Bartonella DNA Research Articles

Wild rodent fleas carrying Bartonella and Rickettsia in an area endemic for vector-borne diseases from Argentina.

Vector-borne diseases account for nearly 20% of all globally recognised infectious diseases. Within the spectrum of flea-borne pathogens, Bartonella and Rickettsia bacteria are prominent, contributing to the emergence and resurgence of diseases on a global scale. This study investigates the presence of species of Bartonella and Rickettsia harboured by fleas collected from wild rodents in northwestern Argentina (NWA). A total of 28 fleas from three genera and seven species were assessed. DNA of Bartonella and Rickettsia spp. was found in 12 fleas (42.8%). Phylogenetic analysis of concatenated sequences of gltA and rpoB genes showed the presence of Bartonella quintana in eight fleas of two species, Craneopsylla minerva minerva and Polygenis acodontis. Phylogenetic analysis of concatenated sequences of gltA, ompA and ompB genes identified Rickettsia felis in ten fleas of five species, C. m. minerva, P. acodontis, Polygenis bohlsi bohlsi, Polygenis byturus and Tiamastus palpalis. These bacterial species mark the first report in all flea species studied. This study represents the first survey of flea-borne bacteria for NWA. The results provide information to address strategies for the control and prevention of bartonellosis and rickettsiosis that could have an impact on public health in one of the geographical areas of Argentina with the highest incidence of infections transmitted to humans by ectoparasites.

Bartonella spp. in Phlebotominae Sand Flies, Brazil.

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.

Prevalence and genetic diversity of Bartonella spp. in wild small mammals from South Africa.

Bartonella spp. are intracellular bacteria associated with several re-emerging human diseases. Small mammals play a significant role in the maintenance and spread of Bartonella spp. Despite the high small mammal biodiversity in South Africa, there is limited epidemiological information regarding Bartonella spp. in these mammals. The main aim of this study was to determine the prevalence and genetic diversity of Bartonella spp. from wild small mammals from 15 localities in 8 provinces of South Africa. Small mammals (n = 183) were trapped in the Eastern Cape, Free State, Gauteng, Limpopo, Mpumalanga, Northern Cape, North West, and Western Cape provinces of South Africa between 2010 and 2018. Heart, kidney, liver, lung, and spleen were harvested for Bartonella DNA screening, and prevalence was determined based on the PCR amplification of partial fragments of the 16S-23S rRNA intergenic spacer (ITS) region, gltA, and rpoB genes. Bartonella DNA was detected in Aethomys chrysophilus, Aethomys ineptus, Gerbillurus spp., Lemniscomys rosalia, Mastomys coucha, Micaelamys namaquensis, Rhabdomys pumilio, and Thallomys paedulcus. An overall prevalence of 16.9% (31/183, 95% CI: 12.2%-23%) was observed. Bartonella elizabethae, Bartonella grahamii, and Bartonella tribocorum were the zoonotic species identified, while the remaining sequences were aligned to uncultured Bartonella spp. with unknown zoonotic potential. Phylogenetic analyses confirmed five distinct Bartonella lineages (I-V), with lineage IV displaying strong M. coucha host specificity. Our results confirm that South African wild small mammals are natural reservoirs of a diverse assemblage of Bartonella spp., including some zoonotic species with high genetic diversity, although prevalence was relatively low.IMPORTANCESmall mammals play a significant role in the maintenance and spread of zoonotic pathogens such as Bartonella spp. Despite the high small mammal biodiversity in southern Africa including South Africa, there is limited epidemiological information regarding Bartonella spp. in these mammals across the country. Results from our study showed the liver and spleen had the highest positive cases for Bartonella spp. DNA among the tested organs. Bartonella elizabethae, B. grahamii, and B. tribocorum were the three zoonotic species identified and five distinct Bartonella lineages (I-V) were confirmed through phylogenetic analyses. To the best of our knowledge, this study presents the first extensive nuclear diversity investigation of Bartonella spp. in South African small mammals in South Africa.

No evidence of Bartonella infections in host-seeking Ixodes scapularis and Ixodes pacificus ticks in the United States

BackgroundBartonella spp. infect a variety of vertebrates throughout the world, with generally high prevalence. Several Bartonella spp. are known to cause diverse clinical manifestations in humans and have been recognized as emerging pathogens. These bacteria are mainly transmitted by blood-sucking arthropods, such as fleas and lice. The role of ticks in the transmission of Bartonella spp. is unclear.MethodsA recently developed quadruplex polymerase chain reaction (PCR) amplicon next-generation sequencing approach that targets Bartonella-specific fragments on gltA, ssrA, rpoB, and groEL was applied to test host-seeking Ixodes scapularis ticks (n = 1641; consisting of 886 nymphs and 755 adults) collected in 23 states of the eastern half of the United States and Ixodes pacificus ticks (n = 966; all nymphs) collected in California in the western United States for the presence of Bartonella DNA. These species were selected because they are common human biters and serve as vectors of pathogens causing the greatest number of vector-borne diseases in the United States.ResultsNo Bartonella DNA was detected in any of the ticks tested by any target.ConclusionsOwing to the lack of Bartonella detection in a large number of host-seeking Ixodes spp. ticks tested across a broad geographical region, our results strongly suggest that I. scapularis and I. pacificus are unlikely to contribute more than minimally, if at all, to the transmission of Bartonella spp.Graphical

Epidemiological survey and genetic diversity of Bartonella in fleas collected from rodents in Fujian Province, Southeast China

BackgroundFleas, considered to be the main transmission vectors of Bartonella, are highly prevalent and show great diversity. To date, no investigations have focused on Bartonella vectors in Southeast China. The aim of this study was to investigate the epidemiological and molecular characteristics of Bartonella in fleas in Southeast China.MethodsFrom 2016 to 2022, flea samples (n = 1119) were collected from 863 rodent individuals in seven inland and coastal cities in Southeast China. Flea species, region, gender, host species and habitat were recorded. The DNA samples from each individual flea were screened by real-time PCR for the Bartonella ssrA gene. All positive samples were confirmed by PCR based on the presence of the gltA gene and sequenced. The factors associated with Bartonella infection were analyzed by the Chi-square test and Fisher's exact test. ANOVA and the t-test were used to compare Bartonella DNA load.ResultsBartonella DNA was detected in 26.2% (293/1119) of the flea samples, including in 27.1% (284/1047) of Xenopsylla cheopis samples, 13.2% (5/38) of Monopsyllus anisus samples, 8.3% (2/24) of Leptopsylla segnis samples and 20.0% (2/10) of other fleas (Nosopsyllus nicanus, Ctenocephalides felis, Stivalius klossi bispiniformis and Neopsylla dispar fukienensis). There was a significant difference in the prevalence of Bartonella among flea species, sex, hosts, regions and habitats. Five species of Bartonella fleas were identified based on sequencing and phylogenetic analyses targeting the gltA gene: B. tribocorum, B. queenslandensis, B. elizabethae, B. rochalimae and B. coopersplainsensis.ConclusionsThere is a high prevalence and diversity of Bartonella infection in the seven species of fleas collected in Southeast China. The detection of zoonotic Bartonella species in this study, including B. tribocorum, B. elizabethae and B. rochalimae, raises public health concerns.Graphical

Molecular detection of Bartonella spp. in bat-associated macronyssid mites (Acari: Macronyssidae) from Southern and Southeastern Brazil.

Despite the worldwide occurrence and high genetic diversity of Bartonella spp. in bats, few studies investigate their occurrence in bat-associated mites. To date, 26 species of Macronyssidae mite species have been reported from Brazil, and 15 of which were found parasitizing bats. The present study aimed to investigate the presence of Bartonella DNA in bat-associated macronyssid mites from Brazil. For this purpose, 393 macronyssid specimens were selected by convenience from the tissue bank of the Acari Collection of the Instituto Butantan (IBSP). These mites were collected from 14 different bat species in three different Brazilian States (Minas Gerais, Paraná, and Rio de Janeiro). Out of 165 mites positive in the PCR for the endogenous 18S rRNA gene, only eight were positive in the qPCR for Bartonella spp. based on the nuoG gene, and we were able to obtain two sequences base in this same gene, and one sequence based on the 16S rRNA gene. The phylogenetic inference based on the nuoG gene grouped the obtained sequences with Bartonella genotypes previously detected in bats and associated bat flies, while the phylogeny based on the 16S rRNA grouped the obtained sequence in the same clade of Bartonella genotypes previously detected in Dermanyssus gallinae. These findings suggest that macronyssid mites might be associated with the maintenance of bartonellae among bats.

Pradofloxacin for Treatment of Bartonella henselae in Experimentally Inoculated Cats.

Bartonella henselae is associated with numerous clinical syndromes in people. Cats are the definitive hosts for B. henselae, develop high levels of bacteremia, and are associated with human infections, particularly in the presence of Ctenocephalides felis. Several antibiotic protocols used for the treatment of B. henselae infection in cats have failed to clear bacteremia. The purpose of this study was to assess the safety and efficacy of a high-dose pradofloxacin protocol to eliminate B. henselae bacteremia. Bartonella henselae infection was initiated in 8 cats by intravenous inoculation of infected feline blood and then pradofloxacin was administered at 7.5 mg/kg, PO, twice daily for 28 days, starting 12 weeks after inoculation. Complete blood cell counts were performed prior to pradofloxacin administration and then every 2 weeks for 10 weeks. Bartonella PCR assay was performed prior to pradofloxacin administration and approximately every 2 weeks for 10 weeks and then weekly for 4 weeks. Methylprednisolone acetate (5 mg/kg) was administered by intramuscular injection to all cats on week 10. The cats remained normal and none developed a hematocrit, platelet count, lymphocyte count, or neutrophil count outside of the normal reference ranges. In the one month prior to pradofloxacin administration, all cats were PCR-positive for Bartonella DNA on at least two of four sample dates; after pradofloxacin administration, all cats were negative for B. henselae DNA in blood on all nine sample dates. The protocol appears to be safe and failure to amplify B. henselae DNA from the blood after the administration of pradofloxacin and one dose of methylprednisolone acetate suggests either an antibiotic effect or the organism was cleared spontaneously.

Diversity of Flea Species (Siphonaptera) and Their Vector-Borne Pathogens from Bats (Chiroptera) in Lithuania

Bat fleas (Insecta: Siphonaptera: Ischnopsyllidae) are highly host-specific ectoparasites distributed worldwide (except Antarctica). The identification of flea species and the detection of flea-borne pathogens plays a significant role not only in veterinary and public health, but also in providing insights into the ecology, behaviour, and geographical distribution of fleas and flea-associated pathogens. The present study aimed to conduct a morphological and molecular identification of fleas collected from Lithuanian bats, as well as to screen for the presence of vector-borne pathogens. A total of 58 flea specimens were collected from bats in Lithuania. Based on morphological analysis, seven flea species were identified (Ischnopsyllus variabilis, I. octactenus, I. simplex, I. elongatus, I. hexactenus, Nycteridopsylla pentactena, and N. eusarca) and screened for the presence of Anaplasma spp., Borrelia spp., Babesia spp., Bartonella spp., Rickettsia spp., and Mycoplasma spp. Molecular analysis of fleas’ COI and 18S rRNA genes showed that the flea species collected from bats are closely genetically related. Bartonella DNA was detected in bat fleas (41.4%). No DNA of Anaplasma, Borrelia, Babesia, Rickettsia, and Mycoplasma was amplified in the tested samples. The Wolbachia sp. was accidentally found in I. elongatus. The phylogenetic analysis of four Bartonella genes (16S-23S rRNA ITS, gltA, rpoB, and groEL) revealed specificity to bats or bats’ ectoparasites. This is the first report about bat flea species and the pathogens in bat fleas in Lithuania. New host records for bat fleas were also reported. This study extends the knowledge about bat fleas and their role as vectors of different pathogens.

Molecular detection and characterization of Bartonella spp. in small mammals in the Amazonia and Cerrado biomes, midwestern Brazil

Although Bartonella spp. have been worldwide described in rodents and bats, few studies have reported these agents in marsupials. The present work aimed to investigate the occurrence and genetic diversity of Bartonella in small mammals (rodents, marsupials, and bats) and associated ectoparasites in two ecoregions (Amazonia and Cerrado biomes) in midwestern Brazil. For this purpose, DNA samples from 378 specimens of small mammals (128 rodents, 111 marsupials, and 139 bats) and 41 fleas (Siphonaptera) were screened for the Bartonella genus employing a quantitative real-time PCR assay (qPCR) based on the nuoG (nicotinamide adenine dinucleotide dehydrogenase gamma subunit) gene. Then, positive samples in qPCR were submitted to conventional PCR (cPCR) assays targeting the gltA, ftsZ, and rpoB genes. One (0.78 %) rodent, 23 (16.54 %) bats, and 3 (7.31 %) fleas showed positive results in the qPCR for Bartonella sp. After cPCR amplification and sequencing, 13 partial Bartonella DNA sequences of the following genes were obtained only from bats´ blood samples: 9 gltA (citrate synthase), 3 ftsZ (cell division protein), and 1 rpoB (RNA polymerase beta subunit). The maximum likelihood inference based on the gltA gene positioned the obtained sequences in three different clades, closely related to Bartonella genotypes previously detected in other bat species and bat flies sampled in Brazil and other countries from Latin America. Similarly, the ftsZ sequences clustered in two different clades with sequences described in bats from Brazil, other countries from Latin America, and Georgia (eastern Europe). Finally, the Bartonella rpoB from a specimen of Lophostoma silvicolum clustered with a Bartonella sp. sequence obtained from a Noctilio albiventris (KP715475) from French Guiana. The present study provided valuable insights into the diversity of Bartonella genotypes infecting bats from two ecoregions (Amazonia and Cerrado) in midwestern Brazil and emphasized that further studies should be conducted regarding the description and evaluation of different lineages of Bartonella in wild small mammals and their ectoparasites in different Brazilian biomes.

Detection and characterization of zoonotic Bartonella spp. in rodents and shrews ectoparasites from Kigoma and Morogoro regions, Tanzania

Abstract Bartonellosis is a vector-borne disease which is increasingly threatening the health of humans and animals worldwide consequent to the growing wildlife-animals-human interactions. Little is known about the epidemiology of this disease in Tanzania. In this study we investigated and characterized Bartonella species in small mammals’ ectoparasites from potentially high-risk areas in the country. A total of 141 ectoparasites pools of mites, fleas, ticks, and lice were analyzed using conventional PCR and sequencing. Bartonella DNA was detected in 34.8 % of the tested ectoparasite pools, with mites at 32.9 %, fleas at 40 %, ticks at 12.5 %, and lice at 50 %. Phylogenetic analysis showed that the Bartonella spp. genotypes were closely related to those found in Uganda, Kenya, and South Africa. Different genotypes with independent haplotypes were observed, although most Bartonella spp. from fleas shared the same haplogroup. The confirmed presence of Bartonella elizabethae and Bartonella tribocorum in field and house rodents emphasizes the prevailing transmission risk of zoonotic infections in the study areas and beyond. Screening of humans, companion animals, and livestock in potentially high-risk areas in Tanzania is necessary in order to inform the development of responsive surveillance and control strategies.

Molecular survey and phylogenetic analysis of Bartonella sp., Coxiella sp., and hemoplamas in pudu (Pudu puda) from Chile: first report of Bartonella henselae in a wild ungulate species.

Recent evidence shows a high diversity of infectious agents in wildlife that represent a threat to human, domestic, and wild animal health. In Chile, wild populations of the most common cervid species, pudu (Pudu puda), have been reported as hosts for novel pathogens such as Mycoplasma ovis-like and a novel ecotype of Anaplasma phagocytophilum. A better understanding of the epidemiology of this group and other intracellular bacteria that might have cervids as hosts would enlighten their population relevance. This study aimed to determine the occurrence and genetic diversity of Bartonella spp., hemotropic mycoplasmas, and Coxiella burnetii in pudus from Chile. The DNA was extracted from the blood samples of 69 wild free-ranging and 30 captive pudus from Chile. A combination of real-time (nouG gene for Bartonella and IS1111 element for C. burnetii) and conventional PCR (16S rRNA for hemotropic Mycoplasma spp. and rpoB, gltA, and ITS for Bartonella spp.) was used for pathogen screening and molecular characterization. DNA of Bartonella spp. was detected in 10.1% [95% CI (5.2-18.2%)] samples, hemotropic Mycoplasma spp. in 1.7% [95% CI (0.08-10.1%)], and C. burnetii in 1.0% [95% CI (0.05-6.3%)] samples. Two sequenced samples were identified as Mycoplasma ovis-like, and one free-ranging pudu was positive for C. burnetii. While one captive and two free-ranging pudus were positive for Bartonella henselae, one wild pudu was co-positive for B. henselae and Bartonella sp., similar to Bartonellae identified in ruminants. To the best of our knowledge, this is the first report of B. henselae in wild ungulate species, and C. burnetii and Bartonella spp. in wild ungulate species in South America. Further research will be necessary to evaluate the potential role of pudu as reservoirs of infection and identify the sources for disease transmission among humans and wild and domestic animals.

Detection of Bartonella in kissing bugs Triatoma rubrofasciata collected from Huizhou City, South China

BackgroundThe blood-feeding behavior of kissing bugs (subfamily Triatominae, family Reduviidae, order Hemiptera) means they are potential vectors of multiple humans pathogens. However, investigations of vector-borne pathogens harbored by kissing bugs are rare. MethodsIn the current study, 22 adult kissing bugs (Triatoma rubrofasciata) were captured in Huizhou City, Guangdong Province, south China. The presence of vector-borne pathogens in the kissing bugs was tested, and the genetic diversity of these potential pathogens was investigated. ResultsAll the kissing bugs were negative for Anaplasmataceae bacteria, Rickettsia, and Coxiella. Bartonella DNA was detected in 36.4% (8/22) of the kissing bugs. The sequences of the Bartonella gltA genes divided into two clades in a phylogenetic tree, with close relationships to B. tribocorum and uncultured Bartonella sp. clone MYR-283, respectively. All the groEL sequences were closely related to those of B. kosoyi (identity 98.75%–100%). The ftsZ and rpoB sequences were most closely related to those of B. elizabethae, a recognized human pathogen, with nucleotide similarities of 98.70%–100% and 99.45%–100%, respectively. ConclusionsWe report the detection of Bartonella DNA in Triatoma kissing bugs in southern China. Although the sample size is limited, the high positive rate of detection of Bartonella DNA, the close relationship of the gene sequences to those of zoonotic Bartonella species, and the distribution of the kissing bugs near human residences, hint at a risk to public health.

Bartonella Infection in Fruit Bats and Bat Flies, Bangladesh.

Bats harbor diverse intracellular Bartonella bacteria, but there is limited understanding of the factors that influence transmission over time. Investigation of Bartonella dynamics in bats could reveal general factors that control transmission of multiple bat-borne pathogens, including viruses. We used molecular methods to detect Bartonella DNA in paired bat (Pteropus medius) blood and bat flies in the family Nycteribiidae collected from a roost in Faridpur, Bangladesh between September 2020 and January 2021. We detected high prevalence of Bartonella DNA in bat blood (35/55, 64%) and bat flies (59/60, 98%), with sequences grouping into three phylogenetic clades. Prevalence in bat blood increased over the study period (33% to 90%), reflecting an influx of juvenile bats in the population and an increase in the prevalence of bat flies. Discordance between infection status and the clade/genotype of detected Bartonella was also observed in pairs of bats and their flies, providing evidence that bat flies take blood meals from multiple bat hosts. This evidence of bat fly transfer between hosts and the changes in Bartonella prevalence during a period of increasing nycteribiid density support the role of bat flies as vectors of bartonellae. The study provides novel information on comparative prevalence and genetic diversity of Bartonella in pteropodid bats and their ectoparasites, as well as demographic factors that affect Bartonella transmission and potentially other bat-borne pathogens.

Detection of Bartonella vinsonii, Anaplasma platys and Bartonella sp. in didelphis marsupialis, Pecari tajacu and Chelonoidis denticulate: Peru

IntroductionEvidence suggest that wildlife Infectious diseases related to wildlife are of most importance because of the agents’ capacity to spill over into humans from the wild reservoir. Among them, the bacteria Bartonella spp. and Anaplasma spp. are related to this zoonotic dynamic.ObjectiveThe primary goal of the present study was to determine the presence of pathogenic bacteria in kidney and liver tissues of Didelphis marsupialis; spleen, liver, and skin of Pecari tajacu; spleen, liver, and skin of Chelonoidis denticulata.MethodologyA PCR using universal and specific primers for 16 S rRNA, of Bartonella spp. with subsequent genetic sequencing were used.ResultsThe results in this study indicate that Bartonella vinsonni was detected in the liver tissue of Didelphis marsupialis using both universal primers and those specific for Bartonella sp. Anaplasma platys was detected at the liver and spleen level using universal primers. Additionally, Bartonella spp. was found at the liver, spleen, and skin level in Pecari tajacu using the specific primers. Finally, using the universal and specific primers at the skin level, Bartonella spp. was evident in Chelonoidis denticulata.ConclusionsThe presence of the DNA of the Bartonella vinsonii was detected at the liver tissue in Didelphis marsupialis. DNA of the Anaplasma platys and Bartonella spp. were identified at the spleen and liver level. This study also identified that DNA Bartonella spp. was detected in Pecari tajacu skin. Finally DNA of Bartonella spp. was evident in Chelonoidis denticulate skin. The findings of this study suggest that these bacteria are present in these animals and may be responsible for outbreaks.

Detection of Bartonella DNA in Yellow Flies, Lone Star Ticks, and a Human Patient with Concurrent Evidence of Borrelia burgdorferi Infection in Northeast Florida, USA.

Background: Bartonella species and Borrelia burgdorferi sensu lato (Bbsl) are emerging zoonotic pathogens. The vectors and frequency of infections with both pathogen groups in the southern United States is understudied. This study describes an investigation of Bartonella and Bbsl in yellow flies collected at a residence in northeast Florida, USA, that led to subsequent discoveries of both organisms in lone star ticks (Amblyomma americanum) and a human patient. Materials and Methods: DNA samples from flies, ticks, and human patient blood specimens were tested via polymerase chain reaction assays for Bartonella or Bbsl species. DNA sequences were compared to reference strains for identification and characterization. Results: An exploratory investigation of arthropod-borne pathogens in yellow flies collected at a residence in northeast Florida revealed the presence of uncharacterized Bartonella species DNA sequences similar to ones previously detected in two lone star ticks from Virginia. Subsequent testing of several lone star ticks from the area detected similar sequences of Bartonella in three ticks. Testing of stored blood samples from a resident of the site, who had experienced chronic relapsing and remitting symptoms for over a decade, identified nearly identical Bartonella DNA sequences in multiple samples collected over a 10-year period. Two lone star ticks and several samples from the same patient and time period also tested positive for Bo. burgdorferi DNA, suggesting possible long-term coinfection of the patient with both organisms. Conclusion: This investigation identified highly similar Bartonella DNA sequences in yellow flies, lone star ticks, and a human patient in northeast Florida. Similarly, Bo. burgdorferi DNA was detected in two lone star ticks and multiple specimens from the patient. Positive PCR results from archived patient blood samples documented the presence of both organisms at multiple time points over more than a decade. More studies on human patients with chronic undefined illness and on the presence of Bartonella and Bbsl in hematophagous arthropods and animal hosts in the southeastern United States are needed.

Molecular pathogen screening of louse flies (Diptera: Hippoboscidae) from domestic and wild ruminants in Austria

BackgroundHippoboscid flies (Diptera: Hippoboscidae), also known as louse flies or keds, are obligate blood-sucking ectoparasites of animals, and accidentally of humans. The potential role of hippoboscids as vectors of human and veterinary pathogens is being increasingly investigated, but the presence and distribution of infectious agents in louse flies is still unknown in parts of Europe. Here, we report the use of molecular genetics to detect and characterize vector-borne pathogens in hippoboscid flies infesting domestic and wild animals in Austria.MethodsLouse flies were collected from naturally infested cattle (n = 25), sheep (n = 3), and red deer (n = 12) across Austria between 2015 and 2019. Individual insects were morphologically identified to species level and subjected to DNA extraction for molecular pathogen screening and barcoding. Genomic DNA from each louse fly was screened for Borrelia spp., Bartonella spp., Trypanosomatida, Anaplasmataceae, Filarioidea and Piroplasmida. Obtained sequences of Trypanosomatida and Bartonella spp. were further characterized by phylogenetic and haplotype networking analyses.ResultsA total of 282 hippoboscid flies corresponding to three species were identified: Hippobosca equina (n = 62) collected from cattle, Melophagus ovinus (n = 100) from sheep and Lipoptena cervi (n = 120) from red deer (Cervus elaphus). Molecular screening revealed pathogen DNA in 54.3% of hippoboscids, including infections with single (63.39%), two (30.71%) and up to three (5.90%) distinct pathogens in the same individual. Bartonella DNA was detected in 36.9% of the louse flies. Lipoptena cervi were infected with 10 distinct and previously unreported Bartonella sp. haplotypes, some closely associated with strains of zoonotic potential. DNA of trypanosomatids was identified in 34% of hippoboscids, including the first description of Trypanosoma sp. in H. equina. Anaplasmataceae DNA (Wolbachia spp.) was detected only in M. ovinus (16%), while < 1% of the louse flies were positive for Borrelia spp. and Filarioidea. All hippoboscids were negative for Piroplasmida.ConclusionsMolecular genetic screening confirmed the presence of several pathogens in hippoboscids infesting domestic and wild ruminants in Austria, including novel pathogen haplotypes of zoonotic potential (e.g. Bartonella spp.) and the first report of Trypanosoma sp. in H. equina, suggesting a potential role of this louse fly as vector of animal trypanosomatids. Experimental transmission studies and expanded monitoring of hippoboscid flies and hippoboscid-associated pathogens are warranted to clarify the competence of these ectoparasites as vectors of infectious agents in a One-Health context.Graphical

Comparison of molecular methods for Bartonella henselae detection in blood donors.

The Bartonella genus consists of neglected pathogens associated with potentially transfusional-transmitted and fatal human diseases. We aimed to evaluate Bartonella sp. prevalence in 500 blood donors and compare the results with the data already published about these samples. We used molecular diagnostic methods to detect Bartonella sp.-DNA from blood and liquid culture samples: (A) conventional PCR for two gene regions, the ITS targeting the genus Bartonella and the specific gltA Bartonella henselae; (B) nested PCR for the ftsZ gene and (C) qualitative real-time PCR for the gltA gene, both B. henselae specific. We obtained 30/500 (6%) DNA detections from the blood samples; 77/500 (15.4%) DNA detections from liquid culture samples and five (1%) samples had DNA detection from both. In total, we detected B. henselae DNA from 102/500 (20.4%) donors. The samples used in this study had already been submitted for Bartonella sp.-DNA detection using only a conventional PCR in liquid culture. Sixteen samples (3.2%) were positive previously, and from these 16 samples, 13 were negative in the new investigation. We concluded that the use of liquid culture combined with different molecular tests increases the possibility of detecting Bartonella sp.-DNA, but the tests do not avoid false-negative results. More than a fifth of blood donors had at least one PCR that detected Bartonella sp.-DNA among the eight molecular reactions performed now (four reactions in whole blood and four in liquid culture). Seven percent had B. henselae-DNA detection for two or more distinct regions. Considering the results obtained previously, the DNA of Bartonella spp. was detected or the agent isolated in 23% of analyzed blood donors. The results establish that the low bacteremia and the fastidious characteristics of the bacterium are challenges to laboratory diagnosis and can make it difficult to confirm the infection in patients with bartonelloses.

Molecular epidemiology of Bartonella species from sympatric mammals collected in urban and rural areas of Punjab, Pakistan

Bartonella can infect a variety of mammals including humans and has been detected in the Americas, Europe, Africa, and Asia. Roughly two-thirds of identified Bartonella species are found and maintained in rodent reservoirs, with some of these species linked to human infections. Rodents (N=236) were caught from the Sahiwal division of Punjab, Pakistan and tested for Bartonella using PCR targeting gltA and rpoB genes, followed by sequencing of rpoB-positive samples. Genetic relatedness to other published Bartonella spp. rpoB gene sequences were examined using BLAST and phylogenetic analysis. Overall, 7.62% (18/236) of rodents were positive for both gltA and rpoB fragments. Rattus rattus and R. norvegicus had 7.94% (12/151) and 7.05% (6/85) positivity rates for Bartonella DNA, respectively. Phylogenetic analysis revealed a close relatedness between Bartonella spp. from Pakistan to Bartonella spp. from China, Nepal, and Malaysia. This study is the first reported detection of Bartonella spp. in R. rattus and R. norvegicus from the Sahiwal area of Punjab, Pakistan.

Molecular identification of Bartonella spp. and Rickettsia felis in fox fleas, Chile

Seventy-five flea pools (one to ten fleas per pool) from 51 Andean foxes (Lycalopex culpaeus) and five South American grey foxes or chillas (Lycalopex griseus) from the Mediterranean region of Chile were analyzed for the presence of DNA of Bartonella spp. and Rickettsia spp. through quantitative real-time PCR for the nouG and gltA genes, respectively. Positive samples were further characterized by conventional PCR protocols, targeting gltA and ITS genes for Bartonella, and gltA, ompA, and ompB genes for Rickettsia. Bartonella was detected in 48 % of the Pulex irritans pools (B. rochalimae in three pools, B. berkhoffii in two pools, B. henselae in one pool), and 8 % of the Ctenocephalides felis felis pools (B. rochalimae, one pool). Rickettsia was confirmed in 11 % of P. irritans pools and 92 % of the Ct. felis pools. Characterization confirmed R. felis in all sequenced Rickettsia-positive pools. All Ct. canis pools were negative. A Ct. felis pool from a wild-found domestic ferret (Mustela putorius furo) also resulted positive for R. felis. Although opportunistic, this survey provides the first description of zoonotic pathogens naturally circulating in fleas parasitizing Chilean free-living carnivores.

Bartonella spp. in households with cats: Risk factors for infection in cats and human exposure

The aim of this study was to estimate the occurrence of Bartonella spp. per household in cats and the risk factors for Bartonella spp. positivity in cats and their owners from Valdivia, Chile. A total of 464 cats (distributed within 324 households) and 326 humans (control group [n = 112] and cat owner [n = 214]) distributed in 262 households were sampled. From the cat owners (n = 214), 128 humans were in households where the cat was also sampled, totaling 84 households with dual sampling. Real-time PCR (qPCR) was used for Bartonella spp. detection in blood from cats and humans, and immunofluorescent immunoassay (IFA) anti-Bartonella henselae was performed in human serum samples. Out of the total of 324 households, 20.43% presented at least one Bartonella positive cat. From the households with dual sampling, 29.7% (25/84) presented at least one qPCR-Bartonella spp. positive cat. However, Bartonella DNA was not amplified in humans, and in 7.3% (6/82) of the households was found at least one of the cat's owners exposed to B. henselae. Cats younger than one year (Odds Ratio (OR) = 5.3), non-neutered (OR 3.46), sampled at home (OR 5.82), and with improper application of tick/flea control products (OR 3.13) showed a higher risk for Bartonella spp. presence. Humans with occupational exposure involving animal contact, were more likely to exhibit B. henselae seropositivity (OR 7.5). Bartonella spp. was present in the cats a moderate number of households, but Bartonella DNA was not detected in owners' blood, inferring that there is a low risk of recent human infection in the studied population.