Gut dysbiosis and microbial translocation are associated with chronic systemic immune activation and inflammation in HIV-1 infection. However, the extent of restoration of gut microbiota in HIV-1 patients with short or long-term antiretroviral therapy (ART) is unclear. To understand the impact of ART on the gut microbiota, we used the rhesus macaque model of SIV infection to characterize and compare the gut microbial community upon SIV infection and during ART. We observed altered taxonomic compositions of gut microbiota communities upon SIV infection and at different time points of ART. SIV-infected animals showed decreased diversity of gut microbiome composition, while the ART group appeared to recover towards the diversity level of the healthy control. Animals undergoing ART for various lengths of time were observed to have differential gut bacterial abundance across different time points. In addition, increased blood lipopolysaccharide (LPS) levels during SIV infection were reduced to near normal upon ART, indicating that microbial translocation and immune activation can be improved during therapy. In conclusion, while short ART may be related to transient increase of certain pathogenic bacterial microbiome, ART may promote microbiome diversity compromised by SIV infection, improve the gut microbiota towards the healthy compositions and alleviate immune activation.

Sensitive and specific blood-based assays for the detection of pulmonary and extrapulmonary tuberculosis would reduce mortality associated with missed diagnoses, particularly in children. Here we report a nanoparticle-enhanced immunoassay read by dark-field microscopy that detects two Mycobacterium tuberculosis virulence factors (the glycolipid lipoarabinomannan and its carrier protein) on the surface of circulating extracellular vesicles. In a cohort study of 147 hospitalized and severely immunosuppressed children living with HIV, the assay detected 58 of the 78 (74%) cases of paediatric tuberculosis, 48 of the 66 (73%) cases that were missed by microbiological assays, and 8 out of 10 (80%) cases undiagnosed during the study. It also distinguished tuberculosis from latent-tuberculosis infections in non-human primates. We adapted the assay to make it portable and operable by a smartphone. With further development, the assay may facilitate the detection of tuberculosis at the point of care, particularly in resource-limited settings.

Background Non-sputum methods are urgently needed to improve tuberculosis diagnosis and treatment monitoring in children. This study evaluated the ability of a serum assay quantifying a species-specific peptide of the Mycobacterium tuberculosis CFP-10 virulence factor via nanotechnology and matrix-assisted laser desorption ionization time-of-flight mass spectrometry to diagnose tuberculosis in HIV-infected and HIV-uninfected infants. Methods Serum CFP-10 peptide signal was blinded evaluated in cryopreserved sera of 519 BCG-immunized, HIV-exposed infants (284 HIV-infected, 235 HIV-uninfected) from a multi-center randomized placebo-controlled isoniazid prophylaxis trial conducted in southern Africa between 2004 and 2008, who were followed up to 192 weeks for Mtb infection and TB. Children were classified as confirmed, unconfirmed, or unlikely tuberculosis cases using 2015 NIH diagnostic criteria for pediatric TB. Results In HIV-infected infants, CFP-10 signal had 100% sensitivity for confirmed TB (5/5, 95% CI, 47.8–100) and 83.7% sensitivity for unconfirmed TB (36/43, 95% CI 69.3–93.2), with 93.1% specificity (203/218, 95% CI 88.9–96.1). In HIV-uninfected infants, CFP-10 signal detected the single confirmed TB case and 75.0% of unconfirmed TB cases (15/20; 95% CI 50.9–91.3), with 96.2% specificity (177/184, 95% CI, 92.3–98.5). Serum CFP-10 achieved 77% diagnostic sensitivity for confirmed and unconfirmed TB (13/17, 95% CI, 50–93%) at ≤ 24 weeks pre-diagnosis, and both CFP-10-positivity and concentration declined following anti-TB therapy initiation. Conclusions Serum CFP-10 signal exhibited high diagnostic sensitivity and specificity for tuberculosis in HIV-infected and HIV-uninfected infants and potential utility for early TB detection and monitoring of anti-TB treatment responses.

Identification of epitopes targeted following virus infection or vaccination can guide vaccine design and development of therapeutic interventions targeting functional sites, but can be laborious. Herein, we employed peptide microarrays to map linear peptide epitopes (LPEs) recognized following SARS-CoV-2 infection and vaccination. LPEs detected by nonhuman primate (NHP) and patient IgMs after SARS-CoV-2 infection extensively overlapped, localized to functionally important virus regions, and aligned with reported neutralizing antibody binding sites. Similar LPE overlap occurred after infection and vaccination, with LPE clusters specific to each stimulus, where strong and conserved LPEs mapping to sites known or likely to inhibit spike protein function. Vaccine-specific LPEs tended to map to sites known or likely to be affected by structural changes induced by the proline substitutions in the mRNA vaccine’s S protein. Mapping LPEs to regions of known functional importance in this manner may accelerate vaccine evaluation and discovery of targets for site-specific therapeutic interventions.

Rationale: Circulating pathogen-derived proteins can serve as useful biomarkers for infections but may be detected with poor sensitivity and specificity by standard immunoassays due to masking effects and cross-reactivity. Mass spectrometry (MS)-read immunoassays for biomarker-derived peptides can resolve these issues, but lack standard workflows to select species-specific peptides with strong MS signal that are suitable for antibody generation. Methods: Using a Mycobacterium tuberculosis ( Mtb ) protein as an example, candidate peptides were selected by length, species-specificity, MS intensity, and antigenicity score. MS data from spiked healthy serum was employed to define MS feature thresholds, including a novel measure of internal MS data correlation, to produce a peak detection algorithm. Results: This algorithm performed better in rejecting false positive signal than each of its criteria, including those currently employed for this purpose. Analysis of an Mtb peptide biomarker (CFP-10pep) by this approach identified tuberculosis cases not detected by microbiologic assays, including extrapulmonary tuberculosis and tuberculosis cases in children infected with HIV-1. Circulating CFP-10pep levels measured in a non-human primate model of tuberculosis distinguished disease from asymptomatic infection and tended to correspond with Mtb granuloma size, suggesting that it could also serve as a surrogate marker for Mtb burden and possibly treatment response. Conclusions: These biomarker selection and analysis approach appears to have strong potential utility for infectious disease diagnosis, including cryptic infections, and possibly to monitor changes in Mtb burden that may reflect disease progression or a response to treatment, which are critical needs for more effective disease control.