Relationship of Pneumocystis jiroveci Humoral Immunity to Prevention of Colonization and Chronic Obstructive Pulmonary Disease in a Primate Model of HIV Infection

Pulmonary colonization by the opportunistic pathogen Pneumocystis jiroveci is common in HIV⁺ subjects and has been associated with development of chronic obstructive pulmonary disease (COPD). Host and environmental factors associated with colonization susceptibility are undefined. Using a simian-human immunodeficiency virus (SHIV) model of HIV infection, the immunologic parameters associated with natural Pneumocystis jiroveci transmission were evaluated. SHIV-infected macaques were exposed to P. jiroveci by cohousing with immunosuppressed, P. jiroveci-colonized macaques in two independent experiments. Serial plasma and bronchoalveolar lavage (BAL) fluid samples were examined for changes in antibody titers to recombinant Pneumocystis-kexin protein (KEX1) and evidence of Pneumocystis colonization by nested PCR of BAL fluid. In experiment 1, 10 of 14 monkeys became Pneumocystis colonized (Pc⁺) by 8 weeks post-SHIV infection, while 4 animals remained Pneumocystis colonization negative (Pc⁻) throughout the study. In experiment 2, 11 of 17 animals became Pneumocystis colonized by 16 weeks post-SHIV infection, while 6 monkeys remained Pc⁻. Baseline plasma KEX1-IgG titers were significantly higher in monkeys that remained Pc⁻, compared to Pc⁺ monkeys, in experiments 1 (P = 0.013) and 2 (P = 0.022). Pc⁻ monkeys had greater percentages of Pneumocystis-specific memory B cells after SHIV infection compared to Pc⁺ monkeys (P = 0.037). After SHIV infection, Pc⁺ monkeys developed progressive obstructive pulmonary disease, whereas Pc⁻ monkeys maintained normal lung function throughout the study. These results demonstrate a correlation between the KEX1 humoral response and the prevention of Pneumocystis colonization and obstructive lung disease in the SHIV model. In addition, these results indicate that an effective Pneumocystis-specific memory B-cell response is maintained despite progressive loss of CD4⁺ T cells during SHIV infection.Despite advances in treatment strategies and the introduction of antiretroviral therapy (ART), pulmonary diseases remain a leading cause of morbidity and mortality in HIV-infected patients (34). Both emphysema and chronic obstructive pulmonary disease (COPD) have been reported at an increased frequency in HIV-infected patients (7, 15) and, unlike many AIDS-associated opportunistic infections, HIV-associated COPD may be increasing due to the prolonged life expectancy of the HIV⁺ population with ART and the high smoking rate in this population (5, 24). Although cigarette smoking is a primary risk factor for the development of COPD, it is interesting that HIV⁺ nonsmokers may also be at increased risk of disease (14). In addition, the observation that most smokers do not develop COPD (33) indicates that other factors may play a role in disease development.Evidence has accumulated suggesting a role for infectious agents as cofactors in the pathogenesis and exacerbation of COPD (58), where pulmonary inflammation due to cigarette smoke may be amplified by the presence of persistent infectious agents. The chronic inflammation associated with infectious agents is also thought to contribute to the development of COPD in HIV⁺ patients (44, 58). This possibility is highlighted by the observation that HIV-infected patients may be more prone to develop subclinical lung infections even if successfully treated with ART (16).Our laboratory and others have accumulated evidence in humans and in animal models that the fungal opportunistic pathogen, Pneumocystis jiroveci (formerly Pneumocystis carinii f. sp. hominis) is an important pathogen in COPD in both HIV⁺ and HIV⁻ populations. COPD-like changes have been reported in HIV⁺ patients with Pneumocystis pneumonia (PcP) (45), and recent studies suggest that low-level asymptomatic carriage of Pneumocystis may be associated with lung damage. An increased frequency of Pneumocystis colonization has been reported in HIV⁺ patients, including those on ART (41), and we have recently shown that HIV⁺ subjects who were Pneumocystis colonized (Pc⁺) have worse airway obstruction than HIV⁺ subjects who were Pc⁻ (40). An association between Pneumocystis colonization and COPD has also been shown in HIV⁻ subjects (3, 52).Animal models also support the role of Pneumocystis colonization in the pathogenesis of COPD. In a model of immunocompetent mice, cigarette exposure and Pneumocystis colonization resulted in greater pulmonary function deficits compared to cigarette exposure alone (4). Our laboratory has reported that in a simian immunodeficiency virus (SIV)-primate model of HIV infection, Pneumocystis colonization results in pulmonary inflammation, pulmonary function deficits, and anatomic emphysema (2, 6, 47, 59). Factors that influence susceptibility to Pneumocystis colonization are not clearly understood.Immunologic control of Pneumocystis infection is strongly correlated with CD4⁺ T-cell responses, although B cells and antibodies also play a role in prevention of PcP (19-23, 25, 38, 62). There is a high frequency of Pneumocystis-specific seroprevalence in immunocompetent adults (1, 9), as well as in nonhuman primates (11, 30), suggesting the persistence of serological memory or Pneumocystis-specific, long-lived plasma cells in response to natural Pneumocystis exposure. Antibodies to the Pneumocystis endoprotease kexin (KEX1) may be particularly important, because immune responses to Pneumocystis KEX1 have been associated with control of Pneumocystis infection in immunosuppressed murine models (62, 63).In the present study, we investigated the capacity of simian/human immunodeficiency virus (SHIV)-infected macaques to generate a humoral immune response to KEX1 in response to natural Pneumocystis exposure and examined the relationship between anti-Pneumocystis humoral immunity, the development of Pneumocystis colonization, and the development of COPD.(This study was presented in part as a poster at the American Thoracic Society International Conference, New Orleans, LA, 14 to 19 May 2010, and the American Thoracic Society International Conference, Toronto, Canada, 16 to 21 May 2008. A portion of the study was also presented as an oral presentation at the International Workshops on Opportunistic Protists in Boston, MA, 28 to 31 May 2008).