Metabolism and development must be closely coupled to meet the changing physiological needs of each stage in the life cycle. The molecular mechanisms that link these pathways, however, remain poorly understood. Here we show that the Drosophila estrogen-related receptor (dERR) directs a transcriptional switch in mid-pupae that promotes glucose oxidation and lipogenesis in young adults. dERR mutant adults are viable but display reduced locomotor activity, susceptibility to starvation, elevated glucose, and an almost complete lack of stored triglycerides. Molecular profiling by RNA-seq, ChIP-seq, and metabolomics revealed that glycolytic and pentose phosphate pathway genes are induced by dERR, and their reduced expression in mutants is accompanied by elevated glycolytic intermediates, reduced TCA cycle intermediates, and reduced levels of long chain fatty acids. Unexpectedly, we found that the central pathways of energy metabolism, including glycolysis, the tricarboxylic acid cycle, and electron transport chain, are coordinately induced at the transcriptional level in mid-pupae and maintained into adulthood, and this response is partially dependent on dERR, leading to the metabolic defects observed in mutants. Our data support the model that dERR contributes to a transcriptional switch during pupal development that establishes the metabolic state of the adult fly.

The proteostasis network is regulated by transcellular communication to promote health and fitness in metazoans. In Caenorhabditis elegans, signals from the germline initiate the decline of proteostasis and repression of cell stress responses at reproductive maturity, indicating that commitment to reproduction is detrimental to somatic health. Here we show that proteostasis and stress resilience are also regulated by embryo-to-mother communication in reproductive adults. To identify genes that act directly in the reproductive system to regulate somatic proteostasis, we performed a tissue targeted genetic screen for germline modifiers of polyglutamine aggregation in muscle cells. We found that inhibiting the formation of the extracellular vitelline layer of the fertilized embryo inside the uterus suppresses aggregation, improves stress resilience in an HSF-1-dependent manner, and restores the heat-shock response in the somatic tissues of the parent. This pathway relies on DAF-16/FOXO activation in vulval tissues to maintain stress resilience in the mother, suggesting that the integrity of the embryo is monitored by the vulva to detect damage and initiate an organismal protective response. Our findings reveal a previously undescribed transcellular pathway that links the integrity of the developing progeny to proteostasis regulation in the parent.

Cell type-specific transcriptional programs that drive differentiation of specialized cell types are key players in development and tissue regeneration. One of the most dramatic changes in the transcription program in Drosophila occurs with the transition from proliferating spermatogonia to differentiating spermatocytes, with >3000 genes either newly expressed or expressed from new alternative promoters in spermatocytes. Here we show that opening of these promoters from their closed state in precursor cells requires function of the spermatocyte-specific tMAC complex, localized at the promoters. The spermatocyte-specific promoters lack the previously identified canonical core promoter elements except for the Inr. Instead, these promoters are enriched for the binding site for the TALE-class homeodomain transcription factors Achi/Vis and for a motif originally identified under tMAC ChIP-seq peaks. The tMAC motif resembles part of the previously identified 14-bp β2UE1 element critical for spermatocyte-specific expression. Analysis of downstream sequences relative to transcription start site usage suggested that ACA and CNAAATT motifs at specific positions can help promote efficient transcription initiation. Our results reveal how promoter-proximal sequence elements that recruit and are acted upon by cell type-specific chromatin binding complexes help establish a robust, cell type-specific transcription program for terminal differentiation.

The emergence of drug resistance is a major obstacle for the success of targeted therapy in melanoma. Additionally, conventional chemotherapy has not been effective as drug-resistant cells escape lethal DNA damage effects by inducing growth arrest commonly referred to as cellular dormancy. We present a therapeutic strategy termed "targeted chemotherapy" by depleting protein phosphatase 2A (PP2A) or its inhibition using a small molecule inhibitor (1,10-phenanthroline-5,6-dione [phendione]) in drug-resistant melanoma. Targeted chemotherapy induces the DNA damage response without causing DNA breaks or allowing cellular dormancy. Phendione treatment reduces tumor growth of BRAF^V600E-driven melanoma patient-derived xenografts (PDX) and diminishes growth of NRAS^Q61R-driven melanoma, a cancer with no effective therapy. Remarkably, phendione treatment inhibits the acquisition of resistance to BRAF inhibition in BRAF^V600E PDX highlighting its effectiveness in combating the advent of drug resistance.

Peripheral somatosensory input is modulated in the dorsal spinal cord by a network of excitatory and inhibitory interneurons. PTF1A is a transcription factor essential in dorsal neural tube progenitors for specification of these inhibitory neurons. Thus, mechanisms regulating Ptf1a expression are key for generating neuronal circuits underlying somatosensory behaviors. Mutations targeted to distinct cis-regulatory elements for Ptf1a in mice, tested the in vivo contribution of each element individually and in combination. Mutations in an autoregulatory enhancer resulted in reduced levels of PTF1A, and reduced numbers of specific dorsal spinal cord inhibitory neurons, particularly those expressing Pdyn and Gal. Although these mutants survive postnatally, at ~3–5 wk they elicit a severe scratching phenotype. Behaviorally, the mutants have increased sensitivity to itch, but acute sensitivity to other sensory stimuli such as mechanical or thermal pain is unaffected. We demonstrate a requirement for positive transcriptional autoregulatory feedback to attain the level of the neuronal specification factor PTF1A necessary for generating correctly balanced neuronal circuits.

Background and objective

Exercise is a cornerstone of management for type 2 diabetes; however, little is known about the cardiovascular (CV) response to submaximal functional exercise in people with type 2 diabetes. The aim of this study was to compare performance and CV response during a 6-minute walk test (6MWT) between people with type 2 diabetes and matched control subjects.

A multidisciplinary endocrinologist-led shared medical appointment (SMA) model showed statistically significant reductions in A1C from baseline over 3 years that were not significantly different from appointments with endocrinologists or primary care providers alone within a resource-poor population. Similarly, the SMA model achieved clinical outcomes on par with endocrinologist-only visits with the added benefit of improving endocrine provider productivity and specialty access for patients. Greater patient engagement with the SMA model was associated with significantly lower A1C.

Background

Advances in information communications technology (ICT) provide opportunities for enhanced diabetes care. Knowledge of the more acceptable communication modalities in patients of different ages will help to inform the direction of future innovations.

Chronic cough is a common complaint in the general population but there are no precise data on the incidence of, and prospectively examined risk factors for chronic cough in a population-based setting. Therefore, we investigated the period prevalence, incidence and risk factors for chronic cough in adult subjects.

In a prospective population-based cohort study among subjects aged ≥45 years, data on chronic cough were collected on two separate occasions using a standardised questionnaire. Chronic cough was defined as daily coughing for at least 3 months duration during the preceding 2 years. Potential risk factors were gathered by interview, physical examination and several investigations.

Of the 9824 participants in this study, 1073 (10.9%) subjects had chronic cough at baseline. The prevalence of chronic cough increased with age and peaked in the eighth decade. In subjects aged <70 years, chronic cough was more common in women. During an average follow-up of 6 years, 439 incident cases of chronic cough occurred with an overall incidence rate of 11.6 per 1000 person-years (95% CI 10.6–12.8). In current smokers, the incidence of chronic cough was higher in men. In the multivariable analysis, current smoking, gastro-oesophageal reflux disease (GORD), asthma and COPD were identified as risk factors for chronic cough.

Chronic cough is common among adults and highly prevalent in the older population. Current smoking, GORD, asthma and COPD are independent risk factors for chronic cough. Individuals at risk of developing chronic cough may benefit from smoking cessation and control of the underlying disease.

Background

COPD patients account for a large proportion of lung transplants; lung transplantation survival benefit for COPD patients is not well established.

Introduction

Immunotherapy has become the standard of care in advanced non-small cell lung cancer (NSCLC). We aimed to quantify the economic impact, in France, of anti-PD-1 therapy for NSCLC.

Background

Positive serology for cytomegalovirus (CMV) has been associated with all-cause mortality risk but its role in COPD mortality is unknown. The objective of the present study was to assess the relationship between CMV serology and COPD mortality.

ABSTRACT

The prevailing paradigm in obstetrics has been the sterile womb hypothesis. However, some are asserting that the placenta, intra-amniotic environment, and fetus harbor microbial communities. The objective of this study was to determine whether the fetal and placental tissues of rhesus macaques harbor bacterial communities. Fetal, placental, and uterine wall samples were obtained from cesarean deliveries without labor (~130/166 days gestation). The presence of bacteria in the fetal intestine and placenta was investigated through culture. The bacterial burden and profiles of the placenta, umbilical cord, and fetal brain, heart, liver, and colon were determined through quantitative real-time PCR and DNA sequencing. These data were compared with those of the uterine wall as well as to negative and positive technical controls. Bacterial cultures of fetal and placental tissues yielded only a single colony of Cutibacterium acnes. This bacterium was detected at a low relative abundance (0.02%) in the 16S rRNA gene profile of the villous tree sample from which it was cultured, yet it was also identified in 12/29 background technical controls. The bacterial burden and profiles of fetal and placental tissues did not exceed or differ from those of background technical controls. By contrast, the bacterial burden and profiles of positive controls exceeded and differed from those of background controls. Among the macaque samples, distinct microbial signals were limited to the uterine wall. Therefore, using multiple modes of microbiologic inquiry, there was not consistent evidence of bacterial communities in the fetal and placental tissues of rhesus macaques.

IMPORTANCE Microbial invasion of the amniotic cavity (i.e., intra-amniotic infection) has been causally linked to pregnancy complications, especially preterm birth. Therefore, if the placenta and the fetus are typically populated by low-biomass microbial communities, current understanding of the role of microbes in reproduction and pregnancy outcomes will need to be fundamentally reconsidered. Could these communities be of benefit by competitively excluding potential pathogens or priming the fetal immune system for the microbial bombardment it will experience upon delivery? If so, what properties (e.g., microbial load and community membership) of these microbial communities preclude versus promote intra-amniotic infection? Given the ramifications of the in utero colonization hypothesis, critical evaluation is required. In this study, using multiple modes of microbiologic inquiry (i.e., culture, quantitative real-time PCR [qPCR], and DNA sequencing) and controlling for potential background DNA contamination, we did not find consistent evidence for microbial communities in the placental and fetal tissues of rhesus macaques.

ABSTRACT

Aspergillus flavus, a ubiquitous and saprophytic fungus, is the second most common cause of aspergillosis worldwide. Several mechanisms contribute to the establishment of the fungal infection. Extracellular vesicles (EVs) have been described as "virulence factor delivery bags" in several fungal species, demonstrating a crucial role during the infection. In this study, we evaluated production of A. flavus EVs and their immunomodulatory functions. We verified that A. flavus EVs induce macrophages to produce inflammatory mediators, such as nitric oxide, tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and IL-1β. Furthermore, the A. flavus EVs enhance phagocytosis and killing by macrophages and induce M1 macrophage polarization in vitro. In addition, a prior inoculation of A. flavus EVs in Galleria mellonella larvae resulted in a protective effect against the fungal infection. Our findings suggest that A. flavus EVs are biologically active and affect the interaction between A. flavus and host immune cells, priming the innate immune system to eliminate the fungal infection. Collectively, our results suggest that A. flavus EVs play a crucial role in aspergillosis.

IMPORTANCE Immunocompromised patients are susceptible to several fungal infections. The genus Aspergillus can cause increased morbidity and mortality. Developing new therapies is essential to understand the fungal biology mechanisms. Fungal EVs carry important virulence factors, thus playing pivotal roles in fungal pathophysiology. No study to date has reported EV production by Aspergillus flavus, a fungus considered to be the second most common cause of aspergillosis and relevant food contaminator found worldwide. In this study, we produced A. flavus EVs and evaluated the in vitro immunomodulatory effects of EVs on bone marrow-derived macrophages (BMDMs) and in vivo effects in a Galleria mellonella model.

Molar element ratio analysis of element concentrations consists of four basic tools that provide substantial insight into the lithogeochemistry (and mineralogy) of rocks under examination. These tools consist of: (1) conserved element ratio analysis; (2) Pearce element ratio analysis; (3) general element ratio analysis; and (4) lithogeochemical mineral mode analysis. Conserved element ratio analysis is useful in creating a chemostratigraphic model for the host rocks to mineral deposits, whereas Pearce element ratio analysis and general element ratio analysis are primarily used to identify mineralogical and metasomatic controls on rock compositions and to investigate and quantify the extent of the material transfers that formed the host rocks and mineralization. Lithogeochemical mineral mode analysis converts element concentrations into mineral concentrations using a matrix-based change-of-basis operation, allowing lithogeochemical data to be interpreted in terms of mineral modes. It can be used to provide proper names to rocks, an important activity for an exploration geologist because of the implications that rock names have on genetic processes and mineral deposit models.

This paper provides a review of the theoretical foundations of each of these four tools and then illustrates how these techniques have been used in a variety of exploration applications to assist in the search for, evaluation and planning of, and the mining of mineral deposits. Examples include the evaluation of total digestion lithogeochemical datasets from mineral deposits hosted by igneous and sedimentary rocks and formed by hydrothermal and igneous processes. In addition, this paper illustrates a more recent geometallurgical application of these methods, whereby the mineral proportions determined by lithogeochemical mineral mode analysis are used to predict rock properties and obtain the ore body knowledge critical for resource evaluation, mine planning, mining and mine remediation.

Thematic collection: This article is part of the Exploration 17 collection available at: https://www.lyellcollection.org/cc/exploration-17

This paper focuses on handheld and top-of-hole techniques which have appeared since 2007 or have undergone major improvements, and discusses their benefits, challenges and pitfalls, why we use them and what to expect from them. There is an ongoing need to be innovative with the way we undertake mineral exploration. Recent technological advances that have been applied to successful mineral exploration include on-site or portable instruments, on-site laboratory technologies, various core scanners, and technologies for fluid analysis. Portable or field technologies such as pXRF, pXRD, pNIR-SWIR, µRaman and LIBS aid in obtaining chemical and mineralogical information. Spectral gamma tools, a well-known technology, recently took advantage of improved ground and airborne (drone) instruments, to complement hyperspectral imagery. At mine and exploration sites, top-of-hole sensing technologies, such as Lab-at-Rig® and various core scanners (both spectral- and XRF-based) have become useful tools to analyse metres of core as it is being drilled. Fluid analyses are not as common as analyses of solid materials, but there are advances in such technologies as anodic stripping voltammetry, polarography and ion-exchange electrodes aiming for analysis of commodity or environmentally important elements.

Field-portable geochemical techniques and on-site technologies now offer instant response and flexibility for most exploration tasks. By providing relevant data within minutes, they allow safer field decisions and focus on the most promising finds, while saving valuable resources in sampling grids or drilling. More efficient laboratory analysis programs are supported by sample screening and homogeneity checking on-site. Field analyses are not always as accurate as laboratory ones, but most of the time can be correlated with them, enabling reliable decisions. The level of confidence in field-made decisions needs to be compared between later and less numerous laboratory analyses, and less precise but more abundant and immediate field analyses. It may be demonstrated that, in many cases, the fit–for-purpose nature of the latter allows a better confidence level. Quality compromises associated with field analyses can be reduced by the application of better sample preparation and quality assurance/quality control (QA/QC) procedures. Most of the further development of on-site chemical analysis is expected to be based on its integration with lab methods and on sound QA/QC practice, allowing a precise evaluation of its confidence level and uncertainties. Mineralogical analyses are constrained by our ability to interpret the data in near-real time but offer promising approaches in both surface and drilling exploration campaigns.

Thematic collection: This article is part of the Exploration 17 collection available at: https://www.lyellcollection.org/cc/exploration-17

After vascular injury and exposure of subendothelial matrix proteins to the intravascular space, mediators of hemostasis are triggered and allow for clot formation and restoration of vascular integrity. Platelets are the mediators of primary hemostasis, creating a platelet plug and allowing for initial cessation of bleeding. Platelet disorders, qualitative and quantitative, may result in bleeding signs and symptoms, particularly mucocutaneous bleeding such as epistaxis, bruising, petechiae, and heavy menstrual bleeding. Increasing evidence suggests that platelets have functional capabilities beyond hemostasis, but this review focuses solely on platelet hemostatic properties. Herein, normal platelet function as well as the effects of abnormal function and thrombocytopenia are reviewed.

Extractables and leachables (E&L) are identified and quantified so that their impact on patient safety can be established and assessed. The uncertainty in the impact assessment is affected by the uncertainty in the substance's experimentally determined identity and concentration. Thus, these experimentally determined quantities must be reported not only in terms of their absolute result but also in terms of the uncertainty in the result, which is based on the amount and rigor of the information on which the result is based. In this way, the impact assessment can be tempered to account for the uncertainty in its input data. To facilitate the assignment and reporting of uncertainty, classification hierarchies are proposed and discussed for both identification and quantitation. Both hierarchies establish levels or degrees of identification and quantitation based on the uncertainty of the result and contain descriptions of the quality and quantity of information required to achieve a certain level within the hierarchy. The minimal levels that must be achieved to support impact assessment are also established.

The bioprocessing industry uses recombinant mammalian cell lines to generate therapeutic biologic drugs. To ensure consistent product quality of the therapeutic proteins, it is imperative to have a controlled production process. Regulatory agencies and the biotechnology industry consider cell line "clonal origin" an important aspect of maintaining process control. Demonstration of clonal origin of the cell substrate, or production cell line, has received considerable attention in the past few years, and the industry has improved methods and devised standards to increase the probability and/or assurance of clonal derivation. However, older production cell lines developed before the implementation of these methods, herein referred to as "legacy cell lines," may not meet current regulatory expectations for demonstration of clonal derivation. In this article, the members of the IQ Consortium Working Group on Clonality present our position that the demonstration of process consistency and product comparability of critical quality attributes throughout the development life cycle should be sufficient to approve a license application without additional genetic analysis to support clonal origin, even for legacy cell lines that may not meet current day clonal derivation standards. With this commentary, we discuss advantages and limitations of genetic testing methods to support clonal derivation of legacy cell lines and wish to promote a mutual understanding with the regulatory authorities regarding their optional use during early drug development, subsequent to Investigational New Drug (IND) application and before demonstration of product and process consistency at Biologics License Applications (BLA) submission.

Capture bioprocessing unit operations were previously shown to clear or kill several log₁₀ of a model mycoplasma Acholeplasma laidlawii in lab-scale spike/removal studies. Here, we confirm this observation with two additional mollicute species relevant to biotechnology products for human use: Mycoplasma orale and Mycoplasma arginini. Clearance of M. orale and M. arginini from protein A column purification was similar to that seen with A. laidlawii, though some between cycle carryover was evident, especially for M. orale. However, on-resin growth studies for all three species revealed that residual mycoplasma in a column slowly die off over time rather than expanding further. Solvent/detergent exposure completely inactivated M. arginini though detectable levels of M. orale remained. A small-scale model of a commercial low-pH hold step did inactivate live M. orale, but this inactivation required a lower pH set point and occurred with slower kinetics than previously seen with A. laidlawii. Additionally, ultraviolet-C irradiation was shown to be effective for A. laidlawii and M. orale inactivation whereas virus-retentive filters for upstream and downstream processes, as expected, cleared A. laidlawii. These data argue that M. orale and M. arginini overall would be largely cleared by early bioprocessing steps as shown previously for A. laidlawii, and that barrier technologies can effectively reduce the risk from media components. For some unit operations, M. orale and M. arginini may be hardier, and require more stringent processing or equipment cleaning conditions to assure effective mycoplasma reduction. By exploring how some of the failure modes in commercial antibody manufacturing processes can still eliminate mycoplasma burden, we demonstrate that required best practices assure biotechnology products will be safe for patients.

Glass is the favorite material for parenteral packaging because of its physico-chemical properties. Type I borosilicate glass is worldwide use at this scope, but it may have some issues related to breakage, corrosion and delamination that might compromise the drug quality, safety and efficacy. These issues can be mitigated and avoided starting from the appropriate selection of the most suitable raw material at the early stage of the glass container design. In this study, Type I borosilicate glass vials manufactured using two glass tubes having different chemical compositions, were studied and compared in terms of their resistance to corrosion. Testing design was applied with the aim to select the best practice approach comparing different storage simulation conditions: ageing treatment through autoclaving and stability testing (real-time and accelerated). Clear differences were found between the different glass types in terms of hydrolytic and corrosion resistance that highlighted the relation between chemical composition and glass chemical durability. Non-negligible differences were also observed using different storage conditions.

Pyrazinamide (PZA) is a cornerstone antimicrobial drug used exclusively for the treatment of tuberculosis (TB). Due to its ability to shorten drug therapy by 3 months and reduce disease relapse rates, PZA is considered an irreplaceable component of standard first-line short-course therapy for drug-susceptible TB and second-line treatment regimens for multidrug-resistant TB. Despite over 60 years of research on PZA and its crucial role in current and future TB treatment regimens, the mode of action of this unique drug remains unclear. Defining the mode of action for PZA will open new avenues for rational design of novel therapeutic approaches for the treatment of TB. In this review, we discuss the four prevailing models for PZA action, recent developments in modulation of PZA susceptibility and resistance, and outlooks for future research and drug development.

Technologies allowing genetic sequencing of the human microbiome are opening new realms to discovery. The host microbiota substantially impacts immune responses both in immune-mediated inflammatory diseases (IMIDs) and in tumors affecting tissues beyond skin and mucosae. However, a mechanistic link between host microbiota and cancer or IMIDs has not been well established. Here, we propose T helper 17 (T_H17) lymphocytes as the connecting factor between host microbiota and rheumatoid or psoriatic arthritides, multiple sclerosis, breast or ovarian cancer, and multiple myeloma. We theorize that similar mechanisms favor the expansion of gut-borne T_H17 cells and their deployment at the site of inflammation in extraborder IMIDs and tumors, where T_H17 cells are driving forces. Thus, from a pathogenic standpoint, tumors may share mechanistic routes with IMIDs. A review of similarities and divergences in microbiota-T_H17 cell interactions in IMIDs and cancer sheds light on previously ignored pathways in either one of the two groups of pathologies and identifies novel therapeutic avenues.

While flagella have been studied extensively as motility organelles, with a focus on internal structures such as the axoneme, more recent research has illuminated the roles of the flagellar surface in a variety of biological processes. Parasitic protists of the order Kinetoplastida, which include trypanosomes and Leishmania species, provide a paradigm for probing the role of flagella in host-microbe interactions and illustrate that this interface between the flagellar surface and the host is of paramount importance. An increasing body of knowledge indicates that the flagellar membrane serves a multitude of functions at this interface: attachment of parasites to tissues within insect vectors, close interactions with intracellular organelles of vertebrate cells, transactions between flagella from different parasites, junctions between the flagella and the parasite cell body, emergence of nanotubes and exosomes from the parasite directed to either host or microbial targets, immune evasion, and sensing of the extracellular milieu. Recent whole-organelle or genome-wide studies have begun to identify protein components of the flagellar surface that must mediate these diverse host-parasite interactions. The increasing corpus of knowledge on kinetoplastid flagella will likely prove illuminating for other flagellated or ciliated pathogens as well.

Acetylation is a conserved modification used to regulate a variety of cellular pathways, such as gene expression, protein synthesis, detoxification, and virulence. Acetyltransferase enzymes transfer an acetyl moiety, usually from acetyl coenzyme A (AcCoA), onto a target substrate, thereby modulating activity or stability. Members of the GCN5-N-acetyltransferase (GNAT) protein superfamily are found in all domains of life and are characterized by a core structural domain architecture. These enzymes can modify primary amines of small molecules or of lysyl residues of proteins. From the initial discovery of antibiotic acetylation, GNATs have been shown to modify a myriad of small-molecule substrates, including tRNAs, polyamines, cell wall components, and other toxins. This review focuses on the literature on small-molecule substrates of GNATs in bacteria, including structural examples, to understand ligand binding and catalysis. Understanding the plethora and versatility of substrates helps frame the role of acetylation within the larger context of bacterial cellular physiology.

The substantial discrepancy between the strong effects of functional foods and various drugs, especially traditional Chinese medicines (TCMs), and the poor bioavailability of these substances remains a perplexing problem. Understanding the gut microbiota, which acts as an effective bioreactor in the human intestinal tract, provides an opportunity for the redefinition of bioavailability. Here, we discuss four different pathways associated with the role of the gut microbiota in the transformation of parent compounds to beneficial or detrimental small molecules, which can enter the body’s circulatory system and be available to target cells, tissues, and organs. We further describe and propose effective strategies for improving bioavailability and alleviating side effects with the help of the gut microbiota. This review also broadens our perspectives for the discovery of new medicinal components.

Escherichia coli ribosomal protein (r-protein) L4 has extraribosomal biological functions. Previously, we described L4 as inhibiting RNase E activity through protein-protein interactions. Here, we report that from stabilized transcripts regulated by L4-RNase E, mRNA levels of tnaA (encoding tryptophanase from the tnaCAB operon) increased upon ectopic L4 expression, whereas TnaA protein levels decreased. However, at nonpermissive temperatures (to inactivate RNase E), tnaA mRNA and protein levels both increased in an rne temperature-sensitive [rne(Ts)] mutant strain. Thus, L4 protein fine-tunes TnaA protein levels independently of its inhibition of RNase E. We demonstrate that ectopically expressed L4 binds with transcribed spacer RNA between tnaC and tnaA and downregulates TnaA translation. We found that deletion of the 5' or 3' half of the spacer compared to the wild type resulted in a similar reduction in TnaA translation in the presence of L4. In vitro binding of L4 to the tnaC-tnaA transcribed spacer RNA results in changes to its secondary structure. We reveal that during early stationary-phase bacterial growth, steady-state levels of tnaA mRNA increased but TnaA protein levels decreased. We further confirm that endogenous L4 binds to tnaC-tnaA transcribed spacer RNA in cells at early stationary phase. Our results reveal the novel function of L4 in fine-tuning TnaA protein levels during cell growth and demonstrate that r-protein L4 acts as a translation regulator outside the ribosome and its own operon.

IMPORTANCE Some ribosomal proteins have extraribosomal functions in addition to ribosome translation function. The extraribosomal functions of several r-proteins control operon expression by binding to own-operon transcripts. Previously, we discovered a posttranscriptional, RNase E-dependent regulatory role for r-protein L4 in the stabilization of stress-responsive transcripts. Here, we found an additional extraribosomal function for L4 in regulating the tna operon by L4-intergenic spacer mRNA interactions. L4 binds to the transcribed spacer RNA between tnaC and tnaA and alters the structural conformation of the spacer RNA, thereby reducing the translation of TnaA. Our study establishes a previously unknown L4-mediated mechanism for regulating gene expression, suggesting that bacterial cells have multiple strategies for controlling levels of tryptophanase in response to varied cell growth conditions.

In bacterial populations, quorum sensing (QS) systems participate in the regulation of specialization processes and regulate collective behaviors that mediate interactions and allow survival of the species. In Gram-positive bacteria, QS systems of the RRNPP family (Rgg, Rap, NprR, PlcR, and PrgX) consist of intracellular receptors and their cognate signaling peptides. Two of these receptors, Rap and NprR, have regained attention in Bacillus subtilis and the Bacillus cereus group. Some Rap proteins, such as RapH and Rap60, are multifunctional and/or redundant in function, linking the specialization processes of sporulation and competence, as well as global expression changes in the transition phase in B. subtilis. NprR, an evolutionary intermediate between Rap and RRNPP transcriptional activators, is a bifunctional regulator that modulates sporulation initiation and activates nutrient scavenging genes. In this review, we discuss how these receptors switch between functions and connect distinct signaling pathways. Based on structural evidence, we propose that RapH and Rap60 should be considered moonlighting proteins. Additionally, we analyze an evolutionary and ecological perspective to understand the multifunctionality and functional redundancy of these regulators in both Bacillus spp. and non-Bacillus Firmicutes. Understanding the mechanistic, structural, ecological, and evolutionary basis for the multifunctionality and redundancy of these QS systems is a key step for achieving the development of innovative technologies for health and agriculture.

Streptococcus pyogenes (Lancefield group A Streptococcus [GAS]) is a β-hemolytic human-selective pathogen that is responsible for a large number of morbid and mortal infections in humans. For efficient infection, GAS requires different types of surface proteins that provide various mechanisms for evading human innate immune responses, thus enhancing pathogenicity of the bacteria. Many such virulence-promoting proteins, including the major surface signature M protein, are translocated after biosynthesis through the cytoplasmic membrane and temporarily tethered to this membrane via a type 1 transmembrane domain (TMD) positioned near the COOH terminus. In these proteins, a sorting signal, LPXTG, is positioned immediately upstream of the TMD, which is cleaved by the membrane-associated transpeptidase, sortase A (SrtA), leading to the covalent anchoring of these proteins to newly emerging l-Ala–l-Ala cross-bridges of the growing peptidoglycan cell wall. Herein, we show that inactivation of the srtA gene in a skin-tropic pattern D GAS strain (AP53) results in retention of the M protein in the cell membrane. However, while the isogenic AP53 srtA strain is attenuated in overall pathogenic properties due to effects on the integrity of the cell membrane, our data show that the M protein nonetheless can extend from the cytoplasmic membrane through the cell wall and then to the surface of the bacteria and thereby retain its important properties of productively binding and activating fluid-phase host plasminogen (hPg). The studies presented herein demonstrate an underappreciated additional mechanism of cell surface display of bacterial virulence proteins via their retention in the cell membrane and extension to the GAS surface.

IMPORTANCE Group A Streptococcus pyogenes (GAS) is a human-specific pathogen that produces many surface factors, including its signature M protein, that contribute to its pathogenicity. M proteins undergo specific membrane localization and anchoring to the cell wall via the transpeptidase sortase A. Herein, we explored the role of sortase A function on M protein localization, architecture, and function, employing, a skin-tropic GAS isolate, AP53, which expresses a human plasminogen (hPg)-binding M (PAM) Protein. We showed that PAM anchored in the cell membrane, due to the targeted inactivation of sortase A, was nonetheless exposed on the cell surface and functionally interacted with host hPg. We demonstrate that M proteins, and possibly other sortase A-processed proteins that are retained in the cell membrane, can still function to initiate pathogenic processes by this underappreciated mechanism.

The capacity of Listeria monocytogenes to adapt to environmental changes is facilitated by a large number of regulatory proteins encoded by its genome. Among these proteins are the uncharacterized LysR-type transcriptional regulators (LTTRs). LTTRs can work as positive and/or negative transcription regulators at both local and global genetic levels. Previously, our group determined by comparative genome analysis that one member of the LTTRs (NCBI accession no. WP_003734782) was present in pathogenic strains but absent from nonpathogenic strains. The goal of the present study was to assess the importance of this transcription factor in the virulence of L. monocytogenes strain F2365 and to identify its regulons. An L. monocytogenes strain lacking lysR (the F2365lysR strain) displayed significant reductions in cell invasion of and adhesion to Caco-2 cells. In plaque assays, the deletion of lysR resulted in a 42.86% decrease in plaque number and a 13.48% decrease in average plaque size. Furthermore, the deletion of lysR also attenuated the virulence of L. monocytogenes in mice following oral and intraperitoneal inoculation. The analysis of transcriptomics revealed that the transcript levels of 139 genes were upregulated, while 113 genes were downregulated in the F2365lysR strain compared to levels in the wild-type bacteria. lysR-repressed genes included ABC transporters, important for starch and sucrose metabolism as well as glycerolipid metabolism, flagellar assembly, quorum sensing, and glycolysis/gluconeogenesis. Conversely, lysR activated the expression of genes related to fructose and mannose metabolism, cationic antimicrobial peptide (CAMP) resistance, and beta-lactam resistance. These data suggested that lysR contributed to L. monocytogenes virulence by broad impact on multiple pathways of gene expression.

IMPORTANCE Listeria monocytogenes is the causative agent of listeriosis, an infectious and fatal disease of animals and humans. In this study, we have shown that lysR contributes to Listeria pathogenesis and replication in cell lines. We also highlight the importance of lysR in regulating the transcription of genes involved in different pathways that might be essential for the growth and persistence of L. monocytogenes in the host or under nutrient limitation. Better understanding L. monocytogenes pathogenesis and the role of various virulence factors is necessary for further development of prevention and control strategies.

The type VI secretion system (T6SS) is a weapon for delivering effectors into target cells that is widespread in Gram-negative bacteria. The T6SS is a highly versatile machine, as it can target both eukaryotic and prokaryotic cells, and it has been proposed that T6SSs are adapted to the specific needs of each bacterium. The expression of T6SS gene clusters and the activation of the secretion apparatus are therefore tightly controlled. In enteroaggregative Escherichia coli (EAEC), the sci1 T6SS gene cluster is subject to a complex regulation involving both the ferric uptake regulator (Fur) and DNA adenine methylase (Dam)-dependent DNA methylation. In this study, an additional, internal, promoter was identified within the sci1 gene cluster using +1 transcriptional mapping. Further analyses demonstrated that this internal promoter is controlled by a mechanism strictly identical to that of the main promoter. The Fur binding box overlaps the –10 transcriptional element and a Dam methylation site, GATC-32. Hence, the expression of the distal sci1 genes is repressed and the GATC-32 site is protected from methylation in iron-rich conditions. The Fur-dependent protection of GATC-32 was confirmed by an in vitro methylation assay. In addition, the methylation of GATC-32 negatively impacted Fur binding. The expression of the sci1 internal promoter is therefore controlled by iron availability through Fur regulation, whereas Dam-dependent methylation maintains a stable ON expression in iron-limited conditions.

IMPORTANCE Bacteria use weapons to deliver effectors into target cells. One of these weapons, the type VI secretion system (T6SS), assembles a contractile tail acting as a spring to propel a toxin-loaded needle. Its expression and activation therefore need to be tightly regulated. Here, we identified an internal promoter within the sci1 T6SS gene cluster in enteroaggregative E. coli. We show that this internal promoter is controlled by Fur and Dam-dependent methylation. We further demonstrate that Fur and Dam compete at the –10 transcriptional element to finely tune the expression of T6SS genes. We propose that this elegant regulatory mechanism allows the optimum production of the T6SS in conditions where enteroaggregative E. coli encounters competing species.

Shigella species, the causal agents of bacillary dysentery, use a type III secretion system (T3SS) to inject two waves of virulence proteins, known as effectors, into the colonic epithelium to subvert host cell machinery. Prior to host cell contact and secretion of the first wave of T3SS effectors, OspD1, an effector and antiactivator protein, prevents premature production of the second wave of effectors. Despite this important role, regulation of the ospD1 gene is not well understood. While ospD1 belongs to the large regulon of VirB, a transcriptional antisilencing protein that counters silencing mediated by the histone-like nucleoid structuring protein H-NS, it remains unclear if VirB directly or indirectly regulates ospD1. Additionally, it is not known if ospD1 is regulated by H-NS. Here, we identify the primary ospD1 transcription start site (+1) and show that the ospD1 promoter is remotely regulated by both VirB and H-NS. Our findings demonstrate that VirB regulation of ospD1 requires at least one of the two newly identified VirB regulatory sites, centered at –978 and –1270 relative to the ospD1 +1. Intriguingly, one of these sites lies on a 193-bp sequence found in three conserved locations on the large virulence plasmids of Shigella. The region required for H-NS-dependent silencing of ospD1 lies between –1120 and –820 relative to the ospD1 +1. Thus, our study provides further evidence that cis-acting regulatory sequences for transcriptional antisilencers and silencers, such as VirB and H-NS, can lie far upstream of the canonical bacterial promoter region (i.e., –250 to +1).

IMPORTANCE Transcriptional silencing and antisilencing mechanisms regulate virulence gene expression in many important bacterial pathogens. In Shigella species, plasmid-borne virulence genes, such as those encoding the type III secretion system (T3SS), are silenced by the histone-like nucleoid structuring protein H-NS and antisilenced by VirB. Previous work at the plasmid-borne icsP locus revealed that VirB binds to a remotely located cis-acting regulatory site to relieve transcriptional silencing mediated by H-NS. Here, we characterize a second example of remote VirB antisilencing at ospD1, which encodes a T3SS antiactivator and effector. Our study highlights that remote transcriptional silencing and antisilencing occur more frequently in Shigella than previously thought, and it raises the possibility that long-range transcriptional regulation in bacteria is commonplace.

Diadenosine tetraphosphate (Ap₄A) is a dinucleotide found in both prokaryotes and eukaryotes. In bacteria, its cellular levels increase following exposure to various stress signals and stimuli, and its accumulation is generally correlated with increased sensitivity to a stressor(s), decreased pathogenicity, and enhanced antibiotic susceptibility. Ap₄A is produced as a by-product of tRNA aminoacylation, and is cleaved to ADP molecules by hydrolases of the ApaH and Nudix families and/or by specific phosphorylases. Here, considering evidence that the recombinant protein YqeK from Staphylococcus aureus copurified with ADP, and aided by thermal shift and kinetic analyses, we identified the YqeK family of proteins (COG1713) as an unprecedented class of symmetrically cleaving Ap₄A hydrolases. We validated the functional assignment by confirming the ability of YqeK to affect in vivo levels of Ap₄A in B. subtilis. YqeK shows a catalytic efficiency toward Ap₄A similar to that of the symmetrically cleaving Ap₄A hydrolases of the known ApaH family, although it displays a distinct fold that is typical of proteins of the HD domain superfamily harboring a diiron cluster. Analysis of the available 3D structures of three members of the YqeK family provided hints to the mode of substrate binding. Phylogenetic analysis revealed the occurrence of YqeK proteins in a consistent group of Gram-positive bacteria that lack ApaH enzymes. Comparative genomics highlighted that yqeK and apaH genes share a similar genomic context, where they are frequently found in operons involved in integrated responses to stress signals.

IMPORTANCE Elevation of Ap₄A level in bacteria is associated with increased sensitivity to heat and oxidative stress, reduced antibiotic tolerance, and decreased pathogenicity. ApaH is the major Ap₄A hydrolase in gamma- and betaproteobacteria and has been recently proposed as a novel target to weaken the bacterial resistance to antibiotics. Here, we identified the orphan YqeK protein family (COG1713) as a highly efficient Ap₄A hydrolase family, with members distributed in a consistent group of bacterial species that lack the ApaH enzyme. Among them are the pathogens Staphylococcus aureus, Streptococcus pneumoniae, and Mycoplasma pneumoniae. By identifying the player contributing to Ap₄A homeostasis in these bacteria, we disclose a novel target to develop innovative antibacterial strategies.

Chitotriosidase (CHIT1) is a highly conserved and regulated chitinase secreted by activated macrophages; it is a member of the 18-glycosylase family (GH18). CHIT1 is the most prominent chitinase in humans, can cleave chitin and participates in the body's immune response and is associated with inflammation, infection, tissue damage and remodelling processes. Recently, CHIT1 has been reported to be involved in the molecular pathogenesis of pulmonary fibrosis, bronchial asthma, COPD and pulmonary infections, shedding new light on the role of these proteins in lung pathophysiology. The potential roles of CHIT1 in lung diseases are reviewed in this article.

An aberrant bi-apical Follicucullus specimen (Albaillellaria, Radiolaria) has been discovered from an upper Guadalupian (Middle Permian) chert block of the Kamiaso Unit of the Mino terrane, central Japan. If this specimen was formed with double malformation, it would be the oldest record of this phenomenon in radiolarians and the first record of its kind in Albaillellaria.

The Henry Buckley Collection of Planktonic Foraminifera at the Natural History Museum in London (NHMUK) consists of 1665 single-taxon slides housing 23 897 individuals from 203 sites in all the major ocean basins, as well as a vast research library of Scanning Electron Microscope (SEM) photomicrographs. Buckley picked the material from the NHMUK Ocean-Bottom Deposit Collection and also from fresh tow samples. However, his collection remains largely unused as he was discouraged by his managers in the Mineralogy Department from working on or publicizing the collection. Nevertheless, Buckley published pioneering papers on isotopic interpretation of oceanographic and climatic change and was one of the first workers to investigate foraminiferal wall structure using the SEM technique. Details of the collection and images of each slide are available via the NHMUK Data Portal (http://dx.doi.org/10.5519/0035055). The Buckley Collection and its associated Ocean-Bottom Deposit Collection have great potential for taxon-specific studies as well as geochemical work, and both collections are available on request.

A new larger benthic porcelaneous foraminifer of soritid affinity is described as Tarburina zagrosiana n. gen., n. sp. from the late Maastrichtian of the Tarbur Formation, Zagros Zone, SW Iran. It occurs in foraminiferal–dasycladalean wackestones and packstones, in association with Loftusia ssp., dicyclinids/cuneolinids, Neobalkhania bignoti Cherchi & Schroeder, Gyroconulina columellifera Schroeder & Darmoian, Spirolina? farsiana Schlagintweit & Rashidi, Broeckina cf. dufrenoyi (d'Archiac), other benthic foraminifers, and dasycladalean algae. Due to its elongate test and marginal chamber subdivision by aligned vertical partitions, Tarburina n. gen. can be compared with representatives of the Praerhapydionininae. The interio-marginal slit-like foramina/aperture of Tarburina represents an outstanding feature in complex porcelaneous taxa. The monospecific genus Tarburina is considered a Maastrichtian newcomer within the Late Cretaceous Global Community Maturation cycle of larger benthic foraminifera. A biostratigraphic and palaeobiogeographical restriction seems possible, as reported for many other Late Cretaceous larger benthic foraminifera.

Planktonic foraminifera are a source of important geochemical, palaeoceanographic, and palaeontological data. However, many aspects of their ecology remain poorly understood, including whether or not gross morphology has an ecological function. Here, we measure the force needed to crush multiple planktonic foraminiferal morphotypes from modern core top and tow samples. We find significant differences in the resistance of different morphotypes to compressional force. Three species, Globorotalia tumida (biconvex, keeled), Menardella menardii (discoidal, keeled), Truncorotalia truncatulinoides (conical, keeled), require on average 59% more force (1.07 v. 0.47 N) to crush than the least resistant species (Orbulina universa and Trilobatus sacculifer) in core-top samples. Towed samples of pre-gametogenic individuals also show significant differences of the same magnitude (0.693 v. 0.53 N) between the conical (T. truncatulinoides) and globular/spherical morphologies (Globoconella inflata and O. universa). We hypothesize that the greater compressional strength of certain shapes confers a fitness advantage against predators and could contribute to the repeated, convergent evolution of keeled, conical and bi-convex forms in planktonic foraminifer lineages.

Supplementary material: Raw data for all crushing experiments, wall thickness measurements, and results for all pair-wise Kolmogorov-Smirnov Tests are available at https://doi.org/10.6084/m9.figshare.c.3725236.v1

ABSTRACTBackground:Some opportunities to routinely capture and improve respectful maternity care (RMC) during facility-based childbirth include quality improvement (QI) initiatives, community-based monitoring efforts through community score cards (CSC), and performance-based financing (PBF) initiatives. But there is limited guidance on which types of RMC indicators are best suited for inclusion in these initiatives. We sought to provide practical evidence-based recommendations on indicators that may be used for routine measurement of RMC in programs.Methods:We used a rapid review approach, which included (1) reviewing existing documents and publications to extract RMC indicators and identify which have or can be used in facility-based QI, CSCs, and PBF schemes; (2) surveying RMC and maternal health experts to rank indicators, and (3) analyzing survey data to select the most recommended indicators.Results:We identified 49 indicators spanning several domains of RMC and mistreatment including dignified/nondignified care, verbal and physical abuse, privacy/confidentiality, autonomy/loss of autonomy, supportive care/lack thereof, communication, stigma, discrimination, trust, facility environment/culture, responsiveness, and nonevidence-based care. Based on the analysis of the survey data, we recommend 33 indicators (between 2 and 6 indicators for each RMC domain) that may be suited for incorporation in both facility-based QI and CSC-related monitoring efforts.Conclusion:Integrating RMC indicators into QI and CSC initiatives, as well as in other maternal and neonatal health programs, could help improve RMC at the facility and community level. More research is needed into whether RMC can be integrated into PBF initiatives. Integration of RMC indicators into programs to improve quality of care and other health system outcomes will facilitate routine monitoring and accountability around experience of care. Measurement and improvement of women's experiences will increase maternal health service utilization and improve quality of care as a means of reducing maternal and neonatal morbidity and mortality.

ABSTRACTThe Standard Days Method (SDM), a modern fertility awareness-based family planning method, has been introduced in 30 countries since its development in 2001. It is still unclear to what extent the SDM was mainstreamed within the family planning method mix, particularly in low- and middle-income country (LMIC) settings, where the SDM had been introduced by donors and implementing partners. This review of implementation science publications on the SDM in LMICs first looked at community pilot studies of the SDM to determine the acceptability of the method; correct use and efficacy rates; demographics of users; and changes to contraceptive prevalence rates and family planning behaviors, especially among men and couples. Then, we examined the status of the SDM in the 16 countries that had attempted to scale up the method within national family planning protocols, training, and service delivery. At the community level, evidence demonstrated a high level of acceptability of the method; efficacy rates comparable to the initial clinical trials; diversity in demographic characteristics of users, including first-time or recently discontinued users of family planning; increased male engagement in family planning; and improved couple's communication. Nationally, few countries had scaled up the SDM due to uneven stakeholder engagement, lackluster political will, and competing resource priorities. Results of this review could help policy makers determine the added value of the SDM in the contraceptive method mix and identify potential barriers to its implementation moving forward.

ABSTRACTBackground:Female sex workers (FSWs) in Cameroon commonly have unmet need for contraception posing a high risk of unintended pregnancy. Unintended pregnancy leads to a range of outcomes, and due to legal restrictions, FSWs often seek unsafe abortions. Aside from the high burden of HIV, little is known about the broader sexual and reproductive health of FSWs in Cameroon.Methods:From December 2015 to October 2016, we recruited FSWs aged ≥18 years through respondent-driven sampling across 5 Cameroonian cities. Cross-sectional data were collected through a behavioral questionnaire. Modified-robust Poisson regression was used to approximate adjusted prevalence ratios (aPR) for TOP and current use of effective nonbarrier contraception.Results:Among 2,255 FSWs (median age 28 years), 57.6% reported history of unintended pregnancy and 40.0% reported prior TOP. In multivariable analysis, TOP history was associated with current nonbarrier contraceptive use (aPR=1.23, 95% confidence interval [CI]=1.07, 1.42); ever using emergency contraception (aPR=1.34, 95% CI=1.17, 1.55); >60 clients in the past month (aPR=1.29, 95% CI= 1.07, 1.54) compared to ≤30; inconsistent condom use with clients (aPR=1.17, 95% CI=1.00, 1.37); ever experiencing physical violence (aPR=1.24, 95% CI=1.09, 1.42); and older age. Most (76.5%) women used male condoms for contraception, but only 33.2% reported consistent condom use with all partners. Overall, 26.4% of women reported currently using a nonbarrier contraceptive method, and 6.2% reported using a long-acting method. Previous TOP (aPR=1.41, 95%CI=1.16, 1.72) and ever using emergency contraception (aPR=2.70, 95% CI=2.23, 3.26) were associated with higher nonbarrier contraceptive use. Recent receipt of HIV information (aPR=0.72, 95% CI=0.59, 0.89) and membership in an FSW community-based organization (aPR=0.73, 95% CI=0.57, 0.92) were associated with lower use nonbarrier contraceptive use.Conclusions:Experience of unintended pregnancies and TOP is common among FSWs in Cameroon. Given the low use of nonbarrier contraceptive methods and inconsistent condom use, FSWs are at risk of repeat unintended pregnancies. Improved integration of client-centered, voluntary family planning within community-led HIV services may better support the sexual and reproductive health and human rights of FSWs consistent with the United Nations Declaration of Human Rights.

ABSTRACTBackground:A significant number of girls are married as children, which negatively impacts their health, education, and development. Given the sheer numbers of girls at risk of child marriage globally, the challenge to eliminate the practice is daunting. Programs to prevent child marriage are typically small-scale and overlook the costs and scalability of the intervention.Implementation:This study tested and costed different approaches to preventing child marriage in rural Burkina Faso and Tanzania. The approaches tested were community dialogue, provision of school supplies, provision of a livestock asset, a model including all components, and a control arm. A quasi-experimental design was employed with surveys undertaken at baseline and after 2 years of intervention. We examined the prevalence of child marriage and school attendance controlling for background characteristics and stratified by age group. Programmatic costs were collected prospectively.Results:Among those in the community dialogue arm in Burkina Faso, girls aged 15 to 17 years had two-thirds less risk (risk ratio [RR]=0.33; 95% confidence interval [CI]=0.19, 0.60) of being married and girls aged 12 to 14 years had a greater chance of being in school (RR=1.18; 95% CI=1.07,1.29) compared to the control site. In Tanzania, girls aged 12 to 14 years residing in the multicomponent arm had two-thirds less risk of being married (RR=0.33; 95% CI=0.11, 0.99), and girls 15 to 17 in the conditional asset location had half the risk (RR=0.52; 95% CI=0.30, 0.91). All the interventions tested in Tanzania were associated with increased risk of girls 12 to 14 years old being in school, and the educational promotion arm was also associated with a 30% increased risk of girls aged 15 to 17 years attending school (RR=1.3; 95% CI=1.01, 1.67). Costs per beneficiary ranged from US$9 to US$117.Conclusion:The study demonstrates that minimal, low-cost approaches can be effective in delaying child marriage and increasing school attendance. However, community dialogues need to be designed to ensure sufficient quality and intensity of messaging. Program managers should pay attention to the cost, quality, and coverage of interventions, especially considering that child marriage persists in the most hard-to-reach rural areas of many countries.

ABSTRACTIntroduction:We evaluated a 2-way short message service (SMS) communication platform to improve continuation of pre-exposure prophylaxis (PrEP) for HIV prevention among Kenyan women who initiated PrEP within routine maternal child health (MCH) and family planning clinics.Methods:We adapted an existing SMS platform (Mobile WACh [mWACh]) to send PrEP-tailored, theory-based SMS and allow clients to communicate with a remote nurse. Women who did not have HIV and who were initiating PrEP at 2 MCH/family planning clinics in Kisumu County, Kenya, from February to October 2018, were offered enrollment into the mWACh-PrEP program; SMS communication was free. We evaluated acceptability, satisfaction, and implementation metrics. In a pre/postevaluation, we compared PrEP continuation at 1-month postinitiation among women who initiated PrEP in the period before (n=166) versus after mWACh-PrEP implementation, adjusting for baseline differences.Results:Of the 334 women who were screened for enrollment into the mWACh-PrEP program; 193 (58%) were eligible and of those, 190 (98%) accepted enrollment. Reasons for ineligibility (n=141) included no phone access (29%) and shared SIM cards (25%). Median age was 25 years (interquartile range=22–30), and 91% were MCH clients. Compared to women who initiated PrEP in the month before mWACh-PrEP implementation, women who enrolled in mWACh-PrEP were more likely to return for their first PrEP follow-up visit (40% vs. 53%; adjusted risk ratio [aRR]=1.26; 95% confidence interval [CI]= 1.06, 1.50; P=.008) and more likely to continue PrEP (22% vs. 43%; aRR=1.75; 95% CI=1.21, 2.55; P=.003). Among those who returned, 99% reported successful receipt of SMS through the mWACh-PrEP system and 94% reported that mWACh-PrEP helped them understand PrEP better. Concerns about PrEP use, how it works, and side effects accounted for the majority (80%) of issues raised by participants using SMS.Conclusions:Two-way SMS expanded support for PrEP and opportunities for dialogue beyond the clinic and enabled women to ask and receive answers in real time regarding PrEP, which facilitated its continued use.

ABSTRACTBackground:The focused assessment with sonography for HIV-associated tuberculosis (TB) (FASH) ultrasound protocol has been increasingly used to help clinicians diagnose TB. We sought to quantify the diagnostic utility of FASH for TB among individuals with HIV in Malawi.Methods:Between March 2016 and August 2017, 210 adults with HIV who had 2 or more signs and symptoms that were concerning for TB (fever, cough, night sweats, weight loss) were enrolled from a public HIV clinic in Lilongwe, Malawi. The treating clinicians conducted a history, physical exam, FASH protocol, and additional TB evaluation (laboratory diagnostics and chest radiography) on all participants. The clinician made a final treatment decision based on all available information. At the 6-month follow-up visit, we categorized participants based on clinical outcomes and diagnostic tests as having probable/confirmed TB or unlikely TB; association of FASH with probable/confirmed TB was calculated using Fisher's exact tests. The impact of FASH on empiric TB treatment was determined by asking the clinicians prospectively about whether they would start treatment at 2 time points in the baseline visit: (1) after the initial history and physical exam; and (2) after history, physical exam, and FASH protocol.Results:A total of 181 participants underwent final analysis, of whom 56 were categorized as probable/confirmed TB and 125 were categorized as unlikely TB. The FASH protocol was positive in 71% (40/56) of participants with probable/confirmed TB compared to 24% (30/125) of participants with unlikely TB (odds ratio=7.9, 95% confidence interval=3.9,16.1; P<.001). Among those classified as confirmed/probable TB, FASH increased the likelihood of empiric TB treatment before obtaining any other diagnostic studies from 9% (5/56) to 46% (26/56) at the point-of-care. For those classified as unlikely TB, FASH increased the likelihood of empiric treatment from 2% to 4%.Conclusion:In the setting of HIV coinfection in Malawi, FASH can be a helpful tool that augments the clinician's ability to make a timely diagnosis of TB.

ABSTRACTIntroduction:Multimonth dispensing (MMD) of antiretroviral therapy (ART) is a differentiated model of care that can help overcome health system challenges and reduce the burden of HIV care on clients. Although 3-month dispensing has been the standard of care, interest has increased in extending refill intervals to 6 months. We explored client and provider experiences with MMD in Malawi as part of a cluster randomized trial evaluating 3- versus 6-month ART dispensing.Methods:Semi-structured in-depth interviews were conducted with 17 ART providers and 62 stable, adult clients with HIV on ART. Clients and providers were evenly divided by arm and were eligible for an interview if they had been participating in the study for 1 year (clients) or 6 months (providers). Questions focused on perceived challenges and benefits of the 3- or 6-month amount of ART dispensing. Interviews were transcribed, and data were coded and analyzed using constant comparison.Results:Both clients and providers reported that the larger medication supply had benefits. Clients reported decreased costs due to less frequent travel to the clinic and increased time for income-generating activities. Clients in the 6-month dispensing arm reported a greater sense of personal freedom and normalcy. Providers felt that the 6-month dispensing interval reduced their workload. They also expressed concerned about clients' challenges with ART storage at home, but clients reported no storage problems. Although providers mentioned the potential risk of clients sharing the larger medication supply with family or friends, clients emphasized the value of ART and reported only rare, short-term sharing, mostly with their spouses. Providers mentioned clients' lack of motivation to seek care for illnesses that might occur between refill appointments.Conclusions:The 6-month ART dispensing arm was particularly beneficial to clients for decreased costs, increased time for income generation, and a greater sense of normalcy. Providers' concerns about storage, sharing, and return visits to the facility did not emerge in client interviews. Further data are needed on the feasibility of implementing a large-scale program with 6-month dispensing.

BACKGROUND

Sex differences have been described in diabetes cardiovascular outcome trials (CVOTs).