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Abstract This study aimed to determine the sequence type (ST) of Bartonella henselae infecting small Indian mongooses from Saint Kitts via multi-locus sequence typing (MLST). This investigation used stored EDTA blood (n = 22) samples from mongooses previously identified as positive for B. henselae. Chocolate agar plates were enriched with Bartonella alpha-Proteobacteria growth medium (BAPGM) to culture and isolate Bartonella from the blood samples. To perform MLST, DNA was extracted and purified from isolates followed by amplification by conventional PCR (300–500 bp) for eight genes (16S rDNA, batR, gltA, groEL, ftsZ, nlpD, ribC, and rpoB). Bartonella henselae STs were deposited in the PubMLST repository. Out of 22 B. henselae-positive blood samples, isolates were obtained from 12 mongooses (54.5%; 12/22). Each mongoose was infected with one ST. The studied mongoose population was infected with sequence types ST2, ST3, ST8, and a novel ST represented by ST38. Bartonella henselae ST2, ST3 and ST8 infecting mongooses are known to circulate in humans and cats, with ST2 and ST8 associated with Cat Scratch Disease (bartonellosis) in humans. The results presented herein denote the circulation of B. henselae STs with zoonotic potential in mongooses with risk of B. henselae transmission to humans.

Case summary A 10-year-old domestic shorthair cat presented for lethargy, anorexia and labored breathing. Significant pleural and pericardial effusions prompted thoracocentesis and pericardiocentesis. Cytologic evaluation of the pericardial effusion revealed a highly cellular hemorrhagic, eosinophilic (12%) effusion, with many markedly atypical suspected mesothelial cells, interpreted as concerning for neoplasia. Thoracoscopic subtotal pericardiectomy and histology of the pericardium revealed predominantly eosinophilic inflammation with multifocal mesothelial hypertrophy and ulceration. A peripheral eosinophilia was not present on serial complete blood counts. Initial infectious disease testing was mostly negative. Toxoplasma gondii titers were most consistent with prior exposure, although reactivation could not be excluded. The owner’s medical history included a prior diagnosis of bartonellosis. Owing to the challenges of definitive Bartonella species exclusion, the cat was treated empirically with pradofloxacin and doxycycline, and a subtotal pericardectomy. There was improvement at first but pleural effusion recurred approximately 3 months after discharge. The cat was euthanized and a necropsy was not performed. Subsequent pericardial effusion Piroplasma/Bartonella/Borrelia droplet digital PCR detected DNA of Bartonella vinsonii subspecies berkhoffii , and peripheral blood culture and sequencing revealed a rare apicomplexan organism (90% homology with Colpodella species) of unknown clinical significance. Testing for filamentous bacteria and fungal pathogens was not performed. Relevance and novel information This case offers several unique entities – eosinophilic pericardial effusion and eosinophilic pericarditis of unknown etiology – and illustrates the well-known marked atypia that may occur in reactive and hyperplastic mesothelial cells, particularly of infrequently sampled and cytologically described feline pericardial effusion, supporting a cautious interpretation of this cytology finding.

Abstract Background Ctenocephalides felis, the cat flea, is the most common ectoparasite of cats and dogs worldwide. As a cause of flea allergy dermatitis and a vector for two genera of zoonotic pathogens (Bartonella and Rickettsia spp.), the effect of the C. felis microbiome on pathogen transmission and vector survival is of substantial medical importance to both human and veterinary medicine. The aim of this study was to assay the pathogenic and commensal eubacterial microbial communities of individual C. felis from multiple geographic locations and analyze these findings by location, qPCR pathogen prevalence, and flea genetic diversity. Methods 16S Next Generation Sequencing (NGS) was utilized to sequence the microbiome of fleas collected from free-roaming cats, and the cox1 gene was used for flea phylogenetic analysis. NGS data were analyzed for 168 individual fleas from seven locations within the US and UK. Given inconsistency in the genera historically reported to constitute the C. felis microbiome, we utilized the decontam prevalence method followed by literature review to separate contaminants from true microbiome members. Results NGS identified a single dominant and cosmopolitan amplicon sequence variant (ASV) from Rickettsia and Wolbachia while identifying one dominant Bartonella clarridgeiae and one dominant Bartonella henselae/Bartonella koehlerae ASV. Multiple less common ASVs from these genera were detected within restricted geographical ranges. Co-detection of two or more genera (Bartonella, Rickettsia, and/or Wolbachia) or multiple ASVs from a single genus in a single flea was common. Achromobacter, Peptoniphilus, and Rhodococcus were identified as additional candidate members of the C. felis microbiome on the basis of decontam analysis and literature review. Ctenocephalides felis phylogenetic diversity as assessed by the cox1 gene fell within currently characterized clades while identifying seven novel haplotypes. NGS sensitivity and specificity for Bartonella and Rickettsia spp. DNA detection were compared to targeted qPCR. Conclusions Our findings confirm the widespread coinfection of fleas with multiple bacterial genera and strains, proposing three additional microbiome members. The presence of minor Bartonella, Rickettsia, and Wolbachia ASVs was found to vary by location and flea haplotype. These findings have important implications for flea-borne pathogen transmission and control. Graphical Abstract

Surveillance of the fleas and flea-borne pathogens infecting cats is important for both human and animal health. Multiple zoonotic Bartonella and Rickettsia species are known to infect the most common flea infesting cats and dogs worldwide: Ctenocephalides felis, the cat flea. The ability of other flea species to transmit pathogens is relatively unexplored. We aimed to determine cat host and flea factors independently associated with flea Bartonella and Rickettsia infection. We also assessed flea and cat infection by flea-host pair and location. To accomplish these aims, we performed qPCR for the detection of Bartonella, hemotropic Mycoplasma, Rickettsia, and Wolbachia DNA using paired cat and flea samples obtained from free-roaming cats presenting for spay or neuter across four locations in the United States. A logistic regression model was employed to identify the effect of cat (sex, body weight, geographic location, and Bartonella, hemotropic Mycoplasma, and Rickettsia spp., infection) and flea (clade and Rickettsia and Wolbachia infection) factors on C. felis Bartonella clarridgeiae infection. From 189 free roaming cats, we collected 84 fleas: Ctenocephalides felis (78/84), Cediopsylla simplex (4/84), Orchopeas howardi (1/84), and Nosopsyllus fasciatus (1/84). Ctenocephalides felis were phylogenetically assigned to Clades 1, 4, and 6 by cox1 gene amplification. Rickettsia asembonensis (52/84) and B. clarridgeiae (16/84) were the most common pathogenic bacteria detected in fleas. Our model identified host cat sex and weight as independently associated with B. clarridgeiae infection in fleas. Rickettsia asembonensis (52/84), Rickettsia felis (7/84) and Bartonella henselae (7/84) were detected in specific clades: R. felis was detected only in Clades 1 and 6 while B. henselae and R. asembonensis were detected only in Clade 4. Wolbachia spp., also displayed clade specificity with strains other than Wolbachia wCfeT only infecting fleas from Clade 6. There was poor flea and host agreement for Bartonella spp., infection; however, there was agreement in the Bartonella species detected in cats and fleas by geographic location. These findings reinforce the importance of considering reservoir host attributes and vector phylogenetic diversity in epidemiological studies of flea-borne pathogens. Widespread sampling is necessary to identify the factors driving flea-borne pathogen presence and transmission.