Diversity of Lyme borreliosis spirochetes isolated from ticks in Serbia.

Spirochetes from the Borrelia burgdorferi sensu lato (s.l.). (Spirochaetales: Spirochaetaceae) species complex, including the causative agents of Lyme borreliosis, have been isolated from ticks, vertebrate reservoirs and humans. Previous analyses based on direct molecular detection in ticks indicated a considerable diversity of B. burgdorferi s.l. complex in Serbia. The present study aimed (a) to isolate borrelia strains from Serbia; (b) to determine their genotypic characteristics; and (c) to establish a collection of viable B. burgdorferi s.l. strains for further biological, ecological and genetic studies. For the present study, 231 adult Ixodes ricinus (Ixodida: Ixodidae) ticks from 16 ecologically different localities in Serbia were individually processed to cultivate B. burgdorferi s.l. This led to the isolation of 36 strains. A hbb gene quantitative real-time polymerase chain reaction (PCR) based on melting temperature determination and ospA gene sequencing were used to genotype the isolated spirochetes. The species identified based on the hbb gene real-time PCR were: Borrelia lusitaniae (44.4%), Borrelia afzelii (36.1%), Borrelia garinii (13.9%) and Borrelia valaisiana (5.6%), whereas the ospA sequence analysis revealed the occurrence of Borrelia bavariensis. This is the first report of the isolation of B. lusitaniae, B. garinii, B. bavariensis and B. valaisiana strains in Serbia.

Patterns of Abundance and Host Specificity of Bat Ectoparasites in the Central Balkans.

Bats are hosts to a number of ectoparasites-acarines (ticks, chiggers, other mites), bat flies, and fleas. Bat ectoparasites might have significant ecological and public health importance as they may be potential vectors of zoonotic agents. It is important to identify their distribution, diversity, and host-parasite associations. Bat ectoparasites in the central Balkans have been largely understudied. The present research was conducted in 45 localities at the territory of Bosnia and Herzegovina, former Yugoslav Republic of Macedonia, Montenegro, and Serbia. In total, 1,143 individuals of 18 species of bats have been examined for the presence and abundance of ectoparasite species during 3 yr of research. In total, 21 ectoparasite species have been identified: three species of ticks, seven species of mites (including one species of chigger), eight species of bat flies, and three species of fleas. In total, 80 host-parasite associations have been identified. The largest number of ectoparasites parasitized primarily only one host species. The highest total number of hosts was identified for ectoparasite species Ixodes vespertilionis Koch, Nycteribia schmidlii Schiner, and Spinturnix myoti Kolenati. The spinturnicid mite Spinturnix psi Kolenati was the most abundant ectoparasite species and together with Penicilidia dufouri Westwood the most widely distributed species of bat ectoparasite, being present at 21 localities in the central Balkans. The presented data include the first systematic records of patterns of prevalence, mean intensity, mean abundance, and host specificity for bat ectoparasites in the central Balkans.

Ticks (Acari: Argasidae, Ixodidae) parasitizing bats in the central Balkans.

Ticks parasitizing bats have been largely understudied, especially in the central part of the Balkan Peninsula, where the last data from the field research date from almost 25 years ago. Bats are hosts to a large number of ectoparasites, including ticks, which can act as vectors of zoonotic agents. For this reason, it is important to identify the distribution of ticks and their relationship to different hosts, including wild animals, bats in particular. The present research was conducted at 16 localities throughout Serbia, Montenegro, Bosnia and Herzegovina, and Former Yugoslav Republic of Macedonia (FYROM). We examined 475 individuals of bats belonging to 13 species. A total of three tick species were identified, I. simplex being the most numerous and widespread, followed by I. vespertilionis and A. vespertilionis. To the best of our knowledge, the presented data include the first records of I. simplex in Serbia and Montenegro, I. vespertilionis for Montenegro and A. vespertilionis in FYROM. Also, we identify a new possible host/parasite association between I. simplex and Rhinolophus euryale.

The impact of postoperative pain on early ambulation after hip fracture.

Hip fractures are the most serious consequence of falling in older people with osteoporosis. Early ambulation is one of the main goals of inpatient rehabilitation aimed at reducing negative effects of immobilization, and promoting functional recovery. Recognizing the role of different factors that are associated with time to ambulation time after hip fracture surgery may help decrease morbidity and mortality rates. The aim of this study was to examine the impact of postoperative pain time to ambulation following hip fracture surgery. A total of 96 patients who consecutively presented at the Clinic for Orthopaedic Surgery and Traumatology, Clinical Center Serbia during a 6 month period were included in the study. Subjects were assessed regarding socioedemographic characteristics, prefracture functional status, cognitive status, general health status, fracture type, type of anaesthesia, and waiting time for surgery. The VAS scale (0-100) was used to measure pain intensity on the first postoperative day. Our results revealed that patients, whose time to ambulation was = 48 h postoperatively were of worse physical health, and had significantly higher VAS score on the first postoperative day. Unrelieved pain in geriatric hip fracture patients is associated with delayed ambulation. Future research should confirm the posi-tive effects of aggressive pain management programs on early functional outcome in elderly patients who sustain hip fractures.

Seasonal and spatial occurrence of glycerol-3-phosphate dehydrogenase variability in Ixodes ricinus (Acari: Ixodidae) populations.

Previous results indicate that glycerol-3-phosphate dehydrogenase variability represents the adaptation of Ixodes ricinus L. (Acari: Ixodidae) to fluctuations of environmental conditions, particularly to temperature. Analysis of crucial polymorphisms in I. ricinus Gpdh gene was done by the restriction method, and three different haplotypes were obtained (GPDH441 1, GPDH441 2, and GPDH441 3), corresponding to GPDH alleles detected by allozyme electrophoresis. Differences in GPDH441 haplotype and genotype frequencies were found between samples from open and forest habitats. Significant seasonal variations of GPDH441 haplotype and genotype frequencies were detected in samples from the open habitats. No seasonal variations were observed at forest localities, probably because of the less pronounced amplitude of environmental factors. The possible role of host availability was discussed as an important factor that affects seasonal dynamics and genetic composition of tick populations.

Coexistence of emerging bacterial pathogens in Ixodes ricinus ticks in Serbia.

The list of tick-borne pathogens is long, varied and includes viruses, bacteria, protozoa and nematodes. As all of these agents can exist in ticks, their co-infections have been previously reported. We studied co-infections of emerging bacterial pathogens (Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum and Francisella tularensis) in Ixodes ricinus ticks in Serbia. Using PCR technique, we detected species-specific sequences, rrf-rrl rDNA intergenic spacer for B. burgdorferi s.l., p44/msp2 paralogs for A. phagocytophilum, and the 17 kDa lipoprotein gene, TUL4, for F. tularensis, respectively, in total DNA extracted from the ticks. Common infections with more than one pathogen were detected in 42 (28.8%) of 146 infected I. ricinus ticks. Co-infections with two pathogens were present in 39 (26.7%) of infected ticks. Simultaneous presence of A. phagocytophilum and different genospecies of B. burgdorferi s.l. complex was recorded in 16 ticks, co-infection with different B. burgdorferi s. l. genospecies was found in 15 ticks and eight ticks harbored mixed infections with F. tularensis and B. burgdorferi s.l. genospecies. Less common were triple pathogen species infections, detected in three ticks, one infected with A. phagocytophilum / B. burgdorferi s.s. / B. lusitaniae and two infected with F. tularensis / B. burgdorferi s.s. / B. lusitaniae. No mixed infections of A. phagocytophilum and F. tularensis were detected.

Exon variability of gene encoding glycerol-3-phosphate dehydrogenase of Ixodes ricinus ticks.

We have previously found apparent differences in Gpdh allele frequences between borrelia infected and uninfected Ixodes ricinus as revealed by native gel electrophoresis of allozyme polymorphisms. The present study deals with the genetic basis of the observed allozyme polymorphism. Multiple sequence alignment of 36 Gpdh open reading frames identified a total of 40 polymorphic nucleotide sites. Of the 40 polymorphic nucleotide sites, 34 were silent (did not result in amino acid residue change), while six were active causing a change in the amino acid chain. All polymorphic amino acid sites were situated within the N-terminal NAD-binding domain, whereas the C-terminal substrate-binding domain was highly conserved. Analysis of the obtained Gpdh sequences and GPDH allozyme polymorphisms for individual ticks pointed to amino acid changes at positions 61 (glycine-to-glutamic acid), 64 (serine-to-cysteine) and 102 (glycine-to-arginine) as a key for differential mobility of GPDH allozymes in an electric field. Our findings are discussed in the context of the molecular basis of I. ricinus host finding behavior.

Assessment of the risk of contracting Lyme disease in areas with significant human presence / Risco de contrair doença de Lyma em áreas com significativa presença humana

In order to describe seasonal changes in Lyme diseases risk rate at three localities in Serbia, during the period of 2003-2005, a total of 1542 Ixodes ricinus ticks (493 nymphs, 525 females and 524 males) were examined. The prevalence of Borrelia burgdorferi in Ixodes ricinus ticks at the Bovan Lake County were higher than the average for European localities (45.9 percent for adults and 18.8 percent for nymphs). In Mt. Avala and Kljajicevo counties adults and nymphs were, respectively, infected at the following percentages: 26.3, 10.7; 16.2 and 7.6 percent. The outcome indicates a relatively high risk of the contracting Lyme disease in all investigates areas.

Para estimar a variação sazonal das taxas de risco para doença de Lyme em três localidades da Sérvia foram examinados, no período de 2003-2005, 1542 espécimes do carrapato Ixodes ricinus (493 ninfas, 525 fêmeas e 524 machos). A prevalência de Borrelia burgdorferi em Ixodes ricinus no município de Bovan Lake foi mais alta que a registrada em outras localidades da Europa. Nos municípios de Mt. Avala e Kljajicevo as porcentagens de adultos e ninfas infectadas foram: 26,3 e 10,7; 16,2 e 7,6, respectivamente. Esses resultados indicam um relativo alto risco de se contrair doença de Lyme nas três localidades estudadas.

[Serologic diagnosis of Crimean hemorrhagic fever in Kosovo and Metohia]. / Seroloska dijagnostika Krimske hemoragijske groznice na Kosovu i Metohiji.

In the period June-November 1995, 292 sera from 159 patients and 80 healthy persons from Kosovo and Metohia were tested for the presence of antibodies against the causative agents of Crimean hemorrhagic fever (CHF) and hemorrhagic fever with renal syndrome (HFRS). Diffusion precipitation test in agar gel and indirect immunofluorescence assay were used. Specific antibodies against CHF virus were found in 6.9% and against HFRS virus in 15.1 patient's sera. Antibodies against CHF virus were proved in sera of patients on the fifth day from the disease onset at the earliest. No wanted antibodies were found in healthy persons' sera. The results of sera testing on antibodies against CHF virus were analyzed in detail.