The motivation for the study rests on a growing recognition that Bartonella, Borrelia, and piroplasmid parasites circulate widely among arthropod vectors and animal reservoirs in Brazil, and that people with weakened immune systems may be particularly vulnerable to their consequences. Bartonella species are fastidious, intracellular bacteria best known for causing cat scratch disease in immunocompetent individuals, but in patients with profound immunosuppression they have been linked to severe vascular lesions such as bacillary angiomatosis, a condition historically described in people with advanced AIDS. Borrelia species, the spirochetes responsible for relapsing fever and Lyme-like illness, and piroplasmids, the protozoan parasites that include Babesia, add further layers of diagnostic complexity because they can produce nonspecific febrile symptoms that overlap with many opportunistic infections.

To investigate these possibilities, the researchers enrolled 75 HIV-positive individuals who were symptomatic and living in Brazil, collecting detailed epidemiological histories and medical records for each participant. The team then took a two-pronged diagnostic approach. The first prong was serological: they used indirect immunofluorescence assays, or IFA, to detect IgG antibodies against six distinct Bartonella antigens, namely Bartonella henselae, Bartonella koehlerae, Bartonella quintana, and two genotypes of Bartonella vinsonii subsp. berkhoffii. All assays were performed in a blinded manner, meaning the laboratory staff did not know the clinical status of the samples they were testing, a design feature intended to reduce bias. A reciprocal antibody titer of 1:64 or greater was defined as seroreactivity, the threshold at which a sample was considered to show evidence of prior exposure.

The second prong was molecular. The researchers extracted DNA from blood samples and searched for genetic signatures of Bartonella, Borrelia, and piroplasmid organisms using both quantitative polymerase chain reaction, or qPCR, and digital PCR, or dPCR. Crucially, they did not rely on a single snapshot of the bloodstream. Because Bartonella bacteria are notorious for circulating at extremely low numbers and in a cyclic fashion, the team enriched blood cultures in Bartonella alpha-Proteobacteria Growth Medium, known as BAPGM, for up to 28 days, and then repeated the qPCR testing on the cultured material. This enrichment step is designed to amplify any sparse organisms that might be present below the detection limit of direct blood testing, making it one of the most sensitive protocols available for hunting these bacteria in human blood.

The serological results were striking. Of the 75 participants, 47, or 62.7 percent, were seroreactive to at least one Bartonella antigen. The most frequently detected antibodies were directed against B. henselae, found in 57.3 percent of the cohort, followed by B. vinsonii subsp. berkhoffii genotype II, detected in 44.0 percent. These figures suggest that exposure to Bartonella species is remarkably common among symptomatic HIV-infected individuals in this Brazilian population, a finding consistent with the documented circulation of these bacteria in fleas, ticks, and animal reservoirs across the country. Whether the antibodies reflect true past infection, cross-reactivity with other bacteria, or persistent low-level colonization that evaded molecular detection remains an open question that the authors themselves acknowledge.

Perhaps the most clinically provocative finding was the statistical association between Bartonella seroreactivity and systemic manifestations of disease. Among the 47 seroreactive patients, 11 exhibited systemic signs and symptoms, compared with only 1 of the 28 non-seroreactive patients. This difference translated into an odds ratio of 8.25, with a 95 percent confidence interval spanning 1.00 to 68.2 and a p value of 0.026. In practical terms, seroreactive patients were roughly eight times more likely to present with systemic manifestations than those without detectable antibodies, although the wide confidence interval reflects the modest sample size and signals considerable statistical uncertainty. The association was exploratory in nature, and the authors were careful to frame it as a hypothesis-generating observation rather than proof of causation.

On the molecular front, the picture was far more definitive, and far more negative. Every one of the 75 participants tested negative for Bartonella and Borrelia DNA by both qPCR and dPCR, both before and after the extended BAPGM enrichment culture period. Two participants did show a low positive partition number for piroplasmid DNA by dPCR, a result that initially raised the possibility of Babesia-like infection, but when the samples were retested the signal could not be reproduced. The authors interpreted these non-repeatable findings as most likely representing technical artifacts or contamination rather than true infection, underscoring a central principle of molecular diagnostics: a single low-level positive result, especially near the limit of detection, demands confirmation before it can be considered evidence of active disease.

The disconnect between high seroprevalence and absent molecular detection is the intellectual heart of the study, and it speaks to a broader challenge in the field of vector-borne pathogen research. Antibodies indicate that the immune system has encountered an organism at some point, but they do not distinguish between a resolved infection years ago and a persistent, sequestered infection hiding in tissue reservoirs such as the vascular endothelium, where Bartonella is known to reside. Conversely, negative blood PCR does not exclude infection in tissues, because these bacteria may only intermittently spill into the bloodstream. The BAPGM enrichment culture was designed specifically to address this intermittency, and its failure to yield any PCR-positive samples after nearly a month of incubation provides reasonably strong evidence that active bloodstream infection with these organisms was absent in this cohort at the time of testing.

The clinical implications are nuanced. On one hand, the study provides reassurance that in this population of symptomatic HIV-infected Brazilians, active Bartonella, Borrelia, and piroplasmid infections were not detectable, suggesting that these pathogens are unlikely to be a common hidden driver of unexplained symptoms in similar patients. On the other hand, the strong association between seroreactivity and systemic manifestations hints that prior Bartonella exposure might somehow shape the clinical course of HIV disease, or that both are correlated with shared risk factors such as living conditions, animal contact, or arthropod exposure that were not fully disentangled in this exploratory analysis. The authors conclude that the clinical relevance of Bartonella seroreactivity in this setting remains to be clarified, a candid acknowledgment that serology alone cannot settle the question.

Looking forward, the study lays groundwork for several important next steps. Larger cohorts with longitudinal follow-up would help determine whether seroreactive patients experience different disease trajectories over time. Tissue-based sampling, rather than blood alone, could reveal whether Bartonella organisms persist in vascular niches that blood PCR cannot reach. And the combination of dPCR technology with extended enrichment culture, as deployed here, offers a template for future investigations of other fastidious vector-borne pathogens in immunocompromised populations worldwide. For now, the message for clinicians is measured: exposure to Bartonella appears frequent among symptomatic HIV patients in Brazil, but confirmed active infection with these three pathogen groups was not found, and the meaning of a positive antibody test in this population remains an open scientific question worthy of continued investigation.

Subject of Research: Prevalence of Bartonella, Borrelia, and piroplasmid exposure and infection in symptomatic HIV-infected individuals in Brazil

Article Title: Prevalence of Bartonella spp., Borrelia spp., and piroplasmid in HIV-infected symptomatic individuals in Brazil: a cross-sectional study

Article References: Robveille, C., Sarris, A. B., Breitschwerdt, E. B., André, M. R., do Amaral, R. B., Maggi, R. G., Mussi-Pinhata, M. M., Dias, C. M., & de Carvalho Santana, R. (2026). Prevalence of Bartonella spp., Borrelia spp., and piroplasmid in HIV-infected symptomatic individuals in Brazil: a cross-sectional study. Parasites & Vectors. https://doi.org/10.1186/s13071-026-07584-x

Image Credits: AI Generated

DOI: 10.1186/s13071-026-07584-x

Keywords: Bartonella, Borrelia, piroplasmid, HIV, vector-borne disease, serology, immunofluorescence assay, qPCR, digital PCR, BAPGM culture, Brazil, opportunistic infection

Bartonella species are vector-borne, Gram-negative bacteria that invade red blood cells and the cells lining blood vessels. More than 45 species have been described, but three dominate human disease: B. henselae, the agent of cat scratch disease, transmitted chiefly by cat fleas and scratches from infected cats; B. quintana, the agent of trench fever, spread by the human body louse; and B. bacilliformis, restricted largely to South America. In immunocompetent people, cat scratch disease typically resolves on its own within two to four months, presenting as swollen lymph nodes, skin lesions, fatigue, and headache. In contrast, B. quintana infection spans a broad clinical spectrum, from recurrent fever with severe leg pain to chronic bacteremia, culture-negative infective endocarditis, bacillary angiomatosis, and bacillary peliosis, conditions that emerge mainly when the immune system cannot contain the organism.

That distinction matters because the host immune response is the central determinant of how Bartonella infection unfolds. In people with intact immunity, the pathological response is typically granulomatous and suppurative, effectively walling off the bacteria. In immunocompromised individuals, including organ transplant recipients and people with HIV, the same organisms can drive proliferative vascular lesions that grow rapidly. Patients with CD4 T-cell counts below 50 cells per microliter are considered especially vulnerable to severe disease and chronic opportunistic infection. Iran has already seen warning signs: the first molecularly confirmed case of B. quintana bacillary angiomatosis in an HIV-infected patient was reported there in 2021, and more than eight Bartonella species have since been identified in insects and livestock reservoirs across the country, five of them known to cause human disease.

To quantify the scale of exposure, the research team enrolled 200 participants between September 2023 and May 2024: 150 patients with HIV infection recruited from the Infectious Diseases Department and HIV clinic at Imam Khomeini Hospital, a referral university hospital in Tehran, and 50 randomly selected healthy controls. Within the HIV-positive group, 39 individuals met the criteria for AIDS, defined by CD4 counts below 200 cells per microliter or clinical AIDS-defining conditions, while 111 had higher counts. Each participant provided five milliliters of venous blood after informed consent, completed a structured questionnaire covering demographics, animal contact, tick and flea exposure, social status, and transfusion history, and had serum separated and stored at minus 20 degrees Celsius under cold chain conditions at the Pasteur Institute.

The serological workhorse of the study was the indirect immunofluorescence assay, or IFA, the technique recommended by the US Centers for Disease Control and Prevention for detecting antibodies against B. henselae and B. quintana. Because the samples came from HIV-positive individuals, all sera were first treated with Triton X-100 for 30 minutes at 37 degrees Celsius to reduce biocontamination risk, a pretreatment shown to inactivate HIV particles without compromising antibody detection. For IgG testing, sera were serially diluted from 1:64 up to 1:2048 and applied to slides coated with fixed bacterial antigens; after incubation and washing, a fluorescein-labeled anti-human IgG conjugate was added, and fluorescence was read at 400-fold magnification. The highest dilution showing distinct apple-green fluorescence defined the endpoint titer, with 1:64 or above scored as positive. IgM testing followed a parallel protocol on a randomly selected subset of 48 participants, using IgG sorbent to strip competing IgG antibodies before dilution, with titers of 1:24 or above considered positive.

The results revealed substantial background exposure. Across all 200 participants, IgG seroprevalence was 24.0 percent for B. henselae and 22.0 percent for B. quintana, while IgM seroprevalence in the tested subset reached 18.8 percent and 22.9 percent respectively. Strikingly, the healthy controls fared no better: B. henselae IgG was found in 30.0 percent of controls and 30.0 percent of AIDS patients, versus 18.3 percent of HIV-positive patients without AIDS, and B. quintana IgG appeared in 26.0 percent of controls, 28.3 percent of AIDS patients, and 17.3 percent of the HIV-positive group. None of these between-group differences reached statistical significance, suggesting that Bartonella circulation in this region extends well beyond the immunocompromised clinic population.

Risk factor analysis sharpened the picture. In multivariable logistic regression, higher age emerged as protective against B. quintana IgG seropositivity, with an odds ratio of 0.48, while a history of drug use nearly tripled the odds, at 3.28. For B. henselae, every body mass index category above underweight carried significantly lower odds of IgG positivity compared with a BMI below 18.5, with odds ratios falling as low as 0.16, and drug use again raised the odds, at 2.46. Underlying chronic diseases were associated with reduced seropositivity, an odds ratio of 0.31, a counterintuitive finding the authors interpret cautiously. Notably, contact with cats, dogs, or other animals, often assumed to be the main route of exposure, showed no significant association with seropositivity for either species, nor did homelessness or documented flea bites.

The most sobering number came from the immunologically weakest participants. Among the 34 patients with CD4 counts below 200 cells per microliter, 28, or 82 percent, were seropositive for at least one Bartonella antibody class. The authors are careful to note that serology alone cannot establish clinically relevant infection or prove a causal link with immunodeficiency, since IgG positivity reflects past exposure and IgM may signal recent infection that molecular testing would be needed to confirm. Still, the figure aligns with international data: seroprevalence among HIV-infected populations has ranged from 17.3 to 40 percent in prior studies, with a Spanish study reporting 22.3 percent, a Brazilian study 40.8 percent, and a South African molecular study detecting bartonellosis in 10 percent of HIV-positive outpatients.

The authors also confront the inherent limits of antibody-based detection. IFA sensitivity ranges from 50 to 95 percent, and cross-reactivity with Coxiella, Chlamydia, and potentially Mycobacterium species, which are common in HIV patients, can blur specificity. In advanced immunosuppression, impaired humoral responses may blunt antibody production entirely, producing false negatives despite active infection, which means negative serology cannot exclude Bartonella in severely immunocompromised patients. The absence of PCR confirmation in this study likewise prevents a clean separation of past from recent exposure and may have led to slight underestimation of recent infections. When treatment is needed, current evidence favors macrolides such as azithromycin, tetracyclines, particularly doxycycline, and rifampin, which carry the most reliable minimum inhibitory concentration data, having displaced the beta-lactams used historically.

The broader message is one of vigilance rather than alarm. Exposure to Bartonella species appears common in both HIV-infected and non-infected Iranians, and the combination of molecular and serological evidence, including a companion PCR study that detected B. quintana exclusively among immunocompromised patients, supports active circulation of these bacteria in vulnerable populations. The authors argue that the healthcare community should pay closer attention to bartonellosis surveillance and management in people with HIV, particularly those with advanced immunosuppression, and that standardized diagnostic approaches combining serology with molecular methods are critically important. As the study’s findings make clear, a bacterium once associated with trench warfare and stray cats has not disappeared; it has simply been waiting for immune systems to falter.

Subject of Research: Seroprevalence of Bartonella quintana and Bartonella henselae antibodies among HIV-positive patients in Iran

Article Title: A cross sectional seroprevalence study of Bartonella quintana and Bartonella henselae among patients with human immunodeficiency virus (HIV) in Iran

Article References: Latifian, M., Hasannezhad, M., Abbasian, L., Tahmasebi Ashtiani, Z., Arabian, M., Bagheri Amiri, F., & Esmaeili, S. (2026). A cross sectional seroprevalence study of Bartonella quintana and Bartonella henselae among patients with human immunodeficiency virus (HIV) in Iran. New Microbes and New Infections, 73, Article 101841. https://doi.org/10.1016/j.nmni.2026.101841

Image Credits: AI Generated

DOI: Not provided

Keywords: Bartonella quintana, Bartonella henselae, HIV, seroprevalence, immunocompromised patients, cat scratch disease, trench fever, immunofluorescence assay, Iran, opportunistic infection, vector-borne bacteria, CD4 count