Pregnancy wastage among HIV infected women in a high HIV prevalence district of India.

BACKGROUND: Bagalkot district in Karnataka state is one of the highest HIV prevalence districts in India. A large proportion of the girls also marry at early age in the district and negative pregnancy outcomes among the HIV positive women likely to have large pregnancy wastages. Therefore, this study examined the pregnancy wastages and the associated factors among HIV positive women in a high prevalent district in India. METHODS: We used data from a cross-sectional survey conducted recently among randomly selected currently married HIV positive women, 15-29 years of age, in one of the high HIV prevalence districts in India. The study used the experience of reported pregnancy wastage as an outcome variable, and both bi-variate and multivariate logistic regression analyses were carried out to understand the factors associated with the pregnancy wastage among HIV infected women. RESULTS: Overall, 17% of the respondents reported pregnancy wastage, of which 81% were due to spontaneous abortions. Respondents who became pregnant since testing HIV positive reported significantly higher level of pregnancy wastage as compared to those were pregnant before they were tested for HIV. (AOR = 1.9; p = 0.00). While a positive association between duration of marriage and pregnancy wastage was noticed (AOR = 7.4; p = 0.01), there was a negative association between number of living children and pregnancy wastage (AOR = 0.24; p = 0.00). Living in a joint family was associated with increased reporting of pregnancy wastage as compared to those living in nuclear families (AOR = 1.7; p = 0.03). CONCLUSIONS: HIV prevention and care programs need to consider the reproductive health needs of HIV infected married women as a priority area since large proportion of these women reported negative pregnancy outcomes. There is also a need to explore ways to raise the age at marriage in order to stop women getting married before the legal age at marriage.

Pathogen espionage: multiple bacterial adrenergic sensors eavesdrop on host communication systems.

The interactions between bacterial pathogens and their eukaryotic hosts are vital in determining the outcome of infections. Bacterial pathogens employ molecular sensors to detect and facilitate adaptation to changes in their niche. The sensing of these extracellular signals enables the pathogen to navigate within mammalian hosts. Intercellular bacterial communication is facilitated by the production and sensing of autoinducer (AI) molecules via quorum sensing. More recently, AI-3 and the host neuroendocrine (NE) hormones adrenaline and noradrenaline were reported to display cross-talk for the activation of the same signalling pathways. Remarkably, there is increasing evidence to suggest that enteric bacteria sense and respond to the host NE stress hormones adrenaline and noradrenaline to modulate virulence. These responses can be inhibited by α and ß-adrenergic receptor antagonists implying a bacterial receptor-based sensing and signalling cascade. In Escherichia coli O157:H7 and Salmonella, QseC has been proposed as the adrenergic receptor. Strikingly, there is an increasing body of evidence that not all the bacterial adrenergic responses require signalling through QseC. Here we provide additional hypotheses to reconcile these observations implicating the existence of alternative adrenergic receptors including BasS, QseE and CpxA and their associated signalling cascades with major roles in interkingdom communication.

The bacterial cytoskeleton modulates motility, type 3 secretion, and colonization in Salmonella.

Although there have been great advances in our understanding of the bacterial cytoskeleton, major gaps remain in our knowledge of its importance to virulence. In this study we have explored the contribution of the bacterial cytoskeleton to the ability of Salmonella to express and assemble virulence factors and cause disease. The bacterial actin-like protein MreB polymerises into helical filaments and interacts with other cytoskeletal elements including MreC to control cell-shape. As mreB appears to be an essential gene, we have constructed a viable ΔmreC depletion mutant in Salmonella. Using a broad range of independent biochemical, fluorescence and phenotypic screens we provide evidence that the Salmonella pathogenicity island-1 type three secretion system (SPI1-T3SS) and flagella systems are down-regulated in the absence of MreC. In contrast the SPI-2 T3SS appears to remain functional. The phenotypes have been further validated using a chemical genetic approach to disrupt the functionality of MreB. Although the fitness of ΔmreC is reduced in vivo, we observed that this defect does not completely abrogate the ability of Salmonella to cause disease systemically. By forcing on expression of flagella and SPI-1 T3SS in trans with the master regulators FlhDC and HilA, it is clear that the cytoskeleton is dispensable for the assembly of these structures but essential for their expression. As two-component systems are involved in sensing and adapting to environmental and cell surface signals, we have constructed and screened a panel of such mutants and identified the sensor kinase RcsC as a key phenotypic regulator in ΔmreC. Further genetic analysis revealed the importance of the Rcs two-component system in modulating the expression of these virulence factors. Collectively, these results suggest that expression of virulence genes might be directly coordinated with cytoskeletal integrity, and this regulation is mediated by the two-component system sensor kinase RcsC.

Multidirectional chemical signalling between Mammalian hosts, resident microbiota, and invasive pathogens: neuroendocrine hormone-induced changes in bacterial gene expression.

Host-pathogen communication appears to be crucial in establishing the outcome of bacterial infections. There is increasing evidence to suggest that this communication can take place by bacterial pathogens sensing and subsequently responding to host neuroendocrine (NE) stress hormones. Bacterial pathogens have developed mechanisms allowing them to eavesdrop on these communication pathways within their hosts. These pathogens can use intercepted communication signals to adjust their fitness to persist and cause disease in their hosts. Recently, there have been numerous studies highlighting the ability of NE hormones to act as an environmental cue for pathogens, helping to steer their responses during host infection. Host NE hormone sensing can take place indirectly or directly via bacterial adrenergic receptors (BARs). The resulting changes in bacterial gene expression can be of strategic benefit to the pathogen. Furthermore, it is intriguing that not only can bacteria sense NE stress hormones but they are also able to produce key signalling molecules known as autoinducers. The rapid advances in our knowledge of the human microbiome, and its impact on health and disease highlights the potential importance of communication between the microbiota, pathogens and the host. It is indeed likely that the microbiota input significantly in the neuroendocrinological homeostasis of the host by catabolic, anabolic, and signalling processes. The arrival of unwanted guests, such as bacterial pathogens, clearly has a major impact on these delicately balanced interactions. Unravelling the pathways involved in interkingdom communication between invading bacterial pathogens, the resident microbiota, and hosts, may provide novel targets in our continuous search for new antimicrobials to control disease.

Revisiting the Clinical Interpretation of CT-Measured Pulmonary Artery-to-Aorta Ratio-The Rotterdam Study.

RATIONALE: The pulmonary artery (PA) diameter-to-aorta ratio (PA:A) ratio is a novel marker in cardiovascular imaging for detecting pulmonary hypertension. However, we question the effect of the varying aorta diameter on the ratio, which complicates the interpretation of the PA:A ratio. OBJECTIVE: Investigate the variability of the PA:A ratio by examining the correlation between PA:A ratio and aorta diameter and by comparing the associations of the PA diameter, aorta diameters, and PA:A ratio. METHODS: We included 2197 participants from the Rotterdam Study who underwent non-contrast multidetector computed tomography to measure the PA and aorta diameters. Pearson correlation coefficient was calculated between the PA:A ratio and aorta diameter. Multiple linear regression analyses were performed to compare the determinants of the individual diameters and PA:A ratio. RESULTS: We found a statistically significant correlation between the PA:A ratio and aorta diameter (r = -0.38, p < 0.001). The PA diameter was statistically significantly associated with, height, weight, diastolic blood pressure, blood pressure medication, prevalence of atrial fibrillation, prevalence of heart failure, and prevalence of stroke (p < 0.05). Except for blood pressure medication, the PA:A ratio had similar determinants compared to the PA diameter but was also statistically significantly associated with sex, and systolic blood pressure (p < 0.05), which were statistically significantly associated with the aorta diameter (p < 0.05). CONCLUSION: The PA:A ratio should not be interpreted without taking into account the variability of the individual components (PA and aorta diameter) according to the anthropomorphic and clinical characteristics.

Salmonella Typhi sense host neuroendocrine stress hormones and release the toxin haemolysin E.

Salmonella enterica serovar Typhi (S. typhi) causes typhoid fever. We show that exposure of S. typhi to neuroendocrine stress hormones results in haemolysis, which is associated with the release of haemolysin E in membrane vesicles. This effect is attributed to increased expression of the small RNA micA and RNA chaperone Hfq, with concomitant downregulation of outer membrane protein A. Deletion of micA or the two-component signal-transduction system, CpxAR, abolishes the phenotype. The hormone response is inhibited by the ß-blocker propranolol. We provide mechanistic insights into the basis of neuroendocrine hormone-mediated haemolysis by S. typhi, increasing our understanding of inter-kingdom signalling.

The changing role of gesture in linguistic development: a developmental trajectory and a cross-cultural comparison between British and Finnish children.

We studied how gesture use changes with culture, age and increased spoken language competence. A picture-naming task was presented to British (N = 80) and Finnish (N = 41) typically developing children aged 2-5 years. British children were found to gesture more than Finnish children and, in both cultures, gesture production decreased after the age of two. Two-year-olds used more deictic than iconic gestures than older children, and gestured more before the onset of speech, rather than simultaneously or after speech. The British 3- and 5-year-olds gestured significantly more when naming praxic (manipulable) items than non-praxic items. Our results support the view that gesture serves a communicative and intrapersonal function, and the relative function may change with age. Speech and language therapists and psychologists observe the development of children's gestures and make predictions on the basis of their frequency and type. To prevent drawing erroneous conclusions about children's linguistic development, it is important to understand developmental and cultural variations in gesture use.

Conjunctival epithelial cells resist productive SARS-CoV-2 infection.

Conjunctival epithelial cells, which express viral-entry receptors angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine type 2 (TMPRSS2), constitute the largest exposed epithelium of the ocular surface tissue and may represent a relevant viral-entry route. To address this question, we generated an organotypic air-liquid-interface model of conjunctival epithelium, composed of basal, suprabasal, and superficial epithelial cells, and fibroblasts, which could be maintained successfully up to day 75 of differentiation. Using single-cell RNA sequencing (RNA-seq), with complementary imaging and virological assays, we observed that while all conjunctival cell types were permissive to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome expression, a productive infection did not ensue. The early innate immune response to SARS-CoV-2 infection in conjunctival cells was characterised by a robust autocrine and paracrine NF-κB activity, without activation of antiviral interferon signalling. Collectively, these data enrich our understanding of SARS-CoV-2 infection at the human ocular surface, with potential implications for the design of preventive strategies and conjunctival transplantation.

Delayed induction of type I and III interferons mediates nasal epithelial cell permissiveness to SARS-CoV-2.

The nasal epithelium is a plausible entry point for SARS-CoV-2, a site of pathogenesis and transmission, and may initiate the host response to SARS-CoV-2. Antiviral interferon (IFN) responses are critical to outcome of SARS-CoV-2. Yet little is known about the interaction between SARS-CoV-2 and innate immunity in this tissue. Here we apply single-cell RNA sequencing and proteomics to a primary cell model of human nasal epithelium differentiated at air-liquid interface. SARS-CoV-2 demonstrates widespread tropism for nasal epithelial cell types. The host response is dominated by type I and III IFNs and interferon-stimulated gene products. This response is notably delayed in onset relative to viral gene expression and compared to other respiratory viruses. Nevertheless, once established, the paracrine IFN response begins to impact on SARS-CoV-2 replication. When provided prior to infection, recombinant IFNß or IFNλ1 induces an efficient antiviral state that potently restricts SARS-CoV-2 viral replication, preserving epithelial barrier integrity. These data imply that the IFN-I/III response to SARS-CoV-2 initiates in the nasal airway and suggest nasal delivery of recombinant IFNs to be a potential chemoprophylactic strategy.

Genome-wide transposon mutagenesis identifies a role for host neuroendocrine stress hormones in regulating the expression of virulence genes in Salmonella.

Bacterial sensing of environmental signals plays a key role in regulating virulence and mediating bacterium-host interactions. The sensing of the neuroendocrine stress hormones epinephrine (adrenaline) and norepinephrine (noradrenaline) plays an important role in modulating bacterial virulence. We used MudJ transposon mutagenesis to globally screen for genes regulated by neuroendocrine stress hormones in Salmonella enterica serovar Typhimurium. We identified eight hormone-regulated genes, including yhaK, iroC, nrdF, accC, yedP, STM3081, and the virulence-related genes virK and mig14. The mammalian alpha-adrenergic receptor antagonist phentolamine reversed the hormone-mediated effects on yhaK, virK, and mig14 but did not affect the other genes. The beta-adrenergic receptor antagonist propranolol had no activity in these assays. The virK and mig14 genes are involved in antimicrobial peptide resistance, and phenotypic screens revealed that exposure to neuroendocrine hormones increased the sensitivity of S. Typhimurium to the antimicrobial peptide LL-37. A virK mutant and a virK mig14 double mutant also displayed increased sensitivity to LL-37. In contrast to enterohemorrhagic Escherichia coli (EHEC), we have found no role for the two-component systems QseBC and QseEF in the adrenergic regulation of any of the identified genes. Furthermore, hormone-regulated gene expression could not be blocked by the QseC inhibitor LED209, suggesting that sensing of hormones is mediated through alternative signaling pathways in S. Typhimurium. This study has identified a role for host-derived neuroendocrine stress hormones in downregulating S. Typhimurium virulence gene expression to the benefit of the host, thus providing further insights into the field of host-pathogen communication.

Salmonella pathogenicity island 1 (SPI-1) type III secretion of SopD involves N- and C-terminal signals and direct binding to the InvC ATPase.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is an important pathogen and a causative agent of gastroenteritis. During infection, S. Typhimurium assembles molecular-needle complexes termed type III secretion (T3S) systems to translocate effector proteins from the bacterial cytoplasm directly into the host cell. The T3S signals that direct the secretion of effectors still remain enigmatic. SopD is a key T3S effector contributing to the systemic virulence of S. Typhimurium and the development of gastroenteritis. We have scrutinized the distribution of the SopD T3S signals using in silico analysis and a targeted deletion approach. We show that amino acid residues 6-10 act as the N-terminal secretion signal for Salmonella pathogenicity island 1 (SPI-1) T3S. Furthermore, we show that two putative C-terminal helical regions of SopD are essential for its secretion and also help prevent erroneous secretion through the flagellar T3S machinery. In addition, using protein-protein interaction assays, we have identified an association between SopD and the SPI-1 T3S system ATPase, InvC. These findings demonstrate that T3S of SopD involves multiple signals and protein interactions, providing important mechanistic insights into effector protein secretion.

LuxS-based quorum sensing does not affect the ability of Salmonella enterica serovar Typhimurium to express the SPI-1 type 3 secretion system, induce membrane ruffles, or invade epithelial cells.

Bacterial species can communicate by producing and sensing small autoinducer molecules by a process known as quorum sensing. Salmonella enterica produces autoinducer 2 (AI-2) via the luxS synthase gene, which is used by some bacterial pathogens to coordinate virulence gene expression with population density. We investigated whether the luxS gene might affect the ability of Salmonella enterica serovar Typhimurium to invade epithelial cells. No differences were found between the wild-type strain of S. Typhimurium, SL1344, and its isogenic luxS mutant with respect to the number and morphology of the membrane ruffles induced or their ability to invade epithelial cells. The dynamics of the ruffling process were also similar in the wild-type strain (SL1344) and the luxS mutant. Furthermore, comparing the Salmonella pathogenicity island 1 (SPI-1) type 3 secretion profiles of wild-type SL1344 and the luxS mutant by Western blotting and measuring the expression of a single-copy green fluorescent protein fusion to the prgH (an essential SPI-1 gene) promoter indicated that SPI-1 expression and activity are similar in the wild-type SL1344 and luxS mutant. Genetic deletion of luxS did not alter the virulence of S. Typhimurium in the mouse model, and therefore, it appears that luxS does not play a significant role in regulating invasion of Salmonella in vitro or in vivo.

First structural characterization of Burkholderia vietnamiensis lipooligosaccharide from cystic fibrosis-associated lung transplantation strains.

This is the first structural elucidation of the lipooligosaccharide (LOS) endotoxin isolated from Burkholderia vietnamiensis, a clinically important member of Burkholderia cepacia complex, a group of over 10 opportunistic species that are highly problematic in cystic fibrosis. We have characterized a novel LOS structure extracted from two clonal strains of B. vietnamiensis isolated from a cystic fibrosis patient who underwent lung transplantation. Strains were selected from the pretransplantation and post-transplantation periods and endotoxin was extracted. Subsequent analysis interestingly revealed identical oligosaccharidic sequences, but variation in lipid A moieties. Further, both LOS fractions were tested for their immunostimulatory activity on human myelomonocytic U937 cells and for signaling on an HEK293 cell line stably expressing both TLR 4 and MD-2. We observed an increase in lipid A acylation and a resultant increase in biological activity in bio-reporter assays of TNF-alpha secretion in the post-transplantation strain.

LuxS affects flagellar phase variation independently of quorum sensing in Salmonella enterica serovar typhimurium.

LuxS catalyzes the synthesis of the quorum-sensing signaling molecule autoinducer 2. We show that in Salmonella enterica serovar Typhimurium, deletion of the luxS gene polarizes flagellar phase variation toward the more immunogenic phase 1 flagellin. This phenotype is complementable by luxS in trans but is independent of quorum-sensing signals.

The structure and proinflammatory activity of the lipopolysaccharide from Burkholderia multivorans and the differences between clonal strains colonizing pre and posttransplanted lungs.

The Burkholderia cepacia complex is a group of Gram-negative bacteria that are opportunistic pathogens for humans especially in cystic fibrosis patients. Lipopolysaccharide (LPS) molecules are potent virulence factors of Gram-negative bacteria organisms essential for bacterial survival. A complete analysis of the bacterial lipopolysaccharide structure to function relationship is required to understand the chemical basis of the inflammatory process. We have therefore investigated the structures of lipopolysaccharides from clonally identical Burkholderia multivorans strains (genomovar II) isolated pre- and post-lung transplantation through compositional analysis, mass spectrometry, and 2D NMR spectroscopy. We tested the LPS proinflammatory activity as a stimulant of human myelomonocytic U937 cell cytokine induction and assessed TLR4/MD2 signaling. Marked changes between the paired strains were found in the lipid A-inner core region. Such structural variations can contribute to the bacterial survival and persistence of infections despite the loss of a CF milieu following lung transplantation.

Adrenaline modulates the global transcriptional profile of Salmonella revealing a role in the antimicrobial peptide and oxidative stress resistance responses.

BACKGROUND: The successful interaction of bacterial pathogens with host tissues requires the sensing of specific chemical and physical cues. The human gut contains a huge number of neurons involved in the secretion and sensing of a class of neuroendocrine hormones called catecholamines. Recently, in Escherichia coli O157:H7, the catecholamines adrenaline and noradrenaline were shown to act synergistically with a bacterial quorum sensing molecule, autoinducer 3 (AI-3), to affect bacterial virulence and motility. We wished to investigate the impact of adrenaline on the biology of Salmonella spp. RESULTS: We have determined the effect of adrenaline on the transcriptome of the gut pathogen Salmonella enterica serovar Typhimurium. Addition of adrenaline led to an induction of key metal transport systems within 30 minutes of treatment. The oxidative stress responses employing manganese internalisation were also elicited. Cells lacking the key oxidative stress regulator OxyR showed reduced survival in the presence of adrenaline and complete restoration of growth upon addition of manganese. A significant reduction in the expression of the pmrHFIJKLM antimicrobial peptide resistance operon reduced the ability of Salmonella to survive polymyxin B following addition of adrenaline. Notably, both phenotypes were reversed by the addition of the beta-adrenergic blocker propranolol. Our data suggest that the BasSR two component signal transduction system is the likely adrenaline sensor mediating the antimicrobial peptide response. CONCLUSION: Salmonella are able to sense adrenaline and downregulate the antimicrobial peptide resistance pmr locus through the BasSR two component signalling system. Through iron transport, adrenaline may affect the oxidative stress balance of the cell requiring OxyR for normal growth. Both adrenaline effects can be inhibited by the addition of the beta-adrenergic blocker propranolol. Adrenaline sensing may provide an environmental cue for the induction of the Salmonella stress response in anticipation of imminent host-derived oxidative stress. However, adrenaline may also serve in favour of the host defences by lowering antimicrobial peptide resistance and hence documenting for the first time such a function for a hormone.

Genetic detoxification of an aroA Salmonella enterica serovar Typhimurium vaccine strain does not compromise protection against virulent Salmonella and enhances the immune responses towards a protective malarial antigen.

Live Salmonella vaccines are limited in use by the inherent toxicity of the lipopolysaccharide. The waaN gene encodes a myristyl transferase required for the secondary acylation of lipid A in lipopolysaccharide. A waaN mutant exhibits reduced induction of the inflammatory cytokines associated with lipopolysaccharide toxicity. Here the characteristics of a Salmonella enterica serovar Typhimurium aroA waaN mutant (SK100) in vitro and in vivo compared with its parent aroA strain (SL3261) were described. Phenotypic analysis of purified lipopolysaccharide obtained from SK100 confirmed that the physical and biological activities of the lipopolysaccharide had been altered. Nevertheless both strains had similar patterns of colonization and persistence in mice and significantly the aroA waaN mutant was equally as effective as the parent at protecting against challenge with wild-type S. Typhimurium. Furthermore, a SK100 strain was constructed expressing both tetanus toxin fragment C and the circumsporozoite protein of a malaria parasite. In marked contrast to its isogenic parent, the new attenuated strain induces significantly enhanced immune responses against the circumsporozoite protein. The waaN mutation enhances the ability of this strain to elicit immune responses towards guest antigens. This study provides important insights into the development of safe and effective multivalent Salmonella vaccines.

Dengue infection in patients presenting with neurological manifestations in a dengue endemic population.

The evaluation of the contribution of neurological dengue in suspected central nervous system (CNS) viral infections is essential to better understand the impact of neurological dengue on morbidity and mortality in dengue endemic regions such as Jamaica. For this study 401 cases of suspected viral CNS infections were investigated for evidence of dengue infection. The frequency of neurological dengue among these CNS cases was found to be 13.5% (54/401). Fifty-three cases were confirmed serologically by haemagglutination inhibition assay (HI) and IgM antibody (ELISA) and the virus was isolated in one case only. Clinical manifestations among dengue positive CNS cases included encephalitis in 51.8% (28/54), meningitis in 33.3% (18/54), seizures in 11.1% (6/54) and acute flaccid paralysis/Guillain-Barré syndrome in 3.7% (2/54). The clinical diagnosis of dengue neurological infection corresponded with laboratory confirmation in 22.2% (12/54) of cases only. Deaths occurred in 3.7% (2/54) of cases and were associated with patients with dengue neurological infection. The high risk of dengue among patients with suspected viral CNS infections in this study supports the need for an increased index of suspicion of dengue in patients presenting with neurological manifestations in dengue endemic countries.

Molecular analysis and implications of neurovirulent circulating vaccine-derived poliovirus in Jamaica. A case report and review of literature.

As the goal to eradicate wild polio virus (WPV) is approached, outbreaks associated with vaccine derived polioviruses (VDPV) with neurovirulent properties have emerged. The relevance for the spread of infection by nonparalytic cVDPV cases, with mutations associated with neurovirulence, is discussed with reference to the molecular analysis of a VDPV isolated from a Jamaican child who presented with aseptic meningitis. Potential risks to the Jamaican community resulting from circulation of cVDPV and critical factors defined by the World Health Organization (WHO) in the global eradication of Polio are analyzed in the context of immunization coverage, and the need to stop all Oral Polio Vaccine (OPV) use once wild polioviruses (WPVs) have been eradicated.

Structural and antigenic polymorphism of the 35- to 48-kilodalton merozoite surface antigen (MSA-2) of the malaria parasite Plasmodium falciparum.

Merozoite surface antigen MSA-2 of the human parasite Plasmodium falciparum is being considered for the development of a malaria vaccine. The antigen is polymorphic, and specific monoclonal antibodies differentiate five serological variants of MSA-2 among 25 parasite isolates. The variants are grouped into two major serogroups, A and B. Genes encoding two different variants from serogroup A have been sequenced, and their DNA together with deduced amino acid sequences were compared with sequences encoded by other alleles. The comparison shows that the serological classification reflects differences in DNA sequences and deduced primary structure of MSA-2 variants and serogroups. Thus, the overall homologies of DNA and amino acid sequences are over 95% among variants in the same serogroup. In contrast, similarities between the group A variants and a group B variant are only 70 and 64% for DNA and amino acid sequences, respectively. We propose that the MSA-2 protein is encoded by two highly divergent groups of alleles, with limited additional polymorphism displayed within each group.