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Background: Changing geographical and seasonal activity patterns of ticks may increase the risk of tick infestation and tick-borne pathogen (TBP) transmission for both humans and animals. Methods: To estimate TBP exposure of dogs and cats, 3000 female I. ricinus from these hosts were investigated for Anaplasma phagocytophilum and Borrelia species. Results: qPCR inhibition, which was observed for ticks of all engorgement stages but not questing ticks, was eliminated at a template volume of 2 µl. In ticks from dogs, A. phagocytophilum and Borrelia spp. prevalence amounted to 19.0% (285/1500) and 28.5% (427/1500), respectively, while ticks from cats showed significantly higher values of 30.9% (464/1500) and 55.1% (827/1500). Accordingly, the coinfection rate with both A. phagocytophilum and Borrelia spp. was significantly higher in ticks from cats (17.5%, 262/1500) than dogs (6.9%, 104/1500). Borrelia prevalence significantly decreased with increasing engorgement duration in ticks from both host species, whereas A. phagocytophilum prevalence decreased only in ticks from dogs. While A. phagocytophilum copy numbers in positive ticks did not change significantly over the time of engorgement, those of Borrelia decreased initially in dog ticks. In ticks from cats, copy numbers of neither A. phagocytophilum nor Borrelia spp. were affected by engorgement. Borrelia species differentiation was successful in 29.1% (365/1254) of qPCR-positive ticks. The most frequently detected species in ticks from dogs were B. afzelii (39.3% of successfully differentiated infections; 70/178), B. miyamotoi (16.3%; 29/178), and B. valaisiana (15.7%; 28/178), while B. afzelii (40.1%; 91/227), B. spielmanii (21.6%; 49/227), and B. miyamotoi (14.1%; 32/227) occurred most frequently in ticks from cats. Conclusions: The differences in pathogen prevalence and Borrelia species distribution between ticks collected from dogs and cats may result from differences in habitat overlap with TBP reservoir hosts. The declining prevalence of A. phagocytophilum with increasing engorgement duration, without a decrease in copy numbers, could indicate transmission to dogs over the time of attachment. The fact that this was not observed in ticks from cats may indicate less efficient transmission. In conclusion, the high prevalence of A. phagocytophilum and Borrelia spp. in ticks collected from dogs and cats underlines the need for effective acaricide tick control to protect both animals and humans from associated health risks. [ABSTRACT FROM AUTHOR]

Background: Hyalomma marginatum and H. rufipes are two-host tick species, which are mainly distributed in southern Europe, Africa to central Asia but may also be found in Central and Northern Europe through introduction by migratory birds., Methods: Ticks were collected while feeding or crawling on animals and humans, or from the environment, in different regions in Germany, between 2019 and 2021 in a citizen science study and from 2022 to 2023 in the wake of this study., Results: From 2019 to 2023, a total of 212 Hyalomma adult ticks were detected in Germany. This included 132 H. marginatum and 43 H. rufipes ticks sent to research institutions and 37 photographic records that were only identified to genus level. The number of detected ticks varied over the years, with the highest number of 119 specimens recorded in 2019, followed by 57 in 2020. Most of the specimens were collected from horses, while some were collected from other animals, humans or found crawling on human clothes or other objects inside or outside houses. The screening of 175 specimens for Crimean-Congo hemorrhagic fever virus and of 132 specimens for Babesia/Theileria spp. by PCR gave negative results, while human-pathogenic Rickettsia were detected in 44% (77/175) of the total samples. Subsequent amplicon sequencing and phylogenetic analysis of representative samples determined the species of 41 Rickettsia aeschlimannii and one R. slovaca sequences., Conclusions: Analysis of climatic factors indicated a significantly higher probability of Hyalomma occurrence at locations with higher average spring temperature during the years 2019 and 2020 compared to randomly generated pseudo-absence locations. Dry and hot conditions probably facilitated Hyalomma nymphs' survival and molting into adults during these years., (© 2024. The Author(s).)

Background: In Europe, canine babesiosis is most frequently caused by Babesia canis and Babesia vogeli, and occasionally by Babesia gibsoni.. In Germany, B. canis is recognized as endemic. The aims of this study were to assess how often Babesia spp. infections were diagnosed in a commercial laboratory in samples from dogs from Germany, and to evaluate potential risk factors for infection., Methods: The database of the LABOKLIN laboratory was screened for Babesia spp.-positive polymerase chain reaction (PCR) tests for dogs for the period January 2007-December 2020. Sequencing was performed for positive tests from 2018 and 2019. Binary logistic regression analysis was performed to determine the effects of sex, season, and year of testing. Questionnaires were sent to the submitting veterinarians to obtain information on travel abroad, tick infestation, and ectoparasite prophylaxis of the respective dogs. Fisher's exact test was used to calculate statistical significance and P < 0.05 was considered statistically significant., Results: In total, 659 out of 20,914 dogs (3.2%) tested positive for Babesia spp. by PCR. Of 172 sequenced samples, B. canis was identified in 156, B. vogeli in nine, B. gibsoni in five, and B. vulpes in two. Season had a statistically significant impact on test results when summer/winter (1.6% tested positive) was compared to spring/autumn (4.7%), with peaks in April (5.2%) and October (7.4%) [P < 0.001, odds ratio (OR) = 3.16]. Sex (male 3.5%, female 2.8%; P = 0.012, OR = 1.49) and age (< 7 years old 4.0%, ≥ 7 years old 2.3%; P < 0.001, OR = 1.76) of the tested dogs also had a statistically significant effect. A statistically significant impact was demonstrated for observed tick attachment (P < 0.001, OR = 7.62) and lack of ectoparasite prophylaxis (P = 0.001, OR = 3.03). The frequency of positive Babesia spp. tests did not significantly differ between the 659 dogs that had never left Germany and the 1506 dogs with known stays abroad (P = 0.088)., Conclusions: The possibility of canine infection with B. canis needs to be especially taken into consideration in spring and autumn in Germany as the activity of the tick Dermacentor reticulatus, a potential vector for canine babesiosis, is highest in these seasons. Travel and importation of dogs are considered major factors associated with canine babesiosis in Germany. However, autochthonous Babesia spp. infections also occur in a considerable number of dogs in Germany., (© 2023. The Author(s).)

Background: Ixodes spp. are vectors of zoonotic pathogens. All three active life stages (larvae, nymphs, adults) need to feed on a host in order to develop. Usually ticks parasitize attached to the external surface of their hosts' skin. Interestingly, in some cases ticks can also be found in the subcutaneous tissue in a variety of hosts, such as red foxes (Vulpes vulpes), raccoon dogs (Nyctereutes procyonoides) and dogs. Methods: The visceral side of 126 red fox-furs from Germany was examined visually searching for ticks. The localization of ticks was recorded and assigned to ten specific body parts. Morphological identification of ticks was performed according to standardized taxonomic protocols. Ticks which could not be further identified were examined genetically via conventional PCR targeting the 16S rRNA and cox1 gene. Hematoxylin and eosin (H&E) staining was used for histopathological examination. Results: In 111 out of 126 (88.1%) examined coats, at least one tick was found in the subcutaneous tissue. A total of 1203 ticks were removed from the subcutaneous tissue. Well-preserved ticks could be identified based on morphological criteria, but most ticks were in a progressed state of decomposition. Here, morphological species identification was not successful. Also, PCR methods did not lead to a successful species identification. The following species and development stages were found by morphological identification: Ixodes ricinus (female, n = 289; male, n = 8; nymph, n = 1), I. hexagonus (female, n = 2), I. canisuga (female, n = 1). Male I. ricinus were found individually or copulating in pairs with females. Subcutaneous ticks were localized at three predominant affected body parts: ears, axillar and inguinal region. Histological examination of subcutaneous ticks revealed a granulomatous panniculitis. Conclusions: To the authors' knowledge, this is the first finding of highly prevalent subcutaneous ticks in red foxes from Germany. Subcutaneous location of ticks seems to be very common in red foxes and the rule rather than the exception. Deep embedment of longirostra and long feeding times of females seem to put the subcutaneous location in favor. Most foxes were infested in the inguinal area, where the skin is thin and less hairy. [ABSTRACT FROM AUTHOR]

Background: Ixodid ticks are important vectors for zoonotic pathogens, with Ixodes ricinus being the most important in Europe. Rodents are hosts of immature life stages of I. ricinus ticks and are considered main reservoirs for tick-borne pathogens, e.g. Borrelia burgdorferi. The aim of this study was to analyse the prevalence as well as genospecies and sequence type (ST) diversity of Borrelia burgdorferi sensu lato in ticks and small mammals from central Germany and to elaborate on the influence of environmental and/or individual host and vector factors on Borrelia prevalence. Methods: After species identification, 1167 small mammal skin samples and 1094 ticks from vegetation were screened by B. burgdorferi sensu lato real-time polymerase chain reaction, and positive samples were characterized by multilocus sequence typing. Generalized linear (mixed) models were used to estimate how seasonality, small mammal species/tick life stage and habitat affect individual infection status. Results: In total, 10 small mammal species and three tick species, Ixodes ricinus, Ixodes inopinatus (both considered members of the I. ricinus complex) and Dermacentor reticulatus, were investigated. Borrelia DNA was detected in eight host species, i.e. the striped field mouse (Apodemus agrarius), the yellow-necked field mouse (Apodemus flavicollis), the wood mouse (Apodemus sylvaticus), the water vole (Arvicola amphibius), the bank vole (Clethrionomys glareolus), the field vole (Microtus agrestis), the common vole (Microtus arvalis), and the common shrew (Sorex araneus). Two species were Borrelia negative, the greater white-toothed shrew (Crocidura russula) and the pygmy shrew (Sorex minutus). The average prevalence was 6.2%, with two genospecies detected, Borrelia afzelii and Borrelia garinii, and at least three STs that had not been previously reported in small mammals. Borrelia prevalence in small mammals did not differ between seasons. Six genospecies of Borrelia—Borrelia afzelii, Borrelia valaisiana, Borrelia garinii, Borrelia lusitaniae, Borrelia spielmanii, and Borrelia burgdorferi sensu stricto—and 25 STs of Borrelia, of which 12 have not been previously described at all and five have not been previously reported in Germany, were detected in 13% of I. ricinus complex ticks. Prevalence was highest in adult females (25.3%) and lowest in nymphs (11.4%). Prevalence was significantly higher in ticks from grassland (16.8%) compared to forests (11.4%). Conclusions: The high level of small mammal diversity in this region of Germany seems to be reflected in a wide variety of genospecies and STs of B. burgdorferi. [ABSTRACT FROM AUTHOR]

Background: Ixodes ricinus is the most common tick species in Europe and the main vector for Borrelia burgdorferi (sensu lato) and tick-borne encephalitis virus (TBEV). It is involved also in the transmission of Borrelia miyamotoi, a relapsing fever spirochete that causes health disorders in humans. Little is known regarding the circulation of Borrelia species and the natural foci of TBEV in north-eastern Germany. The goal of this study was to investigate the infection rates of Borrelia spp. and of TBEV in I. ricinus ticks from north-eastern Germany., Methods: Ticks were collected by flagging from 14 forest sites in Mecklenburg-Western Pomerania between April and October 2018. RNA and DNA extraction was performed from individual adult ticks and from pools of 2-10 nymphs. Real time reverse transcription PCR (RT-qPCR) targeted the 3' non-coding region of TBEV, while DNA of Borrelia spp. was tested by nested PCR for the amplification of 16S-23S intergenic spacer. Multilocus sequence typing (MLST) was performed on B. miyamotoi isolates., Results: In total, 2407 ticks were collected (239 females, 232 males and 1936 nymphs). Female and male I. ricinus ticks had identical infection rates (both 12.1%) for Borrelia spp., while nymphal pools showed a minimum infection rate (MIR) of 3.3%. Sequencing revealed four Borrelia species: B. afzelii, B. garinii, B. valaisiana and B. miyamotoi. Borrelia afzelii had the highest prevalence in adult ticks (5.5%) and nymphs (MIR of 1.8%). Borrelia miyamotoi was identified in 3.0% of adults and registered the MIR of 0.8% in nymphs. Borrelia valaisiana was confirmed in 2.5% adult ticks and nymphs had the MIR of 0.7%, while B. garinii was present in 1.1% of adults and showed a MIR of 0.1% in nymphs. The MLST of B. miyamotoi isolates showed that they belong to sequence type 635. No tick sample was positive after RT-qPCR for TBEV RNA., Conclusions: The prevalence of B. miyamotoi in I. ricinus ticks registered similar levels to other reports from Europe suggesting that this agent might be well established in the local tick population. The detection of B. burgdorferi (s.l.) indicates a constant circulation in tick populations from this region.

Background: The European hedgehog (Erinaceus europaeus) is known for high levels of ectoparasitism that not only represents a health risk for the animals themselves, but also for pet animals and humans as hedgehogs are frequently taken into human care. In the present study, patterns of ectoparasite infestation were assessed in hedgehogs taken into care at northern German animal rehabilitation centres. Methods: Ectoparasites (ticks, fleas and mites) of 498 hedgehogs were collected over a period of 3 years from July 2018 to May 2021. Species were identified based on morphological characteristics and also via amplification and sequencing of the partial cytochrome c oxidase subunit 2 (COX-2) gene for fleas of the family Ceratophyllidae. Seasonal changes in infestation patterns as well as correlations with animal age, body weight and health status were assessed using generalised linear models. Results: Infestation with ticks, fleas and mites occurred throughout the year. Overall, 86.5% (431/498) of the examined hedgehogs were infested with ticks, 91.4% (455/498) with fleas and 17.7% (88/498) with mites.Ixodes ricinus and Ixodes hexagonus/Ixodes canisuga were the most common tick species detected, with the additional occurrence of one Ixodes frontalis. Significant seasonal changes were observed for I. ricinus, but not for I. hexagonus/I. canisuga. Additionally, I. ricinus nymph prevalence declined significantly as of 2020, probably as a consequence of the climate change-related drought as of 2018. In hedgehogs with flea infestations, Archaeopsylla erinacei, Ceratophyllus sciurorum, Nosopsyllus fasciatus and Ctenocephalides felis were identified. In all cases of mite infestation, Caparinia tripilis was detected, in addition to specimens of the family Macronyssidae and free-living mites of the family Acaridae. Statistical analyses showed correlations regarding the factors month, year, body weight and age, but no correlation was evident regarding the health status of the animals. Conclusions: With a detected infestation rate of 98.6%, almost all of the examined hedgehogs were infested with at least one ectoparasite species. The seasonal activity patterns of the different ectoparasite species together with the complex annual cycle of hedgehogs lead to different seasonal patterns in ectoparasite prevalence and infestation intensities. Due to the risk of transmission of zoonotic pathogens as well as the possible negative impact on the host itself, hedgehogs should be treated against ectoparasites when taken into care facilities. [ABSTRACT FROM AUTHOR]

Background: Arthropod-borne diseases remain a major health-threat for humans and animals worldwide. To estimate the distribution of pathogenic agents and especially Bartonella spp., we conducted tick microbiome analysis and determination of the infection status of wild animals, pets and pet owners in the state of Hesse, Germany., Results: In total, 189 engorged ticks collected from 163 animals were tested. Selected ticks were analyzed by next generation sequencing (NGS) and confirmatory PCRs, blood specimens of 48 wild animals were analyzed by PCR to confirm pathogen presence and sera of 54 dogs, one cat and 11 dog owners were analyzed by serology. Bartonella spp. were detected in 9.5% of all ticks and in the blood of 17 roe deer. Further data reveal the presence of the human and animal pathogenic species of genera in the family Spirochaetaceae (including Borrelia miyamotoi and Borrelia garinii), Bartonella spp. (mainly Bartonella schoenbuchensis), Rickettsia helvetica, Francisella tularensis and Anaplasma phagocytophilum in ticks. Co-infections with species of several genera were detected in nine ticks. One dog and five dog owners were seropositive for anti-Bartonella henselae-antibodies and one dog had antibodies against Rickettsia conorii., Conclusions: This study provides a snapshot of pathogens circulating in ticks in central Germany. A broad range of tick-borne pathogens are present in ticks, and especially in wild animals, with possible implications for animal and human health. However, a low incidence of Bartonella spp., especially Bartonella henselae, was detected. The high number of various detected pathogens suggests that ticks might serve as an excellent sentinel to detect and monitor zoonotic human pathogens.

Background: Ticks and tick-borne diseases play a major role in companion animal health. Additionally, the European tick fauna is changing, for instance due to the spread of Dermacentor reticulatus, displaying a higher likelihood of winter activity than Ixodes ricinus. Therefore, we investigated current tick infestations in dogs and cats in Germany and in parts of Austria and the seasonal infestation risk. Methods: Overall, 219 veterinary practices were invited to collect ticks from cats and dogs on a monthly basis. Ticks were morphologically identified and female I. ricinus specimens were measured to estimate attachment duration. Results: In total, 19,514 ticks, 17,789 (91.2%) from Germany and 1506 (7.7%) from Austria, were received between March 2020 and October 2021, with 10,287 specimens (52.7%) detached from dogs, 8005 from cats (41.0%) and 1222 from other species (6.3%). In Germany, the most common tick species collected from dogs were I. ricinus (78.0%) and D. reticulatus (18.8%), while cats mainly harboured I. ricinus (91.3%) and I. hexagonus (5.5%) and only few D. reticulatus (0.6%). In Austria, collected I. ricinus reached similar proportions in dogs (90.4%) and cats (95.3%), followed by D. reticulatus in both dogs (5.2%) and cats (1.5%), with I. hexagonus (0.9%) collected only marginally from cats. The average infestation intensity amounted to 1.62 ticks/dog and 1.88 ticks/cat. The single to multiple infestation ratio was 79.1% to 20.9% in dogs and 69.0% to 31.0% in cats, with cats being significantly more often multiple infested than dogs, while the proportion of mixed-species infestations was 2.0% for both dogs and cats. The average attachment duration of female I. ricinus specimens amounted to 78.76 h for dogs and 82.73 h for cats. Furthermore, year-round tick exposure was confirmed, with 108 D. reticulatus and 70 I. ricinus received on average per month during December 2020 to February 2021. Conclusions: The study shows a year-round tick infestation risk, with activity of both D. reticulatus and I. ricinus during winter, and confirms the widespread occurrence of D. reticulatus in Germany. Additionally, long average attachment durations and frequent multiple infestations underline the need for adequate year-round tick control, even during the winter months. [ABSTRACT FROM AUTHOR]

Background: As the vector-borne diseases tick-borne encephalitis (TBE) and Lyme borreliosis (LB) are common in Germany and transmitted by tick bites, the aim of this study was to assess differences in the number of tick bites in various professions and regions across southern Germany to evaluate the differences in tick-associated risk., Materials and Methods: The analysis is based on three cross-sectional studies that were conducted in 2016 and 2017 in two real-life settings and in one medical setting in Bavaria. All participants filled in a paper-based questionnaire about their history with tick bites. Only adult participants (≥ 18 years) were included in this study., Results: Overall, 3503 individuals (mean age 50.8 ± 15.2 years, median age 53.0 ± 12.2 years, 54.0% female) were included. Of these, 50% worked in an outdoor profession and 56% lived in environs. Around 70% of participants reported at least one previous tick bite. In comparison to indoor workers, forestry workers (OR = 2.50; 95% CI: 1.10-5.68) had the highest risk for a tick bite followed by farmers (OR = 1.22; 95% CI: 1.01-1.47). Furthermore, people living in rural areas (OR = 1.97, 95% CI:1.49-2.59) and environs (OR = 1.98, 95% CI: 1.54-2.55) were twice as likely to have a previous tick bite than people living in urban areas. In general, slightly more tick bites were reported by people living in eastern Bavaria., Conclusion: Rising numbers of TBE and LB indicate the need for further prevention strategies, which should focus on outdoor professions with a higher risk and people living in environs and rural areas., (© 2022. The Author(s).)

Background: Tick-borne encephalitis (TBE) is the most common viral CNS infection with incidences much higher than all other virus infections together in many risk areas of central and eastern Europe. The Odenwald Hill region (OWH) in southwestern Germany is classified as a TBE risk region and frequent case numbers but also more severe infections have been reported within the past decade. The objective of the present study was to survey the prevalence of tick-borne encephalitis virus (TBEV) in Ixodes ricinus and to associate TBEV genetic findings with TBE infections in the OWH. Methods: Ticks were collected by the flagging methods supported by a crowdsourcing project implementing the interested public as collectors to cover completely and collect randomly a 3532 km2 area of the OWH TBE risk region. Prevalence of TBEV in I. ricinus was analysed by reversed transcription quantitative real-time PCR. Phylogeographic analysis was performed to classify OWH TBEV isolates within a European network of known TBEV strains. Mutational sequence analysis including 3D modelling of envelope protein pE was performed and based on a clinical database, a spatial association of TBE case frequency and severity was undertaken. Results: Using the crowd sourcing approach we could analyse a total of 17,893 ticks. The prevalence of TBEV in I. ricinus in the OWH varied, depending on analysed districts from 0.12% to 0% (mean 0.04%). Calculated minimum infection rate (MIR) was one decimal power higher. All TBEV isolates belonged to the European subtype. Sequence analysis revealed a discontinuous segregation pattern of OWH isolates with two putative different lineages and a spatial association of two isolates with increased TBE case numbers as well as exceptional severe to fatal infection courses. Conclusions: TBEV prevalence within the OWH risk regions is comparatively low which is probably due to our methodological approach and may more likely reflect prevalence of natural TBEV foci. As for other European regions, TBEV genetics show a discontinuous phylogeny indicating among others an association with bird migration. Mutations within the pE gene are associated with more frequent, severe and fatal TBE infections in the OWH risk region. [ABSTRACT FROM AUTHOR]

Background: Small mammals are important maintenance hosts of ectoparasites as well as reservoir hosts for many arthropod-borne pathogens. In Germany, only a few studies have investigated ectoparasite communities on small mammals in their natural habitats. The aim of this study was to assess the species diversity and parameters influencing the mean intensity and prevalence of macroscopically visible ectoparasites, such as fleas, predatory mites and ticks., Methods: A total of 779 small mammals and 3383 ticks were available from earlier investigations for the data analysis of the current study from three differently structured study sites. In addition, fleas and predatory mites were collected from the captured rodents and taxonomically identified. Regression analyses were conducted on the group (ticks/mites/fleas) and species levels using hurdle models for the abundance of ectoparasite groups and a negative binomial model for the abundance of species., Results: Nearly 90% of the small mammals analyzed were infested with ectoparasites, with an average of 7.3 specimens per host. Hosts were infested with up to six species of ectoparasites simultaneously. In total, 12 flea, 11 mite and three tick species were detected. Ticks were more prevalent than fleas or mites, with > 80% of the hosts in urban and forest areas hosting ticks and around 60% of hosts presenting fleas, and only 20-40% of hosts presenting mites. Polyparasitism had a statistically significant influence on the prevalence of the investigated tick, mite and flea species, with odds ratios of > 1.0. Trapping location, season and host characteristics had significant influences on some-but not all-of the investigated species., Conclusions: The diversity of flea species was unexpectedly high and higher than that reported in comparable studies, which can be explained by the differently structured habitats and regions examined in this study. Polyparasitism was a key influencing factor and had a positive effect on the prevalence and/or abundance of the predominant tick, flea and mite species occurring on small mammals. Season, trapping location, host species and sex of the host species also had an influence on the prevalence and mean intensity of certain, but not all, ectoparasite species.

Background: Anaplasma phagocytophilum is a Gram-negative obligate intracellular bacterium that replicates in neutrophil granulocytes. It is transmitted by ticks of the Ixodes ricinus complex and causes febrile illness called granulocytic anaplasmosis primarily in humans, horses, dogs, sheep, cattle and goats. In comparison, clinically apparent disease has been described rarely in cats especially compared to dogs and horses. It is currently unknown whether cats are less susceptible to A. phagocytophilum or whether granulocytic anaplasmosis might be underdiagnosed in cats. Methods: To address this question, we examined clinical signs and laboratory findings in seven A. phagocytophilum infected cats from Germany and Switzerland. We then genetically characterized feline A. phagocytophilum strains and compared them to those from other hosts showing clinically apparent disease. For this purpose, ankA-based, groEL-based and multilocus sequence typing (MLST) were applied. Furthermore, the concordance between these typing methods was assessed. Results: Fever, lethargy and anorexia were the most common clinical signs in cats suffering from granulocytic anaplasmosis. The most frequent laboratory finding was thrombocytopenia. All three typing methods consistently indicated that the A. phagocytophilum strains found infecting cats are the same as those that cause disease in humans, dogs and horses. In general, the three typing methods applied exhibited high concordance. Conclusions: The genetic characterization of the feline A. phagocytophilum strains indicates that strain divergence is not the explanation for the fact that granulocytic anaplasmosis is much less frequently diagnosed in cats than in dogs and horses. Otherwise, it may be possible that cats are less susceptible to the same strains than dogs and horse are. However, due to the unspecific clinical signs, it should be considered that granulocytic anaplasmosis may be under-diagnosed in cats. [ABSTRACT FROM AUTHOR]

Background: The aims of this study were to evaluate the host-tick-pathogen interface of Babesia spp. and Anaplasma phagocytophilum in restored areas in both questing and host-attached Ixodes ricinus and Dermacentor reticulatus and their small mammalian hosts., Methods: Questing ticks were collected from 5 sites within the city of Leipzig, Germany, in 2009. Small mammals were trapped at 3 of the 5 sites during 2010 and 2011. DNA extracts of questing and host-attached I. ricinus and D. reticulatus and of several tissue types of small mammals (the majority bank voles and yellow-necked mice), were investigated by PCR followed by sequencing for the occurrence of DNA of Babesia spp. and by real-time PCR for A. phagocytophilum. A selected number of samples positive for A. phagocytophilum were further investigated for variants of the partial 16S rRNA gene. Co-infection with Rickettsia spp. in the questing ticks was additionally investigated., Results: 4.1% of questing I. ricinus ticks, but no D. reticulatus, were positive for Babesia sp. and 8.7% of I. ricinus for A. phagocytophilum. Sequencing revealed B. microti, B. capreoli and Babesia spp. EU1 in Leipzig and sequence analysis of the partial 16S RNA gene of A. phagocytophilum revealed variants either rarely reported in human cases or associated with cervid hosts. The statistical analysis revealed significantly less ticks infected with A. phagocytophilum in a city park in Leipzig as compared to the other sampling sites. A. phagocytophilum-DNA was detected in 2 bank voles, DNA of B. microti in 1 striped field-mouse and of Babesia sp. EU1 in the skin tissue of a mole. Co-infections were detected., Conclusion: Our results show the involvement of small mammals in the natural endemic cycles of tick-borne pathogens. A more thorough understanding of the interactions of ticks, pathogens and hosts is the essential basis for effective preventive control measures.

Background: Hyalomma marginatum and Hyalomma rufipes are two-host tick species, which are mainly distributed in southern Europe, Africa and middle-eastern Asia. They are well-known vectors of Crimean Congo hemorrhagic fever (CCHF) virus and other viruses as well as Rickettsia aeschlimannii. In recent years, these tick species have been found sporadically in Germany, but they do not belong to the autochthonous tick fauna in Germany., Methods: Ticks with unusual morphology were collected and sent from private persons or public health offices to involve institutions for morphological identification and further testing. All ticks identified as Hyalomma spp. were tested using molecular detection methods for CCHF virus, Rickettsia spp., Coxiella burnetii and Coxiella-like organisms, Babesia spp. and Theileria spp., Results: Thirty-five ticks with an unusual appearance or behaviour were reported to us during summer-autumn 2018. For 17 of them, the description or photos implied that they belong to the hard tick genus Hyalomma. The remaining 18 ticks were sent to us and were identified as adult Hyalomma marginatum (10 specimens) or adult Hyalomma rufipes (8 specimens). All ticks tested negative for CCHF virus, Coxiella burnetii, Coxiella-like organisms, Babesia spp. and Theileria spp. The screening for rickettsiae gave positive results in 9 specimens . The Rickettsia species in all cases was identified as R. aeschlimannii., Conclusions: These results show that exotic tick species imported into Germany were able to develop from the nymphal to the adult stage under appropriate weather conditions. Fifty percent of the ticks carried R. aeschlimannii, a human pathogen, while CCHF virus or other pathogens were not detected. Imported Hyalomma ticks may be the source of exotic diseases acquired in Germany.

Background: Tick-borne encephalitis (TBE) virus is transmitted to humans and animals through tick bites and is thought to circulate in very strictly defined natural environments called natural foci. The most common tick serving as a vector for the TBE virus in central Europe is Ixodes ricinus; it is rarely found in other tick species and in Dermacentor reticulatus it has, so far, only been reported in Poland., Methods: Between autumn 2016 and spring 2018 ticks were collected by the flagging method in a new TBE focus in the district of northern Saxony, Germany, outside the known risk areas as defined by the national Robert Koch Institute. Ticks were morphologically identified and tested in pools for the presence of TBE virus using a real-time RT-PCR. TBE virus from positive pools was isolated in A549 cells, and the E gene sequences were determined after conventional RT-PCR, followed by a phylogenetic comparison., Results: TBE virus was detected in 11 pools, 9 times in flagged adults D. reticulatus (n = 1534; MIR: 0.59%, CI: 0.29-11.3%) and only twice in I. ricinus nymphs (n = 349; MIR: 0.57%, CI: 0.02-2.2%). All other ticks, I. ricinus males (n = 33), females (n = 30) and larvae (n = 58), as well as 5 I. inopinatus (2 females, 3 males) and 14 Haemaphysalis concinna (3 females, 11 nymphs), tested negative for TBE virus. TBE virus was not detected in I. ricinus during the summer, when D. reticulatus is not active. Sequence comparison of the entire E gene of the isolated virus strains resembled each other with only 3 nucleotide differences. The most closely related viral sequences belong to TBE virus strains from Poland and Neustadt an der Waldnaab (county of Neustadt an der Waldnaab, Bavaria), approximately 200 km east and 200 km south-west of the new focus, respectively., Conclusions: TBE virus was found in northern Saxony, Germany, with similar MIRs in D. reticulatus and I. ricinus, indicating that D. reticulatus plays an equal role to I. ricinus in virus circulation when both tick species are sympatric.

Background: A spot-on formulation containing fluralaner (280 mg/ml) plus moxidectin (14 mg/ml) (Bravecto® Plus) has been developed to provide broad spectrum parasite protection for cats. The effectiveness and safety of this product against ticks and fleas was assessed in a randomized, controlled, 12-week study in client-owned cats in Germany and Spain., Methods: Eligible households containing at least one cat with at least two fleas and/or two ticks were allocated randomly in a 2:1 ratio to a single treatment with fluralaner plus moxidectin on Day 0, or three 4-weekly treatments with fipronil (Frontline®). Veterinary staff, masked to treatment, completed tick and flea counts on each cat at 14 ± 2 (2 weeks), 28 ± 2 (4 weeks), 56 ± 2 (8 weeks) and 84 ± 2 days (12 weeks) after the initial treatment., Results: In total, 707 cats (257 with ticks) from 332 households (236 with fleas) were included. Ixodes ricinus (78%) and Rhipicephalus sanguineus complex (18%) ticks were the most commonly identified. Tick and flea counts were lower in the fluralaner plus moxidectin group than in the fipronil group throughout the study and the efficacy of fluralaner plus moxidectin exceeded 97 and 98%, respectively. At 12 weeks, 94.1 and 93.3% of cats from the fluralaner plus moxidectin and 92.2 and 60.3% of cats from the fipronil group were free of ticks and fleas, respectively. Fluralaner plus moxidectin was non-inferior to fipronil (P < 0.0001) at all assessments and superior to fipronil at 2 and 8 weeks for the proportion of cats free of ticks (P < 0.0001). Fluralaner plus moxidectin was superior to fipronil for the proportion of both households and cats free of fleas (P < 0.0001). Both products were safe and well tolerated., Conclusions: A single application of fluralaner plus moxidectin spot-on was well tolerated by cats and highly effective for 12 weeks against ticks and fleas. Fluralaner plus moxidectin was non-inferior to fipronil for the proportion of ectoparasite-free and consistently superior to fipronil in controlling fleas.

Background: Tick-borne diseases are a major health risk for humans and dogs. In addition to collection and analysis of questing ticks, analysis of host-associated ticks for the presence of pathogens is a valuable method to gain insight into transmission patterns of tick-borne diseases., Methods: Ticks were collected from dogs living in the Berlin/Brandenburg area. The three tick species Ixodes ricinus, Ixodes hexagonus and Dermacentor reticulatus were examined for the presence of Babesia spp., Borrelia spp., Rickettsia spp. and Anaplasmataceae. Conventional PCR followed by sequencing was used for pathogen detection and characterization., Results: Babesia spp. were found in 2.5% and 3% of I. ricinus and I. hexagonus, respectively. Sequencing revealed the presence of Babesia microti, Babesia capreoli and Babesia venatorum. D. reticulatus were free of Babesia canis. Rickettsia spp. were detected in 61% of I. ricinus, 44% of I. hexagonus and 39% of D. reticulatus. Specifically detected were Rickettsia raoulti in D. reticulatus and I. hexagonus, Rickettsia helvetica in I. ricinus and I. hexagonus and Rickettsia monacensis in I. hexagonus. Anaplasma phagocytophilum and Candidatus Neoehrlichia mikurensis have been reported previously in I. ricinus (6.5% and 4.3%, respectively) and I. hexagonus (3.9% and 5.9%). Borrelia spp. were found in 11.6% of I. ricinus and 11.2% of I. hexagonus. Subsequent genospecies analysis revealed Borrelia afzelii, Borrelia garinii, Borrelia burgdorferi sensu stricto and Borrelia miyamotoi. Simultanous presence of more than one pathogen was found in 20% of I. ricinus and in 59% of I. hexagonus whereas the total frequency of any pathogen was 65% in I. ricinus, 59% in I. hexagonus and 64% in D. reticulatus. Ticks in which A. phagocytophilum was detected had a significantly increased risk of also containing Rickettsia. Ticks harbouring a pathogen had significantly higher scutal indices than ticks without presence of any pathogen., Conclusions: Frequencies of potential human or canine pathogens in ticks were considerable and DNA of all four groups of pathogens was detected. Differences in scutal indices might suggest that pathogens are frequently taken up by ticks when feeding on dogs in Berlin/Brandenburg.

Background: Borrelia burgdorferi sensu lato (sl), the causative agent of Lyme borreliosis, is transmitted by ticks of the genus Ixodes as vector. For identification of Borrelia infections in ticks a TaqMan™ minor groove binder (MGB) probe-based quantitative real time PCR (qPCR) was established targeting the 5S-23S intergenic spacer. Extension to a duplex qPCR included an Ixodes spp. positive control to verify successful DNA isolation. Besides qPCR, an ospAspecific conventional PCR for species-specific identification of B. spielmanii was established. Afterwards 1000 I. ricinus flagged in the city of Hanover, Germany, were investigated for B. burgdorferi sl infections followed by species identification. Furthermore, I. hexagonus ticks were investigated to proof applicability of the PCRs. Results: Quantitative real time PCR (qPCR) identifying B. burgdorferi sl in ticks was able to detect 1-10 copies per reaction. B. spielmanii ospA-specific conventional PCR was also highly specific and showed no cross reactions with the other tested Borrelia species. From 1000 hanoveranian ticks 24.3% were positive compared to only 7.4% positives by dark-field microscopy. Related to tick stage 1.7% larvae, 18.1% nymphs, and 34.6% adults were positive. The most frequent species was B. garinii, followed by B. afzelii, B. spielmanii, B. valaisiana and B. burgdorferi sensu stricto (ss). 70.6% of I. ricinus were mono-infected, whereas 28.0% and 1.4% were infected with two and three Borrelia species, respectively. From 232 I. hexagonus collected from hedgehogs in different sites of Germany, qPCR detected 5.7% to be infected with B. burgdorferi sl, which were identified as B. afzelii, B. garinii and B. spielmanii. Conclusions: The evaluated qPCR to detect B. burgdorferi sl in Ixodes spp. is highly specific and sensitive. As a duplex qPCR including detection of Ixodes spp. DNA it is the first DNA based technique incorporating a control for successful DNA isolation from the vector tick. Establishment of a B. spielmanii specific conventional PCR filled the gap in PCR identification of principal European Borrelia genospecies. Practical application showed that all European pathogenic Borrelia spp. were present in I. ricinus flagged in recreational areas of the city of Hanover and confirmed I. hexagonus as reservoir for pathogenic Borrelia spp. [ABSTRACT FROM AUTHOR]

Background: Candidatus Neoehrlichia mikurensis (CNM) has been described in the hard tick Ixodes ricinus and rodents as well as in some severe cases of human disease. The aims of this study were to identify DNA of CNM in small mammals, the ticks parasitizing them and questing ticks in areas with sympatric existence of Ixodes ricinus and Dermacentor reticulatus in Germany., Methods: Blood, transudate and organ samples (spleen, kidney, liver, skin) of 91 small mammals and host-attached ticks from altogether 50 small mammals as well as questing I. ricinus ticks (n=782) were screened with a real-time PCR for DNA of CNM., Results: 52.7% of the small mammals were positive for CNM-DNA. The majority of the infected animals were yellow-necked mice (Apodemus flavicollis) and bank voles (Myodes glareolus). Small mammals with tick infestation were more often infected with CNM than small mammals without ticks. Compared with the prevalence of ~25% in the questing I. ricinus ticks, twice the prevalence in the rodents provides evidence for their role as reservoir hosts for CNM., Conclusion: The high prevalence of this pathogen in the investigated areas in both rodents and ticks points towards the need for more specific investigation on its role as a human pathogen.

Background Georeferenced locations of ixodid ticks are required to depict the observed distribution of species. Further, they are used as input data for species distribution models also known as niche models. The latter were applied to describe current and future (projected) tick distributions. Beside model assumptions and selected climate parameters, the number of georeferenced tick locations available as a digital dataset is of fundamental importance for the reliability of such models. For Germany, however, no comprehensive dataset of ixodid tick species exists. The goal of this study was to put together all the available information on ixodid tick locations in Germany to produce such a digital dataset and to visualize it in a map. Findings A total of 2,044 georeferenced locations of ixodid ticks in Germany were compiled from two existing datasets (altogether 993 locations) and an extensive literature study (1051 locations). The resulting digital dataset comprises the following tick species: Ixodes ricinus (1,855 locations), Ixodes apronophorus (1), Ixodes frontalis (1), Ixodes hexagonus (1), Ixodes trianguliceps (4), Dermacentor marginatus (77), Dermacentor reticulatus (96), Haemaphysalis concinna (8) and Hyalomma marginatum (1). The data were used to draw a tick map for Germany, showing I. ricinus occurring in the whole federal territory, while D. marginatus has been restricted to the climatically favoured region of the Rhine valley. Clustered locations of D. reticulatus were also documented in the Rhine valley as well as in Berlin and its vicinity. Conclusions The introduced map depicts for the first time the available geographical coordinates of ixodid tick locations in Germany. The digital dataset used to draw the map is provided to the scientific community as a basis for further investigations such as species distribution modelling. [ABSTRACT FROM AUTHOR]

The article focuses on the occurrence of Babesia spp., Rickettsia spp. and Bartonella spp. in Ixodes ricinus in Bavarian public parks, Germany. In 2009, nine different public parks in five Bavarian cities were investigated. A total of 6,593 ticks were screened for the presence of DNA of Babesia spp. Questing ticks were collected monthly from the vegetation by flagging and stored in 70% ethanol.The overall prevalences were 0.4% for 2009 and 0.5 to 0.7% for 2010. differences in infection there was no statistical significance of developmental stages.

Background: Rodents are important in the life-cycle of ticks as hosts for immature developmental stages. Both rodents and ticks are of public health interest as they are reservoirs and vectors for different tick-borne pathogens (TBP). The aim of this study was to reassess the prevalence of TBP in previously studied areas of the city of Leipzig (Saxony, Germany)., Methods: In the years 2015-2017 rodents and ticks were collected in parks and forest areas in Saxony. DNA was extracted from the rodents, attached and questing ticks. Samples were screened for the presence of Anaplasma phagocytophilum, Babesia spp., Borrelia burgdorferi (s.l.), "Candidatus Neoehrlichia mikurensis" (CNM), Bartonella spp., Hepatozoon spp. and Rickettsia spp. using PCR methods. Rodent, attached nymph and questing tick (nymph and adult) samples were tested individually, while attached larvae were further processed in pools., Results: A total of 165 rodents (Apodemus agrarius, n = 1; A. flavicollis, n = 59; Arvicola terrestris, n = 1; Myodes glareolus, n = 104), 1256 attached ticks (Ixodes ricinus, n = 1164; Dermacentor reticulatus, n = 92) and 577 questing ticks (I. ricinus, n = 547; D. reticulatus, n = 30) were collected. The prevalence levels in rodents were 78.2% for Bartonella spp., 58.2% for CNM, 49.1% for B. burgdorferi (s.l.) 29.1% for Rickettsia spp. and 24.2% for Hepatozoon spp. The minimal infection rates (MIR) in attached larvae ticks were 39.8% for Rickettsia spp., 32.7% for Bartonella spp., 7.1% for CNM and 8.8% for B. burgdorferi (s.l.) and the prevalence rates in attached nymphs were 33.7% for Bartonella spp., 52.9% for Rickettsia spp., 13.5% for CNM and 11.3% for B. burgdorferi (s.l.) Both rodents and attached ticks were negative for Babesia spp. The prevalence in questing ticks was 18.2% for Rickettsia spp., 7.3% for CNM, 6.4% for B. burgdorferi (s.l.) and 1.4% for Babesia spp. All tested samples were Anaplasma-negative. Sequencing revealed the occurrence of 14 identified species., Conclusions: This research is the first evaluation of the prevalence for Hepatozoon spp. in rodents from Germany. In comparison to earlier studies, detected pathogens species remained the same; however, the prevalence for particular pathogens differed.

Background: Rickettsioses are caused by pathogenic species of the genus Rickettsia and play an important role as emerging diseases. The bacteria are transmitted to mammal hosts including humans by arthropod vectors. Since detection, especially in tick vectors, is usually based on PCR with genus-specific primers to include different occurring Rickettsia species, subsequent species identification is mainly achieved by Sanger sequencing. In the present study a real-time pyrosequencing approach was established with the objective to differentiate between species occurring in German Ixodes ticks, which are R. helvetica, R. monacensis, R. massiliae, and R. felis. Tick material from a quantitative real-time PCR (qPCR) based study on Rickettsia-infections in I. ricinus allowed direct comparison of both sequencing techniques, Sanger and real-time pyrosequencing., Methods: A sequence stretch of rickettsial citrate synthase (gltA) gene was identified to contain divergent single nucleotide polymorphism (SNP) sites suitable for Rickettsia species differentiation. Positive control plasmids inserting the respective target sequence of each Rickettsia species of interest were constructed for initial establishment of the real-time pyrosequencing approach using Qiagen's PSQ 96MA Pyrosequencing System operating in a 96-well format. The approach included an initial amplification reaction followed by the actual pyrosequencing, which is traceable by pyrograms in real-time. Afterwards, real-time pyrosequencing was applied to 263 Ixodes tick samples already detected Rickettsia-positive in previous qPCR experiments., Results: Establishment of real-time pyrosequencing using positive control plasmids resulted in accurate detection of all SNPs in all included Rickettsia species. The method was then applied to 263 Rickettsia-positive Ixodes ricinus samples, of which 153 (58.2%) could be identified for their species (151 R. helvetica and 2 R. monacensis) by previous custom Sanger sequencing. Real-time pyrosequencing identified all Sanger-determined ticks as well as 35 previously undifferentiated ticks resulting in a total number of 188 (71.5%) identified samples. Pyrosequencing sensitivity was found to be strongly dependent on gltA copy numbers in the reaction setup. Whereas less than 101 copies in the initial amplification reaction resulted in identification of 15.1% of the samples only, the percentage increased to 54.2% at 101-102 copies, to 95.6% at >102-103 copies and reached 100% samples identified for their Rickettsia species if more than 103 copies were present in the template., Conclusions: The established real-time pyrosequencing approach represents a reliable method for detection and differentiation of Rickettsia spp. present in I. ricinus diagnostic material and prevalence studies. Furthermore, the method proved to be faster, more cost-effective as well as more sensitive than custom Sanger sequencing with simultaneous high specificity.

Background: Spotted Fever Group (SFG) Rickettsiae can cause febrile diseases with or without rash in humans worldwide. In Germany only limited data are available about their medical significance. Serological screening tests for antibodies against rickettsiae usually only distinguish between SFG and Typhus Group (TG) Rickettsiae due to the strong cross reactivities within the groups. Seroprevalence rates in dogs, as possible sentinels for tick-borne diseases, could serve as an indicator for the distribution of different Rickettsia species., Methods: In this study, a micro-immunofluorescence assay (micro-IFA) was established for detection and differentiation of antibodies against five Rickettsia species in dogs (R. helvetica, R. raoultii, R. slovaca, R. monacensis and R. felis). Dogs that never left Germany (n = 605) previously investigated with an SFG-ELISA were included in this study and screened at a 1:128 dilution. Endpoint titres of fifty randomly selected seropositive samples of each of the five investigated regions in Germany were determined in order to allow a differentiation of the causative Rickettsia species. Sensitivity and specificity of the micro-IFA were compared with ELISA results of the previous study., Results: A total of 93.9% of the dogs were positive for antibodies of the SFG Rickettsiae at the screening titer of 1:128. Differentiation of SFG Rickettsiae with the micro-IFA was possible in 70.4%, but in 29.6% of the cases the detected antibodies were not differentiable. Considering a clear differentiation by a twofold titre difference between observed reactions, the seroprevalence rates were 66.0% for R. helvetica, 2.8% for R. raoultii, 1.6% for R. slovaca, but no serological reaction could be clearly attributed to R. monacensis or R. felis. No statistically significant regional differences were found for R. helvetica, R. slovaca and R. raoultii comparing the five regions of Germany. Comparison of micro-IFA with ELISA revealed a sensitivity of 82.0% and a specificity of 83.8% for the Rickettsia SFG ELISA., Conclusions: The micro-IFA is a useful serological tool to differentiate antibodies against different Rickettsia species in dogs. Seroprevalence rates in dogs correspond to the prevalence rates and distribution of Rickettsia-carrying tick species.

Background: Fluralaner, a new molecular entity of the isoxazoline class, has potent insecticidal and acaricidal activity and can be safely administered orally to dogs., Methods: A randomized, investigator-blinded, multi-centered field study compared the flea- and tick-control efficacy for dogs over a 12-week period with either a single oral dose of Bravecto (fluralaner) formulated as a chewable tablet or with three sequential topical Frontline (fipronil) treatments. Individual dogs were the experimental unit for ticks and households were the experimental unit for fleas. A total of 108 tick-infested dogs were treated with Bravecto (fluralaner) and 54 tick-infested dogs were treated with Frontline (fipronil). Dogs in 115 flea-infested households received Bravecto (fluralaner) and dogs in 61 flea-infested households received Frontline (fipronil). Flea and tick counts were conducted on all dogs at weeks 2, 4, 8, and 12 following initial treatment and efficacy was calculated as the mean percent reduction in tick or flea count at each time point compared with the mean pretreatment initiation count for each treatment group. Additionally, the percentages of tick-free and flea-free households were determined., Results: At weeks 2, 4, 8, and 12, Bravecto (fluralaner) flea-control efficacy in treated households was 99.2%, 99.8%, 99.8%, and 99.9% respectively, while Frontline (fipronil) efficacy was 94.1%, 93.0%, 96.0%, and 97.3%, respectively. Bravecto (fluralaner) tick-control efficacy on treated dogs at weeks 2, 4, 8, and 12 was 99.9%, 99.9%, 99.7%, and 100%, respectively, and Frontline (fipronil) tick efficacy was 97.6%, 93.8%, 100%, and 100%, respectively. Of dogs showing clinical flea allergy dermatitis (FAD) signs at the study start, 85.7% in the Bravecto™ (fluralaner)-treated group and 55.6% in the Frontline (fipronil)-treated group were evaluated at each time point as showing no clinical signs of FAD until study completion., Conclusions: Bravecto (fluralaner) administered once orally to dogs in a chewable tablet was highly effective for 12 weeks against fleas and ticks on privately-owned dogs and was significantly non-inferior (ticks) and superior (fleas) in comparison with topical Frontline (fipronil) administered 3 times sequentially.

Background: Tularemia re-emerged in Germany starting in 2004 (with 39 human cases from 2004 to 2007) after over 40 years of only sporadic human infections. The reasons for this rise in case numbers are unknown as is the possible reservoir of the etiologic agent Francisella (F.) tularensis. No systematic study on the reservoir situation of F. tularensis has been published for Germany so far., Methods: We investigated three areas six to ten months after the initial tularemia outbreaks for the presence of F. tularensis among small mammals, ticks/fleas and water. The investigations consisted of animal live-trapping, serologic testing, screening by real-time-PCR and cultivation., Results: A total of 386 small mammals were trapped. F. tularensis was detected in five different rodent species with carrier rates of 2.04, 6.94 and 10.87% per trapping area. None of the ticks or fleas (n = 432) tested positive for F. tularensis. We were able to demonstrate F. tularensis-specific DNA in one of 28 water samples taken in one of the outbreak areas., Conclusion: The findings of our study stress the need for long-term surveillance of natural foci in order to get a better understanding of the reasons for the temporal and spatial patterns of tularemia in Germany.

Background: Lyme borreliosis is the most prevalent vector-borne disease in Europe, and numbers might increase due to climate change. However, borreliosis is not notifiable in North Rhine-Westphalia (NRW), Germany. Hence, little is known about the current human seroprevalence in NRW. However, the proportion of Borrelia burgdorferi sensu lato-infected ticks has increased in a NRW nature reserve. The literature suggests increasing age and male sex as risk factors for seropositivity, whereas the influence of socioeconomic status is controversial. Thus, we aimed to determine regional seropositivity for Borrelia burgdorferi sensu lato (B. burgdorferi s.l.) and its risk factors in the Rhineland Study population in Bonn, NRW, and to compare it with previous surveys to evaluate potential effects of climate change. Methods: We assessed seropositivity in 2865 Rhineland Study participants by determining immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies for B. burgdorferi s.l. using a two-step algorithm combining enzyme-linked immunosorbent assay tests and line immunoblots. We calculated the odds of being classified as IgG or IgM positive as a function of age, sex, and educational level using binomial logistic regression models. We applied varying seropositivity classifications and weights considering age, sex and education to compensate for differences between the sample and regional population characteristics. Results: IgG antibodies for B. burgdorferi s.l. were present in 2.4% and IgM antibodies in 0.6% of the participants (weighted: 2.2% [IgG], 0.6% [IgM]). The likelihood of IgG seropositivity increased by 3.0% (95% confidence interval [CI] 1.5–5.2%) per 1 year increase in age. Men had 1.65 times the odds for IgG seropositivity as women (95% CI 1.01–2.73), and highly educated participants had 1.83 times the odds (95% CI 1.10–3.14) as participants with an intermediate level of education. We found no statistically significant link between age, sex, or education and IgM seropositivity. Our weighted and age-standardized IgG seroprevalence was comparable to the preceding serosurvey German Health Interview and Examination Survey for Adults (DEGS) for NRW. Conclusions: We confirmed that increasing age and male sex are associated with increased odds for IgG seropositivity and provide evidence for increased seropositivity in the highly educated group. B. burgdorferi s.l. seropositivity remained constant over the past decade in this regional German population. [ABSTRACT FROM AUTHOR]

The intracellular bacteria Anaplasma spp. and Coxiella burnetii and the tick-borne encephalitis virus (TBEV) are tick-transmitted pathogens circulating in the southern German sheep population. Knowledge of interaction among Anaplasma spp., C. burnetii and TBEV in sheep is lacking, but together they might promote and reinforce disease progression. The current study aimed to identify co-exposure of sheep to Anaplasma spp., C. burnetii and TBEV. For this purpose, 1,406 serum samples from 36 sheep flocks located in both southern German federal states, Baden-Wuerttemberg and Bavaria, were analysed by ELISAs to determine the antibody levels of the three pathogens. Inconclusive and positive results from the TBEV ELISA were additionally confirmed by a serum neutralisation assay. The proportion of sheep with antibodies against Anaplasma spp. (47.2%), C. burnetii (3.7%) and TBEV (4.7%) differed significantly. Significantly more flocks with Anaplasma spp. seropositive sheep (91.7%) were detected than flocks with antibodies against TBEV (58.3%) and C. burnetii (41.7%), but there was no significant difference between the number of flocks which contained TBEV and C. burnetii seropositive sheep. Seropositivity against at least two pathogens was detected in 4.7% of sheep from 20 flocks. Most co-exposed sheep had antibodies against Anaplasma spp./TBEV (n = 36), followed by Anaplasma spp./C. burnetii (n = 27) and Anaplasma spp./C. burnetii/TBEV (n = 2). Only one sheep showed an immune response against C. burnetii and TBEV. Flocks with sheep being positive against more than one pathogen were widely distributed throughout southern Germany. The descriptive analysis revealed no association between the antibody response of the three pathogens at animal level. Taking the flocks as a cluster variable into account, the exposure to TBEV reduced the probability of identifying C. burnetii antibodies in sheep significantly (odds ratio 0.46; 95% confidence interval 0.24–0.85), but the reason for this is unknown. The presence of Anaplasma spp. antibodies did not influence the detection of antibodies against C. burnetii and TBEV. Studies under controlled conditions are necessary to evaluate any possible adverse impact of co-exposure to tick-borne pathogens on sheep health. This can help to clarify rare disease patterns. Research in this field may also support the One Health approach due to the zoonotic potential of Anaplasma spp., C. burnetii and TBEV. [ABSTRACT FROM AUTHOR]

Background: Anaplasma phagocytophilum is a Gram-negative obligate intracellular bacterium that replicates in neutrophil granulocytes. It is transmitted by ticks of the Ixodes ricinus complex and causes febrile illness in humans and animals. We used multilocus sequence typing (MLST) and ankA gene-based typing to study the molecular epidemiology of the A. phagocytophilum strains circulating in a German cattle herd over one pasture season. The aim was to investigate whether co-infection with two distinct variants, reinfection with the same and/or superinfection by a different strain occurred during one pasture season. Eight genetic loci were sequenced in 47 PCR-positive samples from 15 animals. Results: Five different sequence types (ST) and four ankA alleles were detected in the cattle herd. Three different ST caused clinically overt tick-borne fever in primary infected animals. The concordance between ST and ankA allele was 100%. Therefore, the housekeeping genes used for MLST and the highly variable ankA gene were concatenated to increase resolution. Co-infection could be proven because samples of chronologically close collection dates were included. Co-infecting A. phagocytophilum strains differed by 14 to 18 single nucleotide polymorphisms (SNPs). Most superinfecting variants varied by 14 SNPs from the previous strain and appeared in median after a free interval of 31 days. Thus, it is unlikely that superinfecting strains arose by in-animal evolution. Immunity against re- or superinfection was assumed because the cattle developed clinical signs only during primary infection. Conclusions: The tick-pathogen-vertebrate host interaction is probably much more complex than previously thought taking into account the frequently occurring events of co-infection, reinfection and superinfection. This complex situation could not be easily simulated in an experimental infection and underlines the value of field studies. [ABSTRACT FROM AUTHOR]

Background: Tick-borne encephalitis (TBE) virus can cause severe symptoms in humans. The incidence of this vector-borne pathogen in humans is characterised by spatial and temporal heterogeneity. To explain the variation in reported human TBE cases per county in southern Germany, we designed a time-lagged, spatially-explicit model that incorporates ecological, environmental, and climatic factors. Results: We fitted a logistic regression model to the annual counts of reported human TBE cases in each of 140 counties over an eight year period. The model controlled for spatial autocorrelation and unexplained temporal variation. The occurrence of human TBE was found to be positively correlated with the proportions of broad-leafed, mixed and coniferous forest cover. An index of forest fragmentation was negatively correlated with TBE incidence, suggesting that infection risk is higher in fragmented landscapes. The results contradict previous evidence regarding the relevance of a specific spring-time temperature regime for TBE epidemiology. Hunting bag data of roe deer (Capreolus capreolus) in the previous year was positively correlated with human TBE incidence, and hunting bag density of red fox (Vulpes vulpes) and red deer (Cervus elaphus) in the previous year were negatively correlated with human TBE incidence. Conclusions: Our approach suggests that a combination of landscape and climatic variables as well as host-species dynamics influence TBE infection risk in humans. The model was unable to explain some of the temporal variation, specifically the high counts in 2005 and 2006. Factors such as the exposure of humans to infected ticks and forest rodent population dynamics, for which we have no data, are likely to be explanatory factors. Such information is required to identify the determinants of TBE more reliably. Having records of TBE infection sites at a finer scale would also be necessary. [ABSTRACT FROM AUTHOR]

Background: Long-lasting symptoms with a possible relation to psychosomatic comorbidity have been described following COVID-19. However, data is sparse in general practice. The trial's objective was to investigate the time-dependent frequency of disease symptoms and relation to psychosomatic comorbidity and daily life impairment (DLI). Methods: Comparative cohort study of patients reporting a previous SARS-CoV-2 infection and uninfected controls in general practice. Participants were recruited in 14 general practices in the greater Munich area. Data collection was questionnaire based with a 12 months follow-up. Descriptive statistics, multivariable regression and bivariate correlations were used for analysis. Results: A total of n = 204 cases infected up to 42 months ago (n = 141 Omicron, n = 63 earlier variants), and n = 119 controls were included. Disease symptoms were substantially more prevalent in cases (55–79% vs. 43% within one year of infection). This difference also appeared in the multivariable analysis adjusting for socio-demographics and psychosomatic comorbidity with odds ratios (OR) of 4.15 (p < 0.001) and 3.51 (p = 0.054) for the cohorts with Omicron or earlier variants infection (vs. controls), respectively. It was persistent with earlier variants (OR 1.00 per month, p = 0.903), while a decreasing trend was observed for Omicron (OR 0.89 per month, p < 0.001). DLI was especially correlated with fatigue (r = 0.628). Conclusion: DLI, psychosomatic comorbidity and independently increased disease symptoms require holistic treatment of the patient in general practice according to the bio-psycho-social model. A key role in restoring the daily life capability may be attributed to the symptom fatigue. [ABSTRACT FROM AUTHOR]

Background: Lyme borreliosis caused by spirochetes of the Borrelia burgdorferi (sensu lato) complex is still the most common tick-borne disease in Europe, posing a considerable threat to public health. The predominant vector in Europe is the widespread hard tick Ixodes ricinus, which also transmits the relapsing fever spirochete B. miyamotoi as well as pathogenic Rickettsiales (Anaplasma phagocytophilum, Rickettsia spp.). To assess the public health risk, a long-term monitoring of tick infection rates with the named pathogens is indispensable., Methods: The present study is the first German 10-year follow-up monitoring of tick infections with Borrelia spp. and co-infections with Rickettsiales. Furthermore, a specific Reverse Line Blot (RLB) protocol for detection of B. miyamotoi and simultaneous differentiation of B. burgdorferi (s.l.) geno-species was established., Results: Overall, 24.0% (505/2100) of ticks collected in the city of Hanover were infected with Borrelia. In detail, 35.4% (203/573) of adult ticks [38.5% females (111/288) and 32.3% males (92/285)] and 19.8% nymphs (302/1527) were infected, representing consistent infection rates over the 10-year monitoring period. Geno-species differentiation using RLB determined B. miyamotoi in 8.9% (45/505) of positive ticks. Furthermore, a significant decrease in B. afzelii and B. spielmanii infection rates from 2010 to 2015 was observed. Co-infections with Rickettsia spp. and A. phagocytophilum increased between 2010 and 2015 (7.3 vs 10.9% and 0.3 vs 1.1%, respectively)., Conclusions: Long-term monitoring is an essential part of public health risk assessment to capture data on pathogen occurrence over time. Such data will reveal shifts in pathogen geno-species distribution and help to answer the question whether or not climate change influences tick-borne pathogens.

Background: European wildcats (Felis silvestris) are widely distributed in Europe and a strictly protected species in Germany. Lately, anthropogenic protective efforts lead to increasing numbers of wildcats in southwestern Germany. Moreover, in recent years the numbers of domestic cats are increasing. Thus, the contact between domestic and wildcats may lead to the spread of zoonotic pathogens in both animal species. As data on vector-borne pathogens (VBPs) in wildcats from Germany are limited to date, the objective of this study was to investigate the presence and current distribution of VBPs in wildcats from southwestern Germany., Methods: Skin and spleen samples from 117 European wildcats, originating from a regional carcass-monitoring program in southwestern Germany, were examined by real-time and conventional polymerase chain reaction (PCR) for the presence of Anaplasma phagocytophilum, Neoehrlichia mikurensis, Rickettsia spp., Bartonella spp., and Piroplasmida., Results: In total, 6.8% (n = 8) of the wildcats were Rickettsia-positive, specified as R. helvetica. Three wildcats were positive for A. phagocytophilum (2.6%), one for Bartonella spp., namely B. taylorii (0.8%), and 84 for Cytauxzoon spp. (71.8%). Out of these 84 samples, 23 were further sequenced revealing very high identity levels (99.84-100%) to C. europaeus, which is considered to be pathogenic for domestic cats. All wildcats were negative for the presence of N. mikurensis DNA., Conclusions: European wildcats in southwestern Germany are hosting several VBPs. With the exception of Cytauxzoon spp., low prevalence rates of most examined pathogens suggest that wildcats are primarily incidental hosts for sylvatic pathogens associated with rodents, in contrast to domestic cats. However, the high prevalence of the cat-associated pathogen C. europaeus suggests that wildcats in southwestern Germany may serve as reservoirs for this pathogen., (© 2024. The Author(s).)

Background: Mosquito-specific viruses (MSVs) comprise a variety of different virus families, some of which are known to interfere with infections of medically important arboviruses. Viruses belonging to the family Mesoniviridae or taxon Negevirus harbor several insect-specific viruses, including MSVs, which are known for their wide geographical distribution and extensive host ranges. Although these viruses are regularly identified in mosquitoes all over the world, their presence in mosquitoes in Germany had not yet been reported. Methods: A mix of three MSVs (Yichang virus [Mesoniviridae] and two negeviruses [Daeseongdong virus and Dezidougou virus]) in a sample that contained a pool of Coquillettidia richiardii mosquitoes collected in Germany was used to investigate the interaction of these viruses with different arboviruses in Culex-derived cells. In addition, small RNA sequencing and analysis of different mosquito-derived cells infected with this MSV mix were performed. Results: A strain of Yichang virus (Mesoniviridae) and two negeviruses (Daeseongdong virus and Dezidougou virus) were identified in the Cq. richiardii mosquitoes sampled in Germany, expanding current knowledge of their circulation in central Europe. Infection of mosquito-derived cells with these three viruses revealed that they are targeted by the small interfering RNA (siRNA) pathway. In Culex-derived cells, co-infection by these three viruses had varying effects on the representative arboviruses from different virus families (Togaviridae: Semliki forest virus [SFV]; Bunyavirales: Bunyamwera orthobunyavirus [BUNV]; or Flaviviridae: Usutu virus [USUV]). Specifically, persistent MSV co-infection inhibited BUNV infection, as well as USUV infection (but the latter only at specific time points). However, the impact on SFV infection was only noticeable at low multiplicity of infection (MOI 0.1) and at specific time points in combination with the infection status. Conclusions: Taken together, these results are important findings that will lead to a better understanding of the complex interactions of MSVs, mosquitoes and arboviruses. [ABSTRACT FROM AUTHOR]

Background: Serological surveillance, based on the measurement of the presence of specific antibodies in a given population, can be used in addition to traditional and routine disease surveillance methods. The added value of this has been largely documented for vaccine-preventable diseases, but to a lesser extent for vector-borne diseases. This study aimed to evaluate the utility of seroprevalence data as additional source of information on the epidemiology of Lyme borreliosis in Belgium.Methods: In total, 3215 residual blood samples collected in 2013-2015 were analysed with Liaison® Borrelia IgG kit (DiaSorin S.p.A, Saluggia, Italy). Positive and equivocal results were further examined with immunoblotting (recomLine Borrelia IgG kit, Mikrogen, Neuried, Germany). Crude prevalence estimates of equivocal and seropositive results were calculated and further adjusted accounting for clustered sampling and standardized for age, sex and population per province, according to the Belgian population structure in 2014. The effect of age, sex and region on seropositivity was assessed using log-binomial regression.Results: The overall weighted national seroprevalence for Borrelia burgdorferi sensu lato, adjusted for clustered sampling, age, sex and province was 1.06% (95%CI 0.67-1.67). Although not statistically significant, the highest prevalences were observed in men and in those younger than 15 years or older than 59 years of age. At provincial level, the seroprevalence estimates do not follow the geographical distribution of tick bites and diagnoses of Lyme borreliosis as detected through other surveillance systems.Conclusions: Although the use of residual samples for seroprevalence estimates has several advantages, it seems to be a limited tool for serological surveillance of Lyme borreliosis in Belgium, other than follow-up of trends if repeated over time. A population-based sampling strategy might provide a more representative nationwide sample, but would be very time intensive and expensive. Seroprevalence studies within risk groups or risk areas in Belgium could provide a useful alternative approach to complement routine surveillance data of Lyme borreliosis. [ABSTRACT FROM AUTHOR]

Background: Blood-feeding arthropods can transmit parasitic, bacterial, or viral pathogens to domestic animals and wildlife. Vector-borne infections are gaining significance because of increasing travel and import of pets from abroad as well as the changing climate in Europe. The main objective of this study was to assess the percentage of cats with positive test results for selected vector-borne pathogens in Germany and explore any possible association of such results with time spent abroad., Methods: This retrospective study included test results from cats included in the "Feline Travel Profile" established by the LABOKLIN laboratory at the request of veterinarians in Germany between April 2012 and March 2020. This diagnostic panel includes the direct detection of Hepatozoon spp. and Dirofilaria spp. via PCR as well as indirect detection assays (IFAT) for Ehrlichia spp. and Leishmania spp. The panel was expanded to include an IFAT for Rickettsia spp. from July 2015 onwards., Results: A total of 624 cats were tested using the "Feline Travel Profile." Serum for indirect detection assays was available for all 624 cats; EDTA samples for direct detection methods were available from 618 cats. Positive test results were as follows: Ehrlichia spp. IFAT 73 out of 624 (12%), Leishmania spp. IFAT 22 out of 624 (4%), Hepatozoon spp. PCR 53 out of 618 (9%), Dirofilaria spp. PCR 1 out of 618 cats (0.2%), and Rickettsia spp. IFAT 52 out of 467 cats (11%) tested from July 2015 onwards. Three cats had positive test results for more than one pathogen before 2015. After testing for Rickettsia spp. was included in 2015, 19 cats had positive test results for more than one pathogen (Rickettsia spp. were involved in 14 out of these 19 cats)., Conclusions: At least one pathogen could be detected in 175 out of 624 cats (28%) via indirect and/or direct detection methods. Four percent had positive test results for more than one pathogen. These data emphasize the importance of considering the above-mentioned vector-borne infections as potential differential diagnoses in clinically symptomatic cats.

Background: Tick-borne encephalitis (TBE), a viral infectious disease affecting the central nervous system, potentially resulting in prolonged neurological symptoms and other long-term sequelae. Case identification can be challenging as TBE can be associated with non-specific symptoms, and even in cases consistent with typical TBE symptoms, the rate of laboratory testing to confirm cases is unknown. This study assessed real-world TBE laboratory testing rates across Germany. Methods: In this retrospective cross-sectional study, physicians provided data on TBE decision-making, laboratory testing (serological), and diagnostics behavior via in-depth qualitative interviews (N = 12) or a web-based quantitative survey of their patient medical records (N = 166). Hospital-based physicians who specialized in infectious disease, intensive care unit, emergency room, neurology, or pediatrics with experience managing and ordering testing for patients with meningitis, encephalitis, or non-specific central nervous system symptoms in the past 12 months were included. Data were summarized via descriptive statistics. TBE testing and positivity rates were assessed for the aggregate sample of 1400 patient charts and reported by presenting symptoms, region, and tick bite exposure. Results: TBE testing rates ranged from 54.0% (non-specific neurological symptoms only) to 65.6% (encephalitis symptoms only); the percentage of TBE positive results ranged from 5.3% (non-specific neurological symptoms only) to 36.9% (meningitis symptoms only). TBE testing rates were higher among those with a tick bite history and/or who presented with headache, high fever, or flu-like symptoms. Conclusions: The findings of this study suggest that patients with typical TBE symptoms are likely under-tested, thus likely leading to under-diagnosis in Germany. To ensure appropriate case identification, TBE testing should be consistently integrated into routine practice for all patients who present with relevant symptoms or exposure to common risk factors. [ABSTRACT FROM AUTHOR]

Background: TBE is an important tick-borne viral zoonosis in Europe and some parts of Asia. Humans can become infected by tick bite and in some cases also by consumption of nonpasteurized raw milk and raw milk products from ruminants. Serological investigations of milking flocks can help to assess the risk of TBEV infection for humans. 735 blood samples from 50 goat flocks from four federal states of Germany were tested by TBEV-VNT to assess a potential risk for TBEV infection. There are some gaps in the knowledge about immunity in animals, for example with regard to the longevity of TBEV immunity. Two goats and two sheep were immunized and TBEV antibody titers could be detected for up to 7 years. Furthermore, nothing is known about a possible long-lasting immunological memory that could quickly be reactivated by an additional contact to TBEV. Seven years after the first immunization two goats and two sheep as well as two naïve goats and two sheep were boostered and TBEV antibody titers followed., Results: Only one sample in each of the three states was TBEV-antibody positive (VNT), albeit with low titers. However, in Baden-Württemberg seven samples were positive, among them four goats of the same flock. TBEV-antibody positive titers were detected in goats for up to 6 years and 10 months, in sheep for up to 4 years and 7 months. Seven years after immunization a clear immunological recall occurred in response to administration of one dose of vaccine in two goats and two sheep., Conclusion: It can be concluded that in the tested flocks the risk of an alimentary TBEV infection was low. However, in one single flock a considerably higher risk must be assumed. Antibody titers in goats and sheep can last very long after contact to TBEV, albeit at a low level. This should be taken into consideration in cases where the risk of an alimentary infection is assessed in a flock by serological investigations. The immunological recall gives rise to the suspicion that the immunological memory after a first contact to TBEV lasts for many years, probably lifelong.

Background: Rodents are important reservoirs for zoonotic vector-borne agents. Thus, the distribution of rodents and their vicinity to humans and companion animals may have an important impact on human and animal health. However, the reservoir potential of some rodent genera, e.g. Microtus, has not yet been precisely examined concerning tick-borne pathogens in Central Europe. Therefore, we examined small mammals from Germany and the Czech Republic for the following vector-borne pathogens: Babesia spp., Bartonella spp., Anaplasma phagocytophilum, "Candidatus Neoehrlichia mikurensis" (CNM) and Coxiella burnetii. Spleen DNA from 321 small mammals belonging to four genera, Myodes (n = 78), Apodemus (n = 56), Microtus (n = 149), Sorex (n = 38), collected during 2014 in Germany and the Czech Republic were available for this study. DNA samples were examined for the presence of Babesia and Bartonella DNA by conventional PCR targeting the 18S rRNA gene and the 16S–23S rRNA intergenic spacer region, respectively. For the detection of CNM, A. phagocytophilum and C. burnetii real-time PCR assays were performed. Results: Bartonella spp. DNA was detected in 216 specimens (67.3%) with 102/174 (58.6%) positive in Germany and 114/147 (77.6%) in the Czech Republic. The prevalence in each genus was 44.9% for Myodes, 63.2% for Sorex, 77.2% for Microtus and 75% for Apodemus. Four Bartonella species, i.e. Bartonella sp. N40, B. grahamii, B. taylorii and B. doshiae, as well as uncultured bartonellae, were detected. The Bartonella species diversity was higher in rodents than in shrews. In total, 27/321 (8.4%) small mammals were positive for CNM and 3/321 (0.9%) for A. phagocytophilum (S. coronatus and M. glareolus). All samples were negative for Babesia spp. and Coxiella spp. Conclusions: While the detected high prevalence for Bartonella in Apodemus and Myodes spp. is confirmatory with previous findings, the prevalence in Microtus spp. was unexpectedly high. This indicates that individuals belonging to this genus may be regarded as potential reservoirs. Interestingly, only Sorex spp. and M. glareolus were positive for A. phagocytophilum in the present study, suggesting a possible importance of the latter for the maintenance of certain A. phagocytophilum strains in nature. [ABSTRACT FROM AUTHOR]

Background: Recumbent cows are a diagnostic challenge because of a wide range of differential diagnoses, which include trauma, neurological and metabolic disorders, malnutrition and mineral deficiencies. This case report describes recumbent suckler cows that presented as a herd problem. In addition to weakness due to inanition, Cu and Se deficiencies were considered as possible aetiologies of the recumbency. Furthermore, Trypanosoma (T.) theileri, a blood parasite of unknown importance in Germany, was detected in the blood of some cows. Case presentation: Three recumbent cows were referred to the Clinic for Ruminants and Swine, Faculty of Veterinary Medicine of the University of Leipzig. They were unable to rise and had low body condition scores and rough hair coats. Haematological and serum biochemical analyses showed neutrophilia, electrolyte imbalances, increased activities of muscle and liver enzymes and decreased concentrations of trace elements, especially Copper (Cu) and Selenium (Se). T. theileri was detected in a routine blood smear from one cow. The cows did not respond to an intensive care protocol, which included intravenous fluids and electrolytes, mineral substitution, non-steroidal anti-inflammatories and antibiotics, and were therefore euthanized or died. Postmortem examination showed cachexia, subcutaneous and scleral oedema and muscular dystrophy, especially in the hind limbs. Follow-up examination of the herd of origin produced similar findings including the detection of T. theileri in a large proportion of the herd. Ration analysis revealed considerable undersupply of several nutrients. Conclusions: Based on all findings, an aetiological diagnosis of trace mineral and nutrient deficiency with possible involvement of T. theileri was made. [ABSTRACT FROM AUTHOR]

Background: Bats are hosts for a variety of microorganisms, however, little is known about the presence of Chlamydiales and hemotropic mycoplasmas. This study investigated 475 captive and free-living bats from Switzerland, Germany, and Costa Rica for Chlamydiales and hemotropic mycoplasmas by PCR to determine the prevalence and phylogeny of these organisms. Results: Screening for Chlamydiales resulted in a total prevalence of 31.4%. Positive samples originated from captive and free-living bats from all three countries. Sequencing of 15 samples allowed the detection of two phylogenetically distinct groups. These groups share sequence identities to Chlamydiaceae, and to Chlamydia-like organisms including Rhabdochlamydiaceae and unclassified Chlamydiales from environmental samples, respectively. PCR analysis for the presence of hemotropic mycoplasmas resulted in a total prevalence of 0.7%, comprising free-living bats from Germany and Costa Rica. Phylogenetic analysis revealed three sequences related to other unidentified mycoplasmas found in vampire bats and Chilean bats. Conclusions: Bats can harbor Chlamydiales and hemotropic mycoplasmas and the newly described sequences in this study indicate that the diversity of these bacteria in bats is much larger than previously thought. Both, Chlamydiales and hemotropic mycoplasmas are not restricted to certain bat species or countries and captive and free-living bats can be colonized. In conclusion, bats represent another potential host or vector for novel, previously unidentified, Chlamydiales and hemotropic mycoplasmas. [ABSTRACT FROM AUTHOR]

Background: During the last decades, population densities of Ixodes ricinus and prevalences of Borrelia burgdorferi s.l. have increased in different regions in Europe. In the present study, we determined tick abundance and the prevalence of different Borrelia genospecies in ticks from three sites in the Siebengebirge, Germany, which were already examined in the years 1987, 1989, 2001 and 2003. Data from all investigations were compared., Methods: In 2007 and 2008, host-seeking I. ricinus were collected by monthly blanket dragging at three distinct vegetation sites in the Siebengebirge, a nature reserve and a well visited local recreation area near Bonn, Germany. In both years, 702 ticks were tested for B. burgdorferi s.l. DNA by nested PCR, and 249 tick samples positive for Borrelia were further genotyped by reverse line blotting., Results: A total of 1046 and 1591 I. ricinus were collected in 2007 and 2008, respectively. In comparison to previous studies at these sites, the densities at all sites increased from 1987/89 and/or from 2003 until 2008. Tick densities and Borrelia prevalences in 2007 and 2008, respectively, were not correlated for all sites and both years. Overall, Borrelia prevalence of all ticks decreased significantly from 2007 (19.5%) to 2008 (16.5%), thus reaching the same level as in 2001 two times higher than in 1987/89 (7.6%). Since 2001, single infections with a Borrelia genospecies predominated in all collections, but the number of multiple infections increased, and in 2007, for the first time, triple Borrelia infections occurred. Prevalences of Borrelia genospecies differed considerably between the three sites, but B. garinii or B. afzelii were always the most dominant genospecies. B. lusitaniae was detected for the first time in the Siebengebirge, also in co-infections with B. garinii or B. valaisiana., Conclusions: Over the last two centuries tick densities have changed in the Siebengebirge at sites that remained unchanged by human activity since they belong to a nature reserve. Abiotic and biotic conditions most likely favored the host-seeking activity of I. ricinus and the increase of multiple Borrelia infections in ticks. These changes have led to a potential higher risk of humans and animals to be infected with Lyme borreliosis.

Background: Only limited information is available about the occurrence of ticks and tick-borne pathogens in public parks, which are areas strongly influenced by human beings. For this reason, Ixodes ricinus were collected in public parks of different Bavarian cities in a 2-year survey (2009 and 2010) and screened for DNA of Babesia spp., Rickettsia spp. and Bartonella spp. by PCR. Species identification was performed by sequence analysis and alignment with existing sequences in GenBank. Additionally, coinfections with Anaplasma phagocytophilum were investigated., Results: The following prevalences were detected: Babesia spp.: 0.4% (n = 17, including one pool of two larvae) in 2009 and 0.5 to 0.7% (n = 11, including one pool of five larvae) in 2010; Rickettsia spp.: 6.4 to 7.7% (n = 285, including 16 pools of 76 larvae) in 2009. DNA of Bartonella spp. in I. ricinus in Bavarian public parks could not be identified. Sequence analysis revealed the following species: Babesia sp. EU1 (n = 25), B. divergens (n = 1), B. divergens/capreoli (n = 1), B. gibsoni-like (n = 1), R. helvetica (n = 272), R. monacensis IrR/Munich (n = 12) and unspecified R. monacensis (n = 1). The majority of coinfections were R. helvetica with A. phagocytophilum (n = 27), but coinfections between Babesia spp. and A. phagocytophilum, or Babesia spp. and R. helvetica were also detected., Conclusions: I. ricinus ticks in urban areas of Germany harbor several tick-borne pathogens and coinfections were also observed. Public parks are of particularly great interest regarding the epidemiology of tick-borne pathogens, because of differences in both the prevalence of pathogens in ticks as well as a varying species arrangement when compared to woodland areas. The record of DNA of a Babesia gibsoni-like pathogen detected in I. ricinus suggests that I. ricinus may harbor and transmit more Babesia spp. than previously known. Because of their high recreational value for human beings, urban green areas are likely to remain in the research focus on public health issues.

Background: Current research emphasizes the high prevalence and costs of low back pain (LBP). The STarT Back Tool was designed to support primary care decision making for treatment by helping to determine the treatment prognosis of patients with non-specific low back pain. The German version is the STarT-G. The cross-cultural translation of the tool followed a structured and widely accepted process but to date it was only partially validated with a small sample. The aim of the study was to test the psychometric properties construct validity, discriminative ability, internal consistency and test-retest-reliability of the STarT-G and to compare them with values given for the original English version.Methods: A consecutive cohort study with a two-week retest was conducted among patients with non-specific LBP, aged 18 to 60 years, from primary care practices. Questionnaires were collected before the first consultation, and two weeks later by post, using the following reference standards: the Roland and Morris disability questionnaire, the Tampa Scale of Kinesiophobia, the Pain Catastrophizing Scale and the Hospital Anxiety and Depression Scale. Psychometric properties examined included the tool's discriminative abilities, whether the psychosocial subscale was one factor, internal consistency, item redundancy, test-retest reliability and floor and ceiling effects.Results: There were 228 patients recruited with a mean age of 42.2 (SD 11.0) years, and 53 % were female. The areas under the curve (AUC) for discriminative ability ranged from 0.70 (STarT-G Subscale - Pain Catastrophizing Scale; CI95 0.63, 0.78) to 0.77 (STarT-G Total - Composite reference standard, CI95 0.60, 0.94). Factor loadings ranged from 0.49 to 0.74. Cronbach's alpha testing the internal consistency and redundancy for the total/subscale scores were α = 0.52/0.55 respectively. The STarT-G test-retest reliability Kappa values for the total/subscale scores were 0.67/0.68 respectively. No floor or ceiling effects were present.Conclusions: The STarT-G shows acceptable psychometric properties although not in exact agreement with the original English version. The items previously regarded as a psychosocial subscale may be better seen as an index of different individual psychosocial constructs. The relevance of using the tool at the point of consultation should be further examined. [ABSTRACT FROM AUTHOR]

Background Francisella isolates from patients suffering from tularemia in Germany are generally strains of the species F. tularensis subsp. holarctica. To our knowledge, no other Francisella species are known for Germany. Recently, a new Francisella species could be isolated from a water reservoir of a cooling tower in Germany. Results We identified a Francisella sp. (isolate W12-1067) whose 16S rDNA is 99% identical to the respective nucleotide sequence of the recently published strain F. guangzhouensis. The overall sequence identity of the fopA, gyrA, rpoA, groEL, sdhA and dnaK genes is only 89%, indicating that strain W12-1067 is not identical to F. guangzhouensis. W12-1067 was isolated from a water reservoir of a cooling tower of a hospital in Germany. The growth optimum of the isolate is approximately 30°C, it can grow in the presence of 4-5% NaCl (halotolerant) and is able to grow without additional cysteine within the medium. The strain was able to replicate within a mouse-derived macrophage-like cell line. The whole genome of the strain was sequenced (∼1.7 mbp, 32.2% G + C content) and the draft genome was annotated. Various virulence genes common to the genus Francisella are present, but the Francisella pathogenicity island (FPI) is missing. However, another putative type-VI secretion system is present within the genome of strain W12-1067. Conclusions Isolate W12-1067 is closely related to the recently described F. guangzhouensis species and it replicates within eukaryotic host cells. Since W12-1067 exhibits a putative new type-VI secretion system and F. tularensis subsp. holarctica was found not to be the sole species in Germany, the new isolate is an interesting species to be analyzed in more detail. Further research is needed to investigate the epidemiology, ecology and pathogenicity of Francisella species present in Germany. [ABSTRACT FROM AUTHOR]

The article presents a case study of Anaplasma platys infection in a dog from Croatia. Basophilic inclusions in thrombocytes were revealed by haematological examination of a blood smear. The signs of thrombocytopenia, anemia and elevated C-reactive protein (CRP) were noted in the dog. The background, findings and conclusions of the case mentioned above, are discussed in detail.

Background: The study describes the estimation of the spatial distribution of questing nymphal tick densities by investigating Ixodes ricinus in Southwest Germany as an example. The production of high-resolution maps of questing tick densities is an important key to quantify the risk of tick-borne diseases. Previous I. ricinus maps were based on quantitative as well as semi-quantitative categorisations of the tick density observed at study sites with different vegetation types or indices, all compiled on local scales. Here, a quantitative approach on the landscape scale is introduced., Methods: During 2 years, 2013 and 2014, host-seeking ticks were collected each month at 25 sampling sites by flagging an area of 100 square meters. All tick stages were identified to species level to select nymphal ticks of I. ricinus, which were used to develop and calibrate Poisson regression models. The environmental variables height above sea level, temperature, relative humidity, saturation deficit and land cover classification were used as explanatory variables., Results: The number of flagged nymphal tick densities range from zero (mountain site) to more than 1,000 nymphs/100 m(2). Calibrating the Poisson regression models with these nymphal densities results in an explained variance of 72 % and a prediction error of 110 nymphs/100 m(2) in 2013. Generally, nymphal densities (maximum 374 nymphs/100 m(2)), explained variance (46 %) and prediction error (61 nymphs/100 m(2)) were lower in 2014. The models were used to compile high-resolution maps with 0.5 km(2) grid size for the study region of the German federal state Baden-Württemberg. The accuracy of the mapped tick densities was investigated by leave-one-out cross-validation resulting in root-mean-square-errors of 227 nymphs/100 m(2) for 2013 and 104 nymphs/100 m(2) for 2014., Conclusions: The methodology introduced here may be applied to further tick species or extended to other study regions. Finally, the study is a first step towards the spatial estimation of tick-borne diseases in Central Europe.

Background: Tularemia is a zoonotic disease caused by Francisella tularensis that has been found in many different vertebrates. In Germany most human infections are caused by contact with infected European brown hares (Lepus europaeus). The aim of this study was to elucidate the epidemiology of tularemia in hares using phenotypic and genotypic characteristics of F. tularensis., Results: Cultivation of F. tularensis subsp. holarctica bacteria from organ material was successful in 31 of 52 hares that had a positive PCR result targeting the Ft-M19 locus. 17 isolates were sensitive to erythromycin and 14 were resistant. Analysis of VNTR loci (Ft-M3, Ft-M6 and Ft-M24), INDELs (Ftind33, Ftind38, Ftind49, RD23) and SNPs (B.17, B.18, B.19, and B.20) was shown to be useful to investigate the genetic relatedness of Francisella strains in this set of strains. The 14 erythromycin resistant isolates were assigned to clade B.I, and 16 erythromycin sensitive isolates to clade B.IV and one isolate was found to belong to clade B.II. MALDI-TOF mass spectrometry (MS) was useful to discriminate strains to the subspecies level., Conclusions: F. tularensis seems to be a re-emerging pathogen in Germany. The pathogen can easily be identified using PCR assays. Isolates can also be identified within one hour using MALDI-TOF MS in laboratories where specific PCR assays are not established. Further analysis of strains requires genotyping tools. The results from this study indicate a geographical segregation of the phylogenetic clade B.I and B.IV, where B.I strains localize primarily within eastern Germany and B.IV strains within western Germany. This phylogeographical pattern coincides with the distribution of biovar I (erythromycin sensitive) and biovar II (erythromycin resistance) strains. When time and costs are limiting parameters small numbers of isolates can be analysed using PCR assays combined with DNA sequencing with a focus on genetic loci that are most likely discriminatory among strains found in a specific area. In perspective, whole genome data will have to be investigated especially when terrorist attack strains need to be tracked to their genetic and geographical sources.

Background: African swine fever (ASF) is one of the most complex viral diseases affecting both domestic and wild pigs. It is caused by ASF virus (ASFV), the only DNA virus which can be efficiently transmitted by an arthropod vector, soft ticks of the genus Ornithodoros. These ticks can be part of ASFV-transmission cycles, and in Europe, O. erraticus was shown to be responsible for long-term maintenance of ASFV in Spain and Portugal. In 2014, the disease has been reintroduced into the European Union, affecting domestic pigs and, importantly, also the Eurasian wild boar population. In a first attempt to assess the risk of a tick-wild boar transmission cycle in Central Europe that would further complicate eradication of the disease, over 700 pre-existing serum samples from wild boar hunted in four representative German Federal States were investigated for the presence of antibodies directed against salivary antigen of Ornithodoros erraticus ticks using an indirect ELISA format., Results: Out of these samples, 16 reacted with moderate to high optical densities that could be indicative of tick bites in sampled wild boar. However, these samples did not show a spatial clustering (they were collected from distant geographical regions) and were of bad quality (hemolysis/impurities). Furthermore, all positive samples came from areas with suboptimal climate for soft ticks. For this reason, false positive reactions are likely., Conclusion: In conclusion, the study did not provide stringent evidence for soft tick-wild boar contact in the investigated German Federal States and thus, a relevant involvement in the epidemiology of ASF in German wild boar is unlikely. This fact would facilitate the eradication of ASF in the area, although other complex relations (wild boar biology and interactions with domestic pigs) need to be considered.