN ZINC DEFICIENCY

D. Pencheva, A. Sestrimska, A. Dimitrova, L. Borisova, R. Karamfilova, N. Rangelova,Ts. Angelova, N. Georgieva

ABSTRACT
The presence of bactericidal properties against clinical strains of Enterobacteriaceae in synthesized Si02-hydroxypropyl cellulose hybrid materials containing Zn ions is a prerequisite for their use as a feed additive for the prevention of zinc deficiency conditions and treatment of diarrhoea in young children in developing countries.


11. COMPARISON OF THE BACTERICIDAL EFFECT OF TWO SiO2/HYDROXYPROPYL CELLULOSE HYBRID MATERIALS, DOPED WITH Zn IONS AGAINST CLINICAL STRAINS OF SALMONELLA AND SHIGELLA

D. Pencheva, E. Yordanova, V. Susanina, Ts. Angelova, N. Rangelova, N. Georgieva

ABSTRACT
The current study was performed to compare the biological properties of hybrid materials composed of Si02/hydroxypropyl cellulose (HPC) and Zn ions. The concentration of zinc was varied between 0-5 wt% and derived from different precursors - zinc sulphate or zinc nitrate. Antibacterial activity of the materials was tested against selected clinical strains of Salmonella and Shigella with established resistance to two or more antibiotics. The hybrid materials doped with Zn ions showed pronounced bactericidal activity against the tested strains.


12. BULGARIAN GPs AND OBGs ATTITUDE TOWARDS INFLUENZA VACCINATION DURING PREGNANCY

A. Galev

ABSTRACT
Vaccination is considered the best tool for prevention of seasonal influenza distribution. Even though pregnant women are regarded as a high-risk group during the active flu season, small percentage of general practitioners (GPs) and obstetrician-gynaecologists (OBGs) advise on vaccination which is yet available in the list of recommended vaccines. The reason for vaccine refusal both by pregnant women and medical professionals is not fully clarified. The goal of this study is to define the attitude of GPs and OBGs towards influenza vaccination in pregnant women as well as to define a mode of action to increase the vaccine coverage.


 

Editor-in-Chief
Prof. Todor Kantardjiev, MD, PhD, DSc– NCIPD, Sofia, Bulgaria

Editorial Board:
Prof. Iva Christova, MD, DSc – NCIPD, Sofia, Bulgaria
Richard Anthony, PhD – RIVM, The Netherlands
Dale Barnard, PhD – Utah State University, USA
Roger Hewson, PhD – WHO Collaborating Centre for Virus Research & Reference, Porton Down, UK
Judith M. Hübschen – Luxembourg Institute of Health, Luxembourg
Emad Mohareb, PhD– U.S. Naval Medical Research Unit No.3, Egypt
Bruno Gottstein – Institute of Parasitology, University of Berne, Switzerland
Anton Andonov, PhD – Public Health agency, Canada
Anna Papa-Konidari, PhD- Aristotle University, Thessaloniki, Greece
Anna Erdei, DSc - Eötvös L. University, Budapest, Hungary
Lanfranco Fattorini – Instituto Superiore di Sanita, Italy
Hannes Stocklinger, PhD – Medical University, Vienna, Austria
Acad. Bogdan Petrunov, MD, DSc – NCIPD, Sofia, Bulgaria
Assoc. Prof. Iskra Rainova, MD, PhD – NCIPD, Sofia, Bulgaria
Assoc. Prof. Lubomira Nikolaeva-Glomb, MD, PhD – NCIPD, Sofia, Bulgaria

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