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Background Improved understanding of Bartonella spp. serology in dogs may aid clinical decision making. Objective Describe demographic and geographic patterns of Bartonella spp. seroreactivity in dogs, and describe hematologic and serum biochemical abnormalities in Bartonella spp. seroreactive and nonseroreactive dogs. Animals Serum samples from 5957 dogs in the United States, previously submitted to IDEXX Reference Laboratories. Methods Serum was tested using 3 indirect ELISAs for B. henselae, B. vinsonii subsp. berkhoffii, and B. koehlerae. Complete blood count and serum biochemistry panel results were reviewed retrospectively. Results Overall, 6.1% of dogs were Bartonella spp. seroreactive. Toy breeds were less likely to be seroreactive (3.9%) than mixed breeds (7.5%; adjusted odds ratio [aOR], 0.48; 95% confidence interval [CI], 0.32‐0.72), and dogs

Lymphoma is the most common haematological malignancy in dogs and its aetiology is largely unknown. The presence of canine vector-borne agents (CVBD) in lymphoma tissues has been described and its causative effects questioned. We intended to evaluate the presence and extent of Leishmania infantum, Ehrlichia canis, Anaplasma phagocytophilum and Bartonella henselae infection in dogs with lymphoma. Sixty-one dogs, living in the Lisbon metropolitan area, with a diagnosis of lymphoma were enrolled. Immunofluorescence assays were used to detect serum IgG's. The presence of DNA from CVBD agents in tumour tissue was assessed by PCR. All dogs tested negative for B. henselae, A. phagocytophilum and E. canis by both serology and PCR. Regarding L. infantum, 8.2% (n = 5) of the dogs had a positive serologic result. L. infantum DNA was detected in two samples of diffuse large B-cell lymphoma (DLBCL). These results show an increased, but not significant, seropositivity (8.2% vs 7.9%) and molecular detection (3.3% vs 1.2%) for L. infantum in dogs with lymphoma, when compared to the reported canine population in the same geographical area. We could not identify an association between lymphoma and E. canis, A. phagocytophilum, B. henselae or Leishmania infantum infection in the studied population. Nevertheless, further studies, following dogs trough their CVBD disease evolution, are worthwhile and may help clarify a possible role of CVBD agents in lymphomagenesis. Bayer Animal Health- Portugal info:eu-repo/semantics/publishedVersion

Reptiles are reservoirs of a wide range of pathogens, including many protozoa, helminths, pentastomids, and arthropod parasitic species, some of which may be of public health concern. In this review we discuss the zoonotic risks associated with human–reptile interactions. Increased urbanization and introduction of exotic species of reptile may act as drivers for the transmission of zoonotic parasites through the environment. In addition, being a part of human diet, reptiles can be a source of life-threatening parasitoses, such as pentastomiasis or sparganosis. Finally, reptiles kept as pets may represent a risk to owners given the possibility of parasites transmitted by direct contact or fecal contamination. Awareness of reptile-borne zoonotic parasitoses is important to advocate control, prevention, and surveillance of these neglected diseases., Highlights Species of protozoa, helminths, pentastomids, and arthropod vectors exploit reptiles as definitive or paratenic hosts, which may represent a public health concern. The zoonotic risk is associated with human–reptile interactions and includes environmental contamination, reptile consumption, or keeping reptiles as pets. Exotic reptile species may introduce new zoonotic parasites in a previously nonendemic region. Pentastomiasis and sparganosis are life-threatening food-borne parasitoses. In our households, if precautions are not taken, reptiles may transmit zoonotic parasites by direct contact or fecal contamination. Trained veterinarians, physicians, and public health officials are important to advocate for proper diagnostics, parasite identification and treatment, as well as for surveillance strategies and food inspection in areas where reptiles are consumed.

Background Bartonella spp. comprise a genus of bacteria that frequently cause persistent, often subclinical infection. Although many Bartonella spp. have been implicated in a variety of clinical presentations, Bartonella rochalimae has yet to be documented in association with a clinical presentation other than infectious endocarditis (IE) in dogs. Objectives To document a spectrum of clinical presentations accompanied by mild hematological abnormalities in B rochalimae-infected dogs from the United States. Animals Eight dogs with documented B rochalimae infection. Methods Retrospective 10-year study of B rochalimae naturally infected dogs. Clinical and clinicopathologic data, including medical history, CBC, serum biochemistry panel, urinalysis, echocardiogram, and comprehensive vector-borne disease diagnostic panel results, were reviewed. Results Eight dogs were diagnosed with B rochalimae via polymerase chain reaction (PCR) amplification. Five dogs were diagnosed with IE. Three dogs, PCR positive for B rochalimae, were diagnosed with seizures or antibiotic responsive lameness or during routine screening of a military working dog. Conclusions This case series provides support for an association between B rochalimae and IE and provides documentation of dogs infected with B rochalimae with other clinical diagnoses.

Increasing amount of data attest that (in the context of vector-borne infections) birds are not only important as hosts of blood-sucking arthropod vectors, but also as reservoirs of vector-borne pathogens. From 2015 to 2019 cadavers of 100 birds (from 45 species, nine orders) were collected in Hungary, and their organs were screened for DNA from a broad range of vector-borne bacteria with PCR and sequencing. Molecular analyses revealed the presence of Anaplasmataceae, and sequencing identified bacteria closely related to Neorickettsia helminthoeca and Ehrlichia chaffeensis in a Eurasian teal (Anas crecca) and a song thrush (Turdus philomelos), respectively. All samples were PCR negative for rickettsiae, borreliae, Francisella and Coxiella spp., as well as for piroplasms. To our knowledge, this is the first report of a Neorickettsia and an Ehrlichia sp., which belong to the phylogenetic groups of N. helminthoeca and E. chaffeensis, respectively, from Europe. The potential presence of these two vector-borne bacteria needs to be taken into account during future studies on the eco-epidemiology of Anaplasmataceae in Europe.

Background Due to the complexity of retrieving skin-dwelling microfilariae, filarioids of dogs presenting dermal microfilariae (e.g. Cercopithifilaria spp., Onchocerca lupi) are relatively unknown compared to Dirofilaria spp. and Acanthocheilonema spp. whose microfilariae circulate in the blood. Although Cercopithifilaria spp. and O. lupi filarioids are distributed worldwide, there is a paucity of information on their occurrence in Iran. The aim of this study was to investigate these filarioids in a large population of dogs from different regions of Iran. Methods From October 2018 to September 2020, skin biopsies were obtained from dogs housed in shelters (n = 557) and privately owned dogs (n = 26) in seven provinces of Iran (Hamedan, Kermanshah, Yazd, Mazandaran, Khuzestan, Lorestan, Esfahan), as well as from three road-killed jackals (Canis aureus) and three cats (Felis catus) in Hamedan province. The skin biopsies were first soaked in saline solution at room temperature overnight, and examined for dermal microfilariae under the microscope. Positive skin specimens and sediments were tested by PCR for a 304-bp region of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene and amplicons were sequenced. Results Microfilariae of Cercopithifilaria spp. were found in skin biopsies of 32 of the 583 (5.5%) dogs tested, with infection rates of up to 25% in Kermanshah. No microfilariae were recovered from skin biopsy samples collected from dogs in Khorramabad and Ahvaz, nor from the examined jackals and cats. None of the privately owned dogs were found to be infected. Morphologic and morphometric characteristics of the microfilariae were consistent with C. bainae. Eighteen skin samples were positive for the cox1 gene, of which 15 sequences showed a nucleotide identity of 100% and three of 93.4% with the reference sequence of C. bainae available in GenBank (haplotype I; GenBank accession number: JF461457). Conclusions The data from this study broadens current knowledge on the geographical distribution of C. bainae in dogs in Middle Eastern countries. Further studies on different wild canine species in the country (e.g. jackal, fox, wolf) could provide further information on the epidemiology of these filarioids. A particular focus should be put on zoonotic O. lupi given the reports of its presence in human patients from this country. Graphical Abstract

Background The recent detection of the South Asian malaria vector Anopheles stephensi in the Horn of Africa (HOA) raises concerns about the impact of this mosquito on malaria transmission in the region. Analysis of An. stephensi genetic diversity and population structure can provide insight into the history of the mosquito in the HOA to improve predictions of future spread. We investigated the genetic diversity of An. stephensi in eastern Ethiopia, where detection suggests a range expansion into this region, in order to understand the history of this invasive population. Methods We sequenced the cytochrome oxidase subunit I (COI) and cytochrome B gene (CytB) in 187 An. stephensi collected from 10 sites in Ethiopia in 2018. Population genetic, phylogenetic, and minimum spanning network analyses were conducted for Ethiopian sequences. Molecular identification of blood meal sources was also performed using universal vertebrate CytB sequencing. Results Six An. stephensi COI-CytB haplotypes were observed, with the highest number of haplotypes in the northeastern sites (Semera, Bati, and Gewana towns) relative to the southeastern sites (Kebridehar, Godey, and Degehabur) in eastern Ethiopia. We observed population differentiation, with the highest differentiation between the northeastern sites compared to central sites (Erer Gota, Dire Dawa, and Awash Sebat Kilo) and the southeastern sites. Phylogenetic and network analysis revealed that the HOA An. stephensi are more genetically similar to An. stephensi from southern Asia than from the Arabian Peninsula. Finally, molecular blood meal analysis revealed evidence of feeding on cows, goats, dogs, and humans, as well as evidence of multiple (mixed) blood meals. Conclusion We show that An. stephensi is genetically diverse in Ethiopia and with evidence of geographical structure. Variation in the level of diversity supports the hypothesis for a more recent introduction of An. stephensi into southeastern Ethiopia relative to the northeastern region. We also find evidence that supports the hypothesis that HOA An. stephensi populations originate from South Asia rather than the Arabian Peninsula. The evidence of both zoophagic and anthropophagic feeding support the need for additional investigation into the potential for livestock movement to play a role in vector spread in this region. Graphical Abstract

Flaviviruses consist of significant human pathogens responsible for hundreds of millions of infections each year. Their antigenic relationships generate immune responses that are cross-reactive to multiple flaviviruses and their widespread and overlapping geographical distributions, coupled with increases in vaccination coverage, increase the likelihood of exposure to multiple flaviviruses. Depending on the antigenic properties of the viruses to which a person is exposed, flavivirus cross-reactivity can be beneficial or could promote immune pathologies. In this review we describe our knowledge of the functional immune outcomes that arise from varied flaviviral immune statuses. The cross-reactive antibody and T cell immune responses that are protective versus pathological are also addressed.

Land use boundaries represent human&ndash, physical interfaces where risk of vector-borne disease transmission is elevated. Land development practices, coupled with rural and urban land fragmentation, increases the likelihood that immunologically na&iuml, ve humans will encounter infectious vectors at land use interfaces. This research consolidated land use classes from the GLC-SHARE dataset, calculated landscape metrics in linear (edge) density, proportion abundance, and patch density, and derived the incidence rate ratios of the Zika virus occurrence in Colombia, South America during 2016. Negative binomial regression was used to evaluate vector-borne disease occurrence counts in relation to Population Density, Average Elevation, Per Capita Gross Domestic Product, and each of three landscape metrics. Each kilometer of border length per square kilometer of area increase in the linear density of the Cropland and Grassland classes is associated with an increase in Zika virus risk. These spatial associations inform a risk reduction approach to rural and urban morphology and land development that emphasizes simple and compact land use geometry that decreases habitat availability for mosquito vectors of Zika virus.

BACKGROUND Because of poor sensitivity and questionable specificity of immunofluorescent antibody assays (IFAs), serological diagnosis of Bartonella species infections in dogs remains challenging. Despite limitations, IFA testing is the historical "gold standard" for Bartonella serodiagnosis in animals and humans. Because most diagnostic laboratories test against only 1 or 2 Bartonella spp., testing against a broader panel of Bartonella antigens may enhance diagnostic sensitivity and specificity. OBJECTIVE To evaluate the sensitivity and specificity of Bartonella IFA using 8 cell culture-grown Bartonella spp. isolates. ANIMALS Archived serum samples from 34 Bartonella spp. naturally exposed, polymerase chain reaction (PCR)-positive dogs and from 26 PCR-negative and IFA-negative dogs. METHODS Bartonella IFA sensitivity and specificity were assessed using cell culture-grown whole cell antigens derived from 3 Bartonella henselae (Bh) strains (Bh Houston 1, Bh San Antonio Type 2, Bh California 1), 3 Bartonella vinsonii subsp. berkhoffii genotypes (Bvb I, II, and III), Bartonella koehlerae (Bk), and Bartonella quintana (Bq). RESULTS Only 62% of 34 Bartonella spp. PCR-positive dogs were seroreactive to any of the 8 Bartonella IFA antigens, indicating low IFA sensitivity. PCR-positive dogs were most often IFA seroreactive to Bq (n = 15), to Bvb II (n = 13), or to both (n = 9) antigens. Of the 26 previously IFA-negative/PCR-negative dogs, 4 (15%) were seroreactive using the expanded antigen panel. CONCLUSION AND CLINICAL IMPORTANCE Despite IFA testing of dogs against 8 different Bartonella isolates, IFA sensitivity remained poor, and specificity was only 85%. Development of a reliable serological assay is needed to facilitate the diagnosis of Bartonella infection in dogs.

We conducted an investigation of Borrelia miyamotoi infections in humans and ticks in northeastern China. Of 984 patients reporting recent tick bites, 14 (1.4%) were found to be infected with B. miyamotoi by PCR and genomic sequencing. The 14 patients had nonspecific febrile manifestations, including fever, headache, anorexia, asthenia, and arthralgia. Rash, eschar, and regional lymphadenopathy were each observed in 1 patient. Four (28.6%) patients were hospitalized because of severe disease. B. miyamotoi was detected in 3.0% (19/627) of Ixodes persulcatus, 1 (2.8%) of 36 Haemaphysalis concinna, and none of 29 Dermacentor silvarum ticks. Phylogenetic analyses based on sequences of a nearly entire 16s rRNA gene, a partial flagellin gene, and the glycerophosphodiester phosphodiesterase gene revealed that B. miyamotoi identified in patients and ticks were clustered in the group of the Siberian type. These findings indicate that B. miyamotoi is endemic in northeastern China and its public health significance deserves further investigation.

© 2017 Elsevier Inc. Some pesticides are applied directly to aquatic systems to reduce numbers of mosquito larvae (larvicides) and thereby reduce transmission of pathogens that mosquitoes vector to humans and wildlife. Sustained, environmentally-safe control of larval mosquitoes is particularly needed for highly productive waters (e.g., catchment basins, water treatment facilities, septic systems), but also for other habitats to maintain control and reduce inspection costs. Common biorational pesticides include the insect juvenile hormone mimic methoprene and pesticides derived from the bacteria Bacillus thuringiensis israelensis, Lysinibacillus sphaericus and Saccharopolyspora spinosa (spinosad). Health agencies, the public and environmental groups have especially debated the use of methoprene because some studies have shown toxic effects on non-target organisms. However, many studies have demonstrated its apparent environmental safety. This review critically evaluates studies pertinent to the environmental safety of using methoprene to control mosquito larvae, and provides concise assessments of the bacterial larvicides that provide sustained control of mosquitoes. The review first outlines the ecological and health effects of mosquitoes, and distinguishes between laboratory toxicity and environmental effects. The article then interprets non-target toxicity findings in light of measured environmental concentrations of methoprene (as used in mosquito control) and field studies of its non-target effects. The final section evaluates information on newer formulations of bacterially-derived pesticides for sustained mosquito control. Results show that realized environmental concentrations of methoprene were usually 2–5 µg/kg (range 2–45 µg/kg) and that its motility is limited. These levels were not toxic to the vast majority of vertebrates and invertebrates tested in laboratories, except for a few species of zooplankton, larval stages of some other crustaceans, and small Diptera. Studies in natural habitats have not documented population reductions except in small Diptera. Bacterial larvicides showed good results for sustained control with similarly limited environmental effects, except for spinosad, which had broader effects on insects in mesocosms and temporary pools. These findings should be useful to a variety of stakeholders in informing decisions on larvicide use to protect public and environmental health in a ‘One Health’ framework.

Background Anaplasma phagocytophilum is an emerging tick-borne zoonotic pathogen of increased interest worldwide which has been detected in northern Africa. Anaplasma platys is also present in this region and could possibly have a zoonotic potential. However, only one recent article reports on the human esposure to A. phagocytophilum in Morocco and no data are available on canine exposure to both bacteria. Therefore, we conducted a cross-sectional epidemiological study aiming to assess both canine and human exposure to Anaplasma spp. in Morocco. A total of 425 dogs (95 urban, 160 rural and 175 working dogs) and 11 dog owners were sampled from four cities of Morocco. Canine blood samples were screened for Anaplasma spp. antibodies by an enzyme-linked immunosorbent assay (ELISA) and for A. phagocytophilum and A. platys DNA by a real-time polymerase chain reaction (RT-PCR) targeting the msp2 gene. Human sera were tested for specific A. phagocytophilum immunoglobulin G (IgG) using a commercial immunofluorescence assay (IFA) kit. Results Anaplasma spp. antibodies and A. platys DNA were detected in 21.9 and 7.5% of the dogs, respectively. Anaplasma phagocytophilum DNA was not amplified. Anaplasma platys DNA was significantly more frequently amplified for working dogs. No statistically significant differences in the prevalence of Anaplasma spp. antibodies or A. platys DNA detection were observed between sexes, age classes or in relation to exposure to ticks. A total of 348 Rhipicephalus sanguineus (sensu lato) ticks were removed from 35 urban and working dogs. The majority of dog owners (7/10) were seroreactive to A. phagoyctophilum IgG (one sample was excluded because of hemolysis). Conclusions This study demonstrates the occurrence of Anaplasma spp. exposure and A. platys infection in dogs, and A. phagocytophilum exposure in humans in Morocco.

Background Mosquito population dynamics are driven by large-scale (e.g. climatological) and small-scale (e.g. ecological) factors. While these factors are known to independently influence mosquito populations, it remains uncertain how drivers that simultaneously operate under natural conditions interact to influence mosquito populations. We, therefore, developed a well-controlled outdoor experiment to assess the interactive effects of two ecological drivers, predation and nutrient availability, on mosquito life history traits under multiple temperature regimes. Methods We conducted a temperature-controlled mesocosm experiment in Kruger National Park, South Africa, with the yellow fever mosquito, Aedes aegypti. We investigated how larval survival, emergence and development rates were impacted by the presence of a locally-common invertebrate predator (backswimmers Anisops varia Fieber (Notonectidae: Hemiptera), nutrient availability (oligotrophic vs eutrophic, reflecting field conditions), water temperature, and interactions between each driver. Results We observed that the effects of predation and temperature both depended on eutrophication. Predation caused lower adult emergence in oligotrophic conditions but higher emergence under eutrophic conditions. Higher temperatures caused faster larval development rates in eutrophic but not oligotrophic conditions. Conclusions Our study shows that ecological bottom-up and top-down drivers strongly and interactively govern mosquito life history traits for Ae. aegypti populations. Specifically, we show that eutrophication can inversely affect predator–prey interactions and mediate the effect of temperature on mosquito survival and development rates. Hence, our results suggest that nutrient pollution can overrule biological constraints on natural mosquito populations and highlights the importance of studying multiple factors. Electronic supplementary material The online version of this article (10.1186/s13071-019-3431-x) contains supplementary material, which is available to authorized users.

Insects in the orders Hemiptera and Thysanoptera transmit viruses and other pathogens associated with the most serious diseases of plants. Plant viruses transmitted by these insects target similar tissues, genes, and proteins within the insect to facilitate plant-to-plant transmission with some degree of specificity at the molecular level. 'Omics experiments are becoming increasingly important and practical for vector biologists to use towards better understanding the molecular mechanisms and biochemistry underlying transmission of these insect-borne diseases. These discoveries are being used to develop novel means to obstruct virus transmission into and between plants. In this chapter, we summarize 'omics technologies commonly applied in vector biology and the important discoveries that have been made using these methods, including virus and insect proteins involved in transmission, as well as the tri-trophic interactions involved in host and vector manipulation. Finally, we critically examine the limitations and new horizons in this area of research, including the role of endosymbionts and insect viruses in virus-vector interactions, and the development of novel control strategies.

Porcine circovirus 3 (PCV-3) has emerged as a potential threat for swine industry, being consistently reported in the presence of several clinical signs all around the world. Recently, its presence in wild boar has been demonstrated at high prevalence. This evidence is surprising since the lower density of wild populations might not be expected to sustain such efficient viral transmission. Porcine circoviruses were proven to exhibit a certain plasticity in the host tropism and were detected in unrelated species, like mice, dogs and ruminants. However, if this scenario applies also to wild animals remains to be established. Therefore, this study aimed to investigate the presence of PCV-3 in wild ungulates other than wild boar and in related hematophagous ectoparasites. One hundred and nine animals were sampled from different hilly and mountain areas of Friuli Venezia Giulia, including 9 chamois (Rupicapra rupicapra), 17 red deer (Cervus elaphus), 4 mouflons (Ovis musimon), 50 roe deer (Capreolus capreolus) and 29 wild boars (Sus scrofa). Additionally, host-matched ectoparasites were collected when present. Porcine circovirus 3 was diagnosed using molecular techniques and sequencing. This study results confirmed the high PCV-3 occurrence in wild boar and reported for the first time its presence, at low prevalence, in chamois and roe deer. Moreover, two ticks (Ixodes ricinus), one of which non-engorged, collected from PCV-3 negative roe deer, tested PCV-3 positive. The genetic characterization of some of the strains collected from non-swine hosts allowed to prove that, albeit clearly part of PCV-3 species, they were genetically unique, demonstrating the absence of among-samples contamination and thus confirming the actual presence of PCV-3 genome in these new hosts. Therefore, this study highlights an unexpected broad PCV-3 distribution and circulation in the wild, rising further questions on porcine circoviruses infectious cycle, epidemiology and origin, which will deserve additional investigations.

Background Vector-borne pathogens are emerging concerns in multiple regions of Canada. Determining regional prevalence of canine vector-borne pathogens and documenting change will improve clinician awareness, enable targeted prevention, enhance diagnosis and ideally reduce the risk of disease. Study objectives were to: (i) estimate the prevalence of positive canine vector-borne test results from samples submitted in Canada; (ii) assess change in prevalence over time, from baseline (2008) to 2015; and (iii) estimate the prevalence of pathogen co-infections. Methods This repeat cross-sectional study evaluated 753,468 test results for D. immitis antigen and B. burgdorferi, Ehrlichia canis/ewingii/muris serology, and 753,208 test results for Anaplasma phagocytophilum/platys serology using the SNAP® 4Dx®Test and SNAP 4Dx® Plus Test. Results Based on all submitted samples from Canada (2008–2015), the period seroprevalence of B. burgdorferi, Ehrlichia spp., Anaplasma spp. and D. immitis antigen were 2.0%, 0.5%, 0.4% and 0.2%, respectively. Over the 7 years (2008 compared to 2015) we observed a significant increase in seroprevalence for B. burgdorferi (144.4%) and Ehrlichia spp. (150%). Co-infections (positive for two or more pathogens on a single 4 pathogen test kit) were estimated at 5.4% (1162/21,612) of total positive tests. Conclusions The temporal rise and geographical differences in prevalence detected for these pathogens (notably B. burgdorferi) are consistent with anecdotal information on canine illness related to tick-borne pathogen exposure in multiple regions of Canada, particularly canine Lyme disease. Electronic supplementary material The online version of this article (10.1186/s13071-019-3299-9) contains supplementary material, which is available to authorized users.

International audience; Climate change, with warmer temperatures and altered precipitation patterns, has affected the distribution of vectors and vector-borne diseases. In the northern hemisphere, vectors are spreading north, and with them, pathogens of zoonotic and animal health impact. Eurasian moose (Alces alces alces) are physiologically and anatomically adapted for cold climate, and are rarely considered ideal hosts of vectors, apart from deer keds (Lipoptena cervi). To investigate the presence of vector-borne pathogens, spleen samples from 615 moose were collected in southern Sweden from 2008 to 2015. The samples were analyzed with a high-throughput PCR method for 24 bacterial, and 12 parasitic pathogens. Anaplasma (82%), Borrelia (3%), Babesia (3%), and Bartonella (1%) DNA was found, showing that moose are exposed to, and can act as hosts of some of these pathogens, which can have an impact of both animal and human health. These results show that Swedish moose are exposed to pathogens that in some instances are more commonly found in regions with warmer climate, and highlights the importance of also considering moose as sentinels of vector-borne pathogens. Further research is needed to understand the effect of these pathogens on the health of individual moose and to elucidate whether climate change and moose population density interact to create the pattern observed.

Leishmaniasis is a zoonotic disease caused by predominantly vectorborne Leishmania spp. In the United States, canine visceral leishmaniasis is common among hounds, and L. infantum vertical transmission among hounds has been confirmed. We found that L. infantum from hounds remains infective in sandflies, underscoring the risk for human exposure by vectorborne transmission.

Highlights • Information about vector-borne bacteria of camels is scarce. • Anaplasmataceae, SFG rickettsiae, Bartonella and Borrelia were investigated in blood of 200 dromedary camels of Iran. • PCR results revealed 30 animals (15%) to be infected with Anaplasmataceae bacteria. • BlastN® analysis of positive samples revealed identity with “Candidatus Anaplasma camelii” isolates. • This paper reviews the current knowledge on camels’ tick-borne bacteria including microscopy, serology and molecular studies., Despite close association between camels and humans, molecular based studies on vector-borne pathogens infecting camels are scarce compared to other animals in Iran. The current study was carried out to investigate the occurrence of vector-borne bacteria in the blood of dromedaries by molecular tools. A total of 200 peripheral blood samples were collected from apparently healthy animals. Microscopic examination was performed on Giemsa-stained blood smears, and drops of blood were spotted on Whatman FTA® cards for molecular analyses. Genomic DNA was extracted from the cards, and PCR amplification followed by sequencing of positive samples was carried out for the detection of Anaplasmataceae, spotted fever group (SFG) rickettsiae, Bartonella spp. and Borrelia spp. Intra-cytic forms of any blood pathogens could not be detected by light microscopy. PCR results revealed 30 animals (15%) to be infected with Anaplasmataceae bacteria. Analyses of sequences revealed a strain of Anaplasma sp. identical to Candidatus Anaplasma camelii isolated from camels, cattle and deer in Asia and Africa. Neither SFG rickettsiae, nor Borrelia or Bartonella species were found. Further studies for determining epidemiological role of camels and its zoonotic potential are recommended. This paper reviews the current knowledge on camels’ tickborne bacteria including microscopy, serology and molecular studies.

[Background] Haemosporidians are the most important vector-borne parasites due to their cosmopolitan distribution and their wide range of hosts, including humans. Identification of their vectors is critical to highlight ecologically and epidemiologically relevant features such as host specificity or transmission routes. Biting midges of the genus Culicoides are considered the main vectors of Haemoproteus spp., yet important information on aspects such as vector feeding preferences or vector-host specificity involving haemosporidian parasites is frequently missing., [Methods] We assessed the abundance of Culicoides circumscriptus and C. paolae and blood sources of the latter at the nests of cavity-nesting bird species (mainly the European roller Coracias garrulus) and in their surroundings. We also explored the prevalence and genetic diversity of avian haemosporidians in parous females of both species., [Results] Both C. circumscriptus and C. paolae were abundant in the study area and common at European roller nests. Culicoides paolae had a diverse ornithophilic diet, feeding on at least seven bird species. Human DNA was also detected in the blood meal of some individuals. Four Haemoproteus lineages, including a new one reported here for the first time, were isolated from parous females of both biting midges., [Conclusions] Culicoides circumscriptus and C. paolae can play a locally important role in the transmission dynamics of Haemoproteus parasites in a community of cavity-nesting bird species in an arid ecosystem., JV was funded by the Spanish Ministry of Economy, Industry and Competitiveness by means of a predoctoral grant (grant no. BES-2015-075951). FV and JF received financial support from the Spanish Ministry of Economy and Competitiveness, and the European Regional Development Fund (CGL2014-55969-P; CGL2015-65055-P). JMP was partially supported by a 2017 Leonardo Grant for Researchers and Cultural Creators, BBVA Foundation. The Foundation accepts no responsibility for the opinions, statements and contents included in the project and/or the results thereof, which are entirely the responsibility of the authors. We acknowledge support of the publication fees by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI).

Due to the emergence of non-endemic mosquito vectors and the recent outbreaks of mosquito-borne diseases, mosquito-borne pathogens are considered an increasing risk to public and animal health in Europe. To obtain a status quo regarding mosquito-borne viruses and their vectors in Germany, 97,648 mosquitoes collected from 2011 to 2016 throughout the country were screened for arboviruses. Mosquitoes were identified to species, pooled in groups of up to 50 individuals according to sampling location and date, and screened with different PCR assays for Flavi-, Alpha- and Orthobunyavirus RNA. Two pools tested positive for Usutu virus-RNA, two for Sindbis virus-RNA, and 24 for Batai virus-RNA. The pools consisted of Culex pipiens s.l., Culex modestus, Culex torrentium, Culiseta sp., Aedes vexans, Anopheles daciae, and Anopheles messeae mosquitoes and could be assigned to nine different collection sites, with seven of them located in northeastern Germany. Phylogenetic analyses of the viral RNA sequences showed relationships with strains of the viruses previously demonstrated in Germany. These findings confirm continuing mosquito-borne zoonotic arbovirus circulation even though only a rather small percentage of the screened samples tested positive. With respect to sampling sites and periods, virus circulation seems to be particularly intense in floodplains and after flooding events when mosquitoes develop in excessive numbers and where they have numerous avian hosts available to feed on.

International audience; In June 2017, an Anopheles mosquito species was detected in New Caledonia. Morphologic identification and genomic sequencing revealed that the specimens tested belong to An. bancroftii genotype A1. This introduction underscores the risk for local malaria transmission and the vulnerability of New Caledonia to vector introduction.

Human infections by various rickettsial species are frequently reported globally. We investigated a flea-borne rickettsial outbreak infecting 300 people in Western Himalayan region of India. Arthropod vectors (ticks and fleas) and animal and human blood samples from affected households were analysed by gltA and ompB genes based polymerase chain reaction (PCR). Rat flea (Ceratophyllus fasciatus) samples were found harbouring a Rickettsia sp. Phylogenetic analysis based on gltA gene using PHYLIP revealed that the detected Rickettsia sp. has 100% identity with SE313 and RF2125 strains of Rickettsia sp. of flea origin from Egypt and Thai-Myanmar border, respectively and cf1 and 5 strains from fleas and lice from the USA. But, the nucleotide sequence of genetically variable gene ompB of R14 strain was found closely related to cf9 strain, reported from Ctenocephalides felis fleas. These results highlight the public health importance of such newly discovered or less recognised Rickettsia species/strains, harboured by arthropod vectors like fleas.

Background: As there is little data on vector-borne diseases of cats in the Caribbean region and even around the\ud world, we tested feral cats from St Kitts by PCR to detect infections with Babesia, Ehrlichia and spotted fever group\ud Rickettsia (SFGR) and surveyed them for antibodies to Rickettsia rickettsii and Ehrlichia canis.\ud Results: Whole blood was collected from apparently healthy feral cats during spay/ neuter campaigns on St Kitts in\ud 2011 (N = 68) and 2014 (N = 52). Sera from the 52 cats from 2014 were used to detect antibodies to Ehrlichia canis\ud and Rickettsia rickettsii using indirect fluorescent antibody tests and DNA extracted from whole blood of a total of\ud 119 cats (68 from 2011, and 51 from 2014) was used for PCRs for Babesia, Ehrlichia and Rickettsia. We could not\ud amplify DNA of SFG Rickettsia in any of the samples but found DNA of E. canis in 5% (6/119), Babesia vogeli in 13%\ud (15/119), Babesia gibsoni in 4% (5/119), mixed infections with B. gibsoni and B. vogeli in 3% (3/119), and a poorly\ud characterized Babesia sp. in 1% (1/119). Overall, 10% of the 52 cats we tested by IFA for E. canis were positive while\ud 42% we tested by indirect fluorescent antibody (IFA) for R. rickettsii antigens were positive.\ud Conclusions: Our study provides the first evidence that cats can be infected with B. gibsoni and also indicates that\ud cats in the Caribbean may be commonly exposed to other vector-borne agents including SFGR, E. canis and B.\ud vogeli. Animal health workers should be alerted to the possibility of clinical infections in their patients while public\ud health workers should be alerted to the possibility that zoonotic SFGR are likely circulating in the region.

Bartonellae are fastidious, facultative, intracellular vector-borne bacteria distributed among mammalian reservoirs worldwide. The pathogenic potential of many Bartonella spp. has increased the interest in these bacteria and advanced their research. Isolation of Bartonella spp. is laborious using classical bacteriological methods and requires specific conditions and prolonged incubation periods. In contrast, molecular methods for detection of Bartonella DNA are considered as more practical and sensitive than the former. Among the molecular methods, the use of real-time PCR assays for primary screening of Bartonella spp., followed by several molecular confirmatory assays, using either conventional or real-time PCR, is recommended. Although primary isolation of Bartonella is a laborious task, we encourage its application to all PCR-positive samples as this is the most reliable proof for the presence of live bacteria. Moreover, a successful trial will enable a broader molecular characterization and speciation of isolated colonies. The present guideline gathers and summarizes recommendations, including advantages and limitations of isolation and molecular detection of Bartonella from mammalian and arthropod samples.

The purpose of this review is to provide practical information to help researchers intending to perform "from field to laboratory" studies on phleboviruses transmitted by sandflies. This guideline addresses the different steps to be considered starting from the field collection of sandflies to the laboratory techniques aiming at the detection, isolation, and characterization of sandfly-borne phleboviruses. In this guideline article, we address the impact of various types of data for an optimal organization of the field work intending to collect wildlife sandflies for subsequent virology studies. Analysis of different data sets should result in the geographic positioning of the trapping stations. The overall planning, the equipment and tools needed, the manpower to be deployed, and the logistics to be anticipated and set up should be organized according to the objectives of the field study for optimal efficiency. This work was done under the frame of EurNegVec COST Action TD1303. This work was supported, in part, by (1) the European Virus Archive goes Global (EVAg) project that has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement no 653316 and by (2) the EDENext FP7- n 261504 EU project; this article is cataloged by the EDENext Steering Committee as EDENext453 (www.edenext.eu). info:eu-repo/semantics/publishedVersion

International audience; Bluetongue virus serotype 8 (BTV-8) was reported for the first time in Europe in 2006, causing the largest bluetongue outbreak ever recorded. France was mostly impacted in 2007/09. Trade restrictions were implemented all along. Vaccination became available from 2008: a limited number of doses was first administered in an emergency vaccination campaign, followed by two nationwide compulsory vaccination campaigns in 2009 and 2010. France regained a disease-free status in December 2012, but BTV may have kept circulating undetected as infected herds have been reported again since August 2015. We developed a stochastic dynamic compartmental model of BTV transmission in cattle and sheep to analyze the relative importance of vector active flight and host movements in disease spread, and assess the effectiveness of control measures. We represented BTV transmission both within and between French administrative subdivisions called cantons, during the 2007/09 outbreak and until the end of 2010, when compulsory vaccination was interrupted. Within-canton transmission was vector-borne, and between canton transmission could occur through three contact networks that accounted for movements of: (i) vectors between pastures located at close distance; (ii) cattle and sheep between pastures of the same farm; (iii) traded cattle. We estimated the model parameters by approximate Bayesian computation, using data from the 2007 French outbreak. With this framework, we were able to reproduce the BTV-8 epizootic wave. Host movements between distant pastures of the same farm were found to have a major contribution to BTV spread to disease-free areas, thus raising practical questions about herd management during outbreaks. We found that cattle trade restrictions had been well complied with; without them, the whole French territory would have been infected by winter 2007. The 2008 emergency vaccination campaign had little impact on disease spread as almost half vaccine doses had likely been administered to already immune cattle. Alternatively, establishing a vaccination buffer zone would have allowed a better control of BTV in 2008: limiting its spatial expansion and decreasing the number of infected cattle and sheep. We also showed a major role of compulsory vaccination in controlling the outbreak in 2009 and 2010, though we predicted a possible low-level circulation after the last detection.

The role of mammalian skin in harbouring and transmitting arthropod-borne protozoan parasites has been overlooked for decades as these pathogens have been regarded primarily as blood-dwelling organisms. Intriguingly, infections with low or undetected blood parasites are common, particularly in the case of Human African Trypanosomiasis caused by Trypanosoma brucei gambiense. We hypothesise, therefore, the skin represents an anatomic reservoir of infection. Here we definitively show that substantial quantities of trypanosomes exist within the skin following experimental infection, which can be transmitted to the tsetse vector, even in the absence of detectable parasitaemia. Importantly, we demonstrate the presence of extravascular parasites in human skin biopsies from undiagnosed individuals. The identification of this novel reservoir requires a re-evaluation of current diagnostic methods and control policies. More broadly, our results indicate that transmission is a key evolutionary force driving parasite extravasation that could further result in tissue invasion-dependent pathology. DOI: http://dx.doi.org/10.7554/eLife.17716.001

International audience; Equine encephalosis is a midge-borne viral disease of equines caused by equine encephalosis virus (EEV, Orbivirus, Reoviridae), and closely related to African horse sickness virus (AHSV). EEV and AHSV share com- mon vectors and show similar transmission patterns. Until now EEV has caused outbreaks in Africa and Israel. This study aimed to provide insight in the probability of an EEV outbreak in The Netherlands caused by infected vectors or hosts, the contribution of potential source areas (risk regions) to this probability, and the effectiveness of preventive measures (sanitary regimes). A stochastic risk model constructed for risk assessment of AHSV introduction was adapted to EEV. Source areas were categorized in risk regions (high, low, and very low risk) based on EEV history and the presence of competent vectors. Two possible EEV introduction pathways were considered: importation of infected equines and importation of infected vectors along with their vertebrate hosts. The probability of EEV introduction (PEEV ) was calculated by combining the probability of EEV release by either pathway and the probability of EEV establishment. The median current annual probability of EEV introduction by an infected equine was estimated at 0.012 (90% uncertainty interval 0.002–0.020), and by an infected vector at 4.0 10−5 (90% uncertainty interval 5.3 10−6–2.0 10−4). Equines from high risk regions contributed most to the probability of EEV introduction with 74% on the EEV introduction by equines, whereas low and very low risk regions contributed 18% and 8%, respectively. International movements of horses participating in equestrian events contributed most to the probability of EEV introduction by equines from high risk regions (86%), but also contributed substantially for low and very low risk regions with 47% and 56%. The probability of introducing EEV into The Netherlands is much higher than the probability of intro- ducing AHSV with equines from high risk countries contributing most. The introduction by an infected equine is the most likely pathway. Control measures before exportation of equines showed to have a strong mitigating effect on the probability of EEV introduction. The risk of EEV outbreaks should be taken into account when altering these import regulations.

Background The blacklegged tick Ixodes scapularis transmits Borrelia burgdorferi (sensu stricto) in eastern North America; however, the agent of Lyme disease is not the sole pathogen harbored by the blacklegged tick. The blacklegged tick is expanding its range into areas of southern Canada such as Ontario, an area where exposure to blacklegged tick bites and tick-borne pathogens is increasing. We performed a systematic review to evaluate the public health risks posed by expanding blacklegged tick populations and their associated pathogens. Methods We followed PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines for conducting our systematic review. We searched Ovid MEDLINE, Embase, BIOSIS, Scopus and Environment Complete databases for studies published from 2000 through 2015, using subject headings and keywords that included “Ixodes scapularis”, “Rickettsia”, “Borrelia”, “Anaplasma”, “Babesia” and “pathogen.” Two reviewers screened titles and abstracts against eligibility criteria (i.e. studies that included field-collected blacklegged ticks and studies that did not focus solely on B. burgdorferi) and performed quality assessments on eligible studies. Results Seventy-eight studies were included in the final review, 72 were from the US and eight were from Canada (two studies included blacklegged ticks from both countries). Sixty-four (82 %) studies met ≥ 75 % of the quality assessment criteria. Blacklegged ticks harbored 91 distinct taxa, 16 of these are tick-transmitted human pathogens, including species of Anaplasma, Babesia, Bartonella, Borrelia, Ehrlichia, Rickettsia, Theileria and Flavivirus. Organism richness was highest in the Northeast (Connecticut, New York) and Upper Midwest US (Wisconsin); however, organism richness was dependent on sampling effort. The primary tick-borne pathogens of public health concern in Ontario, due to the geographic proximity or historical detection in Ontario, are Anaplasma phagocytophilum, Babesia microti, B. burgdorferi, Borrelia miyamotoi, deer tick virus and Ehrlichia muris-like sp. Aside from B. burgdorferi and to a much lesser concern A. phagocytophilum, these pathogens are not immediate concerns to public health in Ontario; rather they represent future threats as the distribution of vectors and pathogens continue to proliferate. Conclusions Our review is the first systematic assessment of the literature on the human pathogens associated with the blacklegged tick. As Lyme disease awareness continues to increase, it is an opportune time to document the full spectrum of human pathogens transmittable by blacklegged ticks. Electronic supplementary material The online version of this article (doi:10.1186/s13071-016-1529-y) contains supplementary material, which is available to authorized users.

International audience; This study aimed to evaluate the use of routinely collected reproductive and milk production data for the early detection of emerging vector-borne diseases in cattle in the Netherlands and the Flanders region of Belgium (i.e., the northern part of Belgium). Prospective space-time cluster analyses on residuals from a model on milk production were carried out to detect clusters of reduced milk yield. A CUSUM algorithm was used to detect temporal aberrations in model residuals of reproductive performance models on two indicators of gestation length. The Bluetongue serotype-8 (BTV-8) epidemics of 2006 and 2007 and the Schmallenberg virus (SBV) epidemic of 2011 were used as case studies to evaluate the sensitivity and timeliness of these methods. The methods investigated in this study did not result in a more timely detection of BTV-8 and SBV in the Netherlands and BTV-8 in Belgium given the surveillance systems in place when these viruses emerged. This could be due to (i) the large geographical units used in the analyses (country, region and province level), and (ii) the high level of sensitivity of the surveillance systems in place when these viruses emerged. Nevertheless, it might be worthwhile to use a syndromic surveillance system based on non-specific animal health data in real-time alongside regular surveillance, to increase the sense of urgency and to provide valuable quantitative information for decision makers in the initial phase of an emerging disease outbreak.

Sylvatic plague, caused by the bacterium Yersinia pestis, is periodically responsible for large die-offs in rodent populations that can spillover and cause human mortalities. In the western US, prairie dog populations experience nearly 100% mortality during plague outbreaks, suggesting that multiple transmission pathways combine to amplify plague dynamics. Several alternate pathways in addition to flea vectors have been proposed, such as transmission via direct contact with bodily fluids or inhalation of infectious droplets, consumption of carcasses, and environmental sources of plague bacteria, such as contaminated soil. However, evidence supporting the ability of these proposed alternate pathways to trigger large-scale epizootics remains elusive. Here we present a short review of potential plague transmission pathways and use an ordinary differential equation model to assess the contribution of each pathway to resulting plague dynamics in black-tailed prairie dogs (Cynomys ludovicianus) and their fleas (Oropsylla hirsuta). Using our model, we found little evidence to suggest that soil contamination was capable of producing plague epizootics in prairie dogs. However, in the absence of flea transmission, direct transmission, i.e., contact with bodily fluids or inhalation of infectious droplets, could produce enzootic dynamics, and transmission via contact with or consumption of carcasses could produce epizootics. This suggests that these pathways warrant further investigation.

Dengue is a significant mosquito-borne infection in humans, and its worldwide prevalence is rapidly increasing. In 2010, 83 serum samples from febrile travellers returning from dengue-endemic countries to a region in north-eastern Italy, densely infested with Aedes albopictus, were analysed for dengue virus (DENV). DENV RNA was detected in 20.5% of patients. By RT-PCR, DENV serotypes 1 and 3 were the most common. DENV must be identified early in symptomatic travellers returning from high-risk countries, to prevent outbreaks where potential vectors exist.

Background Bats are among the most eco-epidemiologically important mammals, owing to their presence in human settlements and animal keeping facilities. Roosting of bats in buildings may bring pathogens of veterinary-medical importance into the environment of domestic animals and humans. In this context bats have long been studied as carriers of various pathogen groups. However, despite their close association with arthropods (both in their food and as their ectoparasites), only a few molecular surveys have been published on their role as carriers of vector-borne protozoa. The aim of the present study was to compensate for this scarcity of information. Findings Altogether 221 (mostly individual) bat faecal samples were collected in Hungary and the Netherlands. The DNA was extracted, and analysed with PCR and sequencing for the presence of arthropod-borne apicomplexan protozoa. Babesia canis canis (with 99-100 % homology) was identified in five samples, all from Hungary. Because it was excluded with an Ixodidae-specific PCR that the relevant bats consumed ticks, these sequences derive either from insect carriers of Ba. canis, or from the infection of bats. In one bat faecal sample from the Netherlands a sequence having the highest (99 %) homology to Besnoitia besnoiti was amplified. Conclusions These findings suggest that some aspects of the epidemiology of canine babesiosis are underestimated or unknown, i.e. the potential role of insect-borne mechanical transmission and/or the susceptibility of bats to Ba. canis. In addition, bats need to be added to future studies in the quest for the final host of Be. besnoiti.

© Copyright 2015, Mary Ann Liebert, Inc. 2015. We used literature searches and a database of all reported emerging infectious diseases (EIDs) to analyze the most important transmission pathways (e.g., vector-borne, aerosol droplet transmitted) for emerging zoonoses. Our results suggest that at the broad scale, the likelihood of transmission occurring through any one pathway is approximately equal. However, the major transmission pathways for zoonoses differ widely according to the specific underlying drivers of EID events (e.g., land-use change, agricultural intensification). These results can be used to develop better targeting of surveillance for, and more effective control of newly emerged zoonoses in regions under different underlying pressures that drive disease emergence.

Background Newly recognized endemic foci for human babesiosis include Europe, where Ixodes ricinus, a vector for several species of Babesia, is the most commonly identified tick. Vector-based surveillance provides an early warning system for the emergence of human babesiosis, which is likely to be under-reported at emerging sites. In the present study, we set out to screen I. ricinus collected from Danish domestic dogs for Babesia, in order to identify whether humans in Denmark are exposed to the parasite. Findings A total of 661 ticks (Ixodes spp.) were collected from 345 Danish domestic dogs during April-September 2011 and pooled, one sample per dog. DNA was extracted from each sample and examined by PCR and sequencing for Rickettsia spp., Borrelia burgdorferi sensu lato, Bartonella spp., Francisella tularensis, Candidatus Neoehrlichia mikurensis, and Babesia spp. In total, 34% of the samples were positive for tick-borne microorganisms potentially pathogenic to humans: Rickettsia spp. were detected in 16% of the pools, with 79% being R. helvetica. Borrelia burgdorferi sensu lato was found in 15%, with the main species identified as Borrelia afzelii (39%). Likewise, 8% of the samples were positive for Babesia spp. (Babesia microti, 82%; Babesia venatorum (‘EU1’), 18%). Lastly, 1% of the samples tested positive for Candidatus Neoehrlichia mikurensis, and 0.6% for Bartonella spp. No ticks were found to be infected with Francisella tularensis. Conclusions Our data are in support of endemic occurrence of potentially zoonotic Babesia in Denmark and confirms I. ricinus as a vector of multiple pathogens of public health concern. Electronic supplementary material The online version of this article (doi:10.1186/s13071-015-0843-0) contains supplementary material, which is available to authorized users.

Background Bovine besnoitiosis has been recently diagnosed in a three-parted herd of 796 Aubrac and Charolais beef cattle in Hungary. A large scale serological, histological and molecular survey was initiated in order to uncover important factors in the local epidemiology of the disease. Findings Blood samples were collected (three times from the whole herd, and repeatedly from selected animals) for serological screening by ELISA. In addition, various organs from aborted fetuses and newborn calves, skin and colostrum samples of seropositive heifers/cows, and ticks collected from the cattle were histologically and/or molecularly analysed for the presence of Besnoitia besnoiti. All fetal and calf tissues, as well as colostrum and tick samples from cows were PCR negative. Based on ELISA results, only very few local cows seroconverted after mating with imported, infected bulls, and not necessarily as a consequence of this event. Among calves that were born to seropositive, imported cows and stayed with their mother until weaning at seven months of age, seroprevalence decreased significantly, but remained high. At the same time, 28 calves born from seropositive cows, but separated from their dams immediately after receiving colostrum, were successfully reared and remained uninfected. Following a second herd-level screening, all Aubrac cattle (except for heifer calves) and all seropositive Charolais cows and bulls were culled. Manifestation of the disease is currently sporadic. Among chronically affected heifers two types of skin lesions were noted, and histological evaluation indicated marked distension of sweat gland ducts with membrane-bound structures (resembling cystozoites) in their contents. Conclusions Transmission through natural mating, as well as transplacental, colostral and tick-borne transmission of B. besnoiti was either unlikely or did not occur. However, the risk for spreading of the infection was high, when calves stayed with their mother during suckling, and if animals were kept in the same stable (although physically separated) during the main fly season. Herd replacement and generation exchange (i.e. early weaning and artificial feeding) appear to be the successful strategies for the local eradication of bovine besnoitiosis. Adding to the already known mechanical transmission of B. besnoiti by blood-sucking flies, results of the present study suggest that secretophagous flies should also be evaluated as potential vectors of this coccidium species.

West Nile virus (WNV), the most widely distributed flavivirus worldwide, has lately reemerged in Europe, causing worrisome outbreaks in humans and horses. Serological analysis by enzyme-linked immunoassay and plaque reduction neutralization test showed for the first time in Serbia that 12% of 349 horses presented specific neutralizing WNV antibodies, which in one case also cross-neutralized Usutu virus (USUV). This is the first time that anti-USUV high neutralizing antibody titers are reported in horses. All these data indicate that WNV and USUV are circulating in the region and advise on the convenience of implementing surveillance programs. © Copyright 2011, Mary Ann Liebert, Inc.

To the Editor: Bartonellosis, a globally emerging vector-borne zoonotic bacterial disease, is caused by hemotropic, gram-negative, aerobic, facultative intracellular Bartonella spp. (1). Of the 30 Bartonella species/subspecies, 17 have been associated with human infections (2,3). Each species has a reservoir host(s), within which the bacteria can cause intraerythrocytic bacteremia with few or no clinical signs of illness (1,3); the bacteria are transmitted by hematophagous arthropod vectors (1). Various Bartonella spp. have been identified in domestic and wild animals, including canids, deer, cattle, rodents, and marine mammals (1,4). Bartonella DNA from the blood of loggerhead sea turtles (Caretta caretta) has been PCR amplified and sequenced (5); the fact that Bartonella DNA was found suggests the possibility that persistent blood-borne infection can occur in nonmammals and that the host range for Bartonella spp. may be larger than anticipated. Growing evidence suggests that wild birds play key roles in the maintenance and movement of zoonotic pathogens such as tick-borne encephalitis virus and Borrelia and Rickettsia spp. (6–9). Bartonella grahamii DNA was amplified from a bird tick in Korea (10). The substantial mobility, broad distribution, and migrations of birds make them ideal reservoir hosts for dispersal of infectious agents. To investigate whether birds might be a reservoir for Bartonella spp., we screened 86 birds for the presence of Bartonella spp. DNA. The primary study site was a residential backyard in Morehead City, North Carolina, USA (34°43.722′N, 76°43.915′W). Of the 86 birds screened, 78 (16 species) were captured by mist net during March 2010–June 2012 and 8 (3 species) were injured birds that were to be euthanized (Table). Each bird was examined for external abnormalities and ectoparasites, weighed, measured, and tagged with a US Geological Survey–numbered band. A blood sample (0.10–0.25 mL) was collected from each bird by using a 1-mL insulin syringe with a 28-gauge × 1.27-cm needle. Blood remaining after preparation of blood smears was added to an EDTA tube and frozen (−80°C) until processed. Blood smears were examined for hemoparasites. Research was conducted under required state and federal bird banding permits and with the approval of the North Carolina State University Institutional Animal Care and Use Committee. Table Bartonella species detected in birds* Before DNA was extracted from the samples, 10 μL of blood was diluted in 190 µL of phosphate-buffered saline. DNA was automatically extracted by using a BioRobot Symphony Workstation and MagAttract DNA Blood M96 Kit (QIAGEN, Valencia, CA, USA). Bartonella DNA was amplified by using conventional Bartonella genus PCR primers targeting the 16S–23S intergenic spacer region: oligonucleotides, 425s (5′-CCG GGG AAG GTT TTC CGG TTT ATCC-3′) and 1,000as (5′-CTG AGC TAC GGC CCC TAA ATC AGG-3′). Amplification was performed in a 25-μL reaction, as described (3). All PCR reactions were analyzed by 2% agarose gel electrophoresis. Amplicons were sequenced to identify the Bartonella sp. and intergenic spacer region genotype. To compare sequences with those in GenBank, we identified bacterial species and genotypes by using Blast version 2.0 (http://blast.ncbi.nlm.nih.gov/Blast.cgi). DNA extraction and PCR-negative controls remained negative throughout the study. Results are summarized in the Table. None of the screened birds were anemic, but 5 were PCR positive for Bartonella spp. (3 for B. henselae and 2 for B. koehlerae). B. henselae was amplified from 2 Northern Mockingbirds (Mimus polyglottos) and 1 Red-winged Blackbird (Agelaius phoeniceus) (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"KC814161","term_id":"501412833","term_text":"KC814161"}}KC814161). The DNA sequences were identical to each other and had 99.6% (456/457 bp) sequence similarity with B. henselae San Antonio 2 intergenic spacer region genotype (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"AF369529","term_id":"15290601","term_text":"AF369529"}}AF369529). B. koehlerae was amplified from a Red-bellied Woodpecker (Melanerpes carolinus) and a Common Loon (Gavia immer) (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"KC814162","term_id":"501412834","term_text":"KC814162"}}KC814162). The DNA sequences were identical to each other (404/404 bp) and to GenBank sequence {"type":"entrez-nucleotide","attrs":{"text":"AF312490","term_id":"15277539","term_text":"AF312490"}}AF312490. Lice (Mallophaga order) were found on 5 Boat-tailed Grackles (Quiscalus major), but no ectoparasites were observed on Bartonella spp.–positive birds. Hemoparasites (Haemoproteus and Plasmodium spp.) were detected in 7 of 86 birds, indicating exposure to hematophagous ectoparasites, but hemoparasites were not detected in the Bartonella spp.–positive birds. No bacteria were visualized in Bartonella PCR–positive blood smears. Bartonella spp. are increasingly associated with animal and human illnesses; thus, the identification of reservoirs and increased understanding of Bartonella spp. disease ecology are of public health importance. Our finding of 2 pathogenic species not previously reported in birds has expanded the potential sources for zoonotic infection. There is growing evidence that migratory birds serve as reservoirs and/or mechanical vectors for pathogens such as tick-borne encephalitis virus and Rickettsia spp. (6–8). Birds have been implicated as reservoirs for several Borrelia spp. (9,10) and for possible dispersion of other tick-borne pathogens (e.g., Anaplasma and Bartonella spp.) (6,10). Tick transmission of Bartonella spp. to birds should be investigated, and additional studies that investigate the reservoir host range of Bartonella spp. and the transmission of these bacteria to non–host species will improve epidemiologic understanding of bartonellosis and will identify additional risk factors for Bartonella spp. transmission to new hosts, including humans.

To assess environmental and horse-level risk factors associated with West Nile Virus (WNV) circulation in Camargue, Southern France, a serosurvey was conducted on non-vaccinated horses (n = 1159 from 134 stables) in 2007 and 2008. Fifteen Landsat images were examined to quantify areas with open water and flooded vegetation around sampled horses. Mean percentages of areas of open water and flooded vegetation, as well as variations in these percentages between 3 periods (November to February = NOT, March to July = END and August to October = EPI), were calculated for buffers of 2 km radius around the stables. Results of the final logistic regression showed that the risk of WNV seropositivity in horses decreased with their date of acquisition and age. Results also demonstrated the significant role of environmental variables. Horse serological status was associated with variations of open water areas between the NOT (November to February) and END (March to July) periods, as well as between END and EPI (August to October). WNV spillover was found more intense in areas where water level decreased strongly from winter to spring and from spring to summer.

To control West Nile virus (WNV), it is necessary to know which mosquitoes are able to transmit this virus. Therefore, we evaluated the WNV vector potential of several North American mosquito species. Culex restuans and Cx. salinarius, two species from which WNV was isolated in New York in 2000, were efficient laboratory vectors. Cx. quinquefasciatus and Cx. nigripalpus from Florida were competent but only moderately efficient vectors. Coquillettidia perturbans was an inefficient laboratory vector. As WNV extends its range, exposure of additional mosquito species may alter its epidemiology.

We report the arthropod-borne pediatric encephalitic agent La Crosse virus in Aedes albopictus mosquitoes collected in Dallas County, Texas, USA, in August 2009. The presence of this virus in an invasive vector species within a region that lies outside the virus's historically recognized geographic range is of public health concern.