Cíl studie: Rozlišit genotypy a subtypy Borrelia burgdorferi sensu lato a Ehrlichia spp. V kulturách, ve vzorcích mozkomíšního moku a krve pacientů a ve tkáních zvířat. Studie byla zaměřena na střední a východní Čechy a Moravu. Materiál: Zkoumáno bylo 16 kmenů borelií, izolovaných z krve a moku pacientů, dále 49 vzorků mozkomíšního moku a krve pacientů s boreliózou, tkáně 80 kusů lovné zvěře a krev 55 krav. Metody: Diferenciaci a kvantifikaci obou patogenů umožnilo užití LightCycler PCR v reálném čase s příměry 16S rRNA, OspA, recA genů a přímá PCR-sekvenační analýza produktů. Výsledky: Prokázali jsme 11 kmenů Borrelia garinii, OspA-typy 5,6,4 a pět kmenů B. afélii. B. burgdorferi sensu stricto nebyla v kulturách prokázána. Sekvenační analýzou PCR produktů ze 49 vzorků mozkomíšního moku jsme prokázali B. burgdorferi s.s. v 20,4 %, B. afzeJii v 12,2 %, B. garinii, subtypy 6, 5, 4 a 3 v 61,3 %, z nichž 8,2 % vzorků obsahovalo kombinace blízkých OspA-typů. Koinfekce s EhrJichia phagocytophiia byla zjištěna v 6,1 % vzorků. E. phagocytophiia subtyp Frankonia I a II jsme prokázali u 12,5 % lovné zvěře a u 9 % skotu. B. burgdorferi sensu strigo a B. garinii byla zjištěna u 17,5 % lovné zvěře. Light Cycler PCR v reálném čase byla méně sensitivní v tekutinách pacientů než v kulturách a v tkáních zvířat. Závěr: Prokázali jsme, že neuroboreliózu mohou působit všechny tři genospecies B. burgdorferi sensu lato. Průběh infekce a symptomy mohou ovlivnit různé OspA- subtypy.

Purpose of the study: To differentiate the genotypes and subtypes of Borreiia burgdorferi sensu lato and Ehrlichia spp. in cultures, in samples of patients' cerebrospinal fluid and blood and in animal tissues. The study focuses on southern and eastern Bohemia and on Moravia. Material: The investigation involved 16 borrelia strains isolated from the blood and CSP of patients and further 49 samples of CSF and blood of borreliosis patients, tissues from 80 pieces of game animals and blood from 55 cows. Methods: The differentiation and quantification of the two pathogenes was done using real-time LightCycler PCR with the primers loS rRNA, OpA, recA genes and direct PCR sequential analysis of the products. Results: We demonstrated 11 strains of Borreiia garinii, OspA types 5, 6, 4 and 5 strains of B. afzelii. B. burgdorferi sensu stricto was not demonstrated in the cultures. The sequential analysis PCR of the products from 49 CSF samples demonstrated B, burgdorferi s.s. in 20.4 %, B. afzelii in 12.2 %, B. garnii subtypes 6, 5, 4 and 3 in 61.3 % - out of these 8.2 % of the samples contained combinations similar to OspA types. Co-infection with Ehrlichia phagocytophila was found in 6.1 % of samples. E. phagocytophila subtype Frankonia I and II was demonstrated in 12.5 % of game animals and in 9 % of cattle. B. burgdorferi sensu stricto and B. garinii were identified in 17.5 % of game. Real-time LightCycler PCR was less sensitive in patients' fluids than in cultures and animal tissues. Conclusions: We demonstrated that all three genotypes of B. burgdorferi sensu lato are capable of causing neuroborreliosis. The course of the infection and its symptoms may be affected by various OspA subtypes.

• MeSH
• Borrelia genetics   classification   pathogenicity   MeSH
• Ehrlichia genetics   pathogenicity   MeSH
• Ehrlichiosis diagnosis   microbiology   MeSH
• Research Support as Topic MeSH
• Humans MeSH
• Lyme Disease diagnosis   microbiology   MeSH
• Reverse Transcriptase Polymerase Chain Reaction methods   MeSH
• Signs and Symptoms diagnosis   MeSH
• Sequence Analysis methods   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Review MeSH
• Comparative Study MeSH

• MeSH
• Bartonella genetics   isolation & purification   classification   MeSH
• Borrelia burgdorferi Group genetics   isolation & purification   classification   MeSH
• Ehrlichia genetics   isolation & purification   classification   MeSH
• Ehrlichiosis microbiology   MeSH
• Humans MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH

BACKGROUND: Ehrlichia species are the etiological agents of emerging and life-threatening tick-borne human zoonoses that inflict serious and fatal infections in companion animals and livestock. The aim of this paper was to phylogeneticaly characterise a new species of Ehrlichia isolated from Rhipicephalus (Boophilus) microplus from Minas Gerais, Brazil. METHODS: The agent was isolated from the hemolymph of Rhipicephalus (B.) microplus engorged females that had been collected from naturally infested cattle in a farm in the state of Minas Gerais, Brazil. This agent was then established and cultured in IDE8 tick cells. The molecular and phylogenetic analysis was based on 16S rRNA, groEL, dsb, gltA and gp36 genes. We used the maximum likelihood method to construct the phylogenetic trees. RESULTS: The phylogenetic trees based on 16S rRNA, groEL, dsb and gltA showed that the Ehrlichia spp isolated in this study falls in a clade separated from any previously reported Ehrlichia spp. The molecular analysis of the ortholog of gp36, the major immunoreactive glycoproteins in E. canis and ortholog of the E. chaffeensis gp47, showed a unique tandem repeat of 9 amino acids (VPAASGDAQ) when compared with those reported for E. canis, E. chaffeensis and the related mucin-like protein in E. ruminantium. CONCLUSIONS: Based on the molecular and phylogenetic analysis of the 16S rRNA, groEL, dsb and gltA genes we concluded that this tick-derived microorganism isolated in Brazil is a new species, named E. mineirensis (UFMG-EV), with predicted novel antigenic properties in the gp36 ortholog glycoprotein. Further studies on this new Ehrlichia spp should address questions about its transmissibility by ticks and its pathogenicity for mammalian hosts.

• MeSH
• Bacteriological Techniques MeSH
• Arthropod Vectors microbiology   MeSH
• Species Specificity MeSH
• Ehrlichia classification   genetics   isolation & purification   MeSH
• Phylogeny MeSH
• Glycoproteins genetics   isolation & purification   metabolism   MeSH
• Humans MeSH
• Molecular Sequence Data MeSH
• Gene Expression Regulation, Bacterial physiology   MeSH
• Rhipicephalus microbiology   MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Amino Acid Sequence MeSH
• Zoonoses MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Companion animals can be infested by various species of parasitic insects. Cat flea Ctenocephalides felis (C. felis felis) (Bouché, 1835) and dog flea Ctenocephalides canis (Curtis, 1826) belong to multihost external parasites of mammals, which most frequently occur on domestic cats Felis catus Linnaeus and dogs Canis familiaris Linnaeus. The main aim of this study was to investigate the presence of pathogens, such as Anaplasma phagocytophilum (syn. Ehrlichia phagocytophila) and Rickettsia spp., in adult C. felis and C. canis fleas. Flea sampling has been realised from January 2013 to April 2017 in veterinary clinics, animal shelters and pet grooming salons. Fleas were collected from domestic cats and dogs, directly from the pet skin or hair. Then, the DNA was isolated from a single flea by using the alkaline hydrolysis and samples were screened for the presence of pathogens using PCR method. Anaplasma phagocytophilum has occurred in 29% of examined C. felis and 16% of C. canis individuals. In turn, the prevalence of Rickettsia spp. in cat fleas population was only 3%, and the dog fleas 7%. The present study showed the presence of pathogenic agents in cat and dog fleas, which indicates the potential role of these insects in circulation of A. phagocytophilum and Rickettsia spp. in the natural habitat. Furthermore, exposition to these flea species, whose hosts are domestic cats and dogs, can pose a potential risk of infection for humans.

• MeSH
• Anaplasma phagocytophilum isolation & purification   MeSH
• Ctenocephalides microbiology   MeSH
• Rickettsia isolation & purification   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Poland MeSH

Anaplasma phagocytophilum and Rickettsia spp. are vector-borne zoonotic bacteria, which are clinically important especially in immunocompromised patients. There are large gaps in the current knowledge of their geographic distribution and prevalence in both their vectors and hosts. Our aim was to develop reliable and easy detection method for both these pathogens. We made a new hydrolysis probe based duplex Real-Time PCR assay based on previous studies. We optimized the assays and tested them to provide reliable recommended procedures with a sensitivity to a minimum of 10 target DNA copies per sample. The assays were designed to be specific for A. phagocytophilum and in the same reaction detect multiple species of rickettsiae. We designed gBlock quantification standards that provide the option to identify differences in pathogen load among different samples in subsequent studies.

• MeSH
• Anaplasma phagocytophilum isolation & purification   MeSH
• DNA, Bacterial analysis   MeSH
• Hydrolysis MeSH
• Ixodes microbiology   MeSH
• Polymerase Chain Reaction methods   MeSH
• Rickettsia isolation & purification   MeSH
• Sensitivity and Specificity MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Celkem jsme v roce 2007 vyšetřili 1048 jedinců klíštěte Ixodes ricinus z toho 935 ze sběrů vlajkováním v pražských parcích a 114 jedinců ze zákusu na člověku. V roce 2008 jsme vyšetřili 364 jedinců z toho 277 ze sběru a 86 sejmutých z lidí. Promořenost klíšťat původcem lymeské borreliózy v roce 2007 byla 9,7 %, zjištěná v temném poli a v 13 % byla prokázána DNA pomocí PCR. V roce 2008 byla promořenost klíšťat vyšší, 14 % v temném poli a 20 % pomocí PCR. Klíšťata sejmutá po zákusu na lidech byla infikována v 7 %. Přítomnost Anaplasma phagocytophilum, Bartonella sp. a Rickettsia sp byla prokázána pomocí polymerázové řetězové reakce a kontrolována sekvencí geonomu u 11,3 - 11,8 % klíšťat. Spiroplasma rodu Babesia byla zjištěna v 7,4 % u samic I. ricinus. Klíšťata sejmutá z lidí byla infikovaná borrelií v 6,5 % a anaplasmou v 1,5 %. Pravidelnými sběry v jednotlivých měsících r. 2007 a 2008 na jedné lokalitě v Praze 10 jsme ověřili změny výskytu ruzných stádií klíštěte a jejich měnící se nákazu ruznými agens v závislosti na teplotě a vývoji. K růstu bakterií Borrelia a Anaplasma sp. ve střevě klíštěte je zapotřebí určité doby s vyšší teplotou. Nejmenší počet infikovaných klíšťat byl v srpnu 2007 a 2008, kdy se z velké většiny vyskytovaly pouze larvy.

In 2007, 1048 Ixodes ricinus ticks were investigated: 935 of these were collected by flagging in Prague parks and 113 were ticks attached to the human skin. In 2008, 364 ticks were investigated, i.e. 277 and 87 ticks collected by flagging and from humans, respectively. In 2007, the causative agent of lyme borreliosis was detected in 9.7% of ticks in dark field and in 13% of ticks in DNA by PCR. In 2008, higher positivity rates were found, i.e. 14% and 20%, respectively. Seven percent of ticks obtained from humans were infected. Anaplasma phagocytophilum, Bartonella sp. and Rickettsia sp. were detected by PCR and sequence analysis in 11.3% - 11.8% of ticks. Spiroplasma of the genus Babesia was detected in 7.4% of I. ricinus females. The ticks collected from humans were infected by Borrelia in 6.5% and by Anaplasma in 1.5%. Regular monthly flagging was performed in one locality in Prague 10 in 2007 and 2008 to obtain data on the incidence of different stages of ticks and rates of infection by different agents depending on temperature and season. To grow in the tick intestine, Borrelia and Anaplasma need higher temperature for a certain period of time. The lowest numbers of infected ticks were found in August 2007 and 2008 when mostly tick larvae were collected.

• MeSH
• Anaplasma phagocytophilum genetics   isolation & purification   MeSH
• Babesia genetics   isolation & purification   MeSH
• Bartonella genetics   isolation & purification   MeSH
• Borrelia burgdorferi genetics   isolation & purification   MeSH
• Disease Vectors MeSH
• Ixodes microbiology   MeSH
• Lyme Disease epidemiology   transmission   MeSH
• Polymerase Chain Reaction utilization   MeSH
• Publication type
• Chart MeSH

Wild birds are known to be a reservoir of infectious disease agents and disseminatory hosts of ticks. The purpose of this work was to obtain information about the occurrence of rickettsial, anaplasmal, and borrelial infections in some ticks that parasitize wild birds in the Czech Republic. A total of 549 subadult ticks of three species Ixodes arboricola (75.0%), Ixodes ricinus (23.1%), and Haemaphysalis concinna (1.8%) were collected from 20 species of birds (Passeriformes). Rickettsiae were detected in 44.0% larvae and 24.5% nymphs of I. arboricola collected from Parus major, Poecile palustris, and Sitta europaea. Rickettsiae-positive I. ricinus larvae (13.7%) were collected from P. major, Cyanistes caeruleus, and S. europaea, and 2.6% of nymphs from Erithacus rubecula and Prunella modularis. Comparison of sequences of a gltA gene fragment with data available in GenBank identified Rickettsia helvetica, a spotted fever rickettsia associated with human infections, and other Rickettsia spp. Anaplasma phagocytophilum was found only in two I. ricinus nymphs collected from E. rubecula and P. major. Infections with Borrelia burgdorferi sensu lato were recorded in 1.3% larvae of I. arboricola acquired from P. palustris and P. major and in 11.8% larvae and 25.0% nymphs of I. ricinus collected from P. major, P. palustris, C. caeruleus, Acrocephalus schoenobaenus, Turdus merula, Carpodacus erythrinus, Sylvia atricapilla, P. modularis, and Phylloscopus collybita. Reverse-line blot hybridization showed infections with Borrelia garinii and Borrelia valaisiana and mixed infections with these two genospecies. This is the first record of a high rate of rickettsial infection in I. arboricola subadult ticks acquired from birds in the Czech Republic and in central Europe. Our study suggests that I. arboricola, P. major, and P. palustris play important roles in circulating rickettsiae.

• MeSH
• Anaplasma phagocytophilum isolation & purification   MeSH
• Arachnid Vectors microbiology   MeSH
• Borrelia burgdorferi isolation & purification   MeSH
• Databases, Nucleic Acid MeSH
• Ixodes microbiology   MeSH
• Larva microbiology   MeSH
• Nymph microbiology   MeSH
• Polymerase Chain Reaction MeSH
• Birds parasitology   MeSH
• Rickettsia genetics   isolation & purification   MeSH
• Sequence Analysis MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Czech Republic MeSH

Ehrlichióza a anaplazmóza sú zoonózy zapríčinené baktériami z čeľade AnapJasmataceae, kde patria druhy patogénne pre ľudí aj zvieratá. Pre človeka sú patogénne Ehrhchia chaffeensis, pôvodca ľudskej monocytárnej ehrhchiózy (human monocytic ehrlichiosis HME), Anaplasma phagocytophilum, pôvodca ľudskej granulocytárnej ehrlichiózy (anaplazmózy) (human granulocytic anaplasmosis HGA), E. ewingii - granulocytotropická ehrlichia a Neorickettsia sennetsu ďalšia monocytotropická ehrlichia. Ehrlichie sú malé, gram negatívne, obligátne intracelulárne baktérie. Replikujú sa v cytoplazmatických vakuolách hostiteľských buniek, predovšetkým v granulocytoch a monocytoch, v ktorých vytvárajú mikrokolónie, takzvané moruly. Ehrlichie sú prenášané po aktívnom kontakte s infikovaným kliešťom. V USA sú ako vektory známe kliešte Amblyomma americanum, Ixodes scapularis a Ixodes pacificus. V Európe je primárnym vektorom Ixodes ricinus. Prameňom nákazy v Európe môžu byť hlodavce, jelene, srnce, líšky, hovädzí dobytok, ovce, kozy, kone, psy a pravdepodobne aj ďalšie zvieratá. V USA sú to najčastejšie myš bielonohá Peromyscus leucopus a jeleník bielochvostý OdocoiJeus virginianus. Ochorenie sa u človeka prejavuje nešpecifickými príznakmi pripomínajúce chrípku. Laboratórna diagnostika sa najčastejšie opiera o dôkaz protilátok nepriamou imunofluorescenciou (IFA) a o detekciu DNA polymerázovou reťazovou reakciou (PCR), alebo o mikroskopický dôkaz (farbenie krvného náteru podľa Giemsu - nález morúl v granulocytoch alebo monocytoch). V terapii je účinný doxycyklín. Najlepšou prevenciou ochorenia je ochrana pred kliešťom.

Ehrlichiosis and anaplasmosis are zoonoses caused by bacteria from the family Anapiasmataceae, including human and animal pathogens. The human pathogens are EhrUchia chaffeensis, the causative agent of human monocytic ehrlichiosis (HME), Anapiasma phagocytophilum, the pathogen causing human granulocytic anaplasmosis (HGA), E. ewingii and Neorickettsia sennetsu, granulocytotropic and monocytotropic Ehrlichia species, respectively. Ehrlichia spp. are small, gram-negative, obligate intracellular bacteria. They rephcate in the cytoplasmic vacuoles of host cells, especially granulocytes and monocytes, to form microcolonies called morulae. These agents are transmitted through the bite of infected tick. In the United States, the vectors are Ambiyomma americanum, Ixodes scapuiaris and Ixodes pacificus ticks. The primary vector in Europe is Ixodes ricinus. Rodents, deer, roe deer, foxes, cattle, sheep, goats, horses and dogs are reservoirs of these bacteria in Europe. Peromyscus ieucopus, the white-footed mouse, and Odocoileus virginianus, the white-tailed deer, are the most important reservoirs in the United States. Infection in humans is manifested as a nonspecific flu-like illness. The laboratory diagnosis is most frequently serological - evidence of antibody by indirect immunofluorescence assay (IFA) and detection of DN A by polymerase chain reaction (PCR), or microscopy evidence - Giemsa stain of blood smears (morulae in granulocytes or monocytes). Doxycycline is the drug of choice in therapy. Avoiding exposure to ticks is the best method of prevention of infection.

• MeSH
• Anaplasmosis diagnosis   microbiology   therapy   MeSH
• Anti-Bacterial Agents therapeutic use   MeSH
• Ehrlichiosis diagnosis   microbiology   therapy   MeSH
• Enzyme-Linked Immunosorbent Assay utilization   MeSH
• Financing, Organized MeSH
• Disease Vectors MeSH
• Ticks microbiology   MeSH
• Humans MeSH
• Polymerase Chain Reaction utilization   MeSH
• Disease Reservoirs microbiology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Review MeSH

Babesiosis is an emerging zoonotic disease and various wildlife species are reservoir hosts for zoonotic species of Babesia Starcovici, 1893. The objective of the present study was to investigate the presence and prevalence of Babesia spp. in moose Alces alces (Linnaeus) in two regions of Norway. A total of 99 spleen samples were collected from animals of various ages from an area with the occurrence of the tick Ixodes ricinus (Linnaeus, 1758), and from an area where the ticks are known to be absent. Infection was detected by the amplification of different regions of the 18S rRNA gene by using two different PCR primer sets specific of Babesia. Babesia spp. were found in the spleen samples of four moose. All Babesia-infected animals were from an area where ticks occur, with an infection rate of 6% (4 of 70). Babesia-positive samples were obtained from a five-month old moose calf and three adults. Two Babesia species, Babesia capreoli (Enigk et Friedhoff, 1962) and a B. odocoilei-like, were identified. Co-infection with Anaplasma phagocytophilum was obtained in two animals. This is the first report of the occurrence of B. capreoli and B. odocoilei-like species in moose.

• MeSH
• Anaplasma phagocytophilum physiology   MeSH
• Babesia genetics   isolation & purification   MeSH
• Babesiosis diagnosis   epidemiology   parasitology   MeSH
• Ehrlichiosis diagnosis   veterinary   MeSH
• Ixodes physiology   MeSH
• Coinfection MeSH
• RNA, Ribosomal, 18S genetics   MeSH
• Spleen parasitology   MeSH
• Deer parasitology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Norway epidemiology   MeSH

In this study, we screened a total of 2473 questing (years 2011-2014) and 199 engorged (years 2013 and 2014) Ixodes ricinus ticks for the presence of Rickettsia spp., "Candidatus Neoehrlichia mikurensis", Anaplasma phagocytophilum, and Babesia spp. Host-seeking ticks were collected at three study sites corresponding to natural woodland, urban park and pastureland ecosystem, and analyzed using molecular techniques. All pathogens tested were present at all study sites. The prevalence rates for Rickettsia spp., 'Candidatus Neoehrlichia mikurensis', Anaplasma phagocytophilum, and Babesia spp. ranged from 2.6% to 9.2%, 0.8% to 11.6%, 0% to 12.1%, and 0% to 5.2%, respectively. Engorged I. ricinus ticks collected from sheep on pastureland in the years 2013 and 2014 yielded prevalence rates 7.4% and 6.3%, respectively, for Rickettsia spp., 38.5% and 14.1% for 'Candidatus N. mikurensis', 18.5% and 12.5% for A. phagocytophilum, and 4.4% and 0.0% for Babesia spp. Monitoring of neglected tick-borne pathogens within the scope of epidemiological surveillance is an important tool for prevention and control of human tick-borne infections.

• MeSH
• Anaplasma phagocytophilum isolation & purification   MeSH
• Babesia isolation & purification   MeSH
• Time Factors MeSH
• Ixodes microbiology   parasitology   MeSH
• Rickettsia isolation & purification   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Czech Republic MeSH