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In conjunction with efforts to assess pathogen exposure in feral pigs from the southeastern United States, we amplified Bartonella henselae, B. koehlerae, and B. vinsonii subsp. berkhoffii from blood samples. Feral pigs may represent a zoonotic risk for hunters or butchers and pose a potential threat to domesticated livestock. [ABSTRACT FROM AUTHOR]

Domestic and wild felids are frequently parasitized by apicomplexan protozoa in the genus Cytauxzoon . Expanding species diversity has recently been described within this genus, with potential implications for epidemiology and pathogenesis. In light of these findings, this study assessed the genetic diversity of Cytauxzoon spp. in wild felids (n = 66) from different eco-regions of Brazil and Argentina. Of the 66 blood samples analyzed, 53 (80.3%) were 18S rRNA gene PCR-positive for Cytauxzoon spp., including 43 jaguars ( Panthera onca ) and 10 ocelots ( Leopardus pardalis ). Panthera onca specimens (100%, 43/43) were most frequently infected, followed by Leopardus pardalis (76.9%; 10/13). Cytauxzoon spp. were not detected in Leopardus braccatus (n = 1) or Puma concolor (n = 9). Phylogenetic analyses of fragments of the 18S rRNA, cytB , and cox-1 gene sequences from jaguars were closely related to Cytauxzoon felis . In contrast, sequences from ocelots were more closely associated with Cytauxzoon brasiliensis . Distance and haplotype analysis further confirmed the circulation of at least two distinct genovariants of C. felis among jaguars, as evidenced by their close positioning and low genetic divergence (0-0.14% for 18S rRNA, 0.37-0.56% for cytB , and 0.08-0.74% for cox-1 ). Additionally, sequence data from ocelots suggested that multiple genovariants of C. brasiliensis are circulating among these cats in different Brazilian eco-regions. Our study provides evidence of two distinct Cytauxzoon organisms parasitizing free-ranging and captive jaguars and ocelots, respectively, in Brazil and Argentina.

The Bartonella genus of bacteria encompasses ubiquitous species, some of which are pathogenic in humans and animals. Bartonella henselae, the causative agent of Cat Scratch disease, is responsible for a large portion of human Bartonella infections. These bacteria can grow outside of cells, replicate in erythrocytes and invade endothelial and monocytic cells. We have previously reported reduced antibiotic susceptibility of intracellular Bartonella. In this study we performed comparative transcriptomic analyses between the extracellular and intracellular B. henselae phenotypes. Overall, specific genes involved in invasion, virulence, extracellular adhesion of type 4 secretion system were downregulated following intracellular invasion of B. henselae. Downregulation included BadA, a well-characterized adhesin molecule, of critical importance for cell invasion. These studies demonstrate the ability to purify Bartonella RNA from infected cells and offer a repository of gene expression data for future research. The development of novel therapeutics will benefit from the ability to determine target expression by Bartonella in relevant microenvironments., Competing Interests: Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

This study reinforces the value of a One Health approach to infectious disease outbreak investigations. After the onset of neuropsychiatric symptoms in their son, our investigation focused on a family composed of a mother, father, two daughters, the son, two dogs, and a rabbit, all with exposures to vectors (fleas and ticks), rescued dogs, and other animals. Between 2020 and 2022, all family members experienced illnesses that included neurological symptoms. Prolonged menorrhagia (130d) in the youngest daughter ultimately resolved following antibiotic administration. One dog was diagnosed with a splenic hematoma and months later spinal histiocytic sarcoma. The father, both daughters, and one dog were seroreactive to multiple Bartonella spp. antigens, whereas the mother and son were not seroreactive. Bartonella quintana DNA was amplified from specimens obtained from all family members. Based upon DNA sequencing, infection with B. quintana was confirmed for the mother and both pet dogs. Bartonella henselae DNA was amplified and sequenced from the youngest daughter, the son, and one dog (co-infected with B. quintana ), and from Ctenocephalides felis collected from their pet rabbit. All five family members and one dog were infected with Babesia divergens -like MO-1. Both parents were co-infected with Babesia microti. Droplet digital PCR supported potential infection with a Borrelia species in three family members. This study provided additional case-based evidence supporting the role of stealth Babesia , Bartonella , and Borrelia pathogens as a cause or cofactor in neurological and neuropsychiatric symptoms. We conclude that a One Health investigation approach, particularly for stealth vector borne pathogens such as Babesia , Bartonella , and Borrelia spp., will enhance clinical and epidemiological understanding of these organisms for animal and human health. During outbreak investigations it is critical to document travel and vector exposure histories, symptoms, and pathology in pets and human patients, contact with rescued, wild, or feral animals and perform diagnostic testing that includes family members, pets, and vectors.

More than one-hundred Babesia species that affect animals and humans have been described, eight of which have been associated with emerging and underdiagnosed zoonoses. Most diagnostic studies in humans have used serology or molecular assays based on the 18S rRNA gene. Because the 18S rRNA gene is highly conserved, obtaining an accurate diagnosis at the species level is difficult, particularly when the amplified DNA fragment is small. Also, due to its low copy number, sequencing of the product is often unsuccessful. In contrast, because the Babesia internal transcribed regions (ITS), between 18S rRNA and 5.8S rRNA, and between 5.8S rRNA and 28S rRNA, contain highly variable non-coding regions, the sequences in these regions provide a good option for developing molecular assays that facilitate differentiation at the species level. In this study, the complete ITS1 and ITS2 intergenic regions of different Piroplasmida species were sequenced to add to the existing GenBank database. Subsequently, ITS1 and ITS2 sequences were used to develop species-specific PCR assays and specific single-plex and multiplex conventional (c)PCR, quantitative real-time (q)PCR, and digital (d)PCR assays for four zoonotic Babesia species ( Babesia divergens , Babesia odocoilei , Babesia duncani , and Babesia microti ). The efficacy of the assay protocols was confirmed by testing DNA samples extracted from human blood or enrichment blood cultures. Primers were first designed based on the 18S rRNA-5.8S rRNA and 5.8S rRNA-28S rRNA regions to obtain the ITS1 and ITS2 sequences derived from different Piroplasmida species ( B. odocoilei , Babesia vulpes , Babesia canis , Babesia vogeli , Babesia gibsoni , Babesia lengau , Babesia divergens -like, B. duncani , B. microti , Babesia capreoli , Babesia negevi , Babesia conradae , Theileria bicornis , and Cytauxzoon felis ). Subsequently, using these sequences, single-plex or multiplex protocols were optimized targeting the ITS1 region of B. divergens , B. microti , and B. odocoilei . Each protocol proved to be sensitive and specific for the four targeted Babesia sp., detecting 10 -2 (for B. microti and B. odocoilei ) and 10 -1 (for B. divergens and B. duncani ) DNA copies per microliter. There was no cross-amplification among the Babesia species tested. Using 226 DNA extractions from blood or enrichment blood cultures obtained from 82 humans, B. divergens (seven individuals), B. odocoilei (seven individuals), and B. microti (two individuals) were detected and identified as a single infection, whereas co-infection with more than one Babesia sp. was documented by DNA sequencing in six (7.3%) additional individuals (representing a 26.8% overall prevalence). These newly developed protocols proved to be effective in detecting DNA of four Babesia species and facilitated documentation of co-infection with more than one Babesia sp. in the same individual.

Background: Bartonella species are fastidious, intracellular bacteria responsible for an expanding array of human pathologies. Most are considered to be transmitted by direct inoculation with infected bodily fluids from a mammalian reservoir species or vector-transmitted through a variety of arthropod species and their excrement. However, there are mounting reports of infection in the absence of documented animal or vector contact. A variety of Bartonella species have been documented in conditions affecting both the peripheral and central nervous systems. More common conditions, including neuroretinitis, are often associated with Bartonella henselae. However, Bartonella quintana, the agent of trench fever, as well as emerging pathogens related to rodent reservoir species, B. grahamii and B. elizabethae, have also been documented. Encephalitis and encephalopathy, also most often associated with B. henselae, have been reported with B. quintana, B. washoensis (ground squirrels) and B. vinsonii subsp. vinsonii (voles) infections. Bartonella infections have also been associated with peripheral neuropathies, such as cranial nerve paresis and neuropathic pain, including infection with less commonly encountered species such as Bartonella koehlerae. Recently, molecular diagnostic testing revealed that DNA from Bartonella spp. was found to be more prevalent in blood of patients with neuropsychiatric disorders such as schizophrenia and psychoses compared to healthy controls., Methods: A systematic literature search was conducted on PubMed, Google Scholar and Web of Science. Search terms included Bartonella and specific neurological conditions and focused on peer-reviewed case reports published after 2012 pursuant to a prior review, with limited exceptions for conditions not previously covered. Published diagnostic testing, serology, molecular testing or pathology, were necessary for inclusion, except for one case which had clinical and epidemiological evidence consistent with diagnosis along with follow-up., Results: Neurobartonelloses included neuralgic amyotrophy, complex regional pain syndrome, chronic inflammatory demyelinating polyneuropathy, cranial nerve paralysis, Guillain-Barré syndrome, peripheral vasculitic polyneuropathy, acute transverse myelopathy, neuroretinitis, encephalitis/encephalopathy, cerebral vasculitis/aneurysm and neuropsychiatric conditions., Conclusions: The breadth of reported symptoms and clinical syndromes associated with an increasing number of Bartonella species continues to expand. Increased clinical awareness of this important zoonotic pathogen is necessary to advance One Health among the medical and veterinary communities., (© 2024. The Author(s).)

Bartonella spp. are opportunistic, vectorborne bacteria that can cause disease in both animals and humans. We investigated the molecular occurrence of Bartonella spp. in 634 phlebotomine sand fly specimens, belonging to 44 different sand fly species, sampled during 2017-2021 in north and northeastern Brazil. We detected Bartonella sp. DNA in 8.7% (55/634) of the specimens by using a quantitative real-time PCR targeting the 16S-23S internal transcribed spacer intergenic region. Phylogenetic analysis positioned the Lutzomyia longipalpis sand fly-associated Bartonella gltA gene sequence in the same subclade as Bartonella ancashensis sequences and revealed a Bartonella sp. sequence in a Dampfomyia beltrani sand fly from Mexico. We amplified a bat-associated Bartonella nuoG sequence from a specimen of Nyssomyia antunesi sand fly. Our findings document the presence of Bartonella DNA in sand flies from Brazil, suggesting possible involvement of these insects in the epidemiologic cycle of Bartonella species.

Mild Cognitive Impairment (MCI) is a neurological disorder at the transition between normal cognitive decline and dementia. Despite the potential role of neuroinflammation in the pathogenesis of MCI, infectious triggers remain mostly unknown. Infection with Bartonella spp., a zoonotic bacterium, has recently been associated with diffuse neurological and psychiatric symptoms. Given the preferential endothelial localization of Bartonella spp. and the role of vascular changes in neurocognitive decline, we hypothesized that there is an association between Bartonella spp. infection and pathologically accelerated decline in cognitive function in aging. To test this hypothesis, we collected serological and molecular markers of past and present Bartonella spp. infection in a sample of older people with and without MCI. Samples were processed in a blinded way to exclude laboratory biases. Contrary to our hypothesis, people with MCI were not more likely than people without MCI to have an active Bartonella spp. infection as measured by droplet digital PCR (p = 0.735) and quantitative PCR (p = 1). In addition, there was no significant difference in positive serological results between cases and controls (p = 0.461). Overall, higher-than-expected active Bartonella spp. infection (37% by ddPCR) and seroreactivity (71% by indirect fluorescent antibody assay) were found in people without MCI. Conclusions require caution, as our study was limited by the small number of cases with MCI. Overall, our results identified a higher than previously recognized rate of exposure and infection with Bartonella spp. in this older study population but does not support a specific role for such infection in MCI., Competing Interests: In conjunction with Dr. S. Sontakke and North Carolina State University, E. B. Breitschwerdt holds US Patent No. 7,115,385 Media and Methods for Cultivation of Microorganisms, which was issued on October 3rd, 2006. He is a co-founder, shareholder and Chief Scientific Officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella spp. infections. Dr. Ricardo Maggi is a co-founder and the Chief Technical Officer for Galaxy Diagnostics Inc. Drs Breitschwerdt and Maggi are full-time employees of North Carolina State University. Dr. Frohlich is a full-time employee of the University of North Carolina – Chapel Hill. Dr. Frohlich is the lead inventor of IP issued to UNC and licensed to: Electromedical Products International (EPI). In the last twelve months, Dr. Frohlich has received consulting honoraria from the following entities: EPI, Insel Spital, and the University of Michigan. Dr. Frohlich is a shareholder of EPI. In the last twelve months, Dr. Frohlich has received royalties from Academic Press and the University of North Carolina at Chapel Hill. Ms. Guirguis, Pupillo, and Rodrigues report no financial relationships with commercial interests. All other authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. This does not alter our adherence to PLOS ONE policies on sharing data and materials., (Copyright: © 2024 Guirguis et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: In recent years, Babesia and Bartonella species co-infections in patients with chronic, nonspecific illnesses have continued to challenge and change the collective medical understanding of "individual pathogen" vector-borne infectious disease dynamics, pathogenesis and epidemiology. The objective of this case series is to provide additional molecular documentation of Babesia odocoilei infection in humans in the Americas and to emphasize the potential for co-infection with a Bartonella species., Methods: The development of improved and more sensitive molecular diagnostic techniques, as confirmatory methods to assess active infection, has provided increasing clarity to the healthcare community., Results: Using a combination of different molecular diagnostic approaches, infection with Babesia odocoilei was confirmed in seven people suffering chronic non-specific symptoms, of whom six were co-infected with one or more Bartonella species., Conclusions: We conclude that infection with Babesia odocoilei is more frequent than previously documented and can occur in association with co-infection with Bartonella spp., (© 2024. The Author(s).)

Introduction: The potential role of pathogens, particularly vector-transmitted infectious agents, as a cause of psychosis has not been intensively investigated. We have reported a potential link between Bartonella spp. bacteremia and neuropsychiatric symptoms, including pediatric acute onset neuropsychiatric syndrome and schizophrenia. The purpose of this study was to further assess whether Bartonella spp. exposure or infection are associated with psychosis., Methods: In a blinded manner, we assessed the presence of anti- Bartonella antibodies by indirect immunofluorescence assays (IFA), and infection by amplification of bacterial DNA from blood by quantitative polymerase chain reaction (qPCR), digital PCR (dPCR), and droplet digital PCR (ddPCR) in 116 participants. Participants were categorized into one of five groups: 1) controls unaffected by psychosis ( n = 29); 2) prodromal participants ( n = 16); 3) children or adolescents with psychosis ( n = 7); 4) adults with psychosis ( n = 44); and 5) relatives of a participant with psychosis ( n = 20)., Results: There was no significant difference in Bartonella spp. IFA seroreactivity between adults with psychosis and adult controls unaffected by psychosis. There was a higher proportion of adults with psychosis who had Bartonella spp. DNA in the bloodstream (43.2%) compared to adult controls unaffected by psychosis (14.3%, p = 0.021). The Bartonella species was determined for 18 of the 31 bacteremic participants, including infection or co-infection with Bartonella henselae (11/18), Bartonella vinsonii subsp. b erkhoffii (6/18), Bartonella quintana (2/18), Bartonella alsatica (1/18), and Bartonella rochalimae (1/18)., Discussion: In conjunction with other recent research, the results of this study provide justification for a large national or international multi-center study to determine if Bartonella spp. bacteremia is more prevalent in adults with psychosis compared to adults unaffected by psychosis. Expanding the investigation to include a range of vector-borne and other microbial infections with potential CNS effects would enhance knowledge on the relationship between psychosis and infection., Competing Interests: In conjunction with Dr. S. Sontakke and North Carolina State University, EB holds US Patent No. 7,115,385 Media and Methods for Cultivation of Microorganisms, which was issued on October 3rd, 2006. He is a co-founder, shareholder and Chief Scientific Officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella spp. infections. RM is a co-founder and the Chief Technical Officer for Galaxy Diagnostics Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Delaney, Robveille, Maggi, Lashnits, Kingston, Liedig, Murray, Fallon and Breitschwerdt.)

Bartonellosis refers to disease caused by the Bartonella genus of bacteria. The breadth of disease manifestations associated with Bartonella is currently expanding and includes regional lymphadenopathy, rheumatic, ocular, and neurological disorders. The dearth of knowledge regarding diagnosis, treatment and pathogenesis of this disease can be partially attributed to the lack of a reliable small animal model for the disease. For this study, Bartonella henselae, the most common species associated with human disease, was injected into Swiss Webster (SW) mice. When the outcome indicated that productive infection did not occur, SCID/Beige (immune compromised) mice were inoculated. While SW mice may potentially harbor an acute infection, less than 10 days in length, the SCID/Beige model provided a sustained infection lasting up to 30-days. These data indicate that SCID/Beige mice can provide a model to study Bartonella infection, therapeutics, and vector dynamics in the future., Competing Interests: Edward B. Breitschwerdt, DVM holds U.S. Patent No. 7115385; Media and Methods for cultivation of microorganisms, which was issued October 3, 2006. He is a co‐founder, shareholder and Chief Scientific Officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella species infections. All other authors declare no potential conflicts of interest., (Copyright: © 2024 Bullard et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Pathogen environmental stability is an often-neglected research priority for pathogens that are known to be vector-transmitted. Bartonella henselae , the etiologic agent of Cat Scratch Disease, has become a "pathogen of interest" in several serious human illnesses, which include neoplastic, cardiovascular, neurocognitive, and rheumatologic conditions. Survival in the flea gut and feces as well as the association with a biofilm in culture-negative endocarditis provides insight into this organism's ability to adjust to environmental extremes. The detection of B. henselae DNA in blood and tissues from marine mammals also raises questions about environmental stability and modes of pathogen transmission. We investigated the ability of B. henselae to survive in fluid matrices chosen to mimic potential environmental sources of infective materials. Feline whole blood, serum and urine, bovine milk, and physiologic saline inoculated with a laboratory strain of B. henselae San Antonio 2 were subsequently evaluated by culture and qPCR at specified time intervals. Bacterial viability was also assessed following desiccation and reconstitution of each inoculated fluid matrix. Bartonella henselae SA2 was cultured from feline urine up to 24 h after inoculation, and from blood, serum, cow's milk, and physiologic saline for up to 7 days after inoculation. Of potential medical importance, bacteria were cultured following air-desiccation of all fluid inoculates. The viability and stability of Bartonella within biological and non-biological fluids in the environment may represent a previously unrecognized source of infection for animals and human beings.

Bacteria of the genus Bartonella , a member of the Alphaproteobacteria , are fastidious, Gram-negative, aerobic bacilli that comprise numerous species, subspecies, and genotypes. Bartonella henselae, with a worldwide distribution, infects cats, dogs, horses, humans, and other mammals. Diagnostically, direct detection of Bartonella henselae in patient blood specimens by culture or molecular methods is required to confirm infection with this bacterium. Enrichment blood culture combined with quantitative PCR (qPCR) or ddPCR enhances the sensitivity of direct detection. The addition of sheep blood to liquid culture media increased the Bartonella henselae DNA concentration compared to controls, additionally improving PCR direct detection sensitivity. IMPORTANCE This study aims to improve diagnostic detection of Bartonella henselae. Patient samples are combined with enriched bacterial cultures aimed at growing Bartonella henselae for the best possible chance at detection. However, current Bartonella growth methods could be improved. The DNA extraction method used by most laboratories should also be optimized. Sheep blood was added to increase the growth of Bartonella henselae and multiple DNA extraction methods were to be compared to each other., Competing Interests: The authors declare a conflict of interest. Ricardo G. Maggi is a co-founder and the Chief Technical Officer for Galaxy Diagnostics Inc. In conjunction with S. Sontakke and North Carolina State University, E. B. Breitschwerdt holds US Patent No. 7,115,385 Media and Methods for Cultivation of Microorganisms, which was issued on October 3rd, 2006. He is a co-founder, shareholder and Chief Scientific Officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella spp. infections. All other authors claim no competing interests.

Large populations of unowned cats constitute an animal welfare, ecological, societal and public health issue worldwide. Their relocation and homing are currently carried out in many parts of the world with the intention of relieving suffering and social problems, while contributing to ethical and humane population control in these cat populations. An understanding of an individual cat's lifestyle and disease status by veterinary team professionals and those working with cat charities can help to prevent severe cat stress and the spread of feline pathogens, especially vector-borne pathogens, which can be overlooked in cats. In this article, we discuss the issue of relocation and homing of unowned cats from a global perspective. We also review zoonotic and non-zoonotic infectious agents of cats and give a list of practical recommendations for veterinary team professionals dealing with homing cats. Finally, we present a consensus statement consolidated at the 15th Symposium of the Companion Vector-Borne Diseases (CVBD) World Forum in 2020, ultimately to help veterinary team professionals understand the problem and the role they have in helping to prevent and manage vector-borne and other pathogens in relocated cats., (© 2022. The Author(s).)

Among the Ctenocephalides felis felis -borne pathogens, Bartonella henselae , the main aetiological agent of cat scratch disease (CSD), is of increasing comparative biomedical importance. Despite the importance of B. henselae as an emergent pathogen, prevention of the diseases caused by this agent in cats, dogs and humans mostly relies on the use of ectoparasiticides. A vaccine targeting both flea fitness and pathogen competence is an attractive choice requiring the identification of flea proteins/metabolites with a dual effect. Even though recent developments in vector and pathogen -omics have advanced the understanding of the genetic factors and molecular pathways involved at the tick-pathogen interface, leading to discovery of candidate protective antigens, only a few studies have focused on the interaction between fleas and flea-borne pathogens. Taking into account the period of time needed for B. henselae replication in flea digestive tract, the present study investigated flea-differentially abundant proteins (FDAP) in unfed fleas, fleas fed on uninfected cats, and fleas fed on B. henselae -infected cats at 24 hours and 9 days after the beginning of blood feeding. Proteomics approaches were designed and implemented to interrogate differentially expressed proteins, so as to gain a better understanding of proteomic changes associated with the initial B. henselae transmission period (24 hour timepoint) and a subsequent time point 9 days after blood ingestion and flea infection. As a result, serine proteases, ribosomal proteins, proteasome subunit α-type, juvenile hormone epoxide hydrolase 1, vitellogenin C, allantoinase, phosphoenolpyruvate carboxykinase, succinic semialdehyde dehydrogenase, glycinamide ribotide transformylase, secreted salivary acid phosphatase had high abundance in response of C. felis blood feeding and/or infection by B. henselae . In contrast, high abundance of serpin-1, arginine kinase, ribosomal proteins, peritrophin-like protein, and FS-H/FSI antigen family member 3 was strongly associated with unfed cat fleas. Findings from this study provide insights into proteomic response of cat fleas to B. henselae infected and uninfected blood meal, as well as C. felis response to invading B. henselae over an infection time course, thus helping understand the complex interactions between cat fleas and B. henselae at protein levels., Competing Interests: RGM is a co-founder and the Chief Technical Officer for Galaxy Diagnostics Inc. In conjunction with Dr. S. Sontakke and North Carolina State University, EBB holds US Patent No. 7,115,385 Media and Methods for Cultivation of Microorganisms, which was issued on October 3rd, 2006. He is a co-founder, shareholder and Chief Scientific Officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella spp. infections. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 André, Neupane, Lappin, Herrin, Smith, Williams, Collins, Bai, Jorge, Balbuena, Bradley, Maggi and Breitschwerdt.)

Bartonella spp. comprise a genus of Gram-negative alphaproteobacteria that are slow growing, fastidious, and facultative intracellular pathogens with zoonotic potential. Immunofluorescent antibody assays (IFAs), Western blot (WB), and enzyme-linked immunosorbent assays (ELISAs), the frequently used modalities for the serological diagnosis of canine and human Bartonelloses, generate numerous false negative results. Therefore, the development of a reliable serodiagnostic assay for Bartonelloses is of clinical and epidemiological importance. Pap31, a heme binding surface protein of B. henselae, is associated with bacterial adhesion and related to bacterial colonization. To our knowledge, B. henselae Pap31 and its fragments (N-terminal (NTD), middle (MD), and C-terminal (CTD) domains) have not been investigated for the serodiagnosis of canine and human Bartonelloses. In this study, we evaluate the diagnostic utility of B. henselae recombinant whole Pap31 (rPap31) and Pap31 fragments by ELISA using sera from 70 dogs (36 Bartonella spp. IFA-positive (naturally infected), and 34 Bartonella spp. IFA- and PCR-negative (control dogs)) and 36 humans (18 Bartonella spp. IFA-positive (naturally infected) and 18 controls)). In the dogs, the area under the curve (AUC) score of recombinant whole Pap31 was 0.714 with a sensitivity of 42% and specificity of 94% at an OD cutoff value of 0.8955. Among the evaluated recombinant Pap31 proteins for the diagnosis of canine Bartonelloses, rPap31-NTD yielded the highest AUC score of 0.792 (95% CI 0.688-0.895) with a sensitivity of 44% and specificity of 100% at a cutoff value of 1.198. In concordance with this finding, rPap31-NTD also had the highest AUC score of 0.747 (95% CI 0.581-0.913) among the Pap31 recombinant proteins for the diagnosis of human Bartonelloses, with 39% sensitivity and 94% specificity at a cutoff value of 1.366. Recombinant whole Pap31 (rPap31) resulted in 72% sensitivity and 61% specificity at a cutoff value of 0.215 for human Bartonelloses. Due to either low sensitivity or questionable specificity, our findings indicate that recombinant Pap31 and the selected fragments may not be appropriate diagnostic targets in detecting anti- Bartonella antibodies in Bartonella -infected dogs and humans. The findings from this study can be used to further assess the antigenicity and immunogenicity of B. henselae Pap31 as a diagnostic target.

Currently, a gold standard diagnostic test for Bartonella infection in dogs is lacking. This represents a critical limitation for the development and evaluation of new diagnostic tests, as well as for the diagnosis of, and research on, bartonellosis in dogs. This retrospective observational study aims to compare the results of commonly performed and newly-reported Bartonella spp. diagnostic tests in banked clinical specimens from 90 dogs with hemangiosarcoma (HSA) using composite reference standard (CRS) and random effects latent class analysis (RE-LCA) techniques. Samples from each dog were tested using six serological or molecular diagnostic assays, including indirect fluorescent antibody (IFA) and Western blot (WB) for the detection of antibodies in serum, and qPCR and droplet digital PCR (ddPCR) in blood and fresh frozen tissue biopsy samples (mainly splenic HSA tumors and histopathologically normal spleen or skin/adipose tissue). Bartonella infection prevalence was estimated to be 78% based on the CRS (parallel testing with all six assays), and 64% based on the RE-LCA model. The assay with the highest diagnostic accuracy was qPCR performed on fresh frozen tissue biopsy samples (sensitivity: 94% by RE-LCA and 80% by CRS; specificity: 100%). When comparing newly-reported to traditional Bartonella diagnostic assays, ddPCR was more sensitive for the detection of Bartonella DNA than qPCR when testing blood samples (36% vs. 0%, p < 0.0001). Dogs that were positive on serological assays alone with negative molecular assays were highly unlikely (<3%) to be classified as infected by the RE-LCA model. These data indicate that Bartonella spp. DNA can be PCR amplified from fresh frozen tissues from a majority of dogs with HSA using both qPCR and ddPCR, supporting the use of these methods for future controlled studies comparing the prevalence of Bartonella spp. DNA in the tissue of dogs with HSA to that of unaffected controls.

Bartonella bacilliformis (B. bacilliformis) , Bartonella henselae (B. henselae) , and Bartonella quintana (B. quintana) are bacteria known to cause verruga peruana or bacillary angiomatosis, vascular endothelial growth factor (VEGF)-dependent cutaneous lesions in humans. Given the bacteria's association with the dermal niche and clinical suspicion of occult infection by a dermatologist, we determined if patients with melanoma had evidence of Bartonella spp. infection. Within a one-month period, eight patients previously diagnosed with melanoma volunteered to be tested for evidence of Bartonella spp. exposure/infection. Subsequently, confocal immunohistochemistry and PCR for Bartonella spp. were used to study melanoma tissues from two patients. Blood from seven of the eight patients was either seroreactive, PCR positive, or positive by both modalities for Bartonella spp. exposure. Subsequently, Bartonella organisms that co-localized with VEGFC immunoreactivity were visualized using multi-immunostaining confocal microscopy of thick skin sections from two patients. Using a co-culture model, B. henselae was observed to enter melanoma cell cytoplasm and resulted in increased vascular endothelial growth factor C (VEGFC) and interleukin 8 (IL-8) production. Findings from this small number of patients support the need for future investigations to determine the extent to which Bartonella spp. are a component of the melanoma pathobiome.

Bartonella species are globally important emerging pathogens that were not known to infect animals or humans in North America prior to the human immunodeficiency virus (HIV) epidemic. Ongoing improvements in diagnostic testing modalities have allowed for the discovery of Bartonella species (spp.) DNA in blood; cerebrospinal fluid; and the skin of patients with cutaneous lesions, fatigue, myalgia, and neurological symptoms. We describe Bartonella spp. test results for participants reporting neuropsychiatric symptoms, the majority of whom reported the concurrent development of cutaneous lesions. Study participants completed a medical history, a risk factor questionnaire, and provided cutaneous lesion photographs. Bartonella spp. serology and Bartonella alpha proteobacteria enrichment blood culture/PCR were assessed. Within a 14-month period, 33 participants enrolled; 29/33 had serological and/or PCR evidence supporting Bartonella spp. infection, of whom 24 reported concurrent cutaneous lesions since neuropsychiatric symptom onset. We conclude that cutaneous lesions were common among people reporting neuropsychiatric symptoms and Bartonella spp. infection or exposure. Additional studies, using sensitive microbiological and imaging techniques, are needed to determine if, or to what extent, Bartonella spp. might contribute to cutaneous lesions and neuropsychiatric symptoms in patients.

Background: In a warmer and more globally connected Arctic, vector-borne pathogens of zoonotic importance may be increasing in prevalence in native wildlife. Recently, Bartonella henselae, the causative agent of cat scratch fever, was detected in blood collected from arctic foxes (Vulpes lagopus) that were captured and released in the large goose colony at Karrak Lake, Nunavut, Canada. This bacterium is generally associated with cats and cat fleas, which are absent from Arctic ecosystems. Arctic foxes in this region feed extensively on migratory geese, their eggs, and their goslings. Thus, we hypothesized that a nest flea, Ceratophyllus vagabundus vagabundus (Boheman, 1865), may serve as a vector for transmission of Bartonella spp., Methods: We determined the prevalence of Bartonella spp. in (i) nest fleas collected from 5 arctic fox dens and (ii) 37 surrounding goose nests, (iii) fleas collected from 20 geese harvested during arrival at the nesting grounds and (iv) blood clots from 57 adult live-captured arctic foxes. A subsample of fleas were identified morphologically as C. v. vagabundus. Remaining fleas were pooled for each nest, den, or host. DNA was extracted from flea pools and blood clots and analyzed with conventional and real-time polymerase chain reactions targeting the 16S-23S rRNA intergenic transcribed spacer region., Results: Bartonella henselae was identified in 43% of pooled flea samples from nests and 40% of pooled flea samples from fox dens. Bartonella vinsonii berkhoffii was identified in 30% of pooled flea samples collected from 20 geese. Both B. vinsonii berkhoffii (n = 2) and B. rochalimae (n = 1) were identified in the blood of foxes., Conclusions: We confirm that B. henselae, B. vinsonii berkhoffii and B. rochalimae circulate in the Karrak Lake ecosystem and that nest fleas contain B. vinsonii and B. henselae DNA, suggesting that this flea may serve as a potential vector for transmission among Arctic wildlife.

Hemangiosarcoma (HSA), a locally invasive and highly metastatic endothelial cell neoplasm, accounts for two-thirds of all cardiac and splenic neoplasms in dogs. Bartonella spp. infection has been reported in association with neoplastic and non-neoplastic vasoproliferative lesions in animals and humans. The objective of this study was to determine the prevalence of Bartonella spp. in conjunction with two other hemotropic pathogens, Babesia spp. and hemotropic Mycoplasma spp., in tissues and blood samples from 110 dogs with histopathologically diagnosed HSA from throughout the United States. This was a retrospective, observational study using clinical specimens from 110 dogs with HSA banked by the biospecimen repository of the Canine Comparative Oncology and Genomics Consortium. Samples provided for this study from each dog included: fresh frozen HSA tumor tissue (available from n = 100 of the 110 dogs), fresh frozen non-tumor tissue (n = 104), and whole blood and serum samples (n = 108 and 107 respectively). Blood and tissues were tested by qPCR for Bartonella, hemotropic Mycoplasma, and Babesia spp. DNA; serum was tested for Bartonella spp. antibodies. Bartonella spp. DNA was amplified and sequenced from 73% of dogs with HSA (80/110). In contrast, hemotropic Mycoplasma spp. DNA was amplified from a significantly smaller proportion (5%, p<0.0001) and Babesia spp. DNA was not amplified from any dog. Of the 100 HSA tumor samples submitted, 34% were Bartonella PCR positive (32% of splenic tumors, 57% of cardiac tumors, and 17% of other tumor locations). Of 104 non-tumor tissues, 63% were Bartonella PCR positive (56% of spleen samples, 93% of cardiac samples, and 63% of skin/subcutaneous samples). Of dogs with Bartonella positive HSA tumor, 76% were also positive in non-tumor tissue. Bartonella spp. DNA was not PCR amplified from whole blood. This study documented a high prevalence of Bartonella spp. DNA in dogs with HSA from geographically diverse regions of the United States. While 73% of all tissue samples from these dogs were PCR positive for Bartonella DNA, none of the blood samples were, indicating that whole blood samples do not reflect tissue presence of this pathogen. Future studies are needed to further investigate the role of Bartonella spp. in the development of HSA., Competing Interests: In conjunction with Dr. Sushama Sontakke and North Carolina State University, Edward B. Breitschwerdt, DVM holds U.S. Patent No. 7,115,385: “Media and Methods for Cultivation of Microorganisms”, which was issued October 3, 2006. He is a co-founder, shareholder and Chief Scientific Officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella species infections. This does not alter our adherence to PLOS ONE policies on sharing data and materials. All other authors declare no potential conflicts of interest.

Background: Dynamics of infection by Bartonella and Rickettsia species, which are epidemiologically associated in dogs, have not been explored in a controlled setting., Objectives: Describe an outbreak investigation of occult Bartonella spp. infection among a group of dogs, discovered after experimentally induced Rickettsia rickettsii (Rr) infection., Animals: Six apparently healthy purpose-bred Beagles obtained from a commercial vendor., Methods: Retrospective and prospective study. Dogs were serially tested for Bartonella spp. and Rr using serology, culture, and PCR, over 3 study phases: 3 months before inoculation with Rr (retrospective), 6 weeks after inoculation with Rr (retrospective), and 8 months of follow-up (prospective)., Results: Before Rr infection, 1 dog was Bartonella henselae (Bh) immunofluorescent antibody assay (IFA) seroreactive and 1 was Rickettsia spp. IFA seroreactive. After inoculation with Rr, all dogs developed mild Rocky Mountain spotted fever compatible with low-dose Rr infection, seroconverted to Rickettsia spp. within 4-11 days, and recovered within 1 week. When 1 dog developed ear tip vasculitis with intra-lesional Bh, an investigation of Bartonella spp. infection was undertaken. All dogs had seroconverted to 1-3 Bartonella spp. between 7 and 18 days after Rr inoculation. Between 4 and 8 months after Rr inoculation, Bh DNA was amplified from multiple tissues from 2 dogs, and Bartonella vinsonii subsp. berkhoffii (Bvb) DNA was amplified from 4 of 5 dogs' oral swabs., Conclusions and Clinical Importance: Vector-borne disease exposure was demonstrated in research dogs from a commercial vendor. Despite limitations, our results support the possibilities of recrudescence of chronic subclinical Bartonella spp. infection after Rr infection and horizontal direct-contact transmission between dogs., (© 2019 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.)

Blood samples collected from American black bears ( Ursus americanus ) in eastern and western North Carolina, US, were analyzed for piroplasms. Piroplasmids were detected in 17% (23/132) of the animals surveyed. We detected a Babesia spp. previously identified in North American raccoons ( Procyon lotor ) and a maned wolf ( Chrysocyon brachyurus ); prevalence was 22% (14/64) and 13% (9/68) in the mountain and coastal black bear populations, respectively. The presence of the same Babesia species in black bears, raccoons, and a maned wolf suggests piroplasms may not be host specific.

Companion vector-borne diseases (CVBDs) are an important threat for pet life, but may also have an impact on human health, due to their often zoonotic character. The importance and awareness of CVBDs continuously increased during the last years. However, information on their occurrence is often limited in several parts of the world, which are often especially affected. Latin America (LATAM), a region with large biodiversity, is one of these regions, where information on CVBDs for pet owners, veterinarians, medical doctors and health workers is often obsolete, limited or non-existent. In the present review, a comprehensive literature search for CVBDs in companion animals (dogs and cats) was performed for several countries in Central America (Belize, Caribbean Islands, Costa Rica, Cuba, Dominican Republic, El Salvador, Guatemala, Honduras, Mexico, Nicaragua, Panama, Puerto Rico) as well as in South America (Argentina, Bolivia, Brazil, Chile, Colombia, Ecuador, French Guiana, Guyana (British Guyana), Paraguay, Peru, Suriname, Uruguay, Venezuela) regarding the occurrence of the following parasitic and bacterial diseases: babesiosis, heartworm disease, subcutaneous dirofilariosis, hepatozoonosis, leishmaniosis, trypanosomosis, anaplasmosis, bartonellosis, borreliosis, ehrlichiosis, mycoplasmosis and rickettsiosis. An overview on the specific diseases, followed by a short summary on their occurrence per country is given. Additionally, a tabular listing on positive or non-reported occurrence is presented. None of the countries is completely free from CVBDs. The data presented in the review confirm a wide distribution of the CVBDs in focus in LATAM. This wide occurrence and the fact that most of the CVBDs can have a quite severe clinical outcome and their diagnostic as well as therapeutic options in the region are often difficult to access and to afford, demands a strong call for the prevention of pathogen transmission by the use of ectoparasiticidal and anti-feeding products as well as by performing behavioural changes.

Background: With the advent of more sensitive culture and molecular diagnostic testing modalities, Bartonella spp. infections have been documented in blood and/or cerebrospinal fluid specimens from patients with diverse neurological symptoms. Pediatric acute-onset neuropsychiatric syndrome (PANS) is characterized by an unusually abrupt onset of cognitive, behavioral, or neurological symptoms. Between October 2015 and January 2017, a 14-year-old boy underwent evaluation by multiple specialists for sudden-onset psychotic behavior (hallucinations, delusions, suicidal and homicidal ideation)., Methods: In March 2017, Bartonella spp. serology (indirect fluorescent antibody assays) and polymerase chain reaction (PCR) amplification, DNA sequencing, and Bartonella enrichment blood culture were used on a research basis to assess Bartonella spp. exposure and bloodstream infection, respectively. PCR assays targeting other vector-borne infections were performed to assess potential co-infections., Results: For 18 months, the boy remained psychotic despite 4 hospitalizations, therapeutic trials involving multiple psychiatric medication combinations, and immunosuppressive treatment for autoimmune encephalitis. Neurobartonellosis was diagnosed after cutaneous lesions developed. Subsequently, despite nearly 2 consecutive months of doxycycline administration, Bartonella henselae DNA was PCR amplified and sequenced from the patient's blood, and from Bartonella alphaproteobacteria growth medium enrichment blood cultures. B henselae serology was negative. During treatment with combination antimicrobial chemotherapy, he experienced a gradual progressive decrease in neuropsychiatric symptoms, cessation of psychiatric drugs, resolution of Bartonella -associated cutaneous lesions, and a return to all pre-illness activities., Conclusions: This case report suggests that B henselae bloodstream infection may contribute to progressive, recalcitrant neuropsychiatric symptoms consistent with PANS in a subset of patients., Competing Interests: Declaration of Conflicting Interests:The author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: In conjunction with Dr Sushama Sontakke and North Carolina State University, EBB, DVM holds US Patent No. 7115385; Media and Methods for cultivation of microorganisms, which was issued on October 3, 2006. He is a co-founder, shareholder, and chief scientific officer for Galaxy Diagnostics, a company that provides advanced diagnostic testing for the detection of Bartonella species infections. Dr RGM is the chief technical officer for Galaxy Diagnostics and Dr BRM is the chief medical officer. All the other authors have no potential conflicts of interest to report.

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., (© 2018 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.)

Introduction: Systemic Bartonella spp. infections are being increasingly reported in association with complex medical presentations. Individuals with frequent arthropod exposures or animal contact appear to be at risk for acquiring long standing infections with Bartonella spp., Case Report: This case report describes infections with Bartonella koehlerae and Bartonella henselae in a female veterinarian whose symptoms were predominantly rheumatologic in nature. Infection was confirmed by serology, polymerase chain reaction (PCR), enrichment blood culture, and DNA sequencing of amplified B koehlerae and B henselae DNA. Long-term medical management with antibiotics was required to achieve elimination of these infections and was accompanied by resolution of the patient's symptoms. Interestingly, the patient experienced substantial improvement in the acquired joint hypermobility mimicking Ehlers-Danlos Syndrome (EDS) type III., Conclusion: To facilitate early and directed medical interventions, systemic bartonellosis should potentially be considered as a differential diagnosis in patients with incalcitrant rheumatological symptoms and frequent arthropod exposures or extensive animal contact.

Background: Bartonella koehlerae has been recently described as a new cat- and cat fleas-associated agent of culture-negative human endocarditis. It has been also encountered in one dog from Israel and six dogs from the USA, but other clinically relevant reports involving this bacterium are lacking., Results: A 7-year-old intact male mixed dog presented with clinico-pathological signs consistent with mitral endocarditis and cutaneous hemangiosarcoma. Molecular studies revealed the presence of Bartonella koehlerae DNA in samples from blood and mitral valve tissue., Conclusions: This is the first description of B. koehlerae in Spain, corroborating that it can also be detected in dogs. Bartonella koehlerae infection should also be considered in Spain in humans and dogs presenting with clinical disease suggestive of it, such as culture-negative endocarditis.

Background: In Argentina, only very few reports are available for canine tick-borne diseases where most are related to parasitic diseases. The objective of this survey was to investigate the prevalence of tick-borne pathogens in 70 dogs from Santa Fé and Córdoba, Argentina., Methods: Microscopic blood smear examination as well as polymerase chain reaction (PCR) amplification using species-specific markers of Anaplasma, Babesia, Bartonella, Borrelia, Ehrlichia, Francisella, Mycoplasma (hemotropic group) and Rickettsia, followed by DNA sequencing were used to establish the prevalence of each infecting pathogen., Results: Blood smear analysis showed 81% (57/70) prevalence of structures morphologically compatible with hemotropic mycoplasmas. No structures resembling either piroplasms or Anaplasma/Ehrlichia were detected. Hemotropic mycoplasma species (Mycoplasma haematoparvum, Mycoplasma haemocanis and Mycoplasma suis) were the most prevalent pathogens detected with an overall prevalence of 77.1%. Anaplasma platys was detected and identified in 11 of the 70 dogs (15.7%), meanwhile two Bartonella spp. (B. clarridgeiae and an uncharacterized Bartonella sp.) and Babesia vogeli were detected at 3 and 7% prevalence, respectively., Conclusions: The work presented here describes a high molecular prevalence for hemotropic mycoplasma species in each of the five locations selected. Three Mycoplasma spp., including Mycoplasma suis, reported for the first time in dogs have been identified by DNA amplification and sequencing. This study highlights the risk that these bacterial pathogens represent for companion animals and, due to their potential zoonotic nature, also for people.

We detected Anaplasma phagocytophilum by DNA amplification in whole blood from free-ranging, hunter-killed American black bears ( Ursus americanus ) from the east coast of North Carolina, US. Molecular prevalence for Anaplasma phagocytophilum was 3% from 68 black bears. No DNA of other Anaplasma or Ehrlichia spp. was identified.

Bartonella species constitute emerging, vector-borne, intravascular pathogens that produce long-lasting bacteremia in reservoir-adapted (natural host or passive carrier of a microorganism) and opportunistic hosts. With the advent of more sensitive and specific diagnostic tests, there is evolving microbiological evidence supporting concurrent infection with one or more Bartonella spp. in more than one family member; however, the mode(s) of transmission to or among family members remains unclear. In this study, we provide molecular microbiological evidence of Bartonella henselae genotype San Antonio 2 (SA2) infection in four of six Danish family members, including a child who died of unknown causes at 14 months of age., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Ehrlichia sp. DNA was amplified from 4 Ehrlichia-seroreactive horses from Mérida, Nicaragua. Sequencing of 16S rDNA, sodB, and groEL genes indicated that the bacterium is most likely a novel Ehrlichia species. The tick vector and the potential for canine and human infection remain unknown.

This report describes a 2-year-old collie dog with pulmonary nodules, visualized by computed tomographic (CT) scan, with evidence of Bartonella henselae bacteremia and pyogranulomatous lymphadenitis. Clinical signs resolved with antimicrobial therapy.

Anaplasma platys (Apl), 'Candidatus Mycoplasma haemominutum' (CMh), Bartonella henselae (Bh) and Bartonella koehlerae (Bk) were confirmed by polymerase chain reaction (PCR) amplification and DNA sequencing in a cat diagnosed with multiple myeloma. Other inconsistently documented hematologic abnormalities included anemia, thrombocytopenia, eosinophilia and hypoglycemia. Persistent Apl infection was confirmed for the first time in a North American cat by sequencing three bacterial genes (16S rRNA, p44 and GroEL) in peripheral blood samples collected 100 days apart. Following doxycycline treatment for Apl, multiple myeloma was diagnosed based upon a monoclonal gammopathy and splenic plasmacytosis, and the cat was treated with melphalan, chlorambucil and prednisolone. Apl DNA was not amplified from post-treatment blood samples and the hyperglobulinemia resolved temporarily following chemotherapy. Retrospective PCR analysis of stored DNA extracts identified CMh, Bk and Bh infections. Retrospective PCR for antigen receptor rearrangements (PARR) of splenic aspirates did not confirm B- or T-cell clonality. Co-infection with multiple vector-borne pathogens should be a diagnostic consideration in cats with chronic hypergammaglobulinemia, monoclonal gammopathy and splenic plasmacytosis., (© ISFM and AAFP 2014.)

Background: Anaplasmosis, caused by Anaplasma phagocytophilum and Anaplasma platys, and ehrlichiosis, caused by Ehrlichia chaffeensis, Ehrlichia ewingii, the "Panola Mountain Ehrlichia" and Ehrlichia muris-like pathogens have been identified as emerging tick borne infectious diseases in dogs and human patients. Persistent intravascular infection with these bacteria is well documented in dogs, but is less well documented in human beings., Methods: Serology and PCR targeting multiple microbial genes, followed by DNA sequencing, was used to test sequential blood samples. Tissue culture isolation was attempted in two laboratories., Results: A. platys, E. chaffeensis, and E. ewingii DNA was amplified from two Anaplasma and Ehrlichia seronegative family members and their dog, all lacking typical symptoms of anaplasmosis or ehrlichiosis. Following treatment with doxycycline, the dog and mother were Anaplasma and Ehrlichia spp. PCR negative., Conclusions: Sequential PCR testing provided molecular evidence supporting intravascular persistence of A. platys and Ehrlichia spp. in two humans and their dog. Diagnosticians and clinicians should consider the potential for co-infections due to these tick borne organisms.

Background: Canine vector-borne diseases (CVBD) are caused by a diverse array of pathogens with varying biological behaviors that result in a wide spectrum of clinical presentations and laboratory abnormalities. For many reasons, the diagnosis of canine vector-borne infectious diseases can be challenging for clinicians. The aim of the present study was to compare CVBD serological and molecular testing as the two most common methodologies used for screening healthy dogs or diagnosing sick dogs in which a vector-borne disease is suspected., Methods: We used serological (Anaplasma species, Babesia canis, Bartonella henselae, Bartonella vinsonii subspecies berkhoffii, Borrelia burgdorferi, Ehrlichia canis, and SFG Rickettsia) and molecular assays to assess for exposure to, or infection with, 10 genera of organisms that cause CVBDs (Anaplasma, Babesia, Bartonella, Borrelia, Ehrlichia, Francisella, hemotropic Mycoplasma, Neorickettsia, Rickettsia, and Dirofilaria). Paired serum and EDTA blood samples from 30 clinically healthy dogs (Group I) and from 69 sick dogs suspected of having one or more canine vector-borne diseases (Groups II-IV), were tested in parallel to establish exposure to or infection with the specific CVBDs targeted in this study., Results: Among all dogs tested (Groups I-IV), the molecular prevalences for individual CVBD pathogens ranged between 23.3 and 39.1%. Similarly, pathogen-specific seroprevalences ranged from 43.3% to 59.4% among healthy and sick dogs (Groups I-IV). Among these representative sample groupings, a panel combining serological and molecular assays run in parallel resulted in a 4-58% increase in the recognition of exposure to or infection with CVBD., Conclusions: We conclude that serological and PCR assays should be used in parallel to maximize CVBD diagnosis.

Background: Hemotropic mycoplasmas are epicellular erythrocytic bacteria that can cause infectious anemia in some mammalian species. Worldwide, hemotropic mycoplasmas are emerging or re-emerging zoonotic pathogens potentially causing serious and significant health problems in wildlife. The objective of this study was to determine the molecular prevalence of hemotropic Mycoplasma species in little brown bats (Myotis lucifugus) with and without Pseudogymnoascus (Geomyces) destrucans, the causative agent of white nose syndrome (WNS) that causes significant mortality events in bats., Methods: In order to establish the prevalence of hemotropic Mycoplasma species in a population of 68 little brown bats (Myotis lucifugus) with (n = 53) and without (n = 15) white-nose syndrome (WNS), PCR was performed targeting the 16S rRNA gene., Results: The overall prevalence of hemotropic Mycoplasmas in bats was 47%, with similar (p = 0.5725) prevalence between bats with WNS (49%) and without WNS (40%). 16S rDNA sequence analysis (~1,200 bp) supports the presence of a novel hemotropic Mycoplasma species with 91.75% sequence homology with Mycoplasma haemomuris. No differences were found in gene sequences generated from WNS and non-WNS animals., Conclusions: Gene sequences generated from WNS and non-WNS animals suggest that little brown bats could serve as a natural reservoir for this potentially novel Mycoplasma species. Currently, there is minimal information about the prevalence, host-specificity, or the route of transmission of hemotropic Mycoplasma spp. among bats. Finally, the potential role of hemotropic Mycoplasma spp. as co-factors in the development of disease manifestations in bats, including WNS in Myotis lucifugus, remains to be elucidated.

PCR amplification targeting the 16S rRNA gene was used to test individuals with and without extensive arthropod and animal contact for the possibility of hemotropic mycoplasma infection. The prevalence of hemotropic mycoplasma infection (4.7%) was significantly greater in previously reported cohorts of veterinarians, veterinary technicians, spouses of veterinary professionals, and others with extensive arthropod exposure and/or frequent animal contact than in a previously reported cohort of patients examined by a rheumatologist because of chronic joint pain or evidence of small-vessel disease (0.7%). Based upon DNA sequence analysis, a Mycoplasma ovis-like species was the most prevalent organism detected; however, infection with "Candidatus Mycoplasma haematoparvum" and a potentially novel, but incompletely characterized, hemotropic Mycoplasma species was also documented. Historical exposure to animals and arthropod vectors that can harbor hemotropic Mycoplasma spp. should be considered during epidemiological investigations and in the evaluation of individual patients.

Bed bugs (Cimex lectularius L.) have resurged in the United States and globally. Bed bugs are hematophagous ectoparasites of humans and other animals, including domestic pets, chickens, and bats, and their blood feeding habits contribute to their potential as disease vectors. Several species of Bartonella are re-emergent bacterial pathogens that also affect humans, domestic pets, bats and a number of other wildlife species. Because reports of both bed bugs and Bartonella have been increasing in the U.S., and because their host ranges can overlap, we investigated whether the resurgences of these medically important pathogens and their potential vector might be linked, by screening for Bartonella spp. in bed bugs collected from geographic areas where these pathogens are prevalent and from bed bugs that have been in culture in the laboratory for several years. We screened a total of 331 bed bugs: 316 bed bugs from 36 unique collections in 29 geographic locations in 13 states, 10 bed bugs from two colonies maintained in the laboratory for 3 yr, and 5 bed bugs from a colony that has been in culture since before the recent resurgence of bed bugs. Bartonella spp. DNA was screened using a polymerase chain reaction assay targeting the 16S-23S rRNA intergenic transcribed spacer region. Bartonella DNA was not amplified from any bed bug, but five bed bugs from four different apartments of an elderly housing building in North Carolina contained DNA sequences that corresponded to Burkholderia multivorans, an important pathogen in nosocomial infections that was not previously linked to an arthropod vector.

Background: Bartonella alpha-Proteobacteria growth medium (BAPGM) enrichment culture has proven useful for documenting Bartonella species infection and has facilitated growth of other fastidious bacteria from human samples., Purpose: To report non-Bartonella bacterial isolates obtained from canine samples cultured using BAPGM enrichment culture., Animals: Between 2004 and 2008, 695 specimens from 513 dogs were tested by the NCSU-IPRL using the BAPGM enrichment culture. Over the same period of time, blood samples from 270 dogs were cultured by the NCSU-CML using Bactec-Plus Aerobic/F media., Methods: BAPGM isolates were characterized using Bartonella genus primers and 16S rDNA primers followed by DNA sequencing. NCSU medical records were retrospectively reviewed. Blood culture results from the NCSU-CML were compared with BAPGM blood culture results., Results: Seventy-nine non-Bartonella isolates were obtained from 69/513 dogs. The most commonly isolated phylum was Proteobacteria (48.1%) with alpha-Proteobacteria being the most commonly isolated class. Staphylococcus and Sphingomonas were the most commonly isolated genera. The majority of the remaining isolates were bacteria that are rarely isolated from canine samples. Comparison of NCSU-CML and IPRL (BAPGM) blood culture isolates showed alpha-Proteobacteria were isolated more often from BAPGM., Conclusions and Clinical Importance: Use of insect cell culture enrichment medium, such as BAPGM, appears to enhance the growth of alpha-Proteobacteria, but also results in isolation of non-alpha-Proteobacteria from sick dogs. Future studies are needed to elucidate the utility of BAPGM and other "nonconventional" growth media and methods for isolation of fastidious organisms and to determine if these organisms play a causal role in disease development., (Copyright © 2013 by the American College of Veterinary Internal Medicine.)

Patients with infection from bacteremic Bartonella spp., tested using Bartonella Alphaproteobacteria growth medium (BAPGM), were retrospectively categorized into one of two groups that included those whose blood was collected once (group 1; n = 55) or three times (group 2; n = 36) within a 1-week period. Overall, 19 patients (20.8%) were PCR positive for one or more Bartonella spp. using the BAPGM platform. Seven patients (12.7%) in group 1 tested positive, and 12 patients (33.3%) in group 2 tested positive. Detection was improved when the patients were tested three times within a 1-week period (odds ratio, 3.4 [95% confidence interval, 1.2 to 9.8]; P = 0.02). Obtaining three sequential blood samples during a 1-week period should be considered a diagnostic approach when bartonellosis is suspected.

Here, we report latent infections with Bartonella quintana and a hemotropic Mycoplasma sp. in a research colony of cynomolgus monkeys (Macaca fascicularis). Sequence alignments, evolutionary analysis, and signature nucleotide sequence motifs of the hemotropic Mycoplasma 16S rRNA and RNase P genes indicate the presence of a novel organism.

Background: Bartonella species comprise a group of zoonotic pathogens that are usually acquired by vector transmission or by animal bites or scratches., Methods: PCR targeting the Bartonella 16S-23S intergenic spacer (ITS) region was used in conjunction with BAPGM (Bartonella alpha Proteobacteria growth medium) enrichment blood culture to determine the infection status of the family members and to amplify DNA from spiders and woodlice. Antibody titers to B. vinsonii subsp. berkhoffii (Bvb) genotypes I-III, B. henselae (Bh) and B. koehlerae (Bk) were determined using an IFA test. Management of the medical problems reported by these patients was provided by their respective physicians., Results: In this investigation, immediately prior to the onset of symptoms two children in a family experienced puncture-like skin lesions after exposure to and presumptive bites from woodlouse hunter spiders. Shortly thereafter, the mother and both children developed hive-like lesions. Over the ensuing months, the youngest son was diagnosed with Guillain-Barre (GBS) syndrome followed by Chronic Inflammatory Demyelinating Polyradiculoneuropathy (CIDP). The older son developed intermittent disorientation and irritability, and the mother experienced fatigue, headaches, joint pain and memory loss. When tested approximately three years after the woodlouse hunter spider infestation, all three family members were Bartonella henselae seroreactive and B. henselae DNA was amplified and sequenced from blood, serum or Bartonella alpha-proteobacteria (BAPGM) enrichment blood cultures from the mother and oldest son. Also, B. henselae DNA was PCR amplified and sequenced from a woodlouse and from woodlouse hunter spiders collected adjacent to the family's home., Conclusions: Although it was not possible to determine whether the family's B. henselae infections were acquired by spider bites or whether the spiders and woodlice were merely accidental hosts, physicians should consider the possibility that B. henselae represents an antecedent infection for GBS, CIDP, and non-specific neurocognitive abnormalities.

Background: Bartonella henselae is a zoonotic, alpha Proteobacterium, historically associated with cat scratch disease (CSD), but more recently associated with persistent bacteremia, fever of unknown origin, arthritic and neurological disorders, and bacillary angiomatosis, and peliosis hepatis in immunocompromised patients. A family from the Netherlands contacted our laboratory requesting to be included in a research study (NCSU-IRB#1960), designed to characterize Bartonella spp. bacteremia in people with extensive arthropod or animal exposure. All four family members had been exposed to tick bites in Zeeland, southwestern Netherlands. The mother and son were exhibiting symptoms including fatigue, headaches, memory loss, disorientation, peripheral neuropathic pain, striae (son only), and loss of coordination, whereas the father and daughter were healthy., Methods: Each family member was tested for serological evidence of Bartonella exposure using B. vinsonii subsp. berkhoffii genotypes I-III, B. henselae and B. koehlerae indirect fluorescent antibody assays and for bacteremia using the BAPGM enrichment blood culture platform., Results: The mother was seroreactive to multiple Bartonella spp. antigens and bacteremia was confirmed by PCR amplification of B. henselae DNA from blood, and from a BAPGM blood agar plate subculture isolate. The son was not seroreactive to any Bartonella sp. antigen, but B. henselae DNA was amplified from several blood and serum samples, from BAPGM enrichment blood culture, and from a cutaneous striae biopsy. The father and daughter were seronegative to all Bartonella spp. antigens, and negative for Bartonella DNA amplification., Conclusions: Historically, persistent B. henselae bacteremia was not thought to occur in immunocompetent humans. To our knowledge, this study provides preliminary evidence supporting the possibility of persistent B. henselae bacteremia in immunocompetent persons from Europe. Cat or flea contact was considered an unlikely source of transmission and the mother, a physician, reported that clinical symptoms developed following tick exposure. To our knowledge, this is the first time that a B. henselae organism has been visualized in and amplified from a striae lesion. As the tick bites occurred three years prior to documentation of B. henselae bacteremia, the mode of transmission could not be determined.

Background: During a two year period, a 27-year-old female veterinarian experienced migraine headaches, seizures, including status epilepticus, and other neurological and neurocognitive abnormalities. Prior to and during her illness, she had been actively involved in hospital-based work treating domestic animals, primarily cats and dogs, in Grenada and Ireland and anatomical research requiring the dissection of wild animals (including lions, giraffe, rabbits, mongoose, and other animals), mostly in South Africa. The woman reported contact with fleas, ticks, lice, biting flies, mosquitoes, spiders and mites and had also been scratched or bitten by dogs, cats, birds, horses, reptiles, rabbits and rodents. Prior diagnostic testing resulted in findings that were inconclusive or within normal reference ranges and no etiological diagnosis had been obtained to explain the patient's symptoms., Methods: PCR assays targeting Anaplasma sp. Bartonella sp. and hemotopic Mycoplasma sp. were used to test patient blood samples. PCR positive amplicons were sequenced directly and compared to Gen Bank sequences. In addition, Bartonella alpha Proteobacteria growth medium (BAPGM) enrichment blood culture was used to facilitate bacterial growth and Bartonella spp. serology was performed by indirect fluorescent antibody testing., Results: Anaplasma platys, Bartonella henselae and Candidatus Mycoplasma haematoparvum DNA was amplified and sequenced from the woman's blood, serum or blood culture samples. Her serum was variably seroreactive to several Bartonella sp. antigens. Despite symptomatic improvement, six months of doxycycline most likely failed to eliminate the B. henselae infection, whereas A. platys and Candidatus M. haematoparvum DNA was no longer amplified from post-treatment samples., Conclusions: As is typical of many veterinary professionals, this individual had frequent exposure to arthropod vectors and near daily contact with persistently bacteremic reservoir hosts, including cats, the primary reservoir host for B. henselae, and dogs, the presumed primary reservoir host for A. platys and Candidatus Mycoplasma haematoparvum. Physicians caring for veterinarians should be aware of the occupational zoonotic risks associated with the daily activities of these animal health professionals.

Bartonella vinsonii subsp. berkhoffii has not been detected previously in white-tailed deer (Odocoileus virginianus). We tested whole blood from 60 white-tailed deer for Bartonella spp. DNA; three (5%) were positive for Bartonella vinsonii subsp. berkhoffii. This is the first detection of Bartonella vinsonii subsp. berkhoffii in white-tailed deer.