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Certain human leukocyte antigens, by presenting conserved immunogenic epitopes for T cell recognition, may, in part, account for the observed differences in human immunodeficiency virus type 1 (HIV-1) susceptibility. To determine whether HLA polymorphism influences HIV-1 susceptibility, a longitudinal cohort of highly HIV-1-exposed female sex workers based in Nairobi, Kenya, was prospectively analyzed. Decreased HIV-1 infection risk was strongly associated with possession of a cluster of closely related HLA alleles (A2/6802 supertype; incidence rate ratio [IRR], 0.45; 95% confidence interval [CI], 0.27-0.72; P=.0003). The alleles in this supertype are known in some cases to present the same peptide epitopes for T cell recognition. In addition, resistance to HIV-1 infection was independently associated with HLA DRB1*01 (IRR, 0.22; 95% CI, 0.06-0.60; P=.0003), which suggests that anti-HIV-1 class II restricted CD4 effector mechanisms may play an important role in protecting against viral challenge. These data provide further evidence that resistance to HIV-1 infection in this cohort of sex workers is immunologically mediated. [ABSTRACT FROM AUTHOR]

Although nosocomial transmission of human respiratory syncytial virus (HRSV) and its effect on morbidity and mortality among immunocompromised adults are well recognized, few studies have applied molecular techniques to differentiate nosocomial from community-acquired infections. Between January and April 1997, an outbreak of HRSV occurred among adult patients in a leukemia/lymphoma ward. Among 45 hospitalized patients undergoing bronchoscopy for investigation of acute respiratory illness, 8 were identified with HRSV infection. One infected patient developed symptoms before admission and was thought to be the index case. However, subsequent sequencing of 7 HRSV isolates identified 2 distinct genotypes, GA5 (1 case) and GB3 (6 cases). The 6 GB3 isolates could be further differentiated into 2 strains with identical nucleotide sequences that differed from each other and from 14 community HRSV isolates. Instead of a single nosocomial outbreak of HRSV, multiple introductions of HRSV likely occurred with distinct lines of nosocomial transmission. Copyright © 1999 The University of Chicago [ABSTRACT FROM AUTHOR]

Accurate identification of fishes is essential for understanding their biology and to ensure food safety for consumers. DNA barcoding is an important tool because it can verify identifications of both whole and processed fishes that have had key morphological characters removed (e.g., filets, fish meal); however, DNA reference libraries are incomplete, and public repositories for sequence data contain incorrectly identified sequences. During a nine-year sampling program in the Philippines, a global biodiversity hotspot for marine fishes, we developed a verified reference library of cytochrome c oxidase subunit I (COI) sequences for 2,525 specimens representing 984 species. Specimens were primarily purchased from markets, with additional diversity collected using rotenone or fishing gear. Species identifications were verified based on taxonomic, phenotypic, and genotypic data, and sequences are associated with voucher specimens, live-color photographs, and genetic samples catalogued at Smithsonian Institution, National Museum of Natural History. The Biodiversity of Philippine Marine Fishes dataset is released herein to increase knowledge of species diversity and distributions and to facilitate accurate identification of market fishes., (© 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Gardnerella species are associated with bacterial vaginosis (BV) and have been investigated as etiological agents of the condition. Nonetheless, the isolation of this taxon from healthy individuals has raised important questions regarding its etiological role. Recently, using advanced molecular approaches, the Gardnerella genus was expanded to include several different species that exhibit differences in virulence potential. Understanding the significance of these different species with respect to mucosal immunity and the pathogenesis and complications of BV could be crucial to solving the BV enigma. Here, we review key findings regarding the unique genetic and phenotypic diversity within this genus, virulence factors, and effects on mucosal immunity as they stand. We also comment on the relevance of these findings to the proposed role of Gardnerella in BV pathogenesis and in reproductive health and identify key gaps in knowledge that should be explored in the future., Competing Interests: The authors declare no conflict of interest.

Most cervicovaginal microbiome-immunology studies to date have relied on 16S rDNA microbial profiling which does not resolve the molecular subgroups of Gardnerella , believed to be central to the pathogenesis of bacterial vaginosis (BV) and subsequent risk of HIV acquisition. Here we used the cpn 60 universal target which in addition to other microbial taxa, resolves four Gardnerella subgroups, for cervicovaginal microbial profiling in a longitudinal cohort of Kenyan women to examine associations with cellular and soluble markers of inflammation and HIV susceptibility. Participants (N = 41) were sampled, contributing 362 samples for microbiome analysis. All non- Lactobacillus dominant microbial communities were associated with high pro-inflammatory cytokine levels. Divergent associations were observed among different Gardnerella subgroup dominated communities with respect to the chemokine IP-10. Specifically, Gardnerella subgroup A dominant and polymicrobial communities were associated with reduced concentrations of IP-10 in adjusted linear mixed models (p<0.0001), compared to microbial communities dominated by Lactobacillus (non-iners) species. However, these associations did not translate to significant differences in the proportion or absolute number of CCR5, HLA-DR and CD38 expressed on cervical CD4 + T- cells. These findings suggest that some associations between Gardnerella subgroup dominant microbiomes and mucosal immunity differ and are relevant for the study of BV-pathogenesis and understanding the mechanisms of BV-associated HIV risk., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Shvartsman, Perciani, Richmond, Russell, Tough, Vancuren, Hill, KAVI-ICR, Jaoko, McKinnon, Sandstrom and MacDonald.)

Biodiversity assessments are critical for setting conservation priorities, understanding ecosystem function and establishing a baseline to monitor change. Surveys of marine biodiversity that rely almost entirely on sampling adult organisms underestimate diversity because they tend to be limited to habitat types and individuals that can be easily surveyed. Many marine animals have planktonic larvae that can be sampled from the water column at shallow depths. This life stage often is overlooked in surveys but can be used to relatively rapidly document diversity, especially for the many species that are rare or live cryptically as adults. Using DNA barcode data from samples of nemertean worms collected in three biogeographical regions-Northeastern Pacific, the Caribbean Sea and Eastern Tropical Pacific-we found that most species were collected as either benthic adults or planktonic larvae but seldom in both stages. Randomization tests show that this deficit of operational taxonomic units collected as both adults and larvae is extremely unlikely if larvae and adults were drawn from the same pool of species. This effect persists even in well-studied faunas. These results suggest that sampling planktonic larvae offers access to a different subset of species and thus significantly increases estimates of biodiversity compared to sampling adults alone. Spanish abstract is available in the electronic supplementary material.

Background: The microbiota of the lower female genital tract plays an important role in women's health. Microbial profiling using the chaperonin60 (cpn60) universal target (UT) improves resolution of vaginal species associated with negative health outcomes compared to the more commonly used 16S ribosomal DNA target. However, the choice of DNA extraction and PCR product purification methods may bias sequencing-based microbial studies and should be optimized for the sample type and molecular target used. In this study, we compared two commercial DNA extraction kits and two commercial PCR product purification kits for the microbial profiling of cervicovaginal samples using the cpn60 UT., Methods: DNA from cervicovaginal secretions and vaginal lavage samples as well as mock community standards were extracted using either the specialized QIAamp DNA Microbiome Kit, or the standard DNeasy Blood & Tissue kit with enzymatic pre-treatment for enhanced lysis of gram-positive bacteria. Extracts were PCR amplified using well-established cpn60 primer sets and conditions. Products were then purified using a column-based method (QIAquick PCR Purification Kit) or a gel-based PCR clean-up method using the QIAEX II Gel Extraction Kit. Purified amplicons were sequenced with the MiSeq platform using standard procedures. The overall quality of each method was evaluated by measuring DNA yield, alpha diversity, and microbial composition., Results: DNA extracted from cervicovaginal samples using the DNeasy Blood and Tissue kit, pre-treated with lysozyme and mutanolysin, resulted in increased DNA yield, bacterial diversity, and species representation compared to the QIAamp DNA Microbiome kit. The column-based PCR product purification approach also resulted in greater average DNA yield and wider species representation compared to a gel-based clean-up method. In conclusion, this study presents a fast, effective sample preparation method for high resolution cpn60 based microbial profiling of cervicovaginal samples., Competing Interests: The authors have declared that no competing interests exist.

Across plants and animals, genome size is often correlated with life-history traits: large genomes are correlated with larger seeds, slower development, larger body size and slower cell division. Among decapod crustaceans, caridean shrimps are among the most variable both in terms of genome size variation and life-history characteristics such as larval development mode and egg size, but the extent to which these traits are associated in a phylogenetic context is largely unknown. In this study, we examine correlations among egg size, larval development and genome size in two different genera of snapping shrimp, Alpheus and Synalpheus, using phylogenetically informed analyses. In both Alpheus and Synalpheus, egg size is strongly linked to larval development mode: species with abbreviated development had significantly larger eggs than species with extended larval development. We produced the first comprehensive dataset of genome size in Alpheus (n = 37 species) and demonstrated that genome size was strongly and positively correlated with egg size in both Alpheus and Synalpheus. Correlated trait evolution analyses showed that in Alpheus, changes in genome size were clearly dependent on egg size. In Synalpheus, evolutionary path analyses suggest that changes in development mode (from extended to abbreviated) drove increases in egg volume; larger eggs, in turn, resulted in larger genomes. These data suggest that variation in reproductive traits may underpin the high degree of variation in genome size seen in a wide variety of caridean shrimp groups more generally., (© 2021 European Society for Evolutionary Biology.)

Background: The earliest coronavirus disease-2019 (COVID-19) cases in Central Asia were announced in March 2020 by Kazakhstan. Despite the implementation of aggressive measures to curb infection spread, gaps remain in the understanding of the clinical and epidemiologic features of the regional pandemic., Methods: We did a retrospective, observational cohort study of patients with laboratory-confirmed COVID-19 hospitalized in Kazakhstan between February and April 2020. We compared demographic, clinical, laboratory and radiological data of patients with different COVID-19 severities on admission. Logistic regression was used to assess factors associated with disease severity and in-hospital death. Whole-genome SARS-CoV-2 analysis was performed in 53 patients., Findings: Of the 1072 patients with laboratory-confirmed COVID-19 in March-April 2020, the median age was 36 years (IQR 24-50) and 484 (45%) were male. On admission, 683 (64%) participants had asymptomatic/mild, 341 (32%) moderate, and 47 (4%) severe-to-critical COVID-19 manifestation; 20 in-hospital deaths (1•87%) were reported by 5 May 2020. Multivariable regression indicated increasing odds of severe disease associated with older age (odds ratio 1•05, 95% CI 1•03-1•07, per year increase; p<0•001), the presence of comorbidities (2•34, 95% CI 1•18-4•85; p=0•017) and elevated white blood cell count (WBC, 1•13, 95% CI 1•00-1•27; p=0•044) on admission, while older age (1•09, 95% CI 1•06-1•13, per year increase; p<0•001) and male sex (5•63, 95% CI 2•06-17•57; p=0•001) were associated with increased odds of in-hospital death. The SARS-CoV-2 isolates grouped into seven phylogenetic lineages, O/B.4.1, S/A.2, S/B.1.1, G/B.1, GH/B.1.255, GH/B.1.3 and GR/B.1.1.10; 87% of the isolates were O and S sub-types descending from early Asian lineages, while the G, GH and GR isolates were related to lineages from Europe and the Americas., Interpretation: Older age, comorbidities, increased WBC count, and male sex were risk factors for COVID-19 disease severity and mortality in Kazakhstan. The broad SARS-CoV-2 diversity suggests multiple importations and community-level amplification predating travel restriction., Funding: Ministry of Education and Science of the Republic of Kazakhstan., Competing Interests: The authors declare that they have no competing interests., (© 2021 The Author(s).)

Objectives: Ceftobiprole is an advanced-generation cephalosporin with a favourable safety profile. Published data on the clinical use of ceftobiprole are limited. We report use of ceftobiprole in Canadian patients using data captured by the CLEAR registry., Methods: The CLEAR registry uses the web-based research data management program REDCap™ (online survey) to facilitate clinicians entering details associated with their clinical experiences using ceftobiprole., Results: Data were available for 38 patients treated with ceftobiprole. The most common infections treated were endocarditis (42.1% of patients), bone and joint infection (23.7%) and hospital-associated bacterial pneumonia (15.8%). 92.1% of patients had bacteraemia and 21.1% were in intensive care. Ceftobiprole was used because of failure of (71.1%), resistance to (18.4%) or adverse effects from (10.5%) previously prescribed antimicrobial agents. Ceftobiprole was primarily used as directed therapy for methicillin-resistant Staphylococcus aureus (MRSA) infections (94.7% of patients). Ceftobiprole susceptibility testing was performed on isolates from 47.4% of patients. It was used concomitantly with daptomycin in 55.3% of patients and with vancomycin in 18.4% of patients. Treatment duration was primarily >10 days (65.8% of patients) with microbiological success in 97.0% and clinical success in 84.8% of patients. 2.6% of patients had gastrointestinal adverse effects., Conclusion: In Canada to date, ceftobiprole is used as directed therapy to treat a variety of severe infections caused by MRSA. It is primarily used in patients failing previous antimicrobials, is frequently added to, and thus used in combination with daptomycin or vancomycin with high microbiological and clinical cure rates and an excellent safety profile., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Noise-induced hearing loss (NIHL) is known to have significant consequences for temporal, spectral, and spatial resolution. However, much remains to be discovered about their underlying pathophysiology. This report extends the recent development of a nonhuman primate model of NIHL to explore its consequences for hearing in noisy environments, and its correlations with the underlying cochlear pathology. Ten macaques (seven with normal-hearing, three with NIHL) were used in studies of masked tone detection in which the temporal or spatial properties of the masker were varied to assess metrics of temporal and spatial processing. Normal-hearing (NH) macaques showed lower tone detection thresholds for sinusoidally amplitude modulated (SAM) broadband noise maskers relative to unmodulated maskers (modulation masking release, MMR). Tone detection thresholds were lowest at low noise modulation frequencies, and increased as modulation frequency increased, until they matched threshold in unmodulated noise. NH macaques also showed lower tone detection thresholds for spatially separated tone and noise relative to co-localized tone and noise (spatial release from masking, SRM). Noise exposure caused permanent threshold shifts that were verified behaviorally and audiologically. In hearing-impaired (HI) macaques, MMR was reduced at tone frequencies above that of the noise exposure. HI macaques also showed degraded SRM, with no SRM observed across all tested tone frequencies. Deficits in MMR correlated with audiometric threshold changes, outer hair cell loss, and synapse loss, while the differences in SRM did not correlate with audiometric changes, or any measure of cochlear pathophysiology. This difference in anatomical-behavioral correlations suggests that while many behavioral deficits may arise from cochlear pathology, only some are predictable from the frequency place of damage in the cochlea., Competing Interests: Declaration of Competing Interest The authors hereby declare that they have no conflicts of interest., (Copyright © 2020. Published by Elsevier B.V.)

Exposure to loud noise causes damage to the inner ear, including but not limited to outer and inner hair cells (OHCs and IHCs) and IHC ribbon synapses. This cochlear damage impairs auditory processing and increases audiometric thresholds (noise-induced hearing loss, NIHL). However, the exact relationship between the perceptual consequences of NIHL and its underlying cochlear pathology are poorly understood. This study used a nonhuman primate model of NIHL to relate changes in frequency selectivity and audiometric thresholds to indices of cochlear histopathology. Three macaques (one Macaca mulatta and two Macaca radiata) were trained to detect tones in quiet and in noises that were spectrally notched around the tone frequency. Audiograms were derived from tone thresholds in quiet; perceptual auditory filters were derived from tone thresholds in notched-noise maskers using the rounded-exponential fit. Data were obtained before and after a four-hour exposure to a 50-Hz noise centered at 2 kHz at 141 or 146 dB SPL. Noise exposure caused permanent audiometric threshold shifts and broadening of auditory filters at and above 2 kHz, with greater changes observed for the 146-dB-exposed monkeys. The normalized bandwidth of the perceptual auditory filters was strongly correlated with audiometric threshold at each tone frequency. While changes in audiometric threshold and perceptual auditory filter widths were primarily determined by the extent of OHC survival, additional variability was explained by including interactions among OHC, IHC, and ribbon synapse survival. This is the first study to provide within-subject comparisons of auditory filter bandwidths in an animal model of NIHL and correlate these NIHL-related perceptual changes with cochlear histopathology. These results expand the foundations for ongoing investigations of the neural correlates of NIHL-related perceptual changes., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflicts of interest., (Copyright © 2020. Published by Elsevier B.V.)

Background: Varicella-zoster virus (VZV) is under consideration as a promising recombinant viral vector to deliver foreign antigens including HIV. However, new vectors have come under increased scrutiny, since trials with adenovirus serotype 5-vectored (Ad5-vectored) HIV vaccine demonstrated increased HIV risk in individuals with pre-immunity to the vector that was thought to be associated with mucosal immune activation (IA). Therefore, given the prospect of developing an HIV/VZV chimeric vaccine, it is particularly important to define the impact of VZV vaccination on IA., Methods: Healthy VZV-seropositive Kenyan women (n = 44) were immunized with high-dose live attenuated VZV vaccine, and we assessed the expression on CD4+ T cells isolated from blood, cervix, and rectum of IA markers including CD38 and HLA-DR and of markers of cell migration and tissue retention, as well as the concentration of genital and intestinal cytokines. A delayed-start group (n = 22) was used to control for natural variations in these parameters., Results: Although immunogenic, VZV vaccination did not result in significant difference in the frequency of cervical activated (HLA-DR+CD38+) CD4+ T cells (median 1.61%, IQR 0.93%-2.76%) at 12 weeks after vaccination when compared with baseline (median 1.58%, IQR 0.75%-3.04%), the primary outcome for this study. VZV vaccination also had no measurable effect on any of the IA parameters at 4, 8, and 12 weeks after vaccination., Conclusion: This study provides the first evidence to our knowledge about the effects of VZV vaccination on human mucosal IA status and supports further evaluation of VZV as a potential vector for an HIV vaccine., Trial Registration: ClinicalTrials.gov NCT02514018., Funding: Primary support from the Canadian Institutes for Health Research (CIHR). For other sources, see Acknowledgments.

Background: Attenuated varicella zoster virus (VZV) is a promising vector for recombinant vaccines. Because human immunodeficiencyvirus (HIV) vaccines are believed to require mucosal immunogenicity, we characterized mucosal VZV-specific humoral immunity following VZVOka vaccination., Methods: Adult Kenyan VZV-seropositive women (n = 44) received a single dose of the live zoster VZVOka vaccine. The anamnestic responses to the virus were followed longitudinally in both plasma and mucosal secretions using an in-house glycoprotein enzyme-linked immunosorbent assay and safety and reactogenicity monitored. VZV seroprevalence and baseline responses to the virus were also characterized in our cohorts (n = 288)., Results: Besides boosting anti-VZV antibody responses systemically, vaccination also boosted anti-VZV immunity in the cervicovaginal mucosa with a 2.9-fold rise in immunoglobulin G (P < .0001) and 1.6-fold rise in immunoglobulin A (IgA) (P = .004) from the time before immunization and 4 weeks postvaccination. Baseline analysis demonstrated high avidity antibodies at the gastrointestinal and genital mucosa of VZV-seropositive women. Measurement of VZV-specific IgA in saliva is a sensitive tool for detecting prior VZV infection., Conclusions: VZVOka vaccine was safe and immunogenic in VZV-seropositive adult Kenyan women. We provided compelling evidence of VZV ability to induce genital mucosa immunity., Clinical Trials Registration: NCT02514018.

Cell surface expression of α4β7, α4β1 and αEβ7 integrins play a key role in T cell distribution. Understanding the contribution of integrins to the density and ratios of CD4+: CD4negT cell at the portals of entry for HIV is of fundamental importance for the advance of more effective HIV prevention strategies. We therefore set out to characterize and compare the expression of α4β7, α4β1 and αEβ7 integrins on systemic, cervical and rectal CD4+ and CD4negT cells isolated from a cohort of healthy Kenyan women at low risk for sexually transmitted infections (STI) (n = 45). Here we show that blood and cervix were enriched in α4+β1+CD4+T cells and α4+β7hiCD4+T cells, whereas the rectum had an equal frequency of α4+β7hiCD4+T cells and αE+β7hiCD4+T cells. Most cervical and rectal αE+β7hiCD4+T cells expressed CCR5 as well as CD69. Interestingly, αEβ7 was the predominant integrin expressed by CD4negT cells in both mucosal sites, outnumbering αE+β7hiCD4+T cells approximately 2-fold in the cervix and 7-fold in the rectum. The majority of αE+β7hiCD4negT cells expressed CD69 at the mucosa. Taken together, our results show unique tissue-specific patterns of integrin expression. These results can help in guiding vaccine design and also the use of therapeutically targeting integrin adhesion as a means to preventing HIV.

Introduction: A protective HIV vaccine would be expected to induce durable effector immune responses at the mucosa, restricting HIV infection at its portal of entry. We hypothesise that use of varicella-zoster virus (VZV) as an HIV delivery vector could generate sustained and robust tissue-based immunity against HIV antigens to provide long-term protection against HIV. Given that HIV uniquely targets immune-activated T cells, the development of human vaccines against HIV must also involve a specific examination of the safety of the vector. Thus, we aim to evaluate the effects of VZV vaccination on the recipients' immune activation state, and on VZV-specific circulating humoral and cellular responses in addition to those at the cervical and rectal mucosa., Methods and Analysis: This open-label, randomised, longitudinal crossover study includes healthy Kenyan VZV-seropositive women at low risk for HIV infection. Participants receive a single dose of a commercial live-attenuated VZV Oka vaccine at either week 0 (n=22) or at week 12 (n=22) of the study and are followed for 48 and 36 weeks postvaccination, respectively. The primary outcome is the change on cervical CD4 + T-cell immune activation measured by the coexpression of CD38 and HLA-DR 12 weeks postvaccination compared with the baseline (prevaccination). Secondary analyses include postvaccination changes in VZV-specific mucosal and systemic humoral and cellular immune responses, changes in cytokine and chemokine measures, study acceptability and feasibility of mucosal sampling and a longitudinal assessment of the bacterial community composition of the mucosa., Ethics and Dissemination: The study has ethical approval from Kenyatta National Hospital/University of Nairobi Ethics and Research Committee, the University of Toronto Research Ethics Board and by Kenyan Pharmacy and Poisons Board. Results will be presented at conferences, disseminated to participants and stakeholders as well as published in peer-reviewed journals., Trial Registration Number: NCT02514018. Pre-results., Competing Interests: Competing interests: SMM has received grants for unrelated studies from GSK, Merck, Pfizer and Sanofi, and served on advisory boards for GSK and Pfizer., (© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)

Background: This study was done to characterize parameters associated with semen human immunodeficiency virus (HIV)-1 ribonucleic acid (RNA) viral load (VL) variability in HIV-infected, therapy-naive men., Methods: Paired blood and semen samples were collected from 30 HIV-infected, therapy-naive men who have sex with men, and 13 participants were observed longitudinally for up to 1 year. Human immunodeficiency virus RNA, bacterial load by 16S RNA, herpesvirus (Epstein-Barr virus and cytomegalovirus [CMV]) shedding, and semen cytokines/chemokines were quantified, and semen T-cell subsets were assessed by multiparameter flow cytometry., Results: Semen HIV RNA was detected at 93% of visits, with >50% of men shedding high levels of virus (defined as >5000 copies/mL). In the baseline cross-sectional analysis, an increased semen HIV VL correlated with local CMV reactivation, the semen bacterial load, and semen inflammatory cytokines, particularly interleukin (IL)-8. T cells in semen were more activated than blood, and there was an increased frequency of Th17 cells and γδ-T-cells. Subsequent prospective analysis demonstrated striking interindividual variability in HIV and CMV shedding patterns, and only semen IL-8 levels and the blood VL were independently associated with semen HIV levels., Conclusions: Several clinical and immune parameters were associated with increased HIV semen levels in antiretroviral therapy-naive men, with induction of local proinflammatory cytokines potentially acting as a common pathway., (© The Author 2017. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Advances in biodiversity genomic sequencing will increasingly depend on the availability of DNA samples-and their quantifiable metadata-preserved in large institutional biorepositories that are discoverable to the scientific community. Improvements in sequencing technology constantly provide longer reads, such that longer fragment length, higher molecular weight, and overall "genome-quality" DNA (gDNA) will be desirable. Ideally, biorepositories should publish numerical scale measurements of DNA quality useful to the user community. However, the most widely used technique to evaluate DNA quality, the classic agarose gel, has yet to be quantified. Here we propose a simple and economical method using open source image analysis software to make gDNA gel images quantifiable, and propose percentage of gDNA "greater than X kb" as a standard of comparison, where X is a band from any widely used DNA ladder with desirably large band sizes. We employ two metadata standards ("DNA Threshold" and "Percent above Threshold") introduced as part of the Global Genome Biodiversity Network (GGBN) Darwin Core extension. We illustrate the method using the traditionally used Hin dIII ladder and the 9,416 base-pair (bp) band as a standard. We also present data, for two taxa, a vertebrate (fish) and an invertebrate (crab), on how gDNA quality varies with seven tissue preservation methods, time since death, preservation method (i.e. buffers vs. cold temperatures), and storage temperature of various buffers over time. Our results suggest that putting tissue into a buffer prior to freezing may be better than directly into ultra-cold conditions., Competing Interests: Sean G. Brady is an Academic Editor for PeerJ.

Background: Cytomegaloviruses belong to a large, ancient, genus of DNA viruses comprised of a wide array of species-specific strains that occur in diverse array of hosts., Methods: In this study we sequenced the ~217 Kb genome of a cytomegalovirus isolated from a Mauritius cynomolgus macaque, CyCMV Mauritius, and compared it to previously sequenced cytomegaloviruses from a cynomolgus macaque of Filipino origin (CyCMV Ottawa) and two from Indian rhesus macaques (RhCMV 180.92 and RhCMV 68-1)., Results: Though more closely related to CyCMV Ottawa, CyCMV Mauritius is less genetically distant from both RhCMV strains than is CyCMV Ottawa. Several individual genes, including homologues of CMV genes RL11B, UL123, UL83b, UL84 and a homologue of mammalian COX-2, show a closer relationship between homologues of CyCMV Mauritius and the RhCMVs than between homologues of CyCMV Mauritius and CyCMV Ottawa. A broader phylogenetic analysis of 12 CMV strains from eight species recovers evolutionary relationships among viral strains that mirror those amongst the host species, further demonstrating co-evolution of host and virus., Conclusions: Phylogenetic analyses of rhesus and cynomolgus macaque CMV genome sequences demonstrate co-speciation of the virus and host.

The use of a number of non-rhesus macaque species, but especially cynomolgus macaques as a model for HIV-1 vaccine development has increased in recent years. Cynomolgus macaques have been used in the United Kingdom, Europe, Canada and Australia as a model for HIV vaccine development for many years. Unlike rhesus macaques, cynomolgus macaques infected with SIV show a pattern of disease pathogenesis that more closely resembles that of human HIV-1 infection, exhibiting lower peak and set-point viral loads and slower progression to disease with more typical AIDS defining illnesses. Several advances have been made recently in the use of the cynomolgus macaque SIV challenge model that allow the demonstration of vaccine efficacy using attenuated viruses and vectors that are both viral and non-viral in origin. This review aims to probe the details of various vaccination trials carried out in cynomolgus macaques in the context of our modern understanding of the highly diverse immunogenetics of this species with a view to understanding the species-specific immune correlates of protection and the efficacy of vectors that have been used to design vaccines., (Copyright © 2015. Published by Elsevier Ltd.)

Infections by Neisseria gonorrhoeae are increasingly common, are often caused by antibiotic-resistant strains, and can result in serious and lasting sequelae, prompting the reemergence of gonococcal disease as a leading global health concern. N. gonorrhoeae is a human-restricted pathogen that primarily colonizes urogenital mucosal surfaces. Disease progression varies greatly between the sexes: men usually present with symptomatic infection characterized by a painful purulent urethral discharge, while in women, the infection is often asymptomatic, with the most severe pathology occurring when the bacteria ascend from the lower genital tract into the uterus and fallopian tubes. Classical clinical studies demonstrated that clinically infectious strains uniformly express Opa adhesins; however, their specificities were unknown at the time. While in vitro studies have since identified CEACAM proteins as the primary target of Opa proteins, the gonococcal specificity for this human family of receptors has not been addressed in the context of natural infection. In this study, we characterize a collection of low-passage-number clinical-specimen-derived N. gonorrhoeae isolates for Opa expression and assess their CEACAM-binding profiles. We report marked in vivo selection for expression of phase-variable Opa proteins that bind CEACAM1 and CEACAM5 but selection against expression of Opa variants that bind to the neutrophil-restricted decoy receptor CEACAM3. This is the first study showing phenotypic selection for distinct CEACAM-binding phenotypes in vivo, and it supports the opposing functions of CEACAMs that facilitate infection versus driving inflammation within the genital tract., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Cytomegalovirus (CMV) is a highly species-specific virus that has co-evolved with its host over millions of years and thus restricting cross-species infection. To examine the extent to which host restriction may prevent cross-species research between closely related non-human primates, we evaluated experimental infection of cynomolgus macaques with a recombinant rhesus macaque-derived CMV (RhCMV-eGFP). Twelve cynomolgus macaques were randomly allocated to three groups: one experimental group (RhCMV-eGFP) and two control groups (UV-inactivated RhCMV-eGFP or media alone). The animals were given two subcutaneous inoculations at week 0 and week 8, and a subset of animals received an intravenous inoculation at week 23. No overt clinical or haematological changes were observed and PBMCs isolated from RhCMV-eGFP inoculated animals had comparable eGFP- and IE-1-specific cellular responses to the control animals. Following inoculation with RhCMV-eGFP, we were unable to detect evidence of infection in any blood or tissue samples up to 4 years post-inoculation, using sensitive viral co-culture, qPCR, and Western blot assays. Co-culture of urine and saliva samples demonstrated the presence of endogenous cynomolgus CMV (CyCMV) cytopathic effect, however no concomitant eGFP expression was observed. The absence of detectable RhCMV-eGFP suggests that the CyCMV-seropositive cynomolgus macaques were not productively infected with RhCMV-eGFP under these inoculation conditions. In a continued effort to develop CMV as a viral vector for an HIV/SIV vaccine, these studies demonstrate that CMV is highly restricted to its host species and can be highly affected by laboratory cell culture. Consideration of the differences between lab-adapted and primary viruses with respect to species range and cell tropism should be a priority in evaluating CMV as vaccine vector for HIV or other pathogens at the preclinical development stage.

Cynomolgus macaques are widely used as an animal model in biomedical research. We have established an immortalized cynomolgus macaque fibroblast cell line (MSF-T) by transducing primary dermal fibroblasts isolated from a 13-year-old male cynomolgus macaque with a retrovirus vector expressing human telomerase reverse transcriptase (hTERT). The MSF-T cells showed increased telomerase enzyme activity and reached over 200 in vitro passages compared to the non-transduced dermal fibroblasts, which reached senescence after 43 passages. The MSF-T cell line is free of mycoplasma contamination and is permissive to the newly identified cynomolgus macaque cytomegalovirus (CyCMV). CyCMV productively infects MSF-T cells and induces down-regulation of MHC class I expression. The MSF-T cell line will be extremely useful for the propagation of CyCMV and other cynomolgus herspesviruses in host-derived fibroblast cells, allowing for the retention of host-specific viral genes. Moreover, this cell line will be beneficial for many in vitro experiments related to this animal model.

As discussed in the larger review in this special issue (MacDonald and Feifel), intranasal oxytocin (OT) is demonstrating a growing potential as a therapeutic agent in psychiatry. Importantly, research suggests that a variety of individual factors may influence a person's response to OT. In this mini-review, I provide a review of three: (1) sex and hormonal status; (2) genetic variation in aspects of the OT system (i.e., OT receptors); and (3) attachment history. Each of these factors will be important to monitor as we strive to develop a richer understanding of OT's role in human development, brain-based disease, and the potential for individualized, OT-targeted treatments.

The tropical shrimp genus Synalpheus includes the only eusocial marine animals. In much of the Caribbean, eusocial species have dominated the diverse fauna of sponge-dwelling shrimp in coral rubble for at least the past two decades. Here we document a recent, dramatic decline and apparent local extinction of eusocial shrimp species on the Belize Barrier Reef. Our collections from shallow reefs in central Belize in 2012 failed to locate three of the four eusocial species formerly abundant in the area, and showed steep declines in colony size and increases in frequency of queenless colonies prior to their disappearance. Concordant with these declines, several nonsocial, pair-forming Synalpheus species increased in frequency. The decline in eusocial shrimp is explained in part by disappearance of two sponge species on which they specialize. Eusocial shrimp collections from Jamaica in 2012 showed similar patterns of decline in colony size and increased queenlessness compared with prior Jamaican collections. The decline and local extinction of eusocial shrimp happened against a backdrop of changes in coral assemblages during recent decades, and may reflect changes in abundance and quality of dead coral substratum and succession of the diverse cryptic organisms living within it. These changes document potentially worrisome declines in a unique taxon of eusocial marine animals.

Introduction: Agitation has significant consequences for patients and staff. When verbal techniques fail, expert guidelines recommend the use of second-generation antipsychotics (SGAs). Perhaps out of familiarity with haloperidol and benzodiazepines, emergency department (ED) clinicians often pair SGAs with benzodiazepines as well. Use of SGAs such as olanzapine in alcohol-intoxicated (ETOH+) patients or with benzodiazepines is not well studied and may be associated with vital sign abnormalities., Methods: This is a structured chart review of all patient visits who received either oral or intramuscular (i.m.) olanzapine in an academic ED from 2004 to 2010 and who had systolic blood pressure, heart rate, and oxygen saturation documented before medication administration and within 4 hours afterwards., Results: Four hundred eighty-two patient visits received olanzapine; 275 patient visits (225 oral, 50 i.m.) had vital signs documented. Neither route of administration, concurrent benzodiazepines, nor ingestion of ETOH were associated with significant decreases in systolic BP or heart rate (P = ns for all comparisons). Decreases in oxygen saturations, however, were significantly larger in ETOH+ patients who received i.m. olanzapine or i.m. olanzapine + benzodiazepines. Route of administration, concurrent benzodiazepines, nor ingestion of ETOH was associated with significant decreases in systolic blood pressure or heart rate (p = ns for all comparisons). Decreases in oxygen saturations, however, were significantly larger in ETOH+ patients who received i.m. olanzapine or i.m. olanzapine + benzodiazepines., Conclusions: Oral olanzapine was not associated with significant vital sign changes in ED patients. Intramuscular olanzapine also was not associated with vital sign changes in ETOH- patients. In ETOH+ patients, i.m. olanzapine was associated with significant oxygen desaturations. In ETOH+ ED patients, oral olanzapine (with or without benzodiazepines) or haloperidol may be safer choices. ETOH+ patients may have differential effects with the use of i.m. SGAs such as olanzapine and should be studied separately in drug trials., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Varicella-zoster virus (VZV) is a member of the alphaherpesvirus family and the causative agent of chickenpox and shingles. To determine the utility of cynomolgus macaques (Macaca fascicularis) as a nonhuman primate model to evaluate VZV-based simian immunodeficiency virus/human immunodeficiency virus (SIV/HIV) vaccines, we experimentally inoculated 10 animals with the parental Oka (Oka-P) strain of VZV derived from MeWo or Telo-RF cells. VZV DNA could be detected in the lungs as late as 4 days postinfection, with replicating virus detected by shell vial culture assay in one case. Infection did not result in any overt clinical symptoms but was characterized by humoral and cell-mediated immunity in a time frame and at a magnitude similar to those observed following VZV vaccination in humans. The cell line source of VZV inoculum influenced both the magnitude and polyfunctionality of cell-mediated immunity. Animals mounted a vigorous anamnestic antibody response following a second inoculation 12 weeks later. Inoculations resulted in transient increases in CD4(+) T-cell activation and proliferation, as well as a sustained increase in CD4(+) T cells coexpressing CCR5 and α4β7 integrin. In contrast to previous failed attempts to successfully utilize attenuated VZV-Oka as an SIV vaccine vector in rhesus macaques due to suboptimal infectivity and cellular immunogenicity, the ability to infect cynomolgus macaques with Oka-P VZV should provide a valuable tool for evaluating VZV-vectored SIV/HIV vaccines.

Human endogenous retrovirus type K (HERV-K) transcripts are upregulated in the plasma of HIV-infected individuals and have been considered as targets for an HIV vaccine. We evaluated cynomolgus macaque endogenous retrovirus (CyERV) mRNA expression by RT-qPCR in PBMCs isolated from a cohort of animals previously utilized in a live attenuated SIV vaccine trial. CyERV env transcript levels decreased following vaccination (control and vaccine groups) and CyERV env and gag mRNA expression was decreased following acute SIV-infection, whereas during chronic SIV infection, CyERV transcript levels were indistinguishable from baseline. Reduced susceptibility to initial SIV infection, as measured by the number of SIV challenges required for infection, was associated with increased CyERV transcript levels in PBMCs. In vitro analysis revealed that SIV infection of purified CD4(+) T-cells did not alter CyERV gene expression. This study represents the first evaluation of ERV expression in cynomolgus macaques following SIV infection, in an effort to assess the utility of cynomolgus macaques as an animal model to evaluate ERVs as a target for an HIV/SIV vaccine. This non-human primate model system does not recapitulate what has been observed to date in the plasma of HIV-infected humans suggesting that further investigation at the cellular level is required to elucidate the impact of HIV/SIV infection on endogenous retrovirus expression.

Cytomegalovirus (CMV) infection is the most common opportunistic infection in immunosuppressed individuals, such as transplant recipients or people living with HIV/AIDS, and congenital CMV is the leading viral cause of developmental disabilities in infants. Due to the highly species-specific nature of CMV, animal models that closely recapitulate human CMV (HCMV) are of growing importance for vaccine development. Here we present the genomic sequence of a novel nonhuman primate CMV from cynomolgus macaques (Macaca fascicularis; CyCMV). CyCMV (Ottawa strain) was isolated from the urine of a healthy, captive-bred, 4-year-old cynomolgus macaque of Philippine origin, and the viral genome was sequenced using next-generation Illumina sequencing to an average of 516-fold coverage. The CyCMV genome is 218,041 bp in length, with 49.5% G+C content and 84% protein-coding density. We have identified 262 putative open reading frames (ORFs) with an average coding length of 789 bp. The genomic organization of CyCMV is largely colinear with that of rhesus macaque CMV (RhCMV). Of the 262 CyCMV ORFs, 137 are homologous to HCMV genes, 243 are homologous to RhCMV 68.1, and 200 are homologous to RhCMV 180.92. CyCMV encodes four ORFs that are not present in RhCMV strain 68.1 or 180.92 but have homologies with HCMV (UL30, UL74A, UL126, and UL146). Similar to HCMV, CyCMV does not produce the RhCMV-specific viral homologue of cyclooxygenase-2. This newly characterized CMV may provide a novel model in which to study CMV biology and HCMV vaccine development.

Cynomolgus macaques have been widely used as an animal model in preclinical biomedical research and are becoming more popular among HIV/SIV vaccine researchers. Here we report the isolation and characterization of a cytomegalovirus from cynomolgus macaques (CyCMV). CyCMV was isolated from a healthy captive-bred 4-year-old cynomolgus macaque of Filipino origin. The virus was identified by its characteristic growth properties in cell culture, ultrastructural morphology and sequence of viral DNA polymerase and glycoprotein B (gB). CyCMV gB shows 77% identity and 88% homology to rhesus cytomegalovirus (RhCMV) gB and 58% identity and 76% homology to human cytomegalovirus gB at the amino acid level. Phylogenetic analysis using known CMV gB protein sequences show that CyCMV is more closely related to RhCMV than to other primate CMVs. CyCMV down-regulates MHC class I expression on infected cells and we show that the colony-bred cynomolgus macaques have detectable CyCMV-specific humoral and cell-mediated immune responses., (Copyright © 2010 Elsevier Inc. All rights reserved.)

This study's purpose was to determine whether asthma medication use in HIV positive children is associated with human leukocyte antigen (HLA) alleles. We reviewed HLA and medication data collected during the Women and Infants Transmission Study for 124 HIV positive children and their mothers. Analysis revealed that HLA-A68 (P = 0.006) was independent and predictive for time to first asthma medication use. There was a preventive association of Cw6 (P = 0.008) with asthma time. Highly active antiretroviral therapy (HAART) was also associated with time to first asthma medication use (P = 0.05). HLA alleles may modulate risk of developing a need for asthma medications and seem to function independently of the actions of HAART therapy.

Objective: Vaccine-induced CD8(+) T-cell responses in primates have been associated with a reduced simian immunodeficiency virus plasma viral load and enhanced T-cell responses, but cellular vaccines have shown limited success in human trials. We previously described HIV-specific T-cell responses in two groups of highly exposed, persistently seronegative Kenyan female sex workers, and a subset of these participants have subsequently acquired HIV. We examined the impact of pre-existing CD8(+) T-cell responses on post-acquisition outcomes., Design and Methods: HIV-specific CD8(+) T-cell responses had been examined in highly exposed, persistently seronegative participants from the Pumwani and Kibera cohorts, using a combination of virus-specific lysis, proliferation, interferon-gamma production, or all. Plasma viral load set point and HIV-specific T-cell proliferation and cytokine production were now examined post hoc by blinded investigators in the subset of participants who acquired HIV., Results: Pre-acquisition cellular immune assays and post-infection viral load were available for 46 participants, and HIV-specific CD8(+) T-cell responses had been detected in 25 of 46 (54%) participants. Pre-acquisition CD8(+) T-cell responses were associated with a lower post-acquisition HIV viral load set point in both cohorts (pooled analysis, 3.1 vs. 4.1 log(10) RNA copies/ml; P=0.0002) and with enhanced post-acquisition HIV-specific CD8(+) T-cell proliferation (3.8 vs. 1.0%, P=0.03), but with a trend to reduced post-acquisition CD8(+) T-cell interferon-gamma responses., Conclusion: HIV-specific CD8(+) T-cell responses prior to HIV acquisition were associated with a lower HIV viral load and an altered functional profile of post-acquisition CD8(+) T-cell responses.

Background: Direct puncture by a needle is a risk factor for nerve damage. This investigation used scanning electron microscopy (SEM) to attempt to visualize the damage caused by different needles., Method: A 15 cm section of the tibial nerve was removed from the ankle of a patient undergoing below-the-knee amputation. The nerve specimen was punctured perpendicular to the fibers once by each of four needles: an insulated 22 G short-beveled (30 degrees), a 25 G long-beveled Quincke spinal needle, an 18 G Tuohy, and a 25 G Whitacre pencil point. The distal and proximal ends on either side of the needles were marked and the nerve was sectioned into 0.5 cm pieces. Each sample was preserved and then prepared for SEM. The needle tract was observed for evidence of mechanical damage at magnifications between x 47 and x 102 using SEM., Results: The epineurium, perineurium, fascicles, endoneurium, and vessels were identified in each sample. In both the short-beveled and the Whitacre samples, all fascicles along with the surrounding perineurium were intact. In both the Tuohy and the Quincke samples, obvious transection of fascicles and disruption of the perineurium were observed., Conclusions: This investigation suggests that both the Tuohy and the Quincke needles may be more likely to cause trauma to the tibial nerve than either the short-beveled or the Whitacre needles.

Hyperattenuated simian immunodeficiency virus SIVmac239-derived constructs Delta5-CMV and Delta6-CCI are an effort to render SIV incapable of, in practical terms, both reversion and recombination while maintaining the immune features of SIV as a retrovirus. Primary inoculation of cynomolgus macaques with 10(8) 50% tissue culture infective doses (TCID(50)) of Delta5-CMV or Delta6-CCI induced low-level humoral and cellular responses detectable in the absence of measureable in vivo replication. The first of three DNA boosts resulted in elevated gamma interferon (IFN-gamma) enzyme-linked immunospot (ELISPOT) responses to Gag, Pol, and Env in the Delta5-CMV vaccine group compared to the Delta6-CCI vaccine group (P = 0.001). Weekly intrarectal challenge with a low dose of SIVmac239 followed by a dose escalation was conducted until all animals became infected. The mean peak viral load of the Delta5-CMV-vaccinated animals (3.7 x 10(5) copies/ml) was approximately 1 log unit lower than that of the control animals. More dramatically, the viral load set point of these animals was decreased by 3 log units compared to that of the controls (<50 versus 1.64 x 10(4) copies/ml; P < 0.0001). Seventy-five percent (6/8) of vaccine recipients controlled virus below 1,000 copies/ml for at least 6 months, with a subset controlling virus and maintaining substantial CD4 T-cell counts for close to 2 years of follow-up. The correlates of protection from SIV disease progression may lie in the rapidity and protective value of immune responses that occur early in primary SIV infection. Prior immunization with hyperattenuated SIVmac239, even if sterilizing immunity is not achieved, may allow a more advantageous host response.

Eusocial societies present a Darwinian paradox, yet they have evolved independently in insects, mole-rats and symbiotic shrimp. Historically, eusociality has been thought to arise as a response to ecological challenges, mediated by kin selection, but the role of kin selection has recently been questioned. Here we use phylogenetically independent contrasts to test the association of eusociality with ecological performance and genetic structure (via life history) among 20 species of sponge-dwelling shrimp (Synalpheus) in Belize. Consistent with hypotheses that cooperative groups enjoy an advantage in challenging habitats, we show that eusocial species are more abundant, occupy more sponges and have broader host ranges than non-social sister species, and that these patterns are robust to correction for the generally smaller body sizes of eusocial species. In contrast, body size explains less or no variation after accounting for sociality. Despite strong ecological pressures on most sponge-dwellers, however, eusociality arose only in species with non-dispersing larvae, which form family groups subject to kin selection. Thus, superior ability to hold valuable resources may favour eusociality in shrimp but close genetic relatedness is nevertheless key to its origin, as in other eusocial animals.

The presence of interleukin-2 (IL-2)-producing human immunodeficiency virus type 1 (HIV-1)-specific CD4(+) T-cell responses has been associated with the immunological control of HIV-1 replication; however, the causal relationship between these factors remains unclear. Here we show that IL-2-producing HIV-1-specific CD4(+) T cells can be cloned from acutely HIV-1-infected individuals. Despite the early presence of these cells, each of the individuals in the present study exhibited progressive disease, with one individual showing rapid progression. In this rapid progressor, three IL-2-producing HIV-1 Gag-specific CD4(+) T-cell responses were identified and mapped to the following optimal epitopes: HIVWASRELER, REPRGSDIAGT, and FRDYVDRFYKT. Responses to these epitopes in peripheral blood mononuclear cells were monitored longitudinally to >1 year postinfection, and contemporaneous circulating plasma viruses were sequenced. A variant of the FRDYVDRFYKT epitope sequence, FRDYVDQFYKT, was observed in 1/21 plasma viruses sequenced at 5 months postinfection and 1/10 viruses at 7 months postinfection. This variant failed to stimulate the corresponding CD4(+) T-cell clone and thus constitutes an escape mutant. Responses to each of the three Gag epitopes were rapidly lost, and this loss was accompanied by a loss of antigen-specific cells in the periphery as measured by using an FRDYVDRFYKT-presenting major histocompatibility complex class II tetramer. Highly active antiretroviral therapy was associated with the reemergence of FRDYVDRFYKT-specific cells by tetramer. Thus, our data support that IL-2-producing HIV-1-specific CD4(+) T-cell responses can exert immune pressure during early HIV-1 infection but that the inability of these responses to enforce enduring control of viral replication is related to the deletion and/or dysfunction of HIV-1-specific CD4(+) T cells rather than to the fixation of escape mutations at high frequencies.

Cytotoxic T-lymphocyte (CTL) epitopes within the HIV genome are subject to negative and positive selective pressures, the balance of which influences CTL escape at a given epitope. We investigated whether viral fitness requirements dictate conservation of the HLA-A2 restricted immunodominant epitope SLYNTVATL (SL9). Viral clones incorporating changes throughout the SL9 epitope region were compared to consensus SL9 virus in terms of replication kinetics and relative viral fitness. Constructs recapitulating in vivo SL9-CTL escape variants showed markedly little effect on replication and fitness, as did non-natural conservative mutations targeting immunologically relevant positions of the epitope. Although certain residues of the epitope were constrained by viral requirements, our research reveals that there are multiple SL9 variants that are well tolerated virologically but fail to arise in vivo. In light of this data, assumptions regarding the balance of immune and viral selective pressures on this immunodominant epitope sequence need to be reassessed.

Background: Several mucosal innate immune proteins exhibit HIV inhibitory activity and their analogues are potential microbicide candidates. However, their clinical associations and in-vivo role in cervicovaginal host defense against HIV acquisition are poorly defined., Methods: Cervicovaginal secretions (CVSs) were collected from HIV uninfected Kenyan sex workers at enrolment into an HIV prevention trial. After trial completion, CVS from participants acquiring HIV (cases) and matched controls were assessed for levels of innate immune factors and HIV neutralizing capacity, by blinded investigators. Cross-sectional and prospective associations of innate immune factors were examined., Results: CVS contained high levels of defensins (human neutrophil peptide-1-3 and human beta defensin-2-3), LL-37 and secretory leukocyte protease inhibitor. Regulated upon activation normal T-cell expressed and secreted levels were lower, and IFNalpha was undetectable. CVS from 20% of participants neutralized a clade A primary HIV isolate, and 12% neutralized both clade A and C isolates. HIV neutralization was correlated with human neutrophil peptide-1-3 (alpha-defensins) and LL-37 levels. However, alpha-defensin and LL-37 levels were increased in participants with bacterial sexually transmitted infections and were independently associated with increased HIV acquisition in multivariate analysis., Conclusions: Despite significant HIV inhibitory activity, cervicovaginal levels of alpha-defensins and LL-37 were associated with increased HIV acquisition, perhaps due to their association with bacterial sexually transmitted infections.

Background: The host immune response against mucosally acquired pathogens may be influenced by the mucosal immune milieu during acquisition. As Neisseria gonorrhoeae can impair dendritic cell and T-cell immune function, we hypothesized that coinfection during HIV acquisition would impair subsequent systemic T-cell responses., Methods: Monthly screening for sexually transmitted infections was performed in high risk, HIV seronegative Kenyan female sex workers as part of an HIV prevention trial. Early HIV-specific CD8 T-cell responses and subsequent HIV viral load set point were assayed in participants acquiring HIV, and were correlated with the presence of prior genital infections during HIV acquisition., Results: Thirty-five participants acquired HIV during follow-up, and 16 out of 35 (46%) had a classical sexually transmitted infection at the time of acquisition. N. gonorrhoeae coinfection was present during HIV acquisition in 6 out of 35 (17%), and was associated with an increased breadth and magnitude of systemic HIV-specific CD8 T-cell responses, using both interferon-gamma gamma and MIP-1 beta as an output. No other genital infections were associated with differences in HIV-specific CD8 T-cell response, and neither N. gonorrhoeae nor other genital infections were associated with differences in HIV plasma viral load at set point., Conclusion: Unexpectedly, genital N. gonorrhoeae infection during heterosexual HIV acquisition was associated with substantially enhanced HIV-specific CD8 T-cell responses, although not with differences in HIV viral load set point. This may have implications for the development of mucosal HIV vaccines and adjuvants.

Background: Chronic coinfection with herpes simplex virus type 2 (HSV-2) and human immunodeficiency virus (HIV) has been associated with an increased HIV viral load and more rapid disease progression, perhaps related to HSV-2-associated alterations in host immunity., Methods: Studies were nested within (1) a cross-sectional study of men coinfected with HIV and HSV-2 and (2) women not infected with HIV, both before and after HSV-2 acquisition. HSV-2 infection status was determined by ELISA. HIV-specific CD8(+) T cell epitopes were mapped, and proliferation of HIV-specific cells was also assessed. Systemic inflammatory and regulatory T cell populations were assayed by flow cytometry., Results: The breadth of both the HIV-specific CD8(+) T cell interferon-gamma and proliferative responses was reduced in participants coinfected with HIV and HSV-2, independent of the HIV plasma viral load and CD4(+) T cell count, and the magnitude of the responses was also reduced. HSV-2 infection in this group was associated with increased T cell CD38 expression but not with differences in the proportion of CD4(+) FoxP3(+) regulatory T cells. However, in women not infected with HIV, acquisition of HSV-2 was associated with an increase in the proportion of regulatory T cells., Conclusions: HSV-2 coinfection was associated with reduced HIV-specific T cell responses and systemic inflammation. The immune effects of HSV-2 may underlie the negative impact that this coinfection has on the clinical course of HIV infection.

Objectives: HIV-neutralizing immunoglobulin A (IgA) and HIV-specific cellular immunity have been described in highly exposed, persistently seronegative (HEPS) individuals, but well controlled studies have not been performed. We performed a prospective, nested case-control study to examine the association of genital IgA and systemic cellular immune responses with subsequent HIV acquisition in high-risk Kenyan female sex workers (FSWs)., Design and Methods: A randomized trial of monthly antibiotic prophylaxis to prevent sexually transmitted disease/HIV infection was performed from 1998 to 2002 in HIV-uninfected Kenyan FSWs. After the completion of trial, FSWs who had acquired HIV (cases) were matched 1: 4 with persistently uninfected controls based on study arm, duration of HIV-seronegative follow-up, and time of cohort enrolment. Blinded investigators assayed the ability at enrolment of genital IgA to neutralize primary HIV isolates as well as systemic HIV-specific cellular IFNgamma-modified enzyme-linked immunospot and proliferative responses., Results: The study cohort comprised 113 FSWs: 24 cases who acquired HIV and 89 matched controls. Genital HIV-neutralizing IgA was associated with reduced HIV acquisition (P = 0.003), as was HIV-specific proliferation (P = 0.002), and these associations were additive. HIV-specific IFNgamma production did not differ between case and control groups. In multivariable analysis, HIV-neutralizing IgA and HIV-specific proliferation each remained independently associated with lack of HIV acquisition. Genital herpes (HSV2) was associated with increased HIV risk and with reduced detection of HIV-neutralizing IgA., Conclusion: Genital HIV-neutralizing IgA and systemic HIV-specific proliferative responses, assayed by blinded investigators, were prospectively associated with HIV nonacquisition. The induction of these immune responses may be an important goal for HIV vaccines.

Objectives: Female sex workers (FSWs) form a core group at high risk of both sexual HIV acquisition and secondary transmission. The magnitude of these risks may vary by sexual risk taking, partner HIV prevalence, host immune factors and genital co-infections. We examined temporal trends in HIV prevalence and per-act incidence, adjusted for behavioral and other variables, in FSWs from Nairobi, Kenya., Methods: An open cohort of FSWs followed since 1985. Behavioral and clinical data were collected six monthly from 1985 to 2005, and sexually transmitted infection (STI) diagnostics and HIV serology performed. A Cox proportional hazards model with time-dependent covariables was used to estimate infection risk as a function of calendar time., Results: HIV prevalence in new FSW enrollees peaked at 81% in 1986, and was consistently below 50% after 1997. Initially uninfected FSWs remained at high risk of acquiring HIV throughout the study period, but the rate of HIV acquisition during unprotected sex with a casual client declined by over four-fold. This reduction correlated closely with decreases in gonorrhea prevalence, and predated reductions in the Kenyan HIV population prevalence by over a decade., Conclusions: The per-act rate of HIV acquisition in high-risk Nairobi FSWs fell dramatically between 1985 and 2005. This decline may represent the impact of improved STI prevention/therapy, immunogenetic shifts in at-risk women, or changes in the proportion of HIV exposures occurring with clients who had acute HIV infection. Declining HIV incidence in high-risk cohorts may predict and/or be causally related to future reductions in population prevalence.

Background: Prevalent herpes simplex virus type 2 (HSV-2) infection increases human immunodeficiency virus acquisition. We hypothesized that HSV-2 infection might also predispose individuals to acquire other common sexually transmitted infections (STIs)., Methods: We studied the association between prevalent HSV-2 infection and STI incidence in a prospective, randomized trial of periodic STI therapy among Kenyan female sex workers. Participants were screened monthly for infection with Neisseria gonorrhoeae and Chlamydia trachomatis, and at least every 6 months for bacterial vaginosis (BV) and infection with Treponema pallidum, Trichomonas vaginalis, and/or HSV-2., Results: Increased prevalence of HSV-2 infection and increased prevalence of BV were each associated with the other; the direction of causality could not be determined. After stratifying for sexual risk-taking, BV status, and antibiotic use, prevalent HSV-2 infection remained associated with an increased incidence of infection with N. gonorrhoeae (incidence rate ratio [IRR], 4.3 [95% confidence interval {CI}, 1.5-12.2]), T. vaginalis (IRR, 2.3 [95% CI, 1.3-4.2]), and syphilis (IRR, 4.7 [95% CI, 1.1-19.9]). BV was associated with increased rates of infection with C. trachomatis (IRR, 2.1 [95% CI, 1.1-3.8]) and T. vaginalis (IRR, 8.0 [95% CI, 3.2-19.8]). CONCLUSION; Increased prevalences of HSV-2 infection and BV were associated with each other and also associated with enhanced susceptibility to an overlapping spectrum of other STIs. Demonstration of causality will require clinical trials that suppress HSV-2 infection, BV, or both.