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BackgroundAlthough vaccines are largely effective and safe, there is variability in post-vaccination experience regarding antibody response, side effects, and affective outcomes. Vaccine mindsets, specific beliefs about the vaccine, may be associated with post-vaccination experience. This is important since mindsets are malleable and may help decrease vaccine hesitancy and improve post-vaccination experience.MethodsIn a prospective study, we measured overall positive vaccine mindset and specific mindsets regarding efficacy, body response, and side effects. We tested whether vaccine mindsets before vaccination predicted neutralizing antibody response, side effects, vaccine-related stress, and affective outcomes (general stress, sadness, and happiness). Antibody response was assessed one month and six months after participants completed a SARS-CoV-2 vaccination series. Side effect experience and affective reactions were assessed daily on the vaccination day and the subsequent five days.ResultsThere was no significant association between the aggregate vaccine mindset score and neutralizing antibody response; however, people with a more positive vaccine mindset reported fewer side effects, less same-day vaccine-related anxiety, and improved affective outcomes after vaccination. In secondary analyses, when specific mindsets were explored, the mindset that vaccine side effects are a sign of treatment efficacy predicted higher antibodies, but not side effects experience and vaccine-related anxiety. Vaccine efficacy and body-response mindsets predicted fewer side effects, vaccine-related anxiety, and improved affective outcomes after vaccination.ConclusionThese findings underscore the potential of vaccine mindsets in enhancing the overall post-vaccination experience and, in some cases, increasing antibody response.

Abstract: Many herbivorous insect species exhibit phenotypic plasticity when using multiple hosts, which facilitates survival in heterogeneous host environments. Physiological host acclimation is an important part of it, yet the effects of host acclimation on insect feeding behaviour are not well studied, particularly for insect vectors of plant pathogens. We studied the combined effects of host acclimation and infection with a plant pathogenic symbiont on feeding behaviour of Bactericera cockerelli, an oligophagous psyllid widespread in both crop and natural habitats that feed primarily on Solanaceae and transmit an economically important plant pathogen, Candidatus Liberibacter solanacearum (CLso). We used a factorial design and the electrical penetration graphing technique to disentangle the effects of host acclimation, CLso infection and psyllid haplotype on the within‐plant feeding behaviour of B. cockerelli during conspecific and heterospecific host switches. This approach allows to connect phenotypic plasticity with the role of B. cockerelli as a vector by quantifying the frequency and duration of behaviours involved in CLso transmission. We found significant reductions in multiple metrics of B. cockerelli feeding efficiency, exacerbated by infection with CLso, which could lead to reduced transmission of this pathogen. Psyllid genotype was also important; the Central haplotype exhibited less dramatic changes in feeding efficiency than the Western haplotype during heterospecific host switches. Our study shows that host acclimation and heterospecific host switching directly alter feeding behaviours underlying pathogen transmission, and that the magnitude of feeding efficiency reductions depends on both host genotype and infection status.

The presence of HIV in sequestered reservoirs is a central impediment to a functional cure, allowing HIV to persist despite life-long antiretroviral therapy (ART), and driving a variety of comorbid conditions. Our understanding of the latent HIV reservoir in the central nervous system is incomplete, because of difficulties in accessing human central nervous system tissues. Microglia contribute to HIV reservoirs, but the molecular phenotype of HIV-infected microglia is poorly understood. We leveraged the unique "Last Gift" rapid autopsy program, in which people with HIV are closely followed until days or even hours before death. Microglial populations were heterogeneous regarding their gene expression profiles but showed similar chromatin accessibility landscapes. Despite ART, we detected occasional microglia containing cell-associated HIV RNA and HIV DNA integrated into open regions of the host's genome (∼0.005%). Microglia with detectable HIV RNA showed an inflammatory phenotype. These results demonstrate a distinct myeloid cell reservoir in the brains of people with HIV despite suppressive ART. Strategies for curing HIV and neurocognitive impairment will need to consider the myeloid compartment to be successful.

Immunocompromised patients are prone to recurrent Campylobacter infections. We report a case of recurrent multi-drug resistant Campylobactor jejuni bloodstream infections in a Bruton's X-linked agammaglobulinemia patient with prolonged ertapenem treatment. The isolate from the fifth recurrence developed carbapenem resistance, which is associated with mutations in a porin gene porA, and promoter changes and duplication of chromosomal blaOXA-61 gene. Combination therapy using cefepime and doxycycline (later switched to moxifloxacin) cleared the infection.

The rapid evolution of viruses generates proteins that are essential for infectivity and replication but with unknown functions, due to extreme sequence divergence1. Here, using a database of 67,715 newly predicted protein structures from 4,463 eukaryotic viral species, we found that 62% of viral proteins are structurally distinct and lack homologues in the AlphaFold database2,3. Among the remaining 38% of viral proteins, many have non-viral structural analogues that revealed surprising similarities between human pathogens and their eukaryotic hosts. Structural comparisons suggested putative functions for up to 25% of unannotated viral proteins, including those with roles in the evasion of innate immunity. In particular, RNA ligase T-like phosphodiesterases were found to resemble phage-encoded proteins that hydrolyse the host immune-activating cyclic dinucleotides 3',3'- and 2',3'-cyclic GMP-AMP (cGAMP). Experimental analysis showed that RNA ligase T homologues encoded by avian poxviruses similarly hydrolyse cGAMP, showing that RNA ligase T-mediated targeting of cGAMP is an evolutionarily conserved mechanism of immune evasion that is present in both bacteriophage and eukaryotic viruses. Together, the viral protein structural database and analyses presented here afford new opportunities to identify mechanisms of virus-host interactions that are common across the virome.

AimsWidespread use of immune checkpoint inhibitors (ICIs) for treatment of advanced malignancies led to an increase in number of immune-related adverse events such as ICI gastrointestinal (GI) injury (ICIGI). The resulting immune dysregulation of the GI mucosa is believed to predispose patients to viral infections. We characterised the histopathological features of ICIGI and the frequency of viral infections such as cytomegalovirus (CMV), adenovirus, and Epstein-Barr virus (EBV).MethodsSingle-centre retrospective study (2011-2020).Results81 GI biopsies from 31 patients with ICIGI (65% male (20/31), 35% female (11/31)) with advanced malignancies were reviewed. Most patients received ipilimumab and nivolumab (14/31, 45%), followed by pembrolizumab (9/31, 29%), ipilimumab (4/31, 13%), nivolumab (2/31, 6%) and combination of all three medications (2/31, 6%). Average regimen prior to incidence of diarrhea was three cycles. Evidence of colitis or erythema by endoscopy was present in 77% of cases, while 23% showed normal endoscopy. Histologically, the predominant ICIGI findings were active inflammation (84%), including cryptitis (77%), crypt abscesses (65%), lymphocytic colitis-like (LCL) pattern (61%), increase in epithelial apoptosis (74%) and/or surface injury (81%). Only one case showed diffuse CMV positivity (3%) with characteristic CMV viral cytopathic effects present on H&E stain and four cases were positive for rare EBV (13%). Adenovirus infection was not identified.ConclusionWhile our cohort is small, ICIGI generally demonstrates active inflammation including cryptitis and crypt abscesses in the colon, LCL pattern, and an increase in epithelial apoptosis. Upfront immunohistochemistry for viral infection without high-degree of clinical and histologic suspicion is not recommended.

Human papillomavirus (HPV) is the most common sexually transmitted infection on U.S. college campuses. Although HPV vaccination is recommended through age 26, current efforts to improve vaccination rates have predominantly focused on adolescents. Consequently, vaccine uptake remains suboptimal among young adults. This represents a significant missed opportunity, as young adults face the highest risk for new HPV infections. To contextualize the factors impacting decision-making process for this vulnerable population, this study reports key themes that emerged from in-depth interviews with participants (N = 30) who had completed an online intervention study for HPV vaccination among college students. Twelve (40%) of the interviewees vaccinated after exposure to the intervention. Findings centered around empowerment among young adults as the facilitator to get the HPV vaccine: key themes emerged were (1) convenience is critical and empowering; (2) adulthood identity, marked by a heightened sense of autonomy, accountability, and responsibility for self/future self and others, is empowering; (3) equal access to health care and preventive resources is empowering, especially for participants with low socioeconomic status; and (4) accurate knowledge provided in the intervention destigmatized HPV vaccination to empower young adults to make informed decisions. Digital interventions with messages highlighting a newly gained autonomy, future-oriented self and social responsibility, inclusive and accurate knowledge, and providing navigation to improve access may enhance HPV vaccination among young adults.

Infectious keratitis is a leading cause of corneal blindness worldwide with little information known about causative etiologies in Malawi, Africa. This area is resource-limited with ophthalmologist and microbiology services. The Department of Ophthalmology at the Kamuzu College of Health Sciences in Blantyre, Malawi, is a participating site of an international corneal ulcer consortium, capriCORN (Comprehensive Analysis of Pathogens, Resistomes, and Inflammatory-markers in the CORNea). In this study, 50 patients with corneal ulcers were swabbed for pathogen identification using RNA-sequencing. Corneal trauma was reported in 41% and 19% of the patients worked in agriculture. A pathogen was identified in 58% of the cases. Fungal pathogens predominated, followed by viruses and bacteria. Aspergillus, Fusarium, HSV-1, and Gardnerella were the most common pathogens detected. 50% of patients reported treatment with an antibiotic before presentation. Pathogens unusual for infectious keratitis, such as Subramaniula asteroids, Aureobasidium pullulans, and Gardnerella vaginalis, were also detected.

Human African trypanosomiasis, or sleeping sickness, is a neglected tropical disease caused by Trypanosoma brucei rhodesiense and Trypanosoma brucei gambiense and is invariably fatal unless treated. Current therapies present limitations in their application, parasite resistance, or require further clinical investigation for wider use. Our work, informed by previous findings, presents novel 4-[4-(4-methylpiperazin-1-yl)phenyl]-6-arylpyrimidine derivatives with promising antitrypanosomal activity. In particular, 32 exhibits an in vitro EC50 value of 0.5 µM against Trypanosoma brucei rhodesiense, and analogues 29, 30 and 33 show antitrypanosomal activities in the

There is little information available on antibiotic resistance (ABR) within shrimp aquaculture environments. The aim of this study was to investigate the presence of antibiotic resistance genes (ARGs) in shrimp farming operations in Atacames, Ecuador. Water samples (n = 162) and shrimp samples (n = 54) were collected from three shrimp farming operations. Samples were cultured and a subset of isolates that grew in the presence of ceftriaxone, a third-generation cephalosporin, were analyzed using whole-genome sequencing (WGS). Among the sequenced isolates (n = 44), 73% of the isolates contained at least one ARG and the average number of ARGs per isolate was two, with a median of 3.5 ARGs. Antibiotic resistance genes that confer resistance to the β-lactam class of antibiotics were observed in 65% of the sequenced isolates from water (20/31) and 54% of the isolates from shrimp (7/13). We identified 61 different ARGs across the 44 sequenced isolates, which conferred resistance to nine antibiotic classes. Over half of all sequenced isolates (59%, n = 26) carried ARGs that confer resistance to more than one class of antibiotics. ARGs for certain antibiotic classes were more common, including beta-lactams (26 ARGs); aminoglycosides (11 ARGs); chloramphenicol (three ARGs); and trimethoprim (four ARGs). Sequenced isolates consisted of a diverse array of bacterial orders and species, including Escherichia coli (48%), Klebsiella pneumoniae (7%), Aeromonadales (7%), Pseudomonadales (16%), Enterobacter cloacae (2%), and Citrobacter freundii (2%). Many ARGs were shared across diverse species, underscoring the risk of horizontal gene transfer in these environments. This study indicated the widespread presence of extended-spectrum β-lactamase (ESBL) genes in shrimp aquaculture, including blaCTX-M, blaSHV, and blaTEM genes. Increased antibiotic resistance surveillance of shrimp farms and identification of aquaculture operation-level risk factors, such as antibiotic use, will likely be important for mitigating the spread of ARGs of clinical significance.

Salmonella translocate to the gut epithelium via microfold cells lining the follicle-associated epithelium (FAE). How Salmonella localize to the FAE is not well characterized. Here we use live imaging and competitive assays between wild-type and chemotaxis-deficient mutants to show that Salmonella enterica serotype Typhimurium (S. Typhimurium) localize to the FAE independently of chemotaxis in an ex vivo mouse caecum infection model. Electrical recordings revealed polarized FAE with sustained outward current and small transepithelial potential, while the surrounding villus is depolarized with inward current and large transepithelial potential. The distinct electrical potentials attracted S. Typhimurium to the FAE while Escherichia coli (E. coli) localized to the villi, through a process called galvanotaxis. Chloride flux involving the cystic fibrosis transmembrane conductance regulator (CFTR) generated the ionic currents around the FAE. Pharmacological inhibition of CFTR decreased S. Typhimurium FAE localization but increased E. coli recruitment. Altogether, our findings demonstrate that bioelectric cues contribute to S. Typhimurium targeting of specific gut epithelial locations, with potential implications for other enteric bacterial infections.

As demonstrated by the SARS-CoV-2 pandemic, the emergence of novel viral strains with increased transmission rates poses a serious threat to global health. Statistical models of genome sequence evolution may provide a critical tool for early detection of these strains. Using a novel stochastic model that links transmission rates to the entire viral genome sequence, we study the utility of phylogenetic methods that use a phylogenetic tree relating viral samples versus count-based methods that use case counts of variants over time exclusively to detect increased transmission rates and identify candidate causative mutations. We find that phylogenies in particular can detect novel transmission-enhancing variants very soon after their origin and may facilitate the development of early detection systems for outbreak surveillance.

AbstractBabesiosis, caused by protozoan parasites of the genus Babesia, is an emerging tick-borne disease of significance for both human and animal health. Babesia parasites infect erythrocytes of vertebrate hosts where they develop and multiply rapidly to cause the pathological symptoms associated with the disease. The identification of new Babesia species underscores the ongoing risk of zoonotic pathogens capable of infecting humans, a concern amplified by anthropogenic activities and environmental changes. One such pathogen, Babesia MO1, previously implicated in severe cases of human babesiosis in the United States, was initially considered a subspecies of B. divergens, the predominant agent of human babesiosis in Europe. Here we report comparative multiomics analyses of B. divergens and B. MO1 that offer insight into their biology and evolution. Our analysis shows that despite their highly similar genomic sequences, substantial genetic and genomic divergence occurred throughout their evolution resulting in major differences in gene functions, expression and regulation, replication rates and susceptibility to antiparasitic drugs. Furthermore, both pathogens have evolved distinct classes of multigene families, crucial for their pathogenicity and adaptation to specific mammalian hosts. Leveraging genomic information for B. MO1, B. divergens, and other members of the Babesiidae family within Apicomplexa provides valuable insights into the evolution, diversity, and virulence of these parasites. This knowledge serves as a critical tool in preemptively addressing the emergence and rapid transmission of more virulent strains.

Scrub typhus, a vector-borne bacterial infection, is an important but neglected disease globally. Accurately characterizing the burden is challenging because of nonspecific symptoms and limited diagnostics. Prior seroepidemiology studies have struggled to find consensus cutoffs that permit comparisons of estimates across contexts and time. In this study, we present a novel approach that does not require a cutoff and instead uses information about antibody kinetics after infection to estimate seroincidence. We use data from three cohorts of scrub typhus patients in Chiang Rai, Thailand, and Vellore, India, to characterize antibody kinetics after infection and two population serosurveys in the Kathmandu Valley, Nepal, and Tamil Nadu, India, to estimate seroincidence. The samples were tested for IgM and IgG responses to Orientia tsutsugamushi-derived recombinant 56-kDa antigen using commercial enzyme-linked immunosorbent assay kits. We used Bayesian hierarchical models to characterize antibody responses after scrub typhus infection and used the joint distributions of the peak antibody titers and decay rates to estimate population-level incidence rates in the cross-sectional serosurveys. Median responses persisted above an optical density (OD) of 1.8 for 23.6 months for IgG and an OD of 1 for 4.5 months for IgM. Among 18- to 29-year-olds, the seroincidence was 10 per 1,000 person-years (95% CI, 5-19) in Tamil Nadu, India, and 14 per 1,000 person-years (95% CI: 10-20) in the Kathmandu Valley, Nepal. When seroincidence was calculated with antibody decay ignored, the disease burden was underestimated by more than 50%. The approach can be deployed prospectively, coupled with existing serosurveys, or leverage banked samples to efficiently generate scrub typhus seroincidence estimates.

In the pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, epithelial populations in the distal lung expressing Angiotensin-converting enzyme 2 (ACE2) are infrequent, and therefore, the model of viral expansion and immune cell engagement remains incompletely understood. Using human lungs to investigate early host-viral pathogenesis, we found that SARS-CoV-2 had a rapid and specific tropism for myeloid populations. Human alveolar macrophages (AMs) reliably expressed ACE2 allowing both spike-ACE2-dependent viral entry and infection. In contrast to Influenza A virus, SARS-CoV-2 infection of AMs was productive, amplifying viral titers. While AMs generated new viruses, the interferon responses to SARS-CoV-2 were muted, hiding the viral dissemination from specific antiviral immune responses. The reliable and veiled viral depot in myeloid cells in the very early phases of SARS-CoV-2 infection of human lungs enables viral expansion in the distal lung and potentially licenses subsequent immune pathologies.

People with HIV and comorbid substance use problems may be among those who benefit most from long-acting HIV antiretroviral treatment, but they are routinely excluded from Phase 3 clinical trials. Their inclusion would permit an examination of the clinical value of long-acting therapies for people with adherence problems and an exploration of syndemic interactions between HIV, mental health conditions, and substance use problems, which compound into a major challenge in the efforts to end the HIV epidemic.

BACKGROUND: Exposure to antiretrovirals at or early after HIV acquisition can suppress viral replication and blunt antibody (Ab) responses; a reduced HIV detectability could impact diagnosis and blood donation screening. METHODS: We used three antigen (Ag)/Ab assays and one nucleic acid test (NAT) to analyze samples collected in pre-exposure prophylaxis (PrEP) trials (iPrEx; Partners PrEP) before infection detection by Ab-only rapid diagnostic tests (RDTs), and in early antiretroviral treatment (ART) initiation studies (RV254; SIPP). RESULTS: Reactivity using NAT and Ag/Ab assays in samples collected up to 8 weeks prior to the first reactive RDT from 251 PrEP trials participants varied between 49-61% for active PrEP users and between 27-37% for placebo users. Among RV254 participants, reactivity in Ag/Ab assays was

BackgroundDespite national testing guidelines, rates of testing for HIV, sexually transmitted infections, and hepatitis C remain lower than recommended for men who have sex with men (MSM) in the US. To help address this, the TakeMeHome (TMH) program was started in March 2020 by a consortium of public health organizations and dating apps - Building Healthy Online Communities - to work with health departments to increase access to HIV testing for MSM on dating apps.MethodsUsers of participating dating apps were sent messages about opportunities for testing with self-collected specimens through TMH. Program users were eligible to receive test kits if they lived in a participating zip code and were aged at least 18. Users who were interested in testing could order kits to be mailed to them for lab-based testing of HIV, hepatitis C, chlamydia, gonorrhea, and/or syphilis, depending on risk and availability in their zip code. Orders were sent via application programming interface (API) to Molecular Testing Labs (MTL) for fulfillment; kits were provided at no cost to the program user. Within approximately 24 hours of order receipt, MTL mailed program users a kit with required collection supplies, directions, and a link to a video instruction for self-collection. Program users received an automated email after testing was complete with a link to access results through their online account. Individuals with positive results on any of the relevant tests were directed to additional information and supported with linkage to additional testing or treatment, depending on local protocols.ResultsThe positivity rate of specimens processed through TMH was 1.4% for HIV, 0.6% for hepatitis C, and 2.9% for all STIs combined. The per-person positivity rate was 15.3% across all STIs.Conclusions: The TakeMeHome program demonstrates that self-collected lab-processed testing is feasible and effective at identifying new HIV and STI cases.

Z-nucleic acid structures play vital roles in cellular processes and have implications in innate immunity due to their recognition by Zα domains containing proteins (Z-DNA/Z-RNA binding proteins, ZBPs). Although Zα domains have been identified in six proteins, including viral E3L, ORF112, and I73R, as well as, cellular ADAR1, ZBP1, and PKZ, their prevalence across living organisms remains largely unexplored. In this study, we introduce a computational approach to predict Zα domains, leading to the revelation of previously unidentified Zα domain-containing proteins in eukaryotic organisms, including non-metazoan species. Our findings encompass the discovery of new ZBPs in previously unexplored giant viruses, members of the Nucleocytoviricota phylum. Through experimental validation, we confirm the Zα functionality of select proteins, establishing their capability to induce the B-to-Z conversion. Additionally, we identify Zα-like domains within bacterial proteins. While these domains share certain features with Zα domains, they lack the ability to bind to Z-nucleic acids or facilitate the B-to-Z DNA conversion. Our findings significantly expand the ZBP family across a wide spectrum of organisms and raise intriguing questions about the evolutionary origins of Zα-containing proteins. Moreover, our study offers fresh perspectives on the functional significance of Zα domains in virus sensing and innate immunity and opens avenues for exploring hitherto undiscovered functions of ZBPs.

BackgroundHepatitis-E virus (HEV), an etiologic agent of acute inflammatory liver disease, is a significant cause of morbidity and mortality in South Asia. HEV is considered endemic in Nepal; but data on population-level infection transmission is sparse.MethodsWe conducted a longitudinal serosurvey in central Nepal to assess HEV exposure. At each visit, capillary blood samples were collected and analyzed for the presence of anti-HEV IgG antibodies. The study took place between February 2019 and April 2021, with up to 4 visits per participant approximately 6 months apart.ResultsWe collected 2513 samples from 923 participants aged 0-25 years, finding a seroprevalence of 4.8% and a seroincidence rate of 10.9 per 1000 person-years. Young adults and individuals consuming surface water faced the highest incidence of infection. Geospatial analysis identified potential HEV clusters, suggesting a need for targeted interventions.SignificanceOur findings demonstrate that HEV is endemic in Nepal and that the risk of infection increases with age.

ImportanceMost research to understand postacute sequelae of SARS-CoV-2 infection (PASC), or long COVID, has focused on adults, with less known about this complex condition in children. Research is needed to characterize pediatric PASC to enable studies of underlying mechanisms that will guide future treatment.ObjectiveTo identify the most common prolonged symptoms experienced by children (aged 6 to 17 years) after SARS-CoV-2 infection, how these symptoms differ by age (school-age [6-11 years] vs adolescents [12-17 years]), how they cluster into distinct phenotypes, and what symptoms in combination could be used as an empirically derived index to assist researchers to study the likely presence of PASC.Design, setting, and participantsMulticenter longitudinal observational cohort study with participants recruited from more than 60 US health care and community settings between March 2022 and December 2023, including school-age children and adolescents with and without SARS-CoV-2 infection history.ExposureSARS-CoV-2 infection.Main outcomes and measuresPASC and 89 prolonged symptoms across 9 symptom domains.ResultsA total of 898 school-age children (751 with previous SARS-CoV-2 infection [referred to as infected] and 147 without [referred to as uninfected]; mean age, 8.6 years; 49% female; 11% were Black or African American, 34% were Hispanic, Latino, or Spanish, and 60% were White) and 4469 adolescents (3109 infected and 1360 uninfected; mean age, 14.8 years; 48% female; 13% were Black or African American, 21% were Hispanic, Latino, or Spanish, and 73% were White) were included. Median time between first infection and symptom survey was 506 days for school-age children and 556 days for adolescents. In models adjusted for sex and race and ethnicity, 14 symptoms in both school-age children and adolescents were more common in those with SARS-CoV-2 infection history compared with those without infection history, with 4 additional symptoms in school-age children only and 3 in adolescents only. These symptoms affected almost every organ system. Combinations of symptoms most associated with infection history were identified to form a PASC research index for each age group; these indices correlated with poorer overall health and quality of life. The index emphasizes neurocognitive, pain, and gastrointestinal symptoms in school-age children but change or loss in smell or taste, pain, and fatigue/malaise-related symptoms in adolescents. Clustering analyses identified 4 PASC symptom phenotypes in school-age children and 3 in adolescents.Conclusions and relevanceThis study developed research indices for characterizing PASC in children and adolescents. Symptom patterns were similar but distinguishable between the 2 groups, highlighting the importance of characterizing PASC separately for these age ranges.

BackgroundSmall incentives could improve engagement in HIV care. We evaluated the short-term and longer-term effects of financial incentives for visit attendance on viral suppression among adults initiating antiretroviral therapy (ART) in Tanzania.MethodsIn a type 1 hybrid effectiveness-implementation study, we randomised (1:1) 32 primary care HIV clinics in four Tanzanian regions to usual care (control group) or the intervention (usual care plus ≤6 monthly incentives [22 500 Tanzanian Shillings, about US$10, each], conditional on visit attendance). Adults (aged ≥18 years) initiating ART (

ABSTRACT : Introduction:: Gastrointestinal (GI) parasites are major health concerns in both humans and domestic animals. Livestock farming is one of the common livelihood practices in rural Nepal. The proximity at human and domestic animal interface increases the chances of dissemination of enteric parasites, especially those of zoonotic importance. This study was aimed at finding the parasite prevalence and risk factors in both humans and their domestic animals in Jirel community. Materials and Methods:: A field survey was conducted on the Jirel ethnic people and their domestic animals in Dolakha district, where a total of 152 fresh fecal samples from humans and domestic animals (cow, pigs, goats, chickens, ducks, and pigeons) were collected. The feces were examined by wet mounts and concentration techniques. A structured questionnaire survey was carried out among the local people and owners of the domestic animals to gather sociodemographic information, awareness, and hygienic practices in relation to parasite transmission Results:: The enteric parasite prevalence was found to be highest in goats (80.0%;12/15), followed by pigs (55.55%;5/9), cows (45.45%;6/11), chickens (11.7%;4/34), and humans (1.41%;1/71), while the fecal samples of ducks and pigeons did not contain any parasites. The only parasite identified in humans was Ascaris lumbricoides. Similarly, three genera of GI parasites (Eimeria sp., Strongyloides sp, and Trichuris sp.) from goats, two genera each from cow (Eimeria sp. and Strongyloides sp.), pigs (Entamoeba sp. and A. suum), and chickens (Eimeria sp. and Ascaridia galli), were detected Conclusions:: Based on the direct field observation, questionnaire survey and laboratory analysis, it is concluded that the Jirel community people are aware of health and hygiene; however, intervention measures are necessary to prevent parasitic infection in their domestic animals.

BackgroundHIV testing rates among South African men lag behind rates for women and national targets. Community-based HIV self-screening (HIVSS) distribution and follow-up by community health workers (CHWs) is a scalable option to increase testing coverage, diagnosis, and treatment initiation. We provided HIVSS and assisted linkage to care to men not recently tested (within the past 12 months) residing in high-HIV-burden areas of Johannesburg.MethodsCHWs distributed HIVSS in 6 clinic catchment areas. Follow-up to encourage confirmatory testing and antiretroviral therapy initiation was conducted through personal support (PS) or an automated short message service (SMS) follow-up and linkage system in 3 clinic areas each. Using a quasi-experimental pre-post design, we compared differences in the proportion of men testing in the clinic catchment areas during the HIVSS campaign (June-August 2019) to the 3 months prior (March-May 2019) and compared treatment initiations by assisted linkage strategy.ResultsAmong 4793 participants accepting HIVSS, 62% had never tested. Among 3993 participants with follow-up data, 90.6% reported using their HIVSS kit. Testing coverage among men increased by 156%, from under 4% when only clinic-based HIV testing services were available to 9.5% when HIVSS and HIV testing services were available (z = -11.6; P < 0.01). Reported test use was higher for men followed through PS (99% vs. 68% in SMS); however, significantly more men reported reactive self-test results in the SMS group compared with PS (6.4% vs. 2.0%), resulting in more antiretroviral therapy initiations in the SMS group compared with PS (23 vs. 9; P < 0.01).ConclusionsCHW HIVSS distribution significantly increases testing among men. While PS enabled personalized follow-up, reporting differences indicate SMS is more acceptable and better aligned with expectations of privacy associated with HIVSS.

Language barriers are major obstacles that Asian American immigrants face when accessing health care in the USA. This study was conducted to explore the impact of language barriers and facilitators on the health care of Asian Americans. Qualitative, in-depth interviews and quantitative surveys were conducted with 69 Asian Americans (Chinese, Filipino, Japanese, Malaysian, Indonesian, Vietnamese, and mixed Asian backgrounds) living with HIV (AALWH) in three urban areas (New York, San Francisco, and Los Angeles) in 2013 and from 2017 to 2020. The quantitative data indicate that language ability is negatively associated with stigma. Major themes emerged related to communication, including the impact of language barriers on HIV care and the positive impact of language facilitators-family members/friends, case managers, or interpreters-who can communicate with healthcare providers in the AALWH's native language. Language barriers negatively impact access to HIV-related services and thus result in decreased adherence to antiretroviral therapy, increased unmet healthcare needs, and increased HIV-related stigma. Language facilitators enhanced the connection between AALWH and the healthcare system by facilitating their engagement with health care providers. Language barriers experienced by AALWH not only impact their healthcare decisions and treatment choices but also increase levels of external stigma which may influence the process of acculturation to the host country. Language facilitators and barriers to health services for AALWH represent a target for future interventions in this population.

Across sub-Saharan Africa, men are less likely to know their HIV status than women, leading to later treatment initiation. Little is known about how experiences with general health services affect men's use of HIV testing. We used data from a 2019 community-representative survey of men in Malawi to understand frequency and cause of men's negative health service experiences (defined as men reporting they "would not recommend" a facility) and their association with future HIV testing. We conducted univariable and multivariable logistic regressions to determine which aspects of health facility visits were associated with would-not-recommend experiences and to determine if would-not-recommend experiences 12-24 months prior to the survey were associated with HIV testing in the 12 months prior to the survey. Among 1,098 men eligible for HIV testing in the 12 months prior to the survey, median age was 34 years; 9% of men reported at least one would-not-recommend experience, which did not differ by sociodemographics, gender norm beliefs, or HIV stigma beliefs. The factors most strongly associated with would-not-recommend experiences were cost (aOR 5.8, 95%CI 2.9-11.4), cleanliness (aOR 4.2, 95%CI 1.8-9.9), medicine availability (aOR 3.3, 95%CI 1.7-6.4), and wait times (aOR 2.7, 95%CI 1.5-5.0). Reporting a would-not-recommend experience 12-24 months ago was associated with a 59% decrease in likelihood of testing for HIV in the last 12 months (aOR 0.41; 95% CI:0.17-0.96). Dissatisfaction with general health services was strongly associated with reduced HIV testing. Coverage of high-priority screening services like HIV testing may benefit from improving overall health system quality.

Candida parapsilosis is known to cause severe and persistent outbreaks in clinical settings. Patients infected with multidrug-resistant C. parapsilosis (MDR Cp) isolates were identified in a large Turkish hospital from 2017-2020. We subsequently identified three additional patients infected with MDR Cp isolates in 2022 from the same hospital and two echinocandin-resistant (ECR) isolates from a single patient in another hospital. The increasing number of MDR and ECR isolates contradicts the general principle that the severe fitness cost associated with these phenotypes could prevent their dominance in clinical settings. Here, we employed a multidimensional approach to systematically assess the fitness costs of MDR and ECR C. parapsilosis isolates. Whole-genome sequencing revealed a novel MDR genotype infecting two patients in 2022. Despite severe in vitro defects, the levels and tolerances of the biofilms of our ECR and MDR isolates were generally comparable to those of susceptible wild-type isolates. Surprisingly, the MDR and ECR isolates showed major alterations in their cell wall components, and some of the MDR isolates consistently displayed increased tolerance to the fungicidal activities of primary human neutrophils and were more immunoevasive during exposure to primary human macrophages. Our systemic infection mouse model showed that MDR and ECR C. parapsilosis isolates had comparable fungal burden in most organs relative to susceptible isolates. Overall, we observed a notable increase in the genotypic diversity and frequency of MDR isolates and identified MDR and ECR isolates potentially capable of causing persistent outbreaks in the future.

Bile acids (BA) are among the most abundant metabolites produced by the gut microbiome. Primary BAs produced in the liver are converted by gut bacterial 7-α-dehydroxylation into secondary BAs, which can differentially regulate host health via signaling based on their varying affinity for BA receptors. Despite the importance of secondary BAs in host health, the regulation of 7-α-dehydroxylation and the role of diet in modulating this process is incompletely defined. Understanding this process could lead to dietary guidelines that beneficially shift BA metabolism. Dietary fiber regulates gut microbial composition and metabolite production. We tested the hypothesis that feeding mice a diet rich in a fermentable dietary fiber, resistant starch (RS), would alter gut bacterial BA metabolism. Male and female wild-type mice were fed a diet supplemented with RS or an isocaloric control diet (IC). Metabolic parameters were similar between groups. RS supplementation increased gut luminal deoxycholic acid (DCA) abundance. However, gut luminal cholic acid (CA) abundance, the substrate for 7-α-dehydroxylation in DCA production, was unaltered by RS. Further, RS supplementation did not change the mRNA expression of hepatic BA producing enzymes or ileal BA transporters. Metagenomic assessment of gut bacterial composition revealed no change in the relative abundance of bacteria known to perform 7-α-dehydroxylation. P. ginsenosidimutans and P. multiformis were positively correlated with gut luminal DCA abundance and increased in response to RS supplementation. These data demonstrate that RS supplementation enriches gut luminal DCA abundance without increasing the relative abundance of bacteria known to perform 7-α-dehydroxylation.

While the effector cells that mediate anti-tumor immunity have historically been attributed to αβ T cells and natural killer cells, γδ T cells are now being recognized as a complementary mechanism mediating tumor rejection. γδ T cells possess a host of functions ranging from antigen presentation to regulatory function and, importantly, have critical roles in eliciting anti-tumor responses where other immune effectors may be rendered ineffective. Recent discoveries have elucidated how these differing functions are mediated by γδ T cells with specific T cell receptors and spatial distribution. Their relative resistance to mechanisms of dysfunction like T cell exhaustion has spurred the development of therapeutic approaches exploiting γδ T cells, and an improved understanding of these cells should enable more effective immunotherapies.

Bacteriophage ϕ6 is a segmented dsRNA virus with a lipid envelope, which are unusual traits in bacterial viruses but common in eukaryotic viruses. This uniqueness allowed ϕ6 and its Pseudomonad hosts to serve as a molecular model for RNA genetics, mutation, replication, packaging, and reassortment in both bacterial and eukaryotic viruses. However, an additional uniqueness of ϕ6, created by its high mutation rate, was its use as an experimental system to study key questions such as the evolution of sex (segment reassortment), host-pathogen interactions, mutational load, rates of adaptation, genetic and phenotypic complexity, and game theory.

The prevalence of "long COVID" is just one of the conundrums highlighting how little we know about the lung's response to viral infection, particularly to syndromecoronavirus-2 (SARS-CoV-2), for which the lung is the point of entry. We used an in vitro human lung system to enable a prospective, unbiased, sequential single-cell level analysis of pulmonary cell responses to infection by multiple SARS-CoV-2 strains. Starting with human induced pluripotent stem cells and emulating lung organogenesis, we generated and infected three-dimensional, multi-cell-type-containing lung organoids (LOs) and gained several unexpected insights. First, SARS-CoV-2 tropism is much broader than previously believed: Many lung cell types are infectable, if not through a canonical receptor-mediated route (e.g., via Angiotensin-converting encyme 2(ACE2)) then via a noncanonical "backdoor" route (via macropinocytosis, a form of endocytosis). Food and Drug Administration (FDA)-approved endocytosis blockers can abrogate such entry, suggesting adjunctive therapies. Regardless of the route of entry, the virus triggers a lung-autonomous, pulmonary epithelial cell-intrinsic, innate immune response involving interferons and cytokine/chemokine production in the absence of hematopoietic derivatives. The virus can spread rapidly throughout human LOs resulting in mitochondrial apoptosis mediated by the prosurvival protein Bcl-xL. This host cytopathic response to the virus may help explain persistent inflammatory signatures in a dysfunctional pulmonary environment of long COVID. The host response to the virus is, in significant part, dependent on pulmonary Surfactant Protein-B, which plays an unanticipated role in signal transduction, viral resistance, dampening of systemic inflammatory cytokine production, and minimizing apoptosis. Exogenous surfactant, in fact, can be broadly therapeutic.

Bacteroides fragilis is a Gram-negative commensal bacterium commonly found in the human colon, which differentiates into two genomospecies termed divisions I and II. Through a comprehensive collection of 694 B. fragilis whole genome sequences, we identify novel features distinguishing these divisions. Our study reveals a distinct geographic distribution with division I strains predominantly found in North America and division II strains in Asia. Additionally, division II strains are more frequently associated with bloodstream infections, suggesting a distinct pathogenic potential. We report differences between the two divisions in gene abundance related to metabolism, virulence, stress response, and colonization strategies. Notably, division II strains harbor more antimicrobial resistance (AMR) genes than division I strains. These findings offer new insights into the functional roles of division I and II strains, indicating specialized niches within the intestine and potential pathogenic roles in extraintestinal sites.ImportanceUnderstanding the distinct functions of microbial species in the gut microbiome is crucial for deciphering their impact on human health. Classifying division II strains as Bacteroides fragilis can lead to erroneous associations, as researchers may mistakenly attribute characteristics observed in division II strains to the more extensively studied division I B. fragilis. Our findings underscore the necessity of recognizing these divisions as separate species with distinct functions. We unveil new findings of differential gene prevalence between division I and II strains in genes associated with intestinal colonization and survival strategies, potentially influencing their role as gut commensals and their pathogenicity in extraintestinal sites. Despite the significant niche overlap and colonization patterns between these groups, our study highlights the complex dynamics that govern strain distribution and behavior, emphasizing the need for a nuanced understanding of these microorganisms.

ObjectivesSome drugs that augment cell-intrinsic defenses or modulate cell death/survival pathways have been reported to selectively kill cells infected with HIV or Simian Immunodeficiency Virus (SIV), but comparative studies are lacking. We hypothesized that these drugs may differ in their ability to kill cells infected with intact and defective proviruses.DesignTo investigate this hypothesis, drugs were tested ex vivo on peripheral blood mononuclear cells (PBMC) from nine antiretroviral therapy (ART)-suppressed individuals.MethodsWe tested drugs currently in clinical use or human trials, including auranofin (p53 modulator), interferon alpha2A, interferon gamma, acitretin (RIG-I inducer), GS-9620/vesatolimod (TLR7 agonist), nivolumab (PD-1 blocker), obatoclax (Bcl-2 inhibitor), birinapant [inhibitor of apoptosis proteins (IAP) inhibitor], bortezomib (proteasome inhibitor), and INK128/sapanisertib [mammalian target of rapamycin mTOR] [c]1/2 inhibitor). After 6 days of treatment, we measured cell counts/viabilities and quantified levels of total, intact, and defective HIV DNA by droplet digital PCR (Intact Proviral DNA Assay).ResultsObatoclax reduced intact HIV DNA [median = 27-30% of dimethyl sulfoxide control (DMSO)] but not defective or total HIV DNA. Other drugs showed no statistically significant effects.ConclusionObatoclax and other Bcl-2 inhibitors deserve further study in combination therapies aimed at reducing the intact HIV reservoir in order to achieve a functional cure and/or reduce HIV-associated immune activation.

Central nervous system (CNS) infections carry a substantial burden of morbidity and mortality worldwide, and accurate and timely diagnosis is required to optimize management. Metagenomic next-generation sequencing (mNGS) has proven to be a valuable tool in detecting pathogens in patients with suspected CNS infection. By sequencing microbial nucleic acids present in a patient's cerebrospinal fluid, brain tissue, or samples collected outside of the CNS, such as plasma, mNGS can detect a wide range of pathogens, including rare, unexpected, and/or fastidious organisms. Furthermore, its target-agnostic approach allows for the identification of both known and novel pathogens. This is particularly useful in cases where conventional diagnostic methods fail to provide an answer. In addition, mNGS can detect multiple microorganisms simultaneously, which is crucial in cases of mixed infections without a clear predominant pathogen. Overall, clinical mNGS testing can help expedite the diagnostic process for CNS infections, guide appropriate management decisions, and ultimately improve clinical outcomes. However, there are key challenges surrounding its use that need to be considered to fully leverage its clinical impact. For example, only a few specialized laboratories offer clinical mNGS due to the complexity of both the laboratory methods and analysis pipelines. Clinicians interpreting mNGS results must be aware of both false negatives-as mNGS is a direct detection modality and requires a sufficient amount of microbial nucleic acid to be present in the sample tested-and false positives-as mNGS detects environmental microbes and their nucleic acids, despite best practices to minimize contamination. Additionally, current costs and turnaround times limit broader implementation of clinical mNGS. Finally, there is uncertainty regarding the best practices for clinical utilization of mNGS, and further work is needed to define the optimal patient population(s), syndrome(s), and time of testing to implement clinical mNGS.

Virulent infectious agents such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and methicillin-resistant Staphylococcus aureus (MRSA) induce tissue damage that recruits neutrophils, monocyte, and macrophages, leading to T cell exhaustion, fibrosis, vascular leak, epithelial cell depletion, and fatal organ damage. Neutrophils, monocytes, and macrophages recruited to pathogen-infected lungs, including SARS-CoV-2-infected lungs, express phosphatidylinositol 3-kinase gamma (PI3Kγ), a signaling protein that coordinates both granulocyte and monocyte trafficking to diseased tissues and immune-suppressive, profibrotic transcription in myeloid cells. PI3Kγ deletion and inhibition with the clinical PI3Kγ inhibitor eganelisib promoted survival in models of infectious diseases, including SARS-CoV-2 and MRSA, by suppressing inflammation, vascular leak, organ damage, and cytokine storm. These results demonstrate essential roles for PI3Kγ in inflammatory lung disease and support the potential use of PI3Kγ inhibitors to suppress inflammation in severe infectious diseases.

Controlling the principal African malaria vector, the mosquito Anopheles gambiae, is considered essential to curtail malaria transmission. However, existing vector control technologies rely on insecticides, which are becoming increasingly ineffective. Sterile insect technique (SIT) is a powerful suppression approach that has successfully eradicated a number of insect pests, yet the A. gambiae toolkit lacks the requisite technologies for its implementation. SIT relies on iterative mass releases of nonbiting, nondriving, sterile males which seek out and mate with monandrous wild females. Once mated, females are permanently sterilized due to mating-induced refractoriness, which results in population suppression of the subsequent generation. However, sterilization by traditional methods renders males unfit, making the creation of precise genetic sterilization methods imperative. Here, we introduce a vector control technology termed precision-guided sterile insect technique (pgSIT), in A. gambiae for inducible, programmed male sterilization and female elimination for wide-scale use in SIT campaigns. Using a binary CRISPR strategy, we cross separate engineered Cas9 and gRNA strains to disrupt male-fertility and female-essential genes, yielding >99.5% male sterility and >99.9% female lethality in hybrid progeny. We demonstrate that these genetically sterilized males have good longevity, are able to induce sustained population suppression in cage trials, and are predicted to eliminate wild A. gambiae populations using mathematical models, making them ideal candidates for release. This work provides a valuable addition to the malaria genetic biocontrol toolkit, enabling scalable SIT-like confinable, species-specific, and safe suppression in the species.

Although enacted and internalized stigma is a continuing problem for people living with HIV (PLWH) in Southeast Asia, there is little understanding of how PLWH cope with discrimination, exclusion, and other negative outcomes caused by HIV-related stigmatization. This article aims to bridge this gap by analyzing the lived experiences of HIV-related stigmatization and coping strategies among 30 people with HIV in Myanmar, a country heavily influenced by religion, especially Buddhism. Among the 30 study participants, 20 were female and 10 were male, with ages ranging from 18 to 50 years. Through the lens of Bourdieu's concepts of habitus, field, and capital, this article first elucidates the various forms of stigmatization in family, work, social, and other settings as symbolic violence on people with HIV. The present article shows that spirituality serves as a perceptual and action framework for people with HIV to generate reflexivity toward their HIV infection and related stigmatization and to further engage in agentic responses. More importantly, this article demonstrates how people with HIV draw on spirituality to support peers in reclaiming control over their lives and how they are perceived by society. The findings indicate that the local context, especially cultural and religious resources, should be considered when developing interventions to mitigate HIV-related stigmatization in Southeast Asia.

Cluster randomized trials (CRTs) often enroll large numbers of participants; yet due to resource constraints, only a subset of participants may be selected for outcome assessment, and those sampled may not be representative of all cluster members. Missing data also present a challenge: if sampled individuals with measured outcomes are dissimilar from those with missing outcomes, unadjusted estimates of arm-specific endpoints and the intervention effect may be biased. Further, CRTs often enroll and randomize few clusters, limiting statistical power and raising concerns about finite sample performance. Motivated by SEARCH-TB, a CRT aimed at reducing incident tuberculosis infection, we demonstrate interlocking methods to handle these challenges. First, we extend Two-Stage targeted minimum loss-based estimation to account for three sources of missingness: (i) subsampling; (ii) measurement of baseline status among those sampled; and (iii) measurement of final status among those in the incidence cohort (persons known to be at risk at baseline). Second, we critically evaluate the assumptions under which subunits of the cluster can be considered the conditionally independent unit, improving precision and statistical power but also causing the CRT to behave like an observational study. Our application to SEARCH-TB highlights the real-world impact of different assumptions on measurement and dependence; estimates relying on unrealistic assumptions suggested the intervention increased the incidence of TB infection by 18% (risk ratio [RR]=1.18, 95% confidence interval [CI]: 0.85-1.63), while estimates accounting for the sampling scheme, missingness, and within community dependence found the intervention decreased the incident TB by 27% (RR=0.73, 95% CI: 0.57-0.92).

We evaluated the HIV-1 DNA reservoir in peripheral blood mononuclear cells (PBMC) and cerebrospinal fluid (CSF) in people with HIV (PWH) and associations to cognitive dysfunction. Using the intact proviral DNA assay (IPDA), an emerging technique to identify provirus that may be the source of viral rebound, we assessed HIV DNA in CSF and PBMC in PWH regardless of antiretroviral therapy (ART). CSF was used as a sampling surrogate for the central nervous system (CNS) as opposed to tissue. IDPA results (3' defective, 5' defective, and intact HIV DNA) were analyzed by compartment (Wilcoxon signed rank; matched and unmatched pairs). Cognitive performance, measured via a battery of nine neuropsychological (NP) tests, were analyzed for correlation to HIV DNA (Spearman's rho). 11 CSF and 8 PBMC samples from PWH were evaluated both unmatched and matched. Total CSF HIV DNA was detectable in all participants and was significantly higher than in matched PBMCs (p ​= ​0.0039). Intact CSF HIV DNA was detected in 7/11 participants and correlated closely with those in PBMCs but tended to be higher in CSF than in PBMC. CSF HIV DNA did not correlate with global NP performance, but higher values did correlate with worse executive function (p ​= ​0.0440). Intact HIV DNA is frequently present in the CSF of PWH regardless of ART. This further supports the presence of an HIV CNS reservoir and provides a method to study CNS reservoirs during HIV cure studies. Larger studies are needed to evaluate relationships with CNS clinical outcomes.

IntroductionIdentifying the optimal approaches to offering HIV prevention to meet the needs of those at risk is a high priority, particularly given the expanding toolkit of biomedical HIV prevention options. An ongoing study in rural East African communities evaluated the uptake of choices in product, testing mode and location of care delivery through a structured patient-centred HIV prevention delivery model. In this qualitative study, we sought to understand clients' experiences of this "dynamic choice prevention model" (DCP) and highlight pathways of action to inform HIV prevention delivery models.MethodsIn-depth semi-structured interviews were conducted from November 2021 through March 2022 with a purposively selected sample of n = 56 participants in DCP trials (across outpatient departments, antenatal clinics and community settings), and n = 21 healthcare providers (total n = 77). A seven-person multi-regional team translated and inductively coded transcript data. We used a framework analysis approach to identify emergent themes.ResultsIndividuals taking up HIV pre-exposure prophylaxis (PrEP) reported feelings of relief, liberation from fears of acquiring HIV and satisfaction with being able to take action despite partners' behaviours. Couples used a range of approaches afforded by the study to persuade partners to get tested and opt for PrEP. Post-exposure prophylaxis (PEP) use was less common, although women welcomed it in the event of sexual coercion or assault. Participants discussed switching from PEP to PrEP after familiarizing themselves with usage and ascertaining ongoing risk. Participants felt respected by providers, trusted them and appreciated being able to contact them directly for telephone support. Prevention uptake was hindered by stigma, limited experience with and knowledge of prevention methods, gendered and generational power dynamics within intimate partnerships and families, and negative perceptions of methods due to the products themselves. Participants anticipated long-acting injectable PrEP could solve their challenges regarding pill size, daily pill burden and the likelihood of unwanted disclosure.ConclusionsDiverse preferences and barriers to uptake of prevention require a choice of HIV prevention options, locations and delivery modalities-but in addition, flexible, competent and friendly care provision is crucial to promote uptake. Helping clients feel valued, and addressing their unique needs and challenges, enables their agency to prioritize their health.

Antibiotics are vital in combating infectious diseases, yet their increasing use fosters resistance. Antimicrobial resistance (AMR) is rising in Nepal due to factors such as indiscriminate, inappropriate, and inadequate antibiotic usage. This study aims to explore the association between demographic factors and the prevalence of specific bacterial strains within the surveyed population. Additionally, it seeks to determine the antibiotic resistance patterns of these bacteria. Antibiotic susceptibility or resistance data were retrieved from the Medical Records Department (MRD) of the Manmohan Memorial Medical College and Teaching Hospital (MMMCTH) in Kathmandu. Samples from patients with certain types of bacterial infections were included, with 56 from sputum reports, 46 from urine, and 8 from blood samples out of 110 retrieved. Analysis revealed that sputum samples were mostly from older males, while urine samples were mostly from females. Yet, gender did not significantly influence bacterial presence across sample types. Overall, Escherichia coli was the most prevalent bacterium (74%), followed by Salmonella typhi (25%), Staphylococcus aureus (25%), and Klebsiella pneumoniae (23%) isolated from different type of clinical samples. Altogether, 6-15 antibiotics were assessed for sensitivity, with 2–6 antibiotics showing sensitivity to blood bacteria, 1-6 antibiotics demonstrating sensitivity to sputum bacteria, and 3–8 antibiotics exhibiting sensitivity to urine bacteria. Many investigated antibiotics were resistant, only gentamicin exhibited sensitivity for all types of bacteria found in blood, sputum and urine. These findings underscore the importance of discerning bacterial resistance patterns for effective antimicrobial treatment selection.

Antigenically sequence variable M proteins of the major bacterial pathogen Streptococcus pyogenes (Strep A) are responsible for recruiting human C4b-binding protein (C4BP) to the bacterial surface, which enables Strep A to evade destruction by the immune system. The most sequence divergent portion of M proteins, the hypervariable region (HVR), is responsible for binding C4BP. Structural evidence points to the conservation of two C4BP-binding sequence patterns (M2 and M22) in the HVR of numerous M proteins, with this conservation applicable to vaccine immunogen design. These two patterns, however, only partially explain C4BP binding by Strep A. Here, we identified several M proteins that lack these patterns but still bind C4BP and determined the structures of two, M68 and M87 HVRs, in complex with a C4BP fragment. Mutagenesis of these M proteins led to the identification of amino acids that are crucial for C4BP binding, enabling formulation of new C4BP-binding patterns. Mutagenesis was also carried out on M2 and M22 proteins to refine or generate experimentally grounded C4BP-binding patterns. The M22 pattern was the most prevalent among M proteins, followed by the M87 and M2 patterns, while the M68 pattern was rare. These patterns, except for M68, were also evident in numerous M-like Enn proteins. Binding of C4BP via these patterns to Enn proteins was verified. We conclude that C4BP-binding patterns occur frequently in Strep A strains of differing M types, being present in their M or Enn proteins, or frequently both, providing further impetus for their use as vaccine immunogens.

Historically neglected by microbial ecologists, soil viruses are now thought to be critical to global biogeochemical cycles. However, our understanding of their global distribution, activities and interactions with the soil microbiome remains limited. Here we present the Global Soil Virus Atlas, a comprehensive dataset compiled from 2,953 previously sequenced soil metagenomes and composed of 616,935 uncultivated viral genomes and 38,508 unique viral operational taxonomic units. Rarefaction curves from the Global Soil Virus Atlas indicate that most soil viral diversity remains unexplored, further underscored by high spatial turnover and low rates of shared viral operational taxonomic units across samples. By examining genes associated with biogeochemical functions, we also demonstrate the viral potential to impact soil carbon and nutrient cycling. This study represents an extensive characterization of soil viral diversity and provides a foundation for developing testable hypotheses regarding the role of the virosphere in the soil microbiome and global biogeochemistry.

IntroductionThere is little HIV counselling that directly meets the needs of men in Eastern and Southern Africa, limiting men's knowledge about the benefits of HIV treatment and how to overcome barriers to engagement, contributing to poorer HIV-related outcomes than women. Male-specific approaches are needed to improve men's outcomes but may be difficult for healthcare workers (HCWs) to implement with fidelity and quality in low-resource settings. We developed a male-specific counselling curriculum which was implemented by male HCWs and then conducted a mixed-methods quality assessment.MethodsWe audio-recorded counselling sessions to assess the quality of implementation (n = 50) by male HCWs from two cadres (nurse, n = 10 and lay cadre, n = 10) and conducted focus group discussions (FGDs) with HCWs at 6 and 9 months after rollout to understand barriers and facilitators to implementation. Counselling sessions and FGDs were translated, transcribed and analysed using thematic analysis adapted from WHO Quality Counselling Guidelines. We assessed if sessions were respectful, informative, interactive, motivating and included tailored action plans for overcoming barriers to care. All data were collected September 2021-June 2022.ResultsAll sessions used respectful, non-judgemental language. Sessions were highly interactive with most HCWs frequently asking open-ended questions (n = 46, 92%) and often incorporating motivational explanations of how antiretroviral therapy contributes to life goals (n = 42, 84%). Few sessions included individually tailored action plans for clients to overcome barriers to care (n = 9, 18%). New counselling themes were well covered; however, occasionally themes of self-compassion and safe sex were not covered during sessions (n = 16 and n = 11). HCWs believed that having male HCWs conduct counselling, ongoing professional development and keeping detailed counselling notes facilitated quality implementation. Perceived barriers included curriculum length and client hesitancy to participate in action plan development. Findings were similar across cadres.ConclusionsImplementing high-quality male-specific counselling using male nurses and/or lay cadre is feasible. Efforts to utilize lay cadres should be prioritized, particularly in low-resource settings. Programmes should provide comprehensive job aids to support HCWs. Ongoing training and professional development are needed to (1) improve HCWs' skills in tailored action plans, and (2) sensitize HCWs to the need for self-compassion within male clients to promote holistic sexual health.

Sunscreen has been used for thousands of years to protect skin from ultraviolet radiation. However, the use of modern commercial sunscreen containing oxybenzone, ZnO, and TiO2 has raised concerns due to their negative effects on human health and the environment. In this study, we aim to establish an efficient microbial platform for production of shinorine, a UV light absorbing compound with anti-aging properties. First, we methodically selected an appropriate host for shinorine production by analyzing central carbon flux distribution data from prior studies alongside predictions from genome-scale metabolic models (GEMs). We enhanced shinorine productivity through CRISPRi-mediated downregulation and utilized shotgun proteomics to pinpoint potential competing pathways. Simultaneously, we improved the shinorine biosynthetic pathway by refining its design, optimizing promoter usage, and altering the strength of ribosome binding sites. Finally, we conducted amino acid feeding experiments under various conditions to identify the key limiting factors in shinorine production. The study combines meta-analysis of 13C-metabolic flux analysis, GEMs, synthetic biology, CRISPRi-mediated gene downregulation, and omics analysis to improve shinorine production, demonstrating the potential of Pseudomonas putida KT2440 as platform for shinorine production.

ObjectiveHIV treatment regimen during pregnancy was associated with preterm delivery (PTD) in the PROMISE 1077 BF trial. Systemic inflammation among pregnant women with HIV could help explain differences in PTD by treatment regimen. We assessed associations between inflammation, treatment regimen, and PTD.Design/methodsA nested 1 : 1 case-control study ( N  = 362) was conducted within a multicountry randomized trial comparing three HIV regimens in pregnant women: zidovudine alone, or combination antiretroviral therapy (ART) with lopinavir/ritonavir and either zidovudine or tenofovir. Cases were women with PTD (

ImportanceUrinary tract infection (UTI) is the second most common infection leading to hospitalization and is often associated with gram-negative multidrug-resistant organisms (MDROs). Clinicians overuse extended-spectrum antibiotics although most patients are at low risk for MDRO infection. Safe strategies to limit overuse of empiric antibiotics are needed.ObjectiveTo evaluate whether computerized provider order entry (CPOE) prompts providing patient- and pathogen-specific MDRO risk estimates could reduce use of empiric extended-spectrum antibiotics for treatment of UTI.Design, setting, and participantsCluster-randomized trial in 59 US community hospitals comparing the effect of a CPOE stewardship bundle (education, feedback, and real-time and risk-based CPOE prompts; 29 hospitals) vs routine stewardship (n = 30 hospitals) on antibiotic selection during the first 3 hospital days (empiric period) in noncritically ill adults (≥18 years) hospitalized with UTI with an 18-month baseline (April 1, 2017-September 30, 2018) and 15-month intervention period (April 1, 2019-June 30, 2020).InterventionsCPOE prompts recommending empiric standard-spectrum antibiotics in patients ordered to receive extended-spectrum antibiotics who have low estimated absolute risk (

ImportancePneumonia is the most common infection requiring hospitalization and is a major reason for overuse of extended-spectrum antibiotics. Despite low risk of multidrug-resistant organism (MDRO) infection, clinical uncertainty often drives initial antibiotic selection. Strategies to limit empiric antibiotic overuse for patients with pneumonia are needed.ObjectiveTo evaluate whether computerized provider order entry (CPOE) prompts providing patient- and pathogen-specific MDRO infection risk estimates could reduce empiric extended-spectrum antibiotics for non-critically ill patients admitted with pneumonia.Design, setting, and participantsCluster-randomized trial in 59 US community hospitals comparing the effect of a CPOE stewardship bundle (education, feedback, and real-time MDRO risk-based CPOE prompts; n = 29 hospitals) vs routine stewardship (n = 30 hospitals) on antibiotic selection during the first 3 hospital days (empiric period) in non-critically ill adults (≥18 years) hospitalized with pneumonia. There was an 18-month baseline period from April 1, 2017, to September 30, 2018, and a 15-month intervention period from April 1, 2019, to June 30, 2020.InterventionCPOE prompts recommending standard-spectrum antibiotics in patients ordered to receive extended-spectrum antibiotics during the empiric period who have low estimated absolute risk (

Enteric pathogens such as Salmonella enterica serovar Typhimurium experience spatial and temporal changes to the metabolic landscape throughout infection. Host reactive oxygen and nitrogen species non-enzymatically convert monosaccharides to alpha hydroxy acids, including L-tartrate. Salmonella utilizes L-tartrate early during infection to support fumarate respiration, while L-tartrate utilization ceases at later time points due to the increased availability of exogenous electron acceptors such as tetrathionate, nitrate, and oxygen. It remains unknown how Salmonella regulates its gene expression to metabolically adapt to changing nutritional environments. Here, we investigated how the transcriptional regulation for L-tartrate metabolism in Salmonella is influenced by infection-relevant cues. L-tartrate induces the transcription of ttdBAU, genes involved in L-tartrate utilization. L-tartrate metabolism is negatively regulated by two previously uncharacterized transcriptional regulators TtdV (STM3357) and TtdW (STM3358), and both TtdV and TtdW are required for the sensing of L-tartrate. The electron acceptors nitrate, tetrathionate, and oxygen repress ttdBAU transcription via the two-component system ArcAB. Furthermore, the regulation of L-tartrate metabolism is required for optimal fitness in a mouse model of Salmonella-induced colitis. TtdV, TtdW, and ArcAB allow for the integration of two cues, i.e., substrate availability and availability of exogenous electron acceptors, to control L-tartrate metabolism. Our findings provide novel insights into how Salmonella prioritizes the utilization of different electron acceptors for respiration as it experiences transitional nutrient availability throughout infection.ImportanceBacterial pathogens must adapt their gene expression profiles to cope with diverse environments encountered during infection. This coordinated process is carried out by the integration of cues that the pathogen senses to fine-tune gene expression in a spatiotemporal manner. Many studies have elucidated the regulatory mechanisms of how Salmonella sense metabolites in the gut to activate or repress its virulence program; however, our understanding of how Salmonella coordinates its gene expression to maximize the utilization of carbon and energy sources found in transitional nutrient niches is not well understood. In this study, we discovered how Salmonella integrates two infection-relevant cues, substrate availability and exogenous electron acceptors, to control L-tartrate metabolism. From our experiments, we propose a model for how L-tartrate metabolism is regulated in response to different metabolic cues in addition to characterizing two previously unknown transcriptional regulators. This study expands our understanding of how microbes combine metabolic cues to enhance fitness during infection.