f Limited information is available about the effects of HIV and subsequent antiretroviral treatment on host-microbe interactions. This study aimed to determine the salivary microbial composition for 10 HIV-seropositive subjects, before and 6 months after highly active antiretroviral therapy (HAART), compared with that for 10 HIV-seronegative subjects. A conventional culture and two culture-independent analyses were used and consistently demonstrated differences in microbial composition among the three sets of samples. HIV-positive subjects had higher levels of total cultivable microbes, including oral streptococci, lactobacilli, Streptococcus mutans, and Candida, in saliva than did HIV-negative subjects. The total cultivable microbial levels were significantly correlated with CD4 ؉ T cell counts. Denaturing gradient gel electrophoresis (DGGE), which compared the overall microbial profiles, showed distinct fingerprinting profiles for each group. The human oral microbe identification microarray (HOMIM) assay, which compared the 16S rRNA genes, showed clear separation among the three sample groups. Veillonella, Synergistetes, and Streptococcus were present in all 30 saliva samples. Only minor changes or no changes in the prevalence of Neisseria, Haemophilus, Gemella, Leptotrichia, Solobacterium, Parvimonas, and Rothia were observed. Seven genera, Capnocytophaga, Slackia, Porphyromonas, Kingella, Peptostreptococcaceae, Lactobacillus, and Atopobium, were detected only in HIVnegative samples. The prevalences of Fusobacterium, Campylobacter, Prevotella, Capnocytophaga, Selenomonas, Actinomyces, Granulicatella, and Atopobium were increased after HAART. In contrast, the prevalence of Aggregatibacter was significantly decreased after HAART. The findings of this study suggest that HIV infection and HAART can have significant effects on salivary microbial colonization and composition.

Protease inhibitor cocktails are routinely added to clinical samples used for proteomic studies to inactivate proteases. Since these same samples are often used for microbial studies, we determined whether addition of protease inhibitors could affect the quantitative or qualitative assessment of microbial profiles. Twenty-two saliva samples were collected and processed immediately with or without the addition of a protease inhibitor cocktail. Conventional cultivation methods were used to evaluate total bacterial growth. Total genomic DNA was isolated and a specific 16S rRNA gene-targeted region was PCR-amplified and separated by denaturing gradient gel electrophoresis. A combination of 1D SDS-PAGE and LC-MS/MS methods was used to determine the effect of the protease inhibitors on the integrity of salivary proteins and peptides. Interestingly, no significant differences were observed in either the bacterial growth and composition or the integrity of salivary proteins between the two groups. Correlation coefficients between the paired samples for total cultivable microbiota (r2=0.847), total mutans streptococci (r2=0.898), total oral lactobacilli (r2=0.933), and total Streptococcus mutans (r2=0.870) also exceeded expected values. The results suggest that the addition of a protease inhibitor cocktail in saliva samples does not impact the growth of oral microbiota or compromise the ability to characterize its composition.