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Evidence of recombination in Tula virus strains from Serbia
No full textTula hantavirus (TULV) belongs to Bunyaviridae family, with negative sense RNA genome. Segmented nature of hantaviral genome allows for genetic reassortment, but the evidence of homologous recombination also exists. In this study we analyzed TULV sequences isolated in Serbia on different occasions and from different rodent hosts: 1987 strain from Microtus subterraneus and 2007 strain from Microtus arvalis. Phylogenetic analysis of both L and S segment sequences is suggestive of geographically related clustering, as previously shown for majority of hantaviruses. Reconstruction of phylogenetic tree for TULV S segment showed that both sequences from Serbia clustered together with sequences from East Slovakia, which had previously been shown to be recombinants (Kosice strain). Exploratory recombination analysis, supported by phylogenetic and amino acid pattern analysis, revealed the presence of recombination in the S segment sequences from Serbia, resulting in mosaic-like structure of TULV S segment similar to the one of Kosice strain. Although recombination is considered a rare event in molecular evolution of negative strand RNA viruses, obtained molecular data in this study support evidence of recombination in TULV, in geographically distant regions of Europe. (C) 2013 Elsevier B.V. All rights reserved
Changes in genetic diversity of the bordetella pertussis population in Serbia between 1953 and 2011
Get PDFMass vaccination has significantly reduced the incidence of pertussis, however, the disease is re-emerging, even in some countries with high vaccination coverage. In Serbia, whole cell pertussis vaccine was introduced in 1957. To monitor changes in bacterial population, 77 isolates collected from 1953 to 2011 were studied. The methods included serotyping of fimbriae (Fim), genotyping of pertactin (pm) and pertussis toxin SI subunit (ptxA). A shift from ptxA2 to ptxA1 has been observed in isolates since the late of 1960s. In the period 1961-1979, the genotype ptxAl became as common as genotype ptxA2. After that, during the period 1980-1989, the predominant ptx genotype was ptxAl. The reappearance of the ptxA2 allele followed an addition of the two strains harboring ptxA1 in the vaccine in 1985. The allele prn I was predominant among the Serbian isolates, though prn3 and prn11 have been detected since 1981. The prn2 allele was only found in one strain isolated in 1984, two of the four strains isolated in 2000 and in three isolated strains from 2011. Serotype Fim2.3 disappeared before 1980 and serotype Fim2 became predominant thereafter. The results of this study indicate that the B. pertussis population in Serbia is different from other vaccinated populations and that this difference may be related to the vaccine used
The importance of cross-reactivity in grass pollen allergy
Get PDFAccording to the data obtained from in vivo and in vitro testing in Serbia, a significant number of patients have allergic symptoms caused by grass pollen. We examined the protein composition of grass pollens (Dactylis glomerata, Lolium perenne and Phleum pratense) and cross-reactivity in patients allergic to grass pollen from our region. The grass pollen allergen extract was characterized by SDS-PAGE, while cross-reactivity of single grass pollens was revealed by immunoblot analysis. A high degree of cross-reactivity was demonstrated for all three single pollens in the sera of allergic patients compared to the grass pollen extract mixture. Confirmation of the existence of cross-reactivity between different antigenic sources facilitates the use of monovalent vaccines, which are easier to standardize and at the same time prevent further sensitization of patients and reduces adverse reactions
Slučaj humane monocitne erlihioze u Srbiji
Get PDFIntroduction Ehrlichiosis is a bacterial zoonosis transmitted by hematophagous arthropods - ticks. In humans, it occurs as monocytic, granulocytic, and ewingii ehrlichiosis. Pathological process is based on parasitic presence of Ehrlichia organisms within peripheral blood cells - monocytes and granulocytes. Case Outline Fifty-two year old patient was admitted to hospital due to high fever of over 40°C that lasted two days, accompanied with chills, muscle aches, malaise, loss of appetite, headache, confusion, breathing difficulties, and mild dry cough. The history suggested tick bite that occurred seven days before the onset of disease. Doxycycline was introduced and administered for 14 days, causing the disease to subside. Indirect immunofluorescence assay was used to analyze three serum samples obtained from this patient for Ehrlichia chaffeensis antibodies, and peripheral blood smear was evaluated for the presence of Ehrlichia and Ehrlichia aggregation into morulae. Conclusion Ehrlichiosis should be considered in each case where there is a history of tick bite together with the clinical picture (high fever, chills, muscle aches, headache, generalized weakness and malaise, and possible maculopapular rash). The presence of Ehrlichia chaffeensis antibodies was confirmed in a patient with the history of tick bite, appropriate clinical picture and indirect immunofluorescence assay. This confirmed the presence of human monocytotropic ehrlichiosis, a disease that is uncommonly identified in our country.Uvod Erlihioza je bakterijska zoonoza koja se prenosi hematofagnim artropodama - krpeljima. Kod ljudi se javlja kao monocitna, granulocitna i ewingii erlihioza. Patološki proces je posledica unutar ćelijskog parazitiranja erlihije u monocitima i granulocitima periferne krvi. Prikaz bolesnika Bolesnik star 52 godine primljen je na odeljenje zbog visoke dvodnevne febrilnosti (preko 40°C) koja je praćena drhtavicom, bolovima u mišićima, malaksalošću, gubitkom apetita, glavoboljom, konfuznošću, otežanim disanjem i oskudnim suvim kašljem. U anamnezi je dobijen podatak o ujedu krpelja sedam dana pre prijema. Laboratorijski nalazi su ukazali na trombocitopeniju, leukopeniju, anemiju i povećanje aktivnosti transaminaza u serumu. Bolesnik je lečen doksiciklinom 14 dana, nakon čega su se tegobe povukle. Metodom indirektne imunofluorescencije analizirana su tri uzorka seruma ovog bolesnika na prisustvo antitela na bakteriju Ehrlichia chaffeensis i pregledan je uzorak razmaza periferne krvi na prisustvo erlihija i konglomerata erlihije u morule, koje predstavljaju citoplazmatske vakuole. Zaključak Kod bolesnika s podatkom o ubodu krpelja, odgovarajućom kliničkom slikom i serološkim testom indirektne imunofluorescencije dokazana su antitela za bakteriju Ehrlichia chaffeensis, što ukazuje na humanu monocitotropnu erlihiozu, bolest koja se kod nas retko dokazuje. Na erlihiozu treba misliti kada uz kliničku sliku (visoka febrilnost, groznica, bolovi u mišićima, glavobolja, opšta slabost i malaksalost, eventualno makulopapulozna ospa) postoji podatak o ubodu krpelja
Two copies of bla (NDM-1) gene are present in NDM-1 producing Pseudomonas aeruginosa isolates from Serbia
No full textNew Delhi metallo-beta-lactamase producing Pseudomonas aeruginosa isolates are of special interest since P. aeruginosa is a major cause of nosocomial infections, the treatment of which could now be jeopardized, especially in developing countries. Six additional NDM-1 positive P. aeruginosa clinical isolates belonging to two different genotypes were shown to be plasmid-free. PFGE-hybridization experiments revealed the chromosomal location of the bla (NDM-1) gene. Restriction analysis and hybridization revealed that two copies of the bla (NDM-1) gene are present in the genomes of all tested isolates, as in previously characterized P. aeruginosa MMA83. Moreover, it was shown that increasing imipenem concentration did not have the effect on copy number of the bla (NDM-1) gene in the genome of P. aeruginosa MMA83
Defining the Structure and Receptor Binding Domain of the Leaderless Bacteriocin LsbB
Get PDFBackground: The bacteriocin LsbB targets a membrane-bound zinc-dependent peptidase. Results: The structure of LsbB was resolved by NMR. The C-terminal unstructured domains of LsbB and several other related bacteriocins were responsible for receptor binding. Conclusion: A subgroup of leaderless bacteriocins has been found to share a similar mechanism in receptor recognition. Significance: The study highlights the structure-function relationship of LsbB. LsbB is a class II leaderless lactococcal bacteriocin of 30 amino acids. In the present work, the structure and function relationship of LsbB was assessed. Structure determination by NMR spectroscopy showed that LsbB has an N-terminal -helix, whereas the C-terminal of the molecule remains unstructured. To define the receptor binding domain of LsbB, a competition assay was performed in which a systematic collection of truncated peptides of various lengths covering different parts of LsbB was used to inhibit the antimicrobial activity of LsbB. The results indicate that the outmost eight-amino acid sequence at the C-terminal end is likely to contain the receptor binding domain because only truncated fragments from this region could antagonize the antimicrobial activity of LsbB. Furthermore, alanine substitution revealed that the tryptophan in position 25 (Trp(25)) is crucial for the blocking activity of the truncated peptides, as well as for the antimicrobial activity of the full-length bacteriocin. LsbB shares significant sequence homology with five other leaderless bacteriocins, especially at their C-terminal halves where all contain a conserved KXXXGXXPWE motif, suggesting that they might recognize the same receptor as LsbB. This notion was supported by the fact that truncated peptides with sequences derived from the C-terminal regions of two LsbB-related bacteriocins inhibited the activity of LsbB, in the same manner as found with the truncated version of LsbB. Taken together, these structure-function studies provide strong evidence that the receptor-binding parts of LsbB and sequence-related bacteriocins are located in their C-terminal halves
Comparative phylogenetic analysis of Dobrava-Belgrade virus l and s genetic segments isolated from an animal reservoir in Serbia
Get PDFThe Dobrava-Belgrade virus (DOBV) is a member of the Bunyaviridae family, genus Hantavirus, possessing a single-stranded RNA genome consisting of three segments, designated L (large), M (medium) and S (small). In this study, we present phylogenetic analysis of a newly detected DOBV strain isolated from Apodemus agrarius. Analysis was based on partial L and S segment sequences, in comparison to previously published DOBV sequences from Serbia and elsewhere. A phylogenetic tree based on partial S segment revealed local geographical clustering of DOBV sequences from Serbia, unrelated to host (rodent or human). The topology of the phylogenetic tree was confirmed with a high percent of completely or partially resolved quartets in likelihood-mapping analysis, whereas no evidence of possible recombination in the examined S segment data set was found
European survey on laboratory preparedness, response and diagnostic capacity for crimean-congo haemorrhagic fever, 2012
No full textCrimean-Congo haemorrhagic fever (CCHF) is an infectious viral disease that has (re-)emerged in the last decade in south-eastern Europe, and there is a risk for further geographical expansion to western Europe. Here we report the results of a survey covering 28 countries, conducted in 2012 among the member laboratories of the European Network for Diagnostics of 'Imported' Viral Diseases (ENIVD) to assess laboratory preparedness and response capacities for CCHF. The answers of 31 laboratories of the European region regarding CCHF case definition, training necessity, biosafety, quality assurance and diagnostic tests are presented. In addition, we identified the lack of a Regional Reference Expert Laboratory in or near endemic areas. Moreover, a comprehensive review of the biosafety level suitable to the reality of endemic areas is needed. These issues are challenges that should be addressed by European public health authorities. However, all respondent laboratories have suitable diagnostic capacities for the current situation
The ocular conjunctiva and conjunctiva-associated lymphoid tissue as a mucosal immunization route: humoral and cellular immune responses against Salmonella typhimurium Bacterial Ghosts
No full textPurpose: In this work we explored the efficacies of Salmonella typhimurium (ST) Bacterial Ghosts (BGs), using the conjunctival route of immunization, to induce mucosal and systemic immunity against ST.
Methods: BALB/c and C57BL/6 mice were immunized via the conjunctiva with ST BGs in three different doses (50, 25 and 12.5 μg/mouse). Three immunizations were performed at 2 weeks interval and the evaluation of local and systemic immune response was done 2 and 8 weeks after the last immunization. This research was conducted in compliance with the “ARVO Statement of the Use of Animals in Ophthalmic and Visual Research” and the “Declaration of Helsinki”.
Results: ST BGs were well tolerated; non-toxic and successfully uptaken by the mouse conjunctival cells. We have found ST BG-specific IgG and IgA in tears and sera of both mice strains, as well as, IgG positive ST BG-specific cells. It was shown that the amount of ST BG-specific antibodies in mice' sera strongly correlated with the presence of ST BG-specific B cells in draining lymph nodes. 8 weeks after the last immunization, the levels of anti-ST BG IgG and IgA in the sera of BALB/c mice were higher than in C57BL/6. BALB/c mice exhibited also more prominent anti-ST BG IgA concentration in mice’ tears. The strongest IgG and IgA response was obtained with the highest dose used for immunization (50 μg/mouse). The conjunctival application of ST BGs in both mouse strains induced skewing of ST BG-specific immune responses toward a Th1/Th17 profile, as determined by the stimulation of IFNγ and IL-17A secretion and/or the concurrent pronounced reduction of IL-4 secretion.
Conclusions: Conjunctival immunization induced a strong ST BG-specific local and systemic immune response, and a Th1-biased response that is critical for control of Salmonella species. ST BGs, non-living shells of Gram negative ST bacteria, possess excellent safety properties and could be used as a candidate vaccine for an ocular delivery
Identification and characterization of lactic acid bacteria isolated from artisanal white brined Golija cows' milk cheeses
Get PDFThe aim of this study was to identify and characterize the lactic acid bacteria (LAB) of artisanal Golija raw and cooked cows' milk cheeses traditionally manufactured without the addition of starter culture. A total of 188 Gram-positive and catalase-negative isolates of Golija cheeses were obtained from seven samples of different ripening time. Phenotype-based assays as well as rep-PCR and 16S rDNA sequence analysis were undertaken for all 188 LAB strains. The most diverse species were isolated from 20-day-old BGGO8 cheese (Lactobacillus fermentum, Lactobacillus plantarum, Lactobacillus casei/paracasei, Lactobacillus sucicola, Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. lactis by. diacetylactis, Enterococcus faecium, Enterococcus durans and Leuconostoc mesenteroides). In other Golija cheeses Lactobacillus reuteri, Lactobacillus curvatus, Lactobacillus rhamnosus, Lactococcus lactis subsp. cremoris, Lactococcus garvieae, Streptococcus thermophilus and Leuconostoc pseudomesenteroides were found. Pronounced antimicrobial properties showed enterococci (13/42) and lactococci (12/31), while the good proteolytic activity demonstrated lactococci (13/31) and lactobacilli (10/29)