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Human immunodeficiency virus (HIV)-infected patients in Malawi have a high incidence of pneumococcal pneumonia. Since IgG2 is known to be protective against Streptococcus pneumoniae, we measured pneumococcal-specific IgG (IgG-pn), IgG1 (IgG1-pn), and IgG2 (IgG2-pn) in bronchoalveolar lavage (BAL) fluid to see if a quantitative deficit might explain this observation. IgG-pn, IgG1-pn, and IgG2-pn were measured to determine if there was a change in the ratio of immunoglobulin isoforms. Fifty-two BAL samples were collected from 26 HIV-infected HAART-naive Malawi patients and 26 HIV-negative volunteers. IgG-pn, IgG1-pn, and IgG2-pn were determined using sandwich ELISA techniques. ELISA wells were coated with pneumococcal vaccine (Pneumovax 23). Biotinylated IgG, IgG1, and IgG2 were used as the secondary antibodies. Data are expressed as ng/mL BAL ± SEM. Total IgG-pn was elevated in HIV+ patientss compared with HIV− patients (p = .0098), as were IgG2-pn (p = .0002) and IgG1 (p = .0067). HIV-infected patients did not have a statistically significant difference in IgG1/IgG2 ratio compared with HIV-negative patients (p = .3804). In conclusion, despite the high incidence of invasive pneumococcal disease in Malawi, HIV-infected Malawians had higher total IgG-pn-, IgG1-pn-, and IgG2-pn-specific antibodies in BAL than uninfected subjects. We speculate that this may be related to elevated proinflammatory cytokines in HIV BAL, including IFN-γ, which is known to stimulate IgG2 production in particular. These results suggest that the high incidence of pneumococcal pneumonia in Malawi is not due to a quantitative deficit in IgG levels but rather points to potential functional antibody defects.

Background and Objective The incidence of invasive pneumococcal disease among HIV-infected persons has declined in the post HAART era. Changes in the humoral immune responses have been implicated in this change. It has already been shown that HAART is associated with a decline in BAL total IgG and IgM concentrations and a decrease in BAL pneumococcal-specific IgG concentrations. We measured total and pneumococcal-specific IgA concentrations in BAL of 24 HIV-infected patients before and at 4 and 24 weeks after initiation of HAART, anticipating a similar response. Methods BAL pneumococcal-specific IgA levels were measured using ELISA. The IgA concentrations were corrected for dilution by determining epithelial lining fluid (ELF) using the urea dilution technique. Results Data expressed as mean ± SEM μg IgA/mL ELF (* p value ≥ .05). Total and pneumococcal-specific IgA levels did not change after initiation of HAART. No change was detected in the levels of pneumococcal-specific IgA when expressed as a percentage of total IgA. No significant correlation was found between BAL IL-6 and IgA concentration or BAL percentage lymphocytes and IgA concentrations. Conclusion HAART is not associated with significant changes in total or pneumococcal-specific IgA concentrations in the distal respiratory tract. We speculate that this reflects the more dominant role of IgG in protection of the alveolar space against bacterial pathogens.

Background and Objective Mycobacterium tuberculosis (MTB) is a common cause of pulmonary disease in HIV-infected patients. Normal alveolar macrophages (AM) phagocytize MTB and are then activated by cytokines to kill ingested MTB. We have previously shown that HIV alveolar AM are intrinsically resistant to MTB infection compared to normal AM, likely due to an AM-activating factor secreted by alveolar T cells. In this work, we examined the ability of concentrated BAL from HIV-infected subjects to activate AM to kill MTB. Methods BAL from HIV-infected subjects before and after starting antiretroviral therapy were concentrated 5 times using 10 kD spin columns and then cultured with normal AM for 24 hours. Virulent and avirulent MTB at an MOI of 1:1 was then added and subsequent infection determined using a BACTEC methodology. BAL cytokine concentrations were measured using a cytometric bead array assay system. Time to initial and maximal infection of the AM by MTB was assessed. Results Virulent and avirulent MTB grew equally well in AM not treated with HIV BAL. However, pretreatment with HIV BAL significantly delayed AM infection (both initial and maximal) by avirulent MTB compared to virulent MTB (initial infection: 12.2 ± 0.3 days vs 9.8 ± 0.3 days, p

Invasive pneumococcal disease is a significant cause of morbidity and mortality in the HIV-infected Malawian population despite high concentrations of anti-pneumococcal antibody concentrations in bronchoalveolar lavage (BAL). Thus we undertook this study to assess the opsonic function of the antibody. 23 HIV-infected and 26 uninfected subjects underwent bronchoscopy with BAL. IgG was purified from BAL on a protein G column. Serotype 1 anti-pneumoccal specific IgG antibody was determined by ELISA. The ability of purified IgG to bind to serotype 1 pneumococci was determined by mixing pneumococcus, purified IgG, and FITC-labelled anti-IgG and analysis by flow cytometry. The ability of FITC-labelled pneumococci coated with purified IgG to bind to monocyte monolayers was determined in a luminometer before and after quenching surface bound FITC with trypan blue. (Table) Serum and BAL from HIV-infected subjects contained significantly more serotype type 1 specific anti-pneumococcal IgG than normal serum and BAL. Despite this, when equal amounts of purified IgG were added to serotype 1 pneumococci significantly less antibody bound to the organism, both as a percent of pneumococci stained (61 ± 11.8% vs 53 ± 6.6%, p = .001) and as a fluorescent geometric mean indicating intensity of binding. Finally, when serotype 1 specific pneumococci were pretreated with equal amounts of purified BAL IgG, less organisms bound to monocytes after opsonization with HIV IgG compared to normal IgG (60 ± 14% vs 67 ± 14%, p = 0.065). Similar amounts of bound organisms were ingested. These studies suggest that although HIV infection is associated with increased anti-pneumococcal antibody secretion, the antibody demonstrates a significant impairment in its ability to opsonize bacteria. These results may explain the increased susceptibility of HIV-infected subjects to pneumococcal infections. This abstract is funded by NIH RO1 HL62058 (H.L.T.) and the Wellcome Trust (S.B.G.).