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Background There are limited data on how coronavirus disease 2019 (COVID-19) severity, timing of infection, and subsequent vaccination impact transplacental transfer and persistence of maternal and infant antibodies. Methods In a longitudinal cohort of pregnant women with polymerase chain reaction-confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, maternal/infant sera were collected at enrollment, delivery/birth, and 6 months. Anti-SARS-CoV-2 spike immunoglobulin (Ig)G, IgM, and IgA were measured by enzyme-linked immunosorbent assay. Results Two-hundred fifty-six pregnant women and 135 infants were enrolled; 148 maternal and 122 neonatal specimens were collected at delivery/birth; 45 maternal and 48 infant specimens were collected at 6 months. Sixty-eight percent of women produced all anti-SARS-CoV-2 isotypes at delivery (IgG, IgM, IgA); 96% had at least 1 isotype. Symptomatic disease and vaccination before delivery were associated with higher maternal IgG at labor and delivery. Detectable IgG in infants dropped from 78% at birth to 52% at 6 months. In the multivariate analysis evaluating factors associated with detectable IgG in infants at delivery, significant predictors were 3rd trimester infection (odds ratio [OR] = 4.0), mild/moderate disease (OR = 4.8), severe/critical disease (OR = 6.3), and maternal vaccination before delivery (OR = 18.8). No factors were significant in the multivariate analysis at 6 months postpartum. Conclusions Vaccination in pregnancy post-COVID-19 recovery is a strategy for boosting antibodies in mother-infant dyads.

We studied the pharmacokinetics (PK) and safety of 100mg doravirine and doravirine/lamivudine/tenofovir disoproxil fumarate fixed dose combination (100/300/300mg- DOR FDC) treatment in adolescents with HIV-1.Adolescents ages 12-18 years enrolled in two sequential cohorts. Cohort 1 evaluated intensive PK and short-term safety of 100mg single-dose doravirine in adolescents ≥ 35 kg. Cohort 2 participants either initiated treatment with DOR FDC (antiretroviral (ARV)-naïve) or switched to DOR FDC from a previous ARV regimen (virologically-suppressed). The first 10 Cohort 2 participants had intensive PK evaluations, and safety, sparse PK and HIV RNA were assessed through week 24.Fifty-five adolescents, median age 15.0 years and baseline weight 51.5 kg, enrolled. Nine participants completed Cohort 1 PK assessments (8 of the 9 participants weighed ≥45 kg) and 45 initiated study drug in Cohort 2. Doravirine geometric mean (GM) AUC0-∞ was 34.8 μM∙h and GM C24 was 514 nM following a single dose, with a predicted steady state GM C24,ss,pred of 690 nM. Cohort 2 enrolled adolescents weighing ≥ 45 kg. Plasma concentrations of doravirine, tenofovir and lamivudine achieved by Cohort 2 participants were similar to those reported in adults. No drug-related serious or grade 3 or 4 adverse events occurred. Forty-two of 45 participants (93.3%; 95% CI [81.7, 98.6]) achieved or maintained HIV-1 RNA40 copies/mL.Doravirine and DOR FDC achieved target PK in adolescents with HIV-1. DOR FDC was well tolerated and maintained excellent virologic efficacy through 24 weeks, offering a favorable option for adolescents.

There are limited data on how COVID-19 severity, timing of infection, and subsequent vaccination impact transplacental transfer and persistence of maternal and infant antibodies.In a longitudinal cohort of pregnant women with PCR-confirmed SARS-CoV-2 infection, maternal/infant sera were collected at enrollment, delivery/birth, and 6 months. Anti-SARS-CoV-2 spike IgG, IgM and IgA were measured by ELISA.256 pregnant women and 135 infants were enrolled; 148 maternal and 122 neonatal specimens were collected at delivery/birth; 45 maternal and 48 infant specimens were collected at 6 months. Sixty-eight percent of women produced all anti-SARS-CoV-2 isotypes at delivery (IgG, IgM, IgA); 96% had at least one isotype. Symptomatic disease, and vaccination prior to delivery, were associated with higher maternal IgG at LD. Detectable IgG in infants dropped from 78% at birth to 52% at 6 months. In the multivariate analysis evaluating factors associated with detectable IgG in infants at delivery, significant predictors were 3rd trimester infection (OR 4.0), mild/moderate disease (OR 4.8), severe/critical disease (OR 6.3), and maternal vaccination prior to delivery (OR 18.8). No factors were significant in the multivariate analysis at 6 months postpartum.Vaccination in pregnancy post-COVID-19 recovery is a strategy for boosting antibodies in mother-infant dyads.

ObjectiveThe rectal microbiome was examined to assess the relationship between the microbiome and liver disease in HIV-infection.DesignEighty-two HIV-1 mono-infected individuals from the PROSPEC-HIV-study (NCT02542020) were grouped into three liver health categories based on results of controlled attenuation parameter (CAP) and liver stiffness measurement (LSM) of transient elastography: normal (n = 30), steatosis (n = 30), or fibrosis (n = 22).MethodsLiver steatosis and fibrosis were defined by CAP at least 248 dB/m and LSM at least 8.0 kPa, respectively. 16S rRNA gene and whole genome shotgun metagenomic sequencing were performed on rectal swabs. Bacterial differences were assessed using zero-inflated negative binomial regression and random forests modeling; taxonomic drivers of functional shifts were identified using FishTaco.ResultsLiver health status explained four percentage of the overall variation (r2 = 0.04, P = 0.003) in bacterial composition. Participants with steatosis had depletions of Akkermansia muciniphila and Bacteroides dorei and enrichment of Prevotella copri, Finegoldia magna, and Ruminococcus bromii. Participants with fibrosis had depletions of Bacteroides stercoris and Parabacteroides distasonis and enrichment of Sneathia sanguinegens. In steatosis, functional analysis revealed increases in primary and secondary bile acid synthesis encoded by increased Eubacterium rectale, F. magna, and Faecalibacterium prausnitzii and decreased A. muciniphila, Bacteroides fragilis and B. dorei. Decreased folate biosynthesis was driven by similar changes in microbial composition.ConclusionHIV mono-infection with steatosis or fibrosis had distinct microbial profiles. Some taxa are similar to those associated with non-alcoholic fatty liver disease in HIV-negative populations. Further studies are needed to define the role of the gut microbiota in the pathogenesis of liver disease in HIV-infected persons.

The lower efficacy of the COVID-19 mRNA vaccines in 5-11 year old children was unexpected. Neutralizing antibody titers elicited by the vaccines in children, adolescents, and young adults suggest that the lower efficacy is not due to the lower dosage. Confirming the efficacy of these vaccines in children, determining if mRNA vaccination strategies are less effective in younger children, as well as optimizing the dosage, dosing intervals, and number of doses needed in children, adolescents, and young adults are critical to improve vaccination strategies for these populations going forward.

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While pregnancy increases the risk for severe COVID19, the clinical and immunological implications of COVID19 on maternal-fetal health remain unknown. Here, we present the clinical and immunological landscapes of 93 COVID19 mothers and 45 of their SARS-CoV-2-exposed infants through comprehensive serum proteomics profiling for >1400 cytokines of their peripheral and cord blood specimens. Prenatal SARS-CoV-2 infection triggers NF-κB-dependent proinflammatory immune activation. Pregnant women with severe COVID19 show increased inflammation and unique IFNλ antiviral signaling, with elevated levels of IFNL1 and IFNLR1. Furthermore, SARS-CoV-2 infection re-shapes maternal immunity at delivery altering the expression of pregnancy complication-associated cytokines, inducing MMP7, MDK, ESM1, and reducing BGN and CD209. Finally, COVID19-exposed infants exhibit induction of T cell-associated cytokines (IL33, NFATC3 and CCL21), while some undergo IL-1β/IL-18/CASP1 axis-driven neonatal respiratory distress despite birth at term. Our findings demonstrate COVID19-induced immune rewiring in both mothers and neonates, warranting long-term clinical follow-up to mitigate potential health risks., Graphical Abstract, The study performed by Foo et al. unveils distinct immune alterations of mothers and infants induced by SARS-CoV-2 infection during pregnancy. These findings highlight the importance of long-term post-pregnancy clinical follow-up of mother-infant dyads to prevent potential unforeseen health conditions following prenatal SARS-CoV-2 infection.

Non-alcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease in western countries both in children and adults. Metabolic dysregulation associated with gut microbial dysbiosis may influence disease progression from hepatic steatosis to inflammation and subsequent fibrosis. Using a multi-omics approach, we profiled the oral and fecal microbiome and plasma metabolites from 241 predominantly Latino children with non-alcoholic steatohepatitis (NASH), non-alcoholic fatty liver (NAFL), and controls. Children with more severe liver pathology were dysbiotic and had increased gene content associated with lipopolysaccharide biosynthesis and lipid, amino acid and carbohydrate metabolism. These changes were driven by increases in Bacteroides and concomitant decreases of Akkermansia, Anaerococcus, Corynebacterium, and Finegoldia. Non-targeted mass spectrometry revealed perturbations in one-carbon metabolism, mitochondrial dysfunction, and increased oxidative stress in children with steatohepatitis and fibrosis. Random forests modeling of plasma metabolites was highly predictive of non-alcoholic steatohepatitis (NASH) (97% accuracy) and hepatic fibrosis, steatosis and lobular inflammation (93.8% accuracy), and can differentiate steatohepatitis from simple steatosis (90.0% accuracy). Multi-omics predictive models for disease and histology findings revealed perturbations in one-carbon metabolism, mitochondrial dysfunction, and increased oxidative stress in children with steatohepatitis and fibrosis. These results highlight the promise of non-invasive biomarkers for the growing epidemic of fatty liver disease.

Objective:We employed a high-dimensional covariate adjustment method in microbiome analysis to better control for behavioural and clinical confounders, and in doing so examine the effects of HIV on the rectal microbiome.Design:Three hundred and eighty-three MSM were grouped into four HIV viremia cat

Studies of two SARS-CoV-2 mRNA vaccines suggested that they yield ∼95% protection from symptomatic infection at least short-term, but important clinical questions remain. It is unclear how vaccine-induced antibody levels quantitatively compare to the wide spectrum induced by natural SARS-CoV-2 infection. Vaccine response kinetics and magnitudes in persons with prior COVID-19 compared to virus-naı̈ve persons are not well-defined. The relative stability of vaccine-induced versus infection-induced antibody levels is unclear. We addressed these issues with longitudinal assessments of vaccinees with and without prior SARS-CoV-2 infection using quantitative enzyme-linked immunosorbent assay (ELISA) of anti-RBD antibodies. SARS-CoV-2-naı̈ve individuals achieved levels similar to mild natural infection after the first vaccination; a second dose generated levels approaching severe natural infection. In persons with prior COVID-19, one dose boosted levels to the high end of severe natural infection even in those who never had robust responses from infection, increasing no further after the second dose. Antiviral neutralizing assessments using a spike-pseudovirus assay revealed that virus-naı̈ve vaccinees did not develop physiologic neutralizing potency until the second dose, while previously infected persons exhibited maximal neutralization after one dose. Finally, antibodies from vaccination waned similarly to natural infection, resulting in an average of ∼90% loss within 90 days. In summary, our findings suggest that two doses are important for quantity and quality of humoral immunity in SARS-CoV-2-naı̈ve persons, while a single dose has maximal effects in those with past infection. Antibodies from vaccination wane with kinetics very similar to that seen after mild natural infection; booster vaccinations will likely be required.

Background Sentiments of vaccine hesitancy and distrust in public health institutions have complicated the government-led coronavirus disease 2019 (COVID-19) vaccine control strategy in the United States. As the first to receive the vaccine, COVID-19 vaccine attitudes among frontline workers are consequential for COVID-19 control and public opinion of the vaccine. Methods In this study, we used a repeated cross-sectional survey administered at 3 time points between 24 September 2020 and 6 February 2021 to a cohort of employees of the University of California, Los Angeles Health and the Los Angeles County Fire Department. The primary outcome of interest was COVID-19 vaccination intent and vaccine uptake. Results Confidence in COVID-19 vaccines and vaccine uptake rose significantly over time. At survey 1, confidence in vaccine protection was 46.4% among healthcare workers (HCWs) and 34.6% among first responders (FRs); by survey 3, this had risen to 90.0% and 75.7%, respectively. At survey 1, about one-third of participants intended to receive a vaccine as soon as possible. By survey 3, 96.0% of HCWs and 87.5% of FRs had received a COVID-19 vaccine. Conclusions Attitudes toward vaccine uptake increased over the study period, likely a result of increased public confidence in COVID-19 vaccines, targeted communications, a COVID-19 winter surge in Los Angeles County, and ease of access from employer-sponsored vaccine distribution.

BackgroundSARS-CoV-2 infections of infants and toddlers are usually mild but can result in life-threatening disease. SARS-CoV-2 RNA been detected in the breast milk of lactating women, but the potential role of breastfeeding in transmission to infants has remained uncertain.MethodsBreast milk specimens were examined for the presence of the virus by RT-PCR and/or culture. Specimens that contained viral RNA (vRNA) were examined for the presence of subgenomic coronavirus RNA (sgRNA), a putative marker of infectivity. Culture methods were used to determine the thermal stability of SARS-CoV-2 in human milk.ResultsBreast milk samples from 110 women (65 confirmed with a SARS-CoV-2 diagnostic test, 36 with symptoms but without tests, and 9 with symptoms but a negative SARS-CoV-2 diagnostic test) were tested by RT-PCR (285 samples) and/or viral culture (160 samples). Although vRNA of SARS-CoV-2 was detected in the milk of 7 of 110 (6%) women with either a confirmed infection or symptomatic illness, and in 6 of 65 (9%) of women with a positive SARS-CoV-2 diagnostic test, virus was not detected in any culture. None of the 7 milk specimens with detectable vRNA contained sgRNA. Notably, when artificially added to human milk in control experiments, infectious SARS-CoV-2 could be cultured despite several freeze-thaw cycles, as occurs in the storage and usage of human milk.ConclusionsSARS-CoV-2 RNA can be found infrequently in the breastmilk of women with recent infection, but we found no evidence that breastmilk contains infectious virus or that breastfeeding represents a risk factor for transmission of infection to infants.Key PointsQuestionSARS-CoV-2 RNA has been detected in a small number of human milk samples collected from recently infected women. The role of breastfeeding in transmission of the virus to infants has remained uncertain due to the small number of specimens analyzed in any study published thus far.FindingsIn a total study group of 110 women, SARS-CoV-2 RNA was detected in milk from 6 of 65 women (9.2%) with recent confirmed infection. Neither infectious virus nor subgenomic RNA (a marker of virus infectivity) were detected in any of the samples.MeaningWe found no evidence that infectious SARS-CoV-2 is present milk from recently infected women, even if SARS-CoV-2 PCR tests are positive, providing reassurance of the safety of breastfeeding.

Two mRNA vaccines (BNT162b2 and mRNA-1273) against severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) are globally authorized as a two-dose regimen. Understanding the magnitude and duration of protective immune responses is vital to curbing the pandemic. We enrolled 461 high-risk health services workers at the University of California, Los Angeles (UCLA) and first responders in the Los Angeles County Fire Department (LACoFD) to assess the humoral responses in previously infected (PI) and infection naïve (NPI) individuals to mRNA-based vaccines (BNT162b2/Pfizer- BioNTech or mRNA-1273/Moderna). A chemiluminescent microparticle immunoassay was used to detect antibodies against SARS-CoV-2 Spike in vaccinees prior to (n = 21) and following each vaccine dose (n = 246 following dose 1 and n = 315 following dose 2), and at days 31–60 (n = 110) and 61–90 (n = 190) following completion of the 2-dose series. Both vaccines induced robust antibody responses in all immunocompetent individuals. Previously infected individuals achieved higher median peak titers (p = 0.002) and had a slower rate of decay (p = 0.047) than infection-naïve individuals. mRNA-1273 vaccinated infection-naïve individuals demonstrated modestly higher titers following each dose (p = 0.005 and p = 0.029, respectively) and slower rates of antibody decay (p = 0.003) than those who received BNT162b2. A subset of previously infected individuals (25%) required both doses in order to reach peak antibody titers. The biologic significance of the differences between previously infected individuals and between the mRNA-1273 and BNT162b2 vaccines remains uncertain, but may have important implications for booster strategies.

HIV-1 is present in many secretions including oral, intestinal, genital, and breast milk. However, most people exposed to HIV-1 within these mucosal compartments do not become infected despite often frequent and repetitive exposure over prolonged periods of time. In this review, we discuss what is known about the levels of cell-free HIV RNA, cell-associated HIV DNA and cellassociated HIV RNA in external secretions. Levels of virus are usually lower than contemporaneously obtained blood, increased in settings of inflammation and infection, and decreased in response to antiretroviral therapy. Additionally, each mucosal compartment has unique innate and adaptive immune responses that affect the composition and presence of HIV-1 within each external secretion. We discuss the current state of knowledge about the types and amounts of virus present in the various excretions, touch on innate and adaptive immune responses as they affect viral levels, and highlight important areas for further study.

Beginning April 8, 2020, we enrolled 1787 frontline heath workers who were asymptomatic for COVID-19 into a longitudinal surveillance study. During that time 4 healthcare workers and 6 first responders tested positive for SARS-CoV-2 by RT-PCR. Additionally, 43 healthcare workers and 55 first responders had detectable IgG antibodies to SARS-CoV-2.

ImportanceHealthcare workers (HCW) are slated to be early recipients of SARS-CoV-2 vaccines due to increased risk of exposure to patients with COVID-19, and will be tasked with administering approved vaccines to the general population. As lynchpins of the vaccination effort, HCWs’ opinions of a vaccine’s safety and efficacy may affect both public perception and uptake of the vaccine. Therefore, it is crucial to understand and address potential hesitancy prior to vaccine administration.ObjectiveTo understand healthcare workers’ attitudes about vaccine safety, efficacy, and acceptability in the context of the COVID-19 pandemic, including acceptance of a novel coronavirus vaccine.Design, Setting, ParticipantsA cross-sectional survey was distributed to participants enrolled in a longitudinal cohort study surveilling SARS-CoV-2 infection among 1,093 volunteer sampled University of California, Los Angeles (UCLA) Health System employees. Surveys were completed online between September 24 and October 16, 2020. In total, 609 participants completed this supplemental survey.ResultsWe averaged a 9-statement Likert scale matrix scored from 1 (“strongly disagree”) to 5 (“strongly agree”) and found respondents overwhelmingly confident about vaccine safety (4.47); effectiveness (4.44); importance, self-protection, and community health (4.67). Notably, 47.3% of respondents reported unwillingness to participate in a coronavirus vaccine trial, and most (66.5%) intend to delay vaccination. The odds of reporting intent to delay coronavirus vaccine uptake were 4.15 times higher among nurses, 2.45 times higher among other personnel with patient contact roles, and 2.15 times higher among those without patient contact compared to doctors. Evolving SARS-CoV-2 science (76.0%), current political climate (57.6%), and fast-tracked vaccine development timeline (83.4%) were cited as primary variables impacting HCW decisions to undergo vaccination. Of note, these results were obtained prior to release of Phase III data from companies manufacturing vaccines in the U.S.Conclusions and RelevanceDespite overall confidence in vaccines, a majority of HCW expressed concerns over a novel coronavirus vaccine. A large proportion plan to delay vaccine uptake due to concerns about expedited development, emerging scientific discoveries, and the political climate. Forthcoming vaccination campaigns must address these unique points of coronavirus vaccine hesitancy in order to achieve adequate vaccine coverage.

Covid-19 Antibodies after Mild Infection Among 34 volunteers who had recovered from mild Covid-19 illness, antiviral antibodies to the receptor-binding domain of the viral spike protein declined wi...

To The Editor, Currently, the U.S. Centers for Disease Control and Prevention, American Academy of Pediatrics and the World Health Organization advise that women who are infected with SARS-CoV-2 may choose to breastfeed with appropriate protections to prevent transmission of the virus through respiratory droplets. However, the potential for exposure to SARS-CoV-2 through breastfeeding is currently unknown. To date, case reports on breastmilk samples from a total of 24 SARS-CoV-2-infected women have been published. Of those, viral RNA was detected in ten breastmilk samples from four women. In some but not all cases, environmental contamination as the source of the virus or retrograde flow from an infected infant could not be ruled out. We established a quantitative RT-PCR assay for SARS-CoV-2 in breastmilk with a limit of detection of 250 copies per mL and validated it by spiking breastmilk from uninfected women with known amounts of viral RNA. In addition, we established tissue culture methods to detect replication-competent SARS-CoV-2 in breastmilk. No viral RNA nor culturable virus was detected after Holder pasteurization of breastmilk samples that had been spiked with replication-competent SARS-CoV-2 (see Supplement). Between March 27 and May 6, 2020, we collected and analyzed 64 serial breastmilk samples from 18 SARS-CoV-2-infected women residing in the U.S. (see Supplement for clinical characteristics). Breastmilk samples were collected before and after women had a positive SARS-CoV-2 RT-PCR test and all but one woman had symptomatic disease (see Figure). One of the 64 breastmilk samples had detectable SARS-CoV-2 RNA by RT-PCR. The positive sample was collected on the day of symptom onset but one sample 2 days prior to symptom onset and two subsequent samples, collected 12 and 41 days later, tested negative for viral RNA. In addition, a subset of 26 breastmilk samples from nine women were tested for the presence of replication-competent virus using our established culture methods, and all were negative including the one sample that tested positive for viral RNA by RT-PCR. Although SARS-CoV-2 RNA was detected in one milk sample from one of eighteen infected women, the viral culture for that sample was negative. This suggests that SARS-CoV-2 RNA does not represent replication-competent virus and that breastmilk itself is likely not a source of infection for the infant. Furthermore, when control breastmilk samples spiked with replication-competent SARS-CoV-2 virus were treated by Holder pasteurization, a process commonly performed by donor milk banks, no replication-competent virus nor viral RNA was detectable. Further research to confirm these findings is needed, as well as an examination of convalescent milk for the presence of antibodies against SARS-CoV-2.

ObjectivesThe prevalence of obesity is rising among people living with HIV, which may synergistically increase inflammation and the risk of associated diseases. Disruption of gut bacterial communities may be one of the key drivers of this inflammation; however, the combined effects of HIV and obesity on the microbiome have not been explored.MethodsThis study included 381 men who have sex with men. Thirty-nine were HIV-positive and obese (H+O+), 143 were HIV-positive and nonobese, 64 were HIV-negative and obese, and 135 were HIV-negative and nonobese. Microbiome composition was assessed by targeted sequencing of the V4 region of the 16S ribosomal RNA (rRNA) gene using rectal swab samples. Inverse probability of treatment-weighted marginal structural models wereused to investigate differences in microbial composition between groups while controlling for numerous clinical and behavioural confounders.ResultsSignificant variability in microbial composition was explained by the combination of HIV and obesity, over and above each condition alone (R2 for the marginal contribution of the H+/O+ group=0.008; P=0.001). H+O+ participants had the highest ratios of Prevotella to Bacteroides, a pro-inflammatory enterotype that has been described in HIV infection and obesity independently. H+O+ participants had lower levels of Bacteroides and Veillonella than all other groups, suggesting a synergistic effect of HIV and obesity on these genera.ConclusionsOur findings support the hypothesis that HIV and obesity act together to disrupt gut microbial communities, which may help explain higher levels of generalized inflammation among people living with both HIV and obesity.

To The Editor, Currently, the U.S. Centers for Disease Control and Prevention, American Academy of Pediatrics and the World Health Organization advise that women who are infected with SARS-CoV-2 may choose to breastfeed with appropriate protections to prevent transmission of the virus through respiratory droplets. However, the potential for exposure to SARS-CoV-2 through breastfeeding is currently unknown. To date, case reports on breastmilk samples from a total of 24 SARS-CoV-2-infected women have been published. Of those, viral RNA was detected in ten breastmilk samples from four women. In some but not all cases, environmental contamination as the source of the virus or retrograde flow from an infected infant could not be ruled out. We established a quantitative RT-PCR assay for SARS-CoV-2 in breastmilk with a limit of detection of 250 copies per mL and validated it by spiking breastmilk from uninfected women with known amounts of viral RNA. In addition, we established tissue culture methods to detect replication-competent SARS-CoV-2 in breastmilk. No viral RNA nor culturable virus was detected after Holder pasteurization of breastmilk samples that had been spiked with replication-competent SARS-CoV-2 (see Supplement). Between March 27 and May 6, 2020, we collected and analyzed 64 serial breastmilk samples from 18 SARS-CoV-2-infected women residing in the U.S. (see Supplement for clinical characteristics). Breastmilk samples were collected before and after women had a positive SARS-CoV-2 RT-PCR test and all but one woman had symptomatic disease (see Figure). One of the 64 breastmilk samples had detectable SARS-CoV-2 RNA by RT-PCR. The positive sample was collected on the day of symptom onset but one sample 2 days prior to symptom onset and two subsequent samples, collected 12 and 41 days later, tested negative for viral RNA. In addition, a subset of 26 breastmilk samples from nine women were tested for the presence of replication-competent virus using our established culture methods, and all were negative including the one sample that tested positive for viral RNA by RT-PCR. Although SARS-CoV-2 RNA was detected in one milk sample from one of eighteen infected women, the viral culture for that sample was negative. This suggests that SARS-CoV-2 RNA does not represent replication-competent virus and that breastmilk itself is likely not a source of infection for the infant. Furthermore, when control breastmilk samples spiked with replication-competent SARS-CoV-2 virus were treated by Holder pasteurization, a process commonly performed by donor milk banks, no replication-competent virus nor viral RNA was detectable. Further research to confirm these findings is needed, as well as an examination of convalescent milk for the presence of antibodies against SARS-CoV-2.

Author(s): Farr Zuend, Christina; Tobin, Nicole H; Vera, Trisha; Kotyrba, Lani; Noel-Romas, Laura; Birse, Kenzie; Mutch, Sarah; Li, Fan; Lee, David; McCorrister, Stuart; Westmacott, Garrett; Aldrovandi, Grace M; Burgener, Adam D | Abstract: ProblemPregnant women are at increased risk of HIV acquisition, but the biological mechanisms contributing to this observation are not well understood.Method of studyHere, we assessed host immune and microbiome differences in the vaginal mucosa of healthy pregnant and non-pregnant women using a metaproteomics approach. Cervicovaginal lavage (CVL) samples were collected from 23 pregnant and 25 non-pregnant women.ResultsMass spectrometry analysis of CVL identified 550 human proteins and 376 bacterial proteins from 11 genera. Host proteome analysis indicated 56 human proteins (10%) were differentially abundant (Pnln.05) between pregnant and non-pregnant women, including proteins involved in angiogenesis (Pn=n3.36E-3), cell movement of phagocytes (Pn=n1.34E-6), and permeability of blood vessels (Pn=n1.27E-4). The major bacterial genera identified were Lactobacillus, Gardnerella, Prevotella, Megasphaera, and Atopobium. Pregnant women had higher levels of Lactobacillus species (Pn=n.017) compared with non-pregnant women. Functional pathway analysis indicated that pregnancy associated with changes to bacterial metabolic pathway involved in energy metabolism, which were increased in pregnant women (Pn=n.035).ConclusionOverall, pregnant women showed differences in the cervicovaginal proteome and microbiome that may be important for HIV infection risk.

BACKGROUND: Infants with gastroschisis require operations and lengthy hospitalizations due to intestinal dysmotility. Dysbiosis may contribute to these problems. Little is known on the microbiome of gastroschisis infants. METHODS: The purpose of this study was to investigate the fecal microbiome in gastroschisis infants. Microbiome profiling was performed by sequencing the V4 region of the 16S rRNA gene. The microbiome of gastroschisis infants was compared to the microbiome of healthy controls, and the effects of mode of birth delivery, gestational age, antibiotic duration, and nutrition type on microbial composition and diversity were investigated. RESULTS: The microbiome of gastroschisis infants (n=13) was less diverse (Chao1, p < 0.001), lacked Bifidobacterium (p = 0.001) and had increased Staphylococcus (p = 0.007) compared to controls (n=83). Mode of delivery (R(2) = 0.04, p = 0.001), antibiotics duration greater than or equal to seven days (R(2) = 0.03, p = 0.003), age at sample collection (R(2) = 0.03, p = 0.009), and gestational age (R(2) = 0.02, p = 0.035) explained a small portion of microbiome variation. In gastroschisis infants, Escherichia-Shigella was the predominate genus, and those delivered via cesarean section had different microbial communities, predominantly Staphylococcus and Streptococcus, from those delivered vaginally. While antibiotic duration contributed to the variation in microbiome composition, there were no significant differences in taxa distribution or alpha diversity by antibiotic duration or nutrition type. CONCLUSION: The microbiome of gastroschisis infants is dysbiotic, and mode of birth delivery, antibiotic duration, and gestational age appear to contribute to microbial variation.

Combinations of antibiotics have shown promise in off-label treatment of Inflammatory Bowel Disease (IBD) flares in adult and pediatric patients1–6. In a small pediatric cohort of moderate-severe medically refractory Ulcerative Colitis (UC), half achieved remission with a regimen of 3–4 antibiotics6. The rationale is to decrease mucosal inflammation by altering intestinal microbiota, however, to date no study has documented these changes. We report our experience with combination oral antibiotics in symptomatic pediatric patients with steroid-dependent or -resistant UC, and demonstrate significant alterations in gut microbiota during and after antibiotic therapy with reductions in inflammatory markers.

Background Oral daily tenofovir (TFV) disoproxil fumarate/emtricitabine (TDF/FTC) for human immunodeficiency virus (HIV) pre-exposure prophylaxis (PrEP) is highly effective for HIVprevention, yet long-term effects are not fully understood. We investigated the effects of PrEP on the rectal microbiome in a cohort of men who have sex with men (MSM). Methods This cross-sectional analysis included HIV-negative MSM either on PrEP (n = 37) or not (n = 37) selected from an ongoing cohort using propensity score matching. Rectal swabs were used to examine microbiome composition using 16S ribosomal ribonucleic acid gene sequencing, and associations between PrEP use and microbiota abundance were examined. Hair specimens were used to quantify TFV and FTC exposure over the past 6 weeks on a subset of participants (n = 15). Results Pre-exposure prophylaxis use was associated with a significant increase in Streptococcus abundance (adjusted P = .015). Similar associations were identified using least absolute shrinkage and selection operator (LASSO) regression, confirming the increase in Streptococcus and also showing increased Mitsuokella, Fusobacterium, and decreased Escherichia/Shigella. Increased Fusobacterium was significantly associated with increasing TFV exposure. Conclusions Oral TDF/FTC for PrEP is associated with rectal microbiome changes compared to well matched controls, specifically increased Streptococcus and Fusobacterium abundance. This study highlights the need for future investigations of the role of microbiome changes on HIV susceptibility and effectiveness of PrEP.

Methamphetamine (MA) use is a major public health problem in the United States, especially among people living with HIV (PLWH). Many MA-induced neurotoxic effects are mediated by inflammation and gut microbiota may play a role in this process, yet the effects of MA on the microbiome have not been adequately explored. Therefore, we performed 16S rRNA gene sequencing on rectal swab samples from 381 men who have sex with men, 48% of whom were PLWH and 41% of whom used MA. We compared microbiome composition between MA users and non-users while testing for potential interactions with HIV and controlling for numerous confounders using inverse probability of treatment weighting. We found that MA use explained significant variation in overall composition (R2 = 0.005, p = 0.008) and was associated with elevated Finegoldia, Parvimonas, Peptoniphilus, and Porphyromonas and reduced Butyricicoccus and Faecalibacterium, among others. Genera including Actinomyces and Streptobacillus interacted with HIV status, such that they were increased in HIV+ MA users. Finegoldia and Peptoniphilus increased with increasing frequency of MA use, among others. In summary, MA use was associated with a microbial imbalance favoring pro-inflammatory bacteria, including some with neuroactive potential and others that have previously been associated with poor HIV outcomes.

BackgroundHuman immunodeficiency virus type 1 (HIV-1) infection alters the human intestinal microbiome; however, behavioral factors driving these changes remain poorly defined. Here we examine the effects of substance use and sex behavior on the microbiome during HIV-1 infection.MethodsArchival rectal swab specimens, urine drug test results, and responses to substance use and sex behavior questionnaires were obtained from 37 HIV-positive participants at 2 time points, separated by 6 months, in a cohort examining the effects of substance use in men who have sex with men (MSM). Microbiome profiling was performed using 16S ribosomal RNA gene sequencing, and associations with behavioral factors were examined using 0-inflated negative binomial regression. Further analysis of selected variables of interest was performed using propensity scores to account for multiple confounders.ResultsUsing permutational multivariate analysis of variance, we found that receptive anal intercourse, methamphetamine use, and marijuana use were among the most important drivers of microbiome variation. Propensity score–adjusted analyses revealed that methamphetamine use and marijuana use displayed unique associations; methamphetamine use was associated with an increased abundance of Porphyromonas and Granulicatella organisms and a decreased abundance of Ruminococcus, Collinsella, and Parabacteroides organisms, whereas marijuana use was associated with an increased abundance of Ruminococcus, Clostridium cluster IV, Solobacterium, and Fusobacterium organisms and a decreased abundance of Acidaminococcus, Prevotella, Dialister, Anaerostipes, and Dorea organisms.ConclusionsDrug use and sex behavior are important factors associated with intestinal dysbiosis during chronic HIV-1 infection among young MSM.

Studies of microbial communities have become widespread with the development of relatively inexpensive, rapid, and high throughput sequencing. However, as with all these technologies, reproducible results depend on a laboratory workflow that incorporates appropriate precautions and controls. This is particularly important with low-biomass samples where contaminating bacterial DNA can generate misleading results. This article details a semi-automated workflow to identify microbes from human breast milk samples using targeted sequencing of the 16S ribosomal RNA (rRNA) V4 region on a low- to mid-throughput scale. The protocol describes sample preparation from whole milk including: sample lysis, nucleic acid extraction, amplification of the V4 region of the 16S rRNA gene, and library preparation with quality control measures. Importantly, the protocol and discussion consider issues that are salient to the preparation and analysis of low-biomass samples including appropriate positive and negative controls, PCR inhibitor removal, sample contamination by environmental, reagent, or experimental sources, and experimental best practices designed to ensure reproducibility. While the protocol as described is specific to human milk samples, it is adaptable to numerous low- and high-biomass sample types, including samples collected on swabs, frozen neat, or stabilized in a preservation buffer.

Metabolic dysregulation associated with gut microbial dysbiosis can influence the progression of nonalcoholic fatty liver disease (NAFLD) from hepatic steatosis to inflammation and subsequent fibrosis. Using a multi-omics approach, we identified microbiome-derived biochemicals and genes associated with progressive hepatic pathology and fibrosis in Latino children (N=241). Children with more severe liver pathology were dysbiotic and had increased gene content associated with lipopolysaccharide biosynthesis and lipid, amino acid and carbohydrate metabolism. These changes were driven by increases in Bacteroides and concomitant decreases of Akkermansia, Anaerococcus, Corynebacterium, and Finegoldia. Nontargeted mass spectrometry revealed perturbations in one-carbon metabolism, mitochondrial dysfunction, and increased oxidative stress in children with steatohepatitis and fibrosis. Random forests modeling of plasma metabolites were highly predictive of nonalcoholic steatohepatitis (NASH) (97% accuracy) and hepatic fibrosis, steatosis and lobular inflammation (93.8% accuracy), and can differentiate steatohepatitis from simple steatosis (90.0% accuracy). These metabolomic biomarkers of hepatic pathology provide insights for therapeutic targeting of bacterially-mediated and host pathways.

PURPOSE OF REVIEW: Emerging research on the pediatric microbiome implicates the importance of the microbiome on the development of the immune system, nervous system, and growth. Changes to the microbiome during infancy are associated with the development of chronic illnesses such as asthma and inflammatory bowel disease. Additionally, the microbiome provides protection against certain pathogens, affects vaccine responses and alters drug metabolism. This review highlights what is known about the microbiome, the establishment of a healthy microbiome and the significance that changes to the microbiome composition have on growth and health of children and adolescents. RECENT FINDINGS: Vaginal delivery, breastfeeding, maternal health and nutrition help shape a healthy microbiome. Caesarian delivery, formula feeding and antibiotic use perturb the microbiome and are associated with the development of type II diabetes, asthma, allergic diseases and obesity later in life. Specific interventions using pre- and probiotics in multiple settings are under investigation with limited success. SUMMARY: A better understanding of the microbiome and the interaction with the immune system may help guide interventions to alter the microbiome towards a state of lifelong health.

Increasing evidence supports the importance of the breast milk microbiome in seeding the infant gut. However, the origin of bacteria in milk and the process of milk microbe-mediated seeding of infant intestine need further elucidation. Presumed sources of bacteria in milk include locations of mother-infant and mother-environment interactions. We investigate the role of mother-infant interaction on breast milk microbes. Shotgun metagenomics and 16S rRNA gene sequencing identified milk microbes of mother-infant pairs in breastfed infants and in infants that have never latched. Although breast milk has low overall biomass, milk microbes play an important role in seeding the infant gut. Breast milk bacteria were largely comprised of Staphylococcus, Streptococcus, Acinetobacter, and Enterobacter primarily derived from maternal areolar skin and infant oral sites in breastfeeding pairs. This suggests that the process of breastfeeding is a potentially important mechanism for propagation of breast milk microbes through retrograde flux via infant oral and areolar skin contact. In one infant delivered via Caesarian section, a distinct strain of Bifidobacteria breve was identified in maternal rectum, breast milk and the infant’s stool potentially suggesting direct transmission. This may support the existence of microbial translocation of this anaerobic bacteria via the enteromammary pathway in humans, where maternal bacteria translocate across the maternal gut and are transferred to the mammary glands. Modulating sources of human milk microbiome seeding potentially imply opportunities to ultimately influence the development of the infant microbiome and health.

Summary Most infants born to human immunodeficiency virus (HIV)-infected women escape HIV infection. Infants evade infection despite an immature immune system and, in the case of breastfeeding, prolonged repetitive exposure. If infants become infected, the course of their infection and response to treatment differs dramatically depending upon the timing (in utero, intrapartum, or during breastfeeding) and potentially the route of their infection. Perinatally acquired HIV infection occurs during a critical window of immune development. HIV's perturbation of this dynamic process may account for the striking age-dependent differences in HIV disease progression. HIV infection also profoundly disrupts the maternal immune system upon which infants rely for protection and immune instruction. Therefore, it is not surprising that infants who escape HIV infection still suffer adverse effects. In this review, we highlight the unique aspects of pediatric HIV transmission and pathogenesis with a focus on mechanisms by which HIV infection during immune ontogeny may allow discovery of key elements for protection and control from HIV.

Background Mother-to-child transmission of human immunodeficiency virus-type 1 (HIV-1) poses a serious health threat in developing countries, and adequate interventions are as yet unrealized. HIV-1 infection is frequently initiated by a single founder viral variant, but the factors that influence particular variant selection are poorly understood. Results Our analysis of 647 full-length HIV-1 subtype C and G viral envelope sequences from 22 mother–infant pairs reveals unique genotypic and phenotypic signatures that depend upon transmission route. Relative to maternal strains, intrauterine HIV transmission selects infant variants that have shorter, less-glycosylated V1 loops that are more resistant to soluble CD4 (sCD4) neutralization. Transmission through breastfeeding selects for variants with fewer potential glycosylation sites in gp41, are more sensitive to the broadly neutralizing antibodies PG9 and PG16, and that bind sCD4 with reduced cooperativity. Furthermore, experiments with Affinofile cells indicate that infant viruses, regardless of transmission route, require increased levels of surface CD4 receptor for productive infection. Conclusions These data provide the first evidence for transmission route-specific selection of HIV-1 variants, potentially informing therapeutic strategies and vaccine designs that can be tailored to specific modes of vertical HIV transmission. Electronic supplementary material The online version of this article (doi:10.1186/s12977-017-0331-z) contains supplementary material, which is available to authorized users.

More than 1 million HIV-exposed, uninfected infants are born annually to HIV-positive mothers worldwide. This growing population of infants experiences twice the mortality of HIV-unexposed infants. We found that although there were very few differences seen in the microbiomes of mothers with and without HIV infection, maternal HIV infection was associated with changes in the microbiome of HIV-exposed, uninfected infants. Furthermore, we observed that human breast milk oligosaccharides were associated with bacterial species in the infant microbiome. The disruption of the infant's microbiome associated with maternal HIV infection may contribute to the increased morbidity and mortality of HIV-exposed, uninfected infants.

Accumulating evidence suggests that rates of low bone mass are greater in HIV-infected males than females. Of eleven biomarkers assessed by sex and HIV-status, HIV-infected males had increased levels of soluble CD14 which inversely correlated with bone mineral content and bone mineral density measures, suggesting macrophage activation as a possible mechanism of differential bone loss.

Objective: Low-level HIV-1 replication may occur during antiretroviral therapy (ART) that suppresses plasma HIV-1 RNA to less than 50 copies/ml (suppressive ART). Antiretroviral drugs appear less effective in macrophages and monocytes compared with lymphocytes, both in vitro and as implied in vivo by greater viral evolution observed during suppressive ART. Our objective was to examine sputum, which is rich in macrophages, for evidence of increased HIV-1 replication compared with that in the blood during suppressive ART. Design: A cross-sectional study during suppressive ART was performed, and HIV-1 DNA sequences derived from induced sputa and peripheral blood mononuclear cells were compared. Methods: Multiple sequences encoding HIV-1 reverse transcriptase, protease, and envelope were generated using single-genome sequencing. Reverse transcriptase and protease sequences were analyzed for genotypic drug resistance. The evolutionary distances of env sequences from the inferred most recent common ancestor of infection were calculated, and CXCR4 usage was predicted. Results: Nine hundred seventy bidirectional sequences from 11 individuals were analyzed. HIV-1 env and pol derived from sputa had greater frequency of drug-resistance mutations (P = 0.05), evolutionary divergence (P = 0.004), and tendency for CXCR4 usage (P = 0.1) compared with viruses derived from peripheral blood mononuclear cells. Conclusion: The greater frequency of HIV-1 drug-resistance mutations and divergence of HIV-1 env in sputa-derived viruses compared with peripheral blood mononuclear cell-derived viruses suggests greater HIV-1 replication in the respiratory tract compared with the blood. Characterization of viral evolution over time and by cell-type could identify cells that provide a sanctuary for low-level viral replication in the respiratory tract during suppressive ART.