Characterization of a Novel Mycobacterium Tuberculosis-Simian Immunodeficiency Virus Vaccine to Prevent Oral Pediatric HIV Transmission

Over 3.3 million children are living with HIV, infected primarily by mother-to-child transmission (MTCT). Breast milk exposure of HIV accounts for up to 44% of MTCT events. Despite serious efforts to prevent vertical HIV transmission, infant testing is often delayed and access to antiretroviral therapies is still limited. Compared to adults, HIV-infected infants experience enhanced disease progression and more severe co-morbidities with pathogens like Mycobacterium tuberculosis (Mtb). The live attenuated BCG vaccine is the only licensed tuberculosis vaccine but BCG can disseminate in immunosuppressed, HIV-infected infants. Considering the significant geographical overlap of the HIV and TB epidemics and the high vulnerability of infants to both pathogens, a pediatric vaccine to safely protect against HIV and Mtb is urgently needed. We hypothesized that human-adapted attenuated Mtb strains engineered to co-express HIV genes (rAMtb-HIV) would safely induce the development of dually-immunogenic HIV- and Mtb-specific cellular and humoral responses. Three distinct attenuated Mtb-SIV strains were evaluated in the neonatal rhesus macaque model. Oral priming with strain mc26435 plus systemic MVA-SIV boosts successfully induced the development of Mtb- and SIV-specific cellular and humoral responses and was identified to be safe in healthy and immunosuppressed, SIV-infected neonatal macaques. However, despite vaccine-induced immunity, vaccination did not protect infants against low-dose oral SIV challenges designed to mimic oral MTCT during breastfeeding. Two important results emerged from the challenge study. First, higher Env-specific mucosal IgA activities and plasma IgG avidities positively correlated with controlled viremia in a subset of vaccinated animals. These animals also maintained peripheral CD4+ T cell populations and IL-17-expressing lymphocytes in the intestinal mucosa. However, the majority of vaccinated animals required fewer low-dose SIV exposures to become infected than unvaccinated animals. Enhanced viral acquisition was associated with vaccine-induced persistent immune activation. At the time of challenge, CCR5-expressing CD4+ T cells were observed with greater frequencies in blood, oral and intestinal tissues in vaccinated animals only, providing increased frequencies of SIV target cells. Due to the potential impact of these data on BCG vaccine safety and pediatric HIV and TB vaccine development, additional studies are required to confirm these complex and intriguing results.Doctor of Philosoph

Early parafoveal semantic integration in natural reading

Humans can read and comprehend text rapidly, implying that readers might process multiple words per fixation. However, the extent to which parafoveal words are previewed and integrated into the evolving sentence context remains disputed. We investigated parafoveal processing during natural reading by recording brain activity and eye movements using MEG and an eye tracker while participants silently read one-line sentences. The sentences contained an unpredictable target word that was either congruent or incongruent with the sentence context. To measure parafoveal processing, we flickered the target words at 60 Hz and measured the resulting brain responses (i.e., Rapid Invisible Frequency Tagging, RIFT) during fixations on the pre-target words. Our results revealed a significantly weaker tagging response for target words that were incongruent with the sentence context compared to congruent ones, even within 100 ms of fixating the word immediately preceding the target. This reduction in the RIFT response was also found to be predictive of individual reading speed. We conclude that semantic information is not only extracted from the parafovea but can also be integrated with the sentence context before the word is fixated. This early and extensive parafoveal processing supports the rapid word processing required for natural reading. Our study suggests that theoretical frameworks of natural reading should incorporate the concept of deep parafoveal processing

Permissivity of Dipeptidyl Peptidase 4 Orthologs to Middle East Respiratory Syndrome Coronavirus Is Governed by Glycosylation and Other Complex Determinants

ABSTRACT Middle East respiratory syndrome coronavirus (MERS-CoV) utilizes dipeptidyl peptidase 4 (DPP4) as an entry receptor. While bat, camel, and human DPP4 support MERS-CoV infection, several DPP4 orthologs, including mouse, ferret, hamster, and guinea pig DPP4, do not. Previous work revealed that glycosylation of mouse DPP4 plays a role in blocking MERS-CoV infection. Here, we tested whether glycosylation also acts as a determinant of permissivity for ferret, hamster, and guinea pig DPP4. We found that, while glycosylation plays an important role in these orthologs, additional sequence and structural determinants impact their ability to act as functional receptors for MERS-CoV. These results provide insight into DPP4 species-specific differences impacting MERS-CoV host range and better inform our understanding of virus-receptor interactions associated with disease emergence and host susceptibility. IMPORTANCE MERS-CoV is a recently emerged zoonotic virus that is still circulating in the human population with an ∼35% mortality rate. With no available vaccines or therapeutics, the study of MERS-CoV pathogenesis is crucial for its control and prevention. However, in vivo studies are limited because MERS-CoV cannot infect wild-type mice due to incompatibilities between the virus spike and the mouse host cell receptor, mouse DPP4 (mDPP4). Specifically, mDPP4 has a nonconserved glycosylation site that acts as a barrier to MERS-CoV infection. Thus, one mouse model strategy has been to modify the mouse genome to remove this glycosylation site. Here, we investigated whether glycosylation acts as a barrier to infection for other nonpermissive small-animal species, namely, ferret, guinea pig, and hamster. Understanding the virus-receptor interactions for these DPP4 orthologs will help in the development of additional animal models while also revealing species-specific differences impacting MERS-CoV host range

Functional gene analysis of individual response to challenge of SIVmac239 in M. mulatta PBMC culture

AbstractIt has previously been shown in macaques that individual animals exhibit varying responses to challenge with the same strain of SIV. We attempted to elucidate these differences using functional genomics and correlate them to biological response. Unfractionated PBMC from three rhesus macaques were isolated, activated, and infected with SIVmac239. Interestingly, one of the three animals used for these experiments exhibited a completely unique response to infection relative to the other two. After repeated attempts to infect the PBMC from this animal, little or no infectivity was seen across the time points considered, and corresponding to this apparent lack of infection, few genes were seen to be differentially expressed when compared to mock-infected cells. For the remaining two animals, gene expression analysis showed that while they exhibited responses for the same groups of pathways, these responses included differences specific to the individual animal at the gene level. In instances where the patterns of differential gene expression differed between these animals, the genes being differentially expressed were associated with the same categories of biological process, mainly immune response and cell signaling. At the pathway level, these animals again exhibited similar responses that could be predicted based on the experimental conditions. Even in these expected results, the degree of response and the specific genes being regulated differed greatly from animal to animal. The differences in gene expression on an individual level have the potential to be used as markers in identification of animals suitable for lentiviral infection experiments. Our results highlight the importance of individual variation in response to viral challenge

Increasing the translation of mouse models of MERS coronavirus pathogenesis through kinetic hematological analysis

Newly emerging viral pathogens pose a constant and unpredictable threat to human and animal health. Coronaviruses (CoVs) have a penchant for sudden emergence, as evidenced by severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome CoV (MERS-CoV) and most recently, swine acute diarrhea syndrome coronavirus (SADS-CoV). Small animal models of emerging viral pathogenesis are crucial to better understand the virus and host factors driving disease progression. However, rodent models are often criticized for their limited translatability to humans. The complete blood count is the most ordered clinical test in the United States serving as the cornerstone of clinical medicine and differential diagnosis. We recently generated a mouse model for MERS-CoV pathogenesis through the humanization of the orthologous entry receptor dipeptidyl peptidase 4 (DPP4). To increase the translatability of this model, we validated and established the use of an automated veterinary hematology analyzer (VetScan HM5) at biosafety level 3 for analysis of peripheral blood. MERS-CoV lung titer peaked 2 days post infection concurrent with lymphopenia and neutrophilia in peripheral blood, two phenomena also observed in MERS-CoV infection of humans. The fluctuations in leukocyte populations measured by Vetscan HM5 were corroborated by standard flow cytometry, thus confirming the utility of this approach. Comparing a sublethal and lethal dose of MERS-CoV in mice, analysis of daily blood draws demonstrates a dose dependent modulation of leukocytes. Major leukocyte populations were modulated before weight loss was observed. Importantly, neutrophil counts on 1dpi were predictive of disease severity with a lethal dose of MERS-CoV highlighting the predictive value of hematology in this model. Taken together, the inclusion of hematological measures in mouse models of emerging viral pathogenesis increases their translatability and should elevate the preclinical evaluation of MERS-CoV therapeutics and vaccines to better mirror the complexity of the human condition

A mouse model for MERS coronavirus-induced acute respiratory distress syndrome

Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel virus that emerged in 2012, causing acute respiratory distress syndrome (ARDS), severe pneumonia-like symptoms, and multi-organ failure, with a case fatality rate of ~36%. Limited clinical studies indicate that humans infected with MERS-CoV exhibited pathology consistent with late stages of ARDS, which is reminiscent of disease observed in patients infected with SARS coronavirus. Models of MERS-CoV-induced severe respiratory disease have been difficult to achieve, and small animal models traditionally used to investigate viral pathogenesis (mouse, hamster, guinea pig, and ferret) are naturally resistant to MERS-CoV. Therefore, we used CRISPR/Cas9 to modify the mouse genome to encode two human amino acids (288 and 330) in the dipeptidyl peptidase 4 receptor, making mice susceptible to MERS-CoV replication. Serial MERS-CoV passage in these engineered mice was then used to generate a mouse-adapted virus that replicated efficiently within the lungs, and evoked symptoms indicative of severe acute respiratory distress syndrome (ARDS), including decreased survival, extreme weight loss, decreased pulmonary function, pulmonary hemorrhage, and pathological signs indicative of end stage lung disease. Importantly, therapeutic countermeasures comprising MERS-CoV neutralizing antibody treatment or a MERS-CoV spike protein vaccine protected engineered mice against MERS-CoV-induced ARDS

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Horizontal ducting of sound by curved nonlinear internal gravity waves in the continental shelf areas

Author Posting. © Acoustical Society of America, 2013. This article is posted here by permission of Acoustical Society of America for personal use, not for redistribution. The definitive version was published in Journal of the Acoustical Society of America 133 (2013): 37-49, doi:10.1121/1.4770240.The acoustic ducting effect by curved nonlinear gravity waves in shallow water is studied through idealized models in this paper. The internal wave ducts are three-dimensional, bounded vertically by the sea surface and bottom, and horizontally by aligned wavefronts. Both normal mode and parabolic equation methods are taken to analyze the ducted sound field. Two types of horizontal acoustic modes can be found in the curved internal wave duct. One is a whispering-gallery type formed by the sound energy trapped along the outer and concave boundary of the duct, and the other is a fully bouncing type due to continual reflections from boundaries in the duct. The ducting condition depends on both internal-wave and acoustic-source parameters, and a parametric study is conducted to derive a general pattern. The parabolic equation method provides full-field modeling of the sound field, so it includes other acoustic effects caused by internal waves, such as mode coupling/scattering and horizontal Lloyd's mirror interference. Two examples are provided to present internal wave ducts with constant curvature and meandering wavefronts.This work was sponsored by the Office of Naval Research under grants N00014-10-1-0040 and N00014-11- 1-0701

Vaccine-Elicited Mucosal and Systemic Antibody Responses Are Associated with Reduced Simian Immunodeficiency Viremia in Infant Rhesus Macaques

ABSTRACT Despite significant progress in reducing peripartum mother-to-child transmission (MTCT) of human immunodeficiency virus (HIV) with antiretroviral therapy (ART), continued access to ART throughout the breastfeeding period is still a limiting factor, and breast milk exposure to HIV accounts for up to 44% of MTCT. As abstinence from breastfeeding is not recommended, alternative means are needed to prevent MTCT of HIV. We have previously shown that oral vaccination at birth with live attenuated Mycobacterium tuberculosis strains expressing simian immunodeficiency virus (SIV) genes safely induces persistent SIV-specific cellular and humoral immune responses both systemically and at the oral and intestinal mucosa. Here, we tested the ability of oral M. tuberculosis vaccine strains expressing SIV Env and Gag proteins, followed by systemic heterologous (MVA-SIV Env/Gag/Pol) boosting, to protect neonatal macaques against oral SIV challenge. While vaccination did not protect infant macaques against oral SIV acquisition, a subset of immunized animals had significantly lower peak viremia which inversely correlated with prechallenge SIV Env-specific salivary and intestinal IgA responses and higher-avidity SIV Env-specific IgG in plasma. These controller animals also maintained CD4 + T cell populations better and showed reduced tissue pathology compared to noncontroller animals. We show that infants vaccinated at birth can develop vaccine-induced SIV-specific IgA and IgG antibodies and cellular immune responses within weeks of life. Our data further suggest that affinity maturation of vaccine-induced plasma antibodies and induction of mucosal IgA responses at potential SIV entry sites are associated with better control of viral replication, thereby likely reducing SIV morbidity. IMPORTANCE Despite significant progress in reducing peripartum MTCT of HIV with ART, continued access to ART throughout the breastfeeding period is still a limiting factor. Breast milk exposure to HIV accounts for up to 44% of MTCT. Alternative measures, in addition to ART, are needed to achieve the goal of an AIDS-free generation. Pediatric HIV vaccines constitute a core component of such efforts. The results of our pediatric vaccine study highlight the potential importance of vaccine-elicited mucosal Env-specific IgA responses in combination with high-avidity systemic Env-specific IgG in protection against oral SIV transmission and control of viral replication in infant macaques. The induction of potent mucosal IgA antibodies by our vaccine is remarkable considering the age-dependent development of mucosal IgA responses postbirth. A deeper understanding of postnatal immune development may inform the design of improved vaccine strategies to enhance systemic and mucosal SIV/HIV antibody responses

Realising advanced risk-based port state control inspection using data-driven Bayesian networks

In the past decades, maritime transportation not only contributes to economic prosperity, but also renders many threats to the industry, causing huge casualties and losses. As a result, various maritime safety measures have been developed, including Port State Control (PSC) inspections. In this paper, we propose a data-driven Bayesian Network (BN) based approach to analyse risk factors influencing PSC inspections, and predict the probability of vessel detention. To do so, inspection data of bulk carriers in seven major European countries from 2005 to 2008 1 in Paris MoU is collected to identify the relevant risk factors. Meanwhile, the network structure is constructed via TAN learning and subsequently validated by sensitivity analysis. The results reveal two conclusions: first, the key risk factors influencing PSC inspections include number of deficiencies, type of inspection, Recognised Organisation (RO) and vessel age. Second, the model exploits a novel way to predict the detention probabilities under different situations, which effectively help port authorities to rationalise their inspection regulations as well as allocation of the resources. Further effort will be made to conduct contrastive analysis between ‘Pre-NIR’ period and ‘Post-NIR’ period to test the impact of NIR started in 2008. © 2018 Elsevier Lt