Multicenter Comparison of Lung and Oral Microbiomes of HIV-infected and HIV-uninfected Individuals

Abstract

Rationale: Improved understanding of the lung microbiome in HIV-infected individuals could lead to better strategies for diagnosis, therapy, and prophylaxis of HIV-associated pneumonias. Differences in the oral and lung microbiomes in HIV-infected and HIV-uninfected individuals are not well defined. Whether highly active antiretroviral therapy influences these microbiomes is unclear.

Objectives: We determined whether oral and lung microbiomes differed in clinically healthy groups of HIV-infected and HIV-uninfected subjects.

Methods: Participating sites in the Lung HIV Microbiome Project contributed bacterial 16S rRNA sequencing data from oral washes and bronchoalveolar lavages (BALs) obtained from HIV-uninfected individuals (n = 86), HIV-infected individuals who were treatment naive (n = 18), and HIV-infected individuals receiving antiretroviral therapy (n = 38).

Measurements and main results: Microbial populations differed in the oral washes among the subject groups (Streptococcus, Actinomyces, Rothia, and Atopobium), but there were no individual taxa that differed among the BALs. Comparison of oral washes and BALs demonstrated similar patterns from HIV-uninfected individuals and HIV-infected individuals receiving antiretroviral therapy, with multiple taxa differing in abundance. The pattern observed from HIV-infected individuals who were treatment naive differed from the other two groups, with differences limited to Veillonella, Rothia, and Granulicatella. CD4 cell counts did not influence the oral or BAL microbiome in these relatively healthy, HIV-infected subjects.

Conclusions: The overall similarity of the microbiomes in participants with and without HIV infection was unexpected, because HIV-infected individuals with relatively preserved CD4 cell counts are at higher risk for lower respiratory tract infections, indicating impaired local immune function.

Keywords: HIV infection; bronchoalveolar lavage; bronchoscopy; lung; microbiome.

Figure 1.

Application of the neutral model to assess possible contaminant sequences identified in control samples. The bronchoscope, saline solution for sample collection, and extraction reagents were considered as potential sources of contaminating DNA. Sequences consistent with the model prediction (gray dots) and sequences enriched in the control samples (red dots) were considered as potential contaminants and removed from subsequent analyses. Sequences overrepresented in bronchoalveolar lavage (BAL) samples (green dots) were retained (A). The 10 most abundant sequences excluded from subsequent analysis (B) and retained for analysis (C) are shown. Some genus names are repeated in B and C as they reflect different sequences with the same taxonomic classification.

Figure 2.

Nonmetric multidimensional scaling (NMDS) plots for all subjects showing oral wash (A) and bronchoalveolar lavage (BAL) (B). Samples were obtained as detailed from HIV-negative subjects (“Negative,” red circles), HIV-positive but treatment-naive subjects (“Naive,” blue squares), and HIV-positive subjects treated with highly active antiretroviral therapy (“HAART,” green triangles). Analysis is based on organizational taxonomic units (OTUs) from sequences of the V1–3 regions of the16S rRNA. Centroids are indicated by crosses. For oral washes, NMDS analysis demonstrated significant differences in OTUs between Negative and Naive subjects (P < 0.01), and between Negative and HAART subjects (P = 0.01), and between Naive and HAART subjects (P = 0.01). There were no significant differences in the BALs.

Figure 3.

Nonmetric multidimensional scaling (NMDS) plots comparing oral washes (open symbols) and bronchoalveolar lavages (BALs) (solid symbols) within subject groups. Individual subjects are linked by lines. Samples were obtained as detailed in Methods, and V1–3 regions were sequenced. There were significant differences in oral wash and BAL samples within HIV-negative subjects (Negative subjects) (A) (P < 0.01), HIV-positive but treatment-naive subjects (Naive subjects) (B) (P = 0.03), and HIV-positive subjects treated with highly active antiretroviral therapy (HAART subjects) (C) (P < 0.01).

Figure 4.

Relative abundances of the most common organizational taxonomic units (OTUs) identified in oral washes from all subject groups. Samples were obtained as detailed in Methods and V1–3 regions were sequenced. OTUs were compared by the Wilcoxon signed-rank test for all OTUs with average relative abundances of greater than 1% across all samples, and significant differences are demonstrated by asterisks. Streptococcus and Actinomyces were more abundant in oral washes from HIV-positive but treatment-naive subjects (Naive) than in the other groups. Rothia was more abundant in oral washes from HIV-positive subjects treated with highly active antiretroviral therapy (HAART) than in the other groups. Some genus designations appear more than once because multiple OTUs have the same consensus taxonomy. Negative = HIV-negative subjects.

Figure 5.

Relative abundances of the most common organizational taxonomic units (OTUs) identified in bronchoalveolar lavage (BAL) from all subject groups. Samples were obtained as detailed, and V1–3 regions were sequenced. OTUs were compared by the Wilcoxon signed-rank test for all OTUs with average relative abundances of greater than 1% across all samples. Overall, there were no significant differences. Some genus designations appear more than once because multiple OTUs have the same consensus taxonomy. HAART = HIV-positive subjects treated with highly active antiretroviral therapy; Naive = HIV-positive but treatment-naive subjects; Negative = HIV-negative subjects.

Figure 6.

Relative abundances of the most common organizational taxonomic units (OTUs) identified in oral wash versus bronchoalveolar lavage (BAL) for each subject group. Samples were obtained as detailed in Methods, and V1–3 regions were sequenced. OTUs were compared by the Wilcoxon signed-rank test for all OTUs with average relative abundances of greater than 1% across all samples. The OTUs indicated by asterisks demonstrated significant differences between BAL and oral wash in HIV-negative subjects (Negative) (A), HIV-positive but treatment-naive subjects (naive) (B), and HIV-positive subjects treated with highly active antiretroviral therapy (HAART) (C). Some genus designations appear more than once because multiple OTUs have the same consensus taxonomy.