Paenibacillus infection with frequent viral coinfection contributes to postinfectious hydrocephalus in Ugandan infants

Postinfectious hydrocephalus (PIH), which often follows neonatal sepsis, is the most common cause of pediatric hydrocephalus worldwide, yet the microbial pathogens underlying this disease remain to be elucidated. Characterization of the microbial agents causing PIH would enable a shift from surgical palliation of cerebrospinal fluid (CSF) accumulation to prevention of the disease. Here, we examined blood and CSF samples collected from 100 consecutive infant cases of PIH and control cases comprising infants with non-postinfectious hydrocephalus in Uganda. Genomic sequencing of samples was undertaken to test for bacterial, fungal, and parasitic DNA; DNA and RNA sequencing was used to identify viruses; and bacterial culture recovery was used to identify potential causative organisms. We found that infection with the bacterium Paenibacillus, together with frequent cytomegalovirus (CMV) coinfection, was associated with PIH in our infant cohort. Assembly of the genome of a facultative anaerobic bacterial isolate recovered from cultures of CSF samples from PIH cases identified a strain of Paenibacillus thiaminolyticus. This strain, designated Mbale, was lethal when injected into mice in contrast to the benign reference Paenibacillus strain. These findings show that an unbiased pan-microbial approach enabled characterization of Paenibacillus in CSF samples from PIH cases, and point toward a pathway of more optimal treatment and prevention for PIH and other proximate neonatal infections

The Role of Capping Protein in Migrating Cells and Neurons

Actin capping protein (CP) binds to the barbed ends of actin filaments and inhibits further polymerization. CP is thought to be essential for cell motility, but its role in mammalian migration has not been directly tested. Moreover, CP is widely believed to be absent from filopodia, and a role for CP in filopodia has remained uninvestigated. I have explored the role of CP in migrating cells and in primary hippocampal neurons. I begin this dissertation by reviewing the relevant literature on CP and on filopodia (Chapter One). I then investigate the effects of silencing CP in migrating B16F10 melanoma cells (Chapter Two). I find that depleting CP impairs cell migration. Moreover, CP is unexpectedly detected in filopodia, and CP depletion has dramatic effects on filopodial morphology and dynamics.These effects may be mediated by unchecked actin polymerization resulting from severely reduced capping activity and consequent depletion of monomeric actin. My novel findings suggest that CP may be an important player in filopodial form and function. I also report that silencing CP has dramatic effects on neuronal development (Chapter Three). CP depletion in primary hippocampal neurons accelerates neuronal maturation in vitro without affecting neurite length or number of Stage III neurons; these effects may possibly be mediated by changes in filopodial number. Finally, I outline the outstanding questions raised by my results in Chapters Two and Three and discuss future experiments that could help to address these questions (Chapter Four

The Role of Capping Protein in Migrating Cells and Neurons

Actin capping protein (CP) binds to the barbed ends of actin filaments and inhibits further polymerization. CP is thought to be essential for cell motility, but its role in mammalian migration has not been directly tested. Moreover, CP is widely believed to be absent from filopodia, and a role for CP in filopodia has remained uninvestigated. I have explored the role of CP in migrating cells and in primary hippocampal neurons. I begin this dissertation by reviewing the relevant literature on CP and on filopodia (Chapter One). I then investigate the effects of silencing CP in migrating B16F10 melanoma cells (Chapter Two). I find that depleting CP impairs cell migration. Moreover, CP is unexpectedly detected in filopodia, and CP depletion has dramatic effects on filopodial morphology and dynamics.These effects may be mediated by unchecked actin polymerization resulting from severely reduced capping activity and consequent depletion of monomeric actin. My novel findings suggest that CP may be an important player in filopodial form and function. I also report that silencing CP has dramatic effects on neuronal development (Chapter Three). CP depletion in primary hippocampal neurons accelerates neuronal maturation in vitro without affecting neurite length or number of Stage III neurons; these effects may possibly be mediated by changes in filopodial number. Finally, I outline the outstanding questions raised by my results in Chapters Two and Three and discuss future experiments that could help to address these questions (Chapter Four

Altered Testicular Gene Expression Patterns in Mice Lacking the Polyubiquitin Gene Ubb

Ubiquitin (Ub) is an essential protein found in all eukaryotic cells and plays important roles in a variety of cellular functions including germ cell development. We have previously reported that targeted disruption of the polyubiquitin gene Ubb results in male and female infertility in Ubb −/− mice, with germ cells arrested at meiotic prophase I. Although reduced Ub levels in germ cells are believed to be responsible for the fertility defect in Ubb −/− mice, it is still unclear how reduced Ub levels result in sterility. Here we describe the results of a microarray analysis of the murine testicular transcriptome, which demonstrates dramatically altered gene expression patterns in Ubb −/− mice, possibly related to reduced levels of histone 2A (H2A) ubiquitylation. We find that large numbers of genes related to fertility, metabolism, transcription, and the ubiquitin–proteasome system (UPS) are misregulated in Ubb −/− mice. Such wide-ranging alterations in gene expression suggest that loss of the Ubb gene does not mimic a single-gene defect phenotype, but instead may affect gene expression more globally. These dramatic changes in gene expression could, at least in part, contribute to the complex fertility and metabolic phenotypes seen in these mice

The Mouse Polyubiquitin Gene Ubb Is Essential for Meiotic Progression▿ †

Ubiquitin is encoded in mice by two polyubiquitin genes, Ubb and Ubc, that are considered to be stress inducible and two constitutively expressed monoubiquitin (Uba) genes. Here we report that targeted disruption of Ubb results in male and female infertility due to failure of germ cells to progress through meiosis I and hypogonadism. In the absence of Ubb, spermatocytes and oocytes arrest during meiotic prophase, before metaphase of the first meiotic division. Although cellular ubiquitin levels are believed to be maintained by a combination of functional redundancy among the four ubiquitin genes, stress inducibility of the two polyubiquitin genes, and ubiquitin recycling by proteasome-associated isopeptidases, our results indicate that ubiquitin is required for and consumed during meiotic progression. The striking similarity of the meiotic phenotype in Ubb−/− germ cells to the sporulation defect in fission yeast (Schizosaccharomyces pombe) lacking a polyubiquitin gene suggests that a meiotic role of the polyubiquitin gene has been conserved throughout eukaryotic evolution

Pan-African evolution of within- and between-country COVID-19 dynamics

The coronavirus disease 2019 (COVID-19) pandemic is heterogeneous throughout Africa and threatening millions of lives. Surveillance and short-term modeling forecasts are critical to provide timely information for decisions on control strategies. We created a strategy that helps predict the country-level case occurrences based on cases within or external to a country throughout the entire African continent, parameterized by socioeconomic and geoeconomic variations and the lagged effects of social policy and meteorological history. We observed the effect of the Human Development Index, containment policies, testing capacity, specific humidity, temperature, and landlocked status of countries on the local within-country and external between-country transmission. One-week forecasts of case numbers from the model were driven by the quality of the reported data. Seeking equitable behavioral and social interventions, balanced with coordinated country-specific strategies in infection suppression, should be a continental priority to control the COVID-19 pandemic in Africa

Vaginal microbiome topic modeling of laboring Ugandan women with and without fever

The composition of the maternal vaginal microbiome influences the duration of pregnancy, onset of labor, and even neonatal outcomes. Maternal microbiome research in sub-Saharan Africa has focused on non-pregnant and postpartum composition of the vaginal microbiome. Here we aimed to illustrate the relationship between the vaginal microbiome of 99 laboring Ugandan women and intrapartum fever using routine microbiology and 16S ribosomal RNA gene sequencing from two hypervariable regions (V1–V2 and V3–V4). To describe the vaginal microbes associated with vaginal microbial communities, we pursued two approaches: hierarchical clustering methods and a novel Grades of Membership (GoM) modeling approach for vaginal microbiome characterization. Leveraging GoM models, we created a basis composed of a preassigned number of microbial topics whose linear combination optimally represents each patient yielding more comprehensive associations and characterization between maternal clinical features and the microbial communities. Using a random forest model, we showed that by including microbial topic models we improved upon clinical variables to predict maternal fever. Overall, we found a higher prevalence of Granulicatella, Streptococcus, Fusobacterium, Anaerococcus, Sneathia, Clostridium, Gemella, Mobiluncus, and Veillonella genera in febrile mothers, and higher prevalence of Lactobacillus genera (in particular L. crispatus and L. jensenii), Acinobacter, Aerococcus, and Prevotella species in afebrile mothers. By including clinical variables with microbial topics in this model, we observed young maternal age, fever reported earlier in the pregnancy, longer labor duration, and microbial communities with reduced Lactobacillus diversity were associated with intrapartum fever. These results better defined relationships between the presence or absence of intrapartum fever, demographics, peripartum course, and vaginal microbial topics, and expanded our understanding of the impact of the microbiome on maternal and potentially neonatal outcome risk

Cytomegalovirus Infections in Ugandan Infants:Newborn-Mother Pairs, Neonates with Sepsis, and Infants with Hydrocephalus

Objective To estimate the prevalence of cytomegalovirus (CMV) infections in newborn-mother pairs, neonates with sepsis, and infants with hydrocephalus in Uganda. Design and Methods Three populations: (1) newborn-mother pairs, (2) neonates with sepsis, and (3) infants (≤ 3 months) with non-postinfectious (NPIH) or postinfectious (PIH) hydrocephalus, were evaluated for CMV infection at three medical centers in Uganda. Quantitative PCR (qPCR) was used to characterize the prevalence of CMV. Results The overall CMV prevalence in 2498 samples in duplicate across all groups was 9%. In newborn-mother pairs, there was a 3% prevalence of cord blood CMV positivity and 33% prevalence of maternal vaginal shedding. In neonates with clinical sepsis there was a 2% CMV prevalence. Maternal HIV seropositivity (aOR, 25.20; 95% CI, 4.43-134.26; p= 0.0001), residence in Eastern Uganda (aOR, 11.06; 95% CI, 2.30-76.18; p=0.003), maternal age < 25 years (aOR, 4.54; 95% CI, 1.40-19.29; p=0.02), and increasing neonatal age (aOR, 1.08 for each day older; 95% CI, 1.00-1.16; p= 0.05), were associated risk factors for CMV in neonates with clinical sepsis. We found a two-fold higher maternal vaginal shedding in Eastern (45%) vs Western (22%) Uganda during parturition (n=22/49 vs. 11/50, Fisher's exact test, p=0.02). In infants with PIH, the prevalence in blood was 24% and in infants with NPIH it was 20%. CMV was present in the CSF of 13% of infants with PIH compared to 0.5% of infants with NPIH (n=26/205 vs. 1/194, p<0.0001). Conclusion Our findings highlight that congenital and postnatal CMV prevalence is substantial in this African setting and the long-term consequences are uncharacterized