National Institute of Allergy and Infectious Diseases

Kenyan guidelines recommend syndromic management for sexually transmitted infections (STIs), which may miss many infections. We used nucleic acid amplification testing to screen for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) at 3 anatomic sites among adolescents and adults with behavioral risk factors in Western Kenya. From December 2021 to April 2023, we enrolled participants without HIV who were aged 14 to 55 years and reported a recent history of an STI, transactional sex, condomless sex with multiple partners, injection drug use, and/or anal sex with male partners. Urine, oropharyngeal swabs, and anorectal swabs were tested using Xpert CT/NG (Cepheid). Multivariable robust Poisson regression models were used to estimate prevalence ratios and 95% confidence intervals for factors potentially associated with the prevalence of both STIs combined. Among 405 participants tested for CT/NG at enrollment, 324 (80.0%) were females, and 324 (80.0%) were aged 14 to 24 years. CT and/or NG was diagnosed in 143 participants (35.3%), including CT in 111 (27.4%) and NG in 76 (18.8%). Infections were most prevalent at the urogenital site, with 124 (30.6%) participants diagnosed with urogenital CT and/or NG. Anorectal CT/NG was observed in 117 (28.9%) and oropharyngeal CT/NG in 14 (3.5%). Females had higher prevalences than males of both urogenital (33.6% vs. 18.5%, P = 0.012) and anorectal CT/NG (31.8% vs. 17.3%, P = 0.015). A theoretical screening approach that only tested for urogenital infections would have missed 18.0% of CT cases and 18.4% of NG cases. In multivariable analyses, there were no statistically significant associations between sociodemographic variables and CT/NG prevalence. Urogenital and anorectal CT/NG infections were common in 2 communities in Western Kenya with behavioral vulnerability but difficult to predict based on sociodemographic variables. Nucleic acid amplification testing-based urogenital and anorectal screening of sexually active adolescents and young adults could reduce CT/NG burden in this population.

Excessive cell death in the epithelium due to prolonged immune activation is associated with intestinal diseases such as Crohn’s disease. Mice with mutations in the Crohn’s disease susceptibility gene Atg16l1 are susceptible to inflammation associated with intestinal epithelial cell (IEC) death, including loss of antimicrobial Paneth cells triggered by infection with murine norovirus (MNV). Here, we show that intestinal disease downstream of MNV depends on IFN-α/β and IFN-λ signaling in IECs. In mouse organoids, IFNs synergize with TNF to induce RIPK1-dependent cell death, amplified by ATG16L1 deficiency. We further show that human intestinal organoids harboring the ATG16L1 risk allele exhibit heightened sensitivity to TNF and IFN co-stimulation and to serum from severe COVID-19 patients. Our findings reveal that virally triggered cytokines including IFNs exacerbate epithelial vulnerability in genetically predisposed hosts.

Understanding the genetic evolution of HIV-1 Transmitted/Founder (T/F) virus is crucial for developing effective treatment and prevention strategies due to its rapid mutation and recombination rates. This study compared the genetic diversity of 24 contemporary T/F viruses collected between 2016 and 2021 in Uganda and Kenya with 29 historical T/F sequences sampled between 2006 and 2011. Subtype analysis based on near-full-length (NFL) HIV-1 T/F genomes revealed that 57.1% (12/21) of contemporary viruses were recombinants, predominantly involving Subtype A1, D, and increasing Subtype C, with 33.3% (7/21) being A1D recombinants (A1 > D) and 19% (4/21) classified as complex recombinants involving three or more subtypes. Historical viruses showed a similar overall proportion (69%) but were mainly A1D mosaics (D > A1) with recombination confined primarily to the envelope region. In contrast, contemporary viruses shifted towards more complex recombinant patterns affecting additional genomic regions, including pol and accessory genes. Phylogenetic analysis demonstrated that contemporary viruses clustered into distinct, well-supported (98% bootstrap) sub-branches, suggesting divergency attributed to an imbalance in their proportions of subtype A1 and D sequences as well as a different content of A1 and D segments in the A1/D mosaic recombinants. These findings underscore the dynamic and shifting nature of HIV-1 genetic diversity in East Africa, highlighting the need for continuous molecular surveillance and region-specific treatment guidelines.

Transmission of highly pathogenic avian influenza from H5 clade 2.3.4.4b has expanded in recent years to infect large populations of birds and mammals, heightening the risk of a human pandemic. Influenza viruses that are adapted to transmission in birds and a variety of mammals tend to have a less stable hemagglutinin (HA) than seasonal influenza viruses, enabling membrane fusion at comparatively higher pH levels. Here, we combined five mutations in the H5 HA that increased its melting temperature and promoted stable closure of the HA trimer. Structural analysis by cryo–electron microscopy revealed that the stabilizing mutations create several new hydrophobic interactions while maintaining the local HA structure. We found that vaccinating mice with stabilized H5 HA immunogens resulted in higher hemagglutination inhibition and neutralization titers than nonstabilized comparators. Epitope mapping of vaccine-elicited polyclonal antibody responses using negative-stain electron microscopy and deep mutational scanning showed that site E on the side of the HA receptor binding domain was immunodominant across all groups; however, the stabilized immunogens shifted responses toward the receptor binding site, which elicited a higher proportion of neutralizing antibodies. Consistent with these findings, stabilized H5 HA immunogens delivered as messenger RNA–lipid nanoparticle (mRNA-LNP) vaccines protected mice against H5N1 challenge. These findings highlight that H5 HA–stabilizing mutations enhance the quality of antibody responses across different vaccine formats, underscoring their potential to improve pandemic preparedness vaccines targeting viruses from this widely circulating clade.

The biogenesis of integral membrane proteins is complex, as revealed by an ever-growing number of cellular components shown to be dedicated to the insertion, folding, surveillance, rectification, or quality control of specific client membrane proteins. The zinc metalloprotease ZMPSTE24 and its yeast homolog Ste24 have well-established roles in the proteolytic maturation of the nuclear scaffold protein lamin A and yeast a-factor, respectively. Additionally, Ste24 has been implicated through yeast genetic screens in a variety of membrane processes, including ER- associated degradation (ERAD), Sec61 translocon “unclogging,” the unfolded protein response (UPR), and potentially as a membrane protein topology determinant. Recently, an interaction was demonstrated between ZMPSTE24 and the antiviral interferon induced transmembrane protein IFITM3, although the functional significance of this interaction is not well-understood. IFITM3 is a tail-anchored protein with a cytoplasmic N-terminus, a single transmembrane span, and a lumenal/exocellular C-terminus. Here, we show that a catalytic-dead version of ZMPSTE24, ZMPSTE24E336A, exhibits enhanced binding to IFITM3, and this bound species of IFITM3 is hypo-palmitoylated. Using a split fluorescence topology reporter, we demonstrate that ZMPSTE24E336A “traps” and stabilizes a subpopulation of IFITM3 molecules with an atypical membrane topology, whose C-terminus is cytosolic instead of lumenal. Such inverted forms of IFITM3 are also detected in the presence of ERAD inhibitors when ZMPSTE24E336A is absent. We hypothesize the ZMPSTE24E336A trap mutant reveals a normally transient isoform of IFITM3 whose transmembrane span is inverted and that ZMPSTE24 is involved in the quality control of IFITM3 topology, either inverting, correcting or assisting in removal of aberrant IFITM3 molecules.

The discovery of novel chemical matter with antitubercular activity that could feed into the tuberculosis (TB) drug discovery pipeline addresses the need to develop novel drugs that inhibit growth of both drug sensitive and drug resistant strains of Mycobacterium tuberculosis (Mtb). Whole cell-based screening yielded a 2-amino-3,4-dihydroquinazoline core as a novel hit and preliminary structure-activity relationship (SAR) study around the structure was performed with 16 analogs derivatized in diverse structural points to determine the pharmacophore. The promising antitubercular activity of L16, the lead compound of this series was shown to be non-toxic to eukaryotic cells and had a novel mechanism of action since it lacked activity against known promiscuous targets of the pathogen. Additionally, L16 retained activity against different drug-resistant clinical strains. Although the MIC was improved significantly without any cytotoxicity, flat SAR, no identifiable target and equivalent activities of the two stereoisomers of the hit compound led us to abandon further optimization of this series.