Transmission from vaccinated individuals in a large SARS-CoV-2 Delta variant outbreak.

An outbreak of over 1,000 COVID-19 cases in Provincetown, Massachusetts (MA), in July 2021-the first large outbreak mostly in vaccinated individuals in the US-prompted a comprehensive public health response, motivating changes to national masking recommendations and raising questions about infection and transmission among vaccinated individuals. To address these questions, we combined viral genomic and epidemiological data from 467 individuals, including 40% of outbreak-associated cases. The Delta variant accounted for 99% of cases in this dataset; it was introduced from at least 40 sources, but 83% of cases derived from a single source, likely through transmission across multiple settings over a short time rather than a single event. Genomic and epidemiological data supported multiple transmissions of Delta from and between fully vaccinated individuals. However, despite its magnitude, the outbreak had limited onward impact in MA and the US overall, likely due to high vaccination rates and a robust public health response.

Virus-derived gene transfer agents benefit host cells by providing templates for DNA repair.

The potential benefits of producing gene transfer agents (GTAs) have long been speculated. A new study in PLOS Biology shows that DNA transfer by these phage-like elements allows cells with DNA damage to perform DNA repair and survive.

Anthropogenic contamination sources drive differences in antimicrobial-resistant Escherichia coli in three urban lakes.

Antimicrobial resistance (AMR) is an ever-present threat to the treatment of infectious diseases. However, the potential relevance of this phenomenon in environmental reservoirs still raises many questions. Detection of antimicrobial-resistant bacteria in the environment is a critical aspect for understanding the prevalence of resistance outside of clinical settings, as detection in the environment indicates that resistance is likely already widespread. We isolated antimicrobial-resistant Escherichia coli from three urban waterbodies over a 15-month time series, determined their antimicrobial susceptibilities, investigated their population structure, and identified genetic determinants of resistance. We found that E. coli populations at each site were composed of different dominant phylotypes and showed distinct patterns of antimicrobial and multidrug resistance, despite close geographic proximity. Many strains that were genome-sequenced belonged to sequence types of international concern, particularly the ST131 clonal complex. We found widespread resistance to clinically important antimicrobials such as amoxicillin, cefotaxime, and ciprofloxacin, but found that all strains were susceptible to amikacin and the last-line antimicrobials meropenem and fosfomycin. Resistance was most often due to acquirable antimicrobial resistance genes, while chromosomal mutations in gyrA, parC, and parE conferred resistance to quinolones. Whole-genome analysis of a subset of strains further revealed the diversity of the population of E. coli present, with a wide array of AMR and virulence genes identified, many of which were present on the chromosome, including blaCTX-M. Finally, we determined that environmental persistence, transmission between sites, most likely mediated by wild birds, and transfer of mobile genetic elements likely contributed significantly to the patterns observed.IMPORTANCEA One Health perspective is crucial to understand the extent of antimicrobial resistance (AMR) globally, and investigation of AMR in the environment has been increasing in recent years. However, most studies have focused on waterways that are directly polluted by sewage, industrial manufacturing, or agricultural activities. Therefore, there remains a lack of knowledge about more natural, less overtly impacted environments. Through phenotypic and genotypic investigation of AMR in Escherichia coli, this study adds to our understanding of the extent and patterns of resistance in these types of environments, including over a time series, and showed that complex biotic and abiotic factors contribute to the patterns observed. Our study further emphasizes the importance of incorporating the surveillance of microbes in freshwater environments in order to better comprehend potential risks for both human and animal health and how the environment may serve as a sentinel for potential future clinical infections.

Patterns of abundance, chromosomal localization, and domain organization among c-di-GMP-metabolizing genes revealed by comparative genomics of five alphaproteobacterial orders.

BACKGROUND: Bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP) is a bacterial second messenger that affects diverse processes in different bacteria, including the cell cycle, motility, and biofilm formation. Its cellular levels are controlled by the opposing activities of two types of enzymes, with synthesis by diguanylate cyclases containing a GGDEF domain and degradation by phosphodiesterases containing either an HD-GYP or an EAL domain. These enzymes are ubiquitous in bacteria with up to 50 encoded in some genomes, the specific functions of which are mostly unknown. RESULTS: We used comparative analyses to identify genomic patterns among genes encoding proteins with GGDEF, EAL, and HD-GYP domains in five orders of the class Alphaproteobacteria. GGDEF-containing sequences and GGDEF-EAL hybrids were the most abundant and had the highest diversity of co-occurring auxiliary domains while EAL and HD-GYP containing sequences were less abundant and less diverse with respect to auxiliary domains. There were striking patterns in the chromosomal localizations of the genes found in two of the orders. The Rhodobacterales' EAL-encoding genes and Rhizobiales' GGDEF-EAL-encoding genes showed opposing patterns of distribution compared to the GGDEF-encoding genes. In the Rhodobacterales, the GGDEF-encoding genes showed a tri-modal distribution with peaks mid-way between the origin (ori) and terminus (ter) of replication and at ter while the EAL-encoding genes peaked near ori. The patterns were more complex in the Rhizobiales, but the GGDEF-encoding genes were biased for localization near ter. CONCLUSIONS: The observed patterns in the chromosomal localizations of these genes suggest a coupling of synthesis and hydrolysis of c-di-GMP with the cell cycle. Moreover, the higher proportions and diversities of auxiliary domains associated with GGDEF domains and GGDEF-EAL hybrids compared to EAL or HD-GYP domains could indicate that more stimuli affect synthesis compared to hydrolysis of c-di-GMP.

Prolactin and prolactin receptor expression in the HPG axis and crop during parental care in both sexes of a biparental bird (Columba livia).

During breeding, multiple circulating hormones, including prolactin, facilitate reproductive transitions in species that exhibit parental care. Prolactin underlies parental behaviors and related physiological changes across many vertebrates, including birds and mammals. While circulating prolactin levels often fluctuate across breeding, less is known about how relevant target tissues vary in their prolactin responsiveness via prolactin receptor (PRLR) expression. Recent studies have also investigated prolactin (PRL) gene expression outside of the pituitary (i.e., extra-pituitary PRL), but how PRL gene expression varies during parental care in non-pituitary tissue (e.g., hypothalamus, gonads) remains largely unknown. Further, it is unclear if and how tissue-specific PRL and PRLR vary between the sexes during biparental care. To address this, we measured PRL and PRLR gene expression in tissues relevant to parental care, the endocrine reproductive hypothalamic-pituitary- gonadal (HPG) axis and the crop (a tissue with a similar function as the mammalian mammary gland), across various reproductive stages in both sexes of a biparental bird, the rock dove (Columba livia). We also assessed how these genes responded to changes in offspring presence by adding chicks mid-incubation, simulating an early hatch when prolactin levels were still moderately low. We found that pituitary PRL expression showed similar increases as plasma prolactin levels, and detected extra-pituitary PRL in the hypothalamus, gonads and crop. Hypothalamic and gonadal PRLR expression also changed as birds began incubation. Crop PRLR expression correlated with plasma prolactin, peaking when chicks hatched. In response to replacing eggs with a novel chick mid-incubation, hypothalamic and gonadal PRL and PRLR gene expression differed significantly compared to mid-incubation controls, even when plasma prolactin levels did not differ. We also found sex differences in PRL and PRLR that suggest gene expression may allow males to compensate for lower levels in prolactin by upregulating PRLR in all tissues. Overall, this study advances our understanding of how tissue-specific changes in responsiveness to parental hormones may differ across key reproductive transitions, in response to offspring cues, and between the sexes.

A SARS-CoV-2 Cluster in an Acute Care Hospital.

BACKGROUND: Little is known about clusters of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in acute care hospitals. OBJECTIVE: To describe the detection, mitigation, and analysis of a large cluster of SARS-CoV-2 infections in an acute care hospital with mature infection control policies. DESIGN: Descriptive study. SETTING: Brigham and Women's Hospital, Boston, Massachusetts. PARTICIPANTS: Patients and staff with cluster-related SARS-CoV-2 infections. INTERVENTION: Close contacts of infected patients and staff were identified and tested every 3 days, patients on affected units were preemptively isolated and repeatedly tested, affected units were cleaned, room ventilation was measured, and specimens were sent for whole-genome sequencing. A case-control study was done to compare clinical interactions, personal protective equipment use, and breakroom and workroom practices in SARS-CoV-2-positive versus negative staff. MEASUREMENTS: Description of the cluster, mitigation activities, and risk factor analysis. RESULTS: Fourteen patients and 38 staff members were included in the cluster per whole-genome sequencing and epidemiologic associations. The index case was a symptomatic patient in whom isolation was discontinued after 2 negative results on nasopharyngeal polymerase chain reaction testing. The patient subsequently infected multiple roommates and staff, who then infected others. Seven of 52 (13%) secondary infections were detected only on second or subsequent tests. Eight of 9 (89%) patients who shared rooms with potentially contagious patients became infected. Potential contributing factors included high viral loads, nebulization, and positive pressure in the index patient's room. Risk factors for transmission to staff included presence during nebulization, caring for patients with dyspnea or cough, lack of eye protection, at least 15 minutes of exposure to case patients, and interactions with SARS-CoV-2-positive staff in clinical areas. Whole-genome sequencing confirmed that 2 staff members were infected despite wearing surgical masks and eye protection. LIMITATION: Findings may not be generalizable. CONCLUSION: SARS-CoV-2 clusters can occur in hospitals despite robust infection control policies. Insights from this cluster may inform additional measures to protect patients and staff. PRIMARY FUNDING SOURCE: None.

Novel Divergent Members of the Kitrinoviricota Discovered through Metagenomics in the Intestinal Contents of Red-Backed Voles (Clethrionomys gapperi).

Metagenomic methods are powerful tools to investigate viral diversity in biological or environmental samples and to identify previously unknown viruses. We used RNA metagenomics to identify, in the gut of red-backed voles, the nearly complete genomes of two novel members of the Kitrinoviricota, a phylum including viruses with positive-sense ssRNA genomes encoding an RNA-directed RNA polymerase. The genome of a novel member of the Tombusviridae presented four open reading frames (ORFs); a -1 frameshift is potentially involved in generating the viral replicase. This sequence was part of a phylogenetic clade that did not include any officially classified species. The second genome presented a large ORF coding for a viral polyprotein containing the typical protein domains common to flexiviruses. The sequence clustered with currently known members of the Deltaflexiviridae. Both viruses appear to represent the first members of novel species in yet undefined genera. The identified viruses likely originated from the vole diet as members of the two viral families are known to infect plants and fungi, respectively. Investigating public databases demonstrated that a much higher richness than currently recognized exists for these two viral families, highlighting the need to update taxonomy systems and possibly also include genomes identified through metagenomics.

The CckA-ChpT-CtrA Phosphorelay Controlling Rhodobacter capsulatus Gene Transfer Agent Production Is Bidirectional and Regulated by Cyclic di-GMP.

Protein phosphorylation is a universal mechanism for transducing cellular signals in prokaryotes and eukaryotes. The histidine kinase CckA, the histidine phosphotransferase ChpT, and the response regulator CtrA are conserved throughout the alphaproteobacteria. In Rhodobacter capsulatus, these proteins are key regulators of the gene transfer agent (RcGTA), which is present in several alphaproteobacteria. Using purified recombinant R. capsulatus proteins, we show in vitro autophosphorylation of CckA protein, and phosphotransfer to ChpT and thence to CtrA, to demonstrate biochemically that they form a phosphorelay. The secondary messenger cyclic di-GMP changed CckA from a kinase to a phosphatase, resulting in reversal of the phosphotransfer flow in the relay. The substitutions of two residues in CckA greatly affected the kinase or phosphatase activity of the protein in vitro, and production of mutant CckA proteins in vivo confirmed the importance of kinase but not phosphatase activity for the lytic release of RcGTA. However, phosphatase activity was needed to produce functional RcGTA particles. The binding of cyclic di-GMP to the wild-type and mutant CckA proteins was evaluated directly using a pulldown assay based on biotinylated cyclic di-GMP and streptavidin-linked beads.IMPORTANCE The CckA, ChpT, and CtrA phosphorelay proteins are widespread in the alphaproteobacteria, and there are two groups of organisms that differ in terms of whether this pathway is essential for cell viability. Little is known about the biochemical function of these proteins in organisms where the pathway is not essential, a group that includes Rhodobacter capsulatus This work demonstrates biochemically that CckA, ChpT, and CtrA also form a functional phosphorelay in the latter group and that the direction of phosphotransfer is reversed by cyclic di-GMP. It is important to improve understanding of more representatives of this pathway in order to obtain deeper insight into the function, composition, and evolutionary significance of a wider range of bacterial regulatory networks.

Transmission of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) From Asymptomatic and Presymptomatic Individuals in Healthcare Settings Despite Medical Masks and Eye Protection.

We describe 3 instances of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission despite medical masks and eye protection, including transmission despite the source person being masked, transmission despite the exposed person being masked, and transmission despite both parties being masked. Whole genome sequencing confirmed perfect homology between source and exposed persons' viruses in all cases.

ICTV Virus Taxonomy Profile: Marnaviridae 2021.

The family Marnaviridae comprises small non-enveloped viruses with positive-sense RNA genomes of 8.6-9.6 kb. Isolates infect marine single-celled eukaryotes (protists) that come from diverse lineages. Some members are known from metagenomic studies of ocean virioplankton, with additional unclassified viruses described from metagenomic datasets derived from marine and freshwater environments. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Marnaviridae, which is available at ictv.global/report/marnaviridae.

Teaching Cheminformatics through a Collaborative Intercollegiate Online Chemistry Course (OLCC).

While cheminformatics skills necessary for dealing with an ever-increasing amount of chemical information are considered important for students pursuing STEM careers in the age of big data, many schools do not offer a cheminformatics course or alternative training opportunities. This paper presents the Cheminformatics Online Chemistry Course (OLCC), which is organized and run by the Committee on Computers in Chemical Education (CCCE) of the American Chemical Society (ACS)'s Division of Chemical Education (CHED). The Cheminformatics OLCC is a highly collaborative teaching project involving instructors at multiple schools who teamed up with external chemical information experts recruited across sectors, including government and industry. From 2015 to 2019, three Cheminformatics OLCCs were offered. In each program, the instructors at participating schools would meet face-to-face with the students of a class, while external content experts engaged through online discussions across campuses with both the instructors and students. All the material created in the course has been made available at the open education repositories of LibreTexts and CCCE Web sites for other institutions to adapt to their future needs.

Remdesivir and Coronavirus Disease 2019 (COVID-19): Essential Questions and Answers for Pharmacists and Pharmacy Technicians.

Objective: To conduct a review of the investigational drug remdesivir and its therapeutic potential for treatment of COVID-19, in the form of a series of questions and answers. The purpose of the review is to narrow gaps in knowledge, clarify concepts, and to investigate research advancements for health care professionals. Data Sources: From June 2020 to August 2020, we conducted comprehensive searches of MEDLINE-PubMed, Scopus, and Google Scholar databases with no time limitations. Search terms were included that contained the terms "remdesivir," "COVID-19," "novel coronavirus" and "evidence," "therapy," "safety," "effectiveness," "efficacy," "clinical trial." Study Selection and Data Extraction: The sources of information include all publicly available data from previously published research reports. Reports must have at least one reference to remdesivir as a treatment modality for COVID-19 with no specified outcomes. Data Synthesis: Major research findings on the efficacy and safety of remdesivir are summarized in tabular format and presented in chronological order. Results of this review reveal remdesivir to be an effective therapy in specific clinical contexts; however, in several areas, available data are insufficient to support evidence-based guidance for remdesivir in the treatment of COVID-19. Conclusions: Clinical trials on remdesivir are ongoing, yet questions remain and further research is needed as to the selection of patients, effectiveness, and duration of treatment in the use of remdesivir for treatment of COVID-19.

A Comparison of Tertiary Drug Resources' Consistency Regarding Drug-Drug Interactions of Adjunctive Analgesics.

Background: Health care providers routinely rely on tertiary drug information resources to affirm knowledge or proactively verify the safety and efficacy of medications. Though all patient care areas are affected, the reliability of these resources is perhaps nowhere as poignant as it is in high-acuity settings, including the emergency department and the intensive care unit. As providers seek to identify adjunctive analgesics for acute pain in these areas, they must be able to rely on the integrity to whichever resource their institution has granted access. Objective: To determine the congruency of drug-drug interaction information found on 3 tertiary drug resources. Methods: A drug-drug interaction analysis was conducted on Micromedex, Lexicomp, and Medscape. Adjunctive analgesics included dexmedetomidine and ketamine, which were compared with the intravenous opioid products morphine, fentanyl, and hydromorphone. Results: Significant discrepancies were appreciated with regard to the severity of drug-drug interactions. In addition, the heterogeneity in which reaction severity and likelihood are described by each respective resource makes direct comparisons difficult. Interaction warnings for dexmedetomidine and fentanyl included a "major interaction" from Micromedex, whereas Lexicomp did not identify a risk and Medscape only recommended increased monitoring on the grounds of respiratory and central nervous system depression. Conclusions: Health care providers must remain vigilant when reviewing tertiary drug information resources. Pharmacists possess the training and skills necessary to assist interdisciplinary medical teams in providing optimal patient care through evaluating and applying the information gleaned from these resources.

Cyclic di-GMP-Mediated Regulation of Gene Transfer and Motility in Rhodobacter capsulatus.

Gene transfer agents (GTAs) are bacteriophage-like particles produced by several bacterial and archaeal lineages that contain small pieces of the producing cells' genomes that can be transferred to other cells in a process similar to transduction. One well-studied GTA is RcGTA, produced by the alphaproteobacterium Rhodobacter capsulatus RcGTA gene expression is regulated by several cellular regulatory systems, including the CckA-ChpT-CtrA phosphorelay. The transcription of multiple other regulator-encoding genes is affected by the response regulator CtrA, including genes encoding putative enzymes involved in the synthesis and hydrolysis of the second messenger bis-(3'-5')-cyclic dimeric GMP (c-di-GMP). To investigate whether c-di-GMP signaling plays a role in RcGTA production, we disrupted the CtrA-affected genes potentially involved in this process. We found that disruption of four of these genes affected RcGTA gene expression and production. We performed site-directed mutagenesis of key catalytic residues in the GGDEF and EAL domains responsible for diguanylate cyclase (DGC) and c-di-GMP phosphodiesterase (PDE) activities and analyzed the functions of the wild-type and mutant proteins. We also measured RcGTA production in R. capsulatus strains where intracellular levels of c-di-GMP were altered by the expression of either a heterologous DGC or a heterologous PDE. This adds c-di-GMP signaling to the collection of cellular regulatory systems controlling gene transfer in this bacterium. Furthermore, the heterologous gene expression and the four gene disruptions had similar effects on R. capsulatus flagellar motility as found for gene transfer, and we conclude that c-di-GMP inhibits both RcGTA production and flagellar motility in R. capsulatusIMPORTANCE Gene transfer agents (GTAs) are virus-like particles that move cellular DNA between cells. In the alphaproteobacterium Rhodobacter capsulatus, GTA production is affected by the activities of multiple cellular regulatory systems, to which we have now added signaling via the second messenger dinucleotide molecule bis-(3'-5')-cyclic dimeric GMP (c-di-GMP). Similar to the CtrA phosphorelay, c-di-GMP also affects R. capsulatus flagellar motility in addition to GTA production, with lower levels of intracellular c-di-GMP favoring increased flagellar motility and gene transfer. These findings further illustrate the interconnection of GTA production with global systems of regulation in R. capsulatus, providing additional support for the notion that the production of GTAs has been maintained in this and related bacteria because it provides a benefit to the producing organisms.

Stress-mediated convergence of splicing landscapes in male and female rock doves.

BACKGROUND: The process of alternative splicing provides a unique mechanism by which eukaryotes are able to produce numerous protein products from the same gene. Heightened variability in the proteome has been thought to potentiate increased behavioral complexity and response flexibility to environmental stimuli, thus contributing to more refined traits on which natural and sexual selection can act. While it has been long known that various forms of environmental stress can negatively affect sexual behavior and reproduction, we know little of how stress can affect the alternative splicing associated with these events, and less still about how splicing may differ between sexes. Using the model of the rock dove (Columba livia), our team previously uncovered sexual dimorphism in the basal and stress-responsive gene transcription of a biological system necessary for facilitating sexual behavior and reproduction, the hypothalamic-pituitary-gonadal (HPG) axis. In this study, we delve further into understanding the mechanistic underpinnings of how changes in the environment can affect reproduction by testing the alternative splicing response of the HPG axis to an external stressor in both sexes. RESULTS: This study reveals dramatic baseline differences in HPG alternative splicing between males and females. However, after subjecting subjects to a restraint stress paradigm, we found a significant reduction in these differences between the sexes. In both stress and control treatments, we identified a higher incidence of splicing activity in the pituitary in both sexes as compared to other tissues. Of these splicing events, the core exon event is the most abundant form of splicing and more frequently occurs in the coding regions of the gene. Overall, we observed less splicing activity in the 3'UTR (untranslated region) end of transcripts than the 5'UTR or coding regions. CONCLUSIONS: Our results provide vital new insight into sex-specific aspects of the stress response on the HPG axis at an unprecedented proximate level. Males and females uniquely respond to stress, yet exhibit splicing patterns suggesting a convergent, optimal splicing landscape for stress response. This information has the potential to inform evolutionary theory as well as the development of highly-specific drug targets for stress-induced reproductive dysfunction.

Linking Epidemiology and Whole-Genome Sequencing to Investigate Salmonella Outbreak, Massachusetts, USA, 2018.

Cross-discipline collaboration among state and local health departments improved foodborne illness surveillance for a 2018 Salmonella enterica serovar Enteritidis outbreak in Massachusetts, USA. Prompt linking of epidemiologic and laboratory data and implementation of in-state whole-genome sequencing and analysis improved public health surveillance capacity for outbreak detection and control.

Influenza A viruses remain infectious for more than seven months in northern wetlands of North America.

In this investigation, we used a combination of field- and laboratory-based approaches to assess if influenza A viruses (IAVs) shed by ducks could remain viable for extended periods in surface water within three wetland complexes of North America. In a field experiment, replicate filtered surface water samples inoculated with duck swabs were tested for IAVs upon collection and again after an overwintering period of approximately 6-7 months. Numerous IAVs were molecularly detected and isolated from these samples, including replicates maintained at wetland field sites in Alaska and Minnesota for 181-229 days. In a parallel laboratory experiment, we attempted to culture IAVs from filtered surface water samples inoculated with duck swabs from Minnesota each month during September 2018-April 2019 and found monthly declines in viral viability. In an experimental challenge study, we found that IAVs maintained in filtered surface water within wetlands of Alaska and Minnesota for 214 and 226 days, respectively, were infectious in a mallard model. Collectively, our results support surface waters of northern wetlands as a biologically important medium in which IAVs may be both transmitted and maintained, potentially serving as an environmental reservoir for infectious IAVs during the overwintering period of migratory birds.

Hippocampal abnormalities and seizures: a 16-year single center review of sudden unexpected death in childhood, sudden unexpected death in epilepsy and SIDS.

Sudden Unexpected Death in Childhood (SUDC) is the unexplained death of children aged between 1 and 18 years old. Hippocampal abnormalities have previously been described in Sudden Unexpected Death in Epilepsy (SUDEP) and it is possible that SUDC shares similar pathogenic mechanisms with SUDEP. Our aim was to determine the prevalence of hippocampal abnormalities, history of seizures and demographic features in our caseload of SUDC, SUDEP and SIDS cases. A review of post-mortem reports from 2003 to 2018 was carried out to identify cases of SUDC, SUDEP and SIDS. Histological evidence of hippocampal abnormalities, patient demographics (age, gender), sleeping position, and past medical history (history of seizures and illness 72 hours prior to death) were recorded. Statistical analysis was performed to compare the three groups. 48 SUDC, 18 SUDEP and 358 SIDS cases were identified. Hippocampal abnormalities associated with temporal lobe epilepsy were found in 44.4% of SUDC cases. 5/15 SUDC cases with a history of seizures demonstrated hippocampal abnormalities. SUDC cases were also more likely to be found prone compared to SIDS cases. In comparison with SIDS, both SUDC and SUDEP cases were more likely to demonstrate hippocampal abnormalities (SUDC: (OR = 9.4, 95% CI: 3.1-29.1, p < 0.001; SUDEP: OR = 35.4, 95% CI: 8.3-151.5, p < 0.001). We found a potential link between hippocampal abnormalities and epileptic seizures in SUDC. A concerted effort should be directed towards consistent sampling and standardized description of the hippocampus and clinical correlation with a history of seizures/epilepsy in postmortem reports.

Induction of Rhodobacter capsulatus Gene Transfer Agent Gene Expression Is a Bistable Stochastic Process Repressed by an Extracellular Calcium-Binding RTX Protein Homologue.

Bacteriophage-like gene transfer agents (GTAs) have been discovered in both of the prokaryotic branches of the three-domain phylogenetic tree of life. The production of a GTA (RcGTA) by the phototrophic alphaproteobacterium Rhodobacter capsulatus is regulated by quorum sensing and a phosphorelay homologous to systems in other species that control essential functions such as the initiation of chromosome replication and cell division. In wild-type strains, RcGTA is produced in <3% of cells in laboratory cultures. Mutants of R. capsulatus that exhibit greatly elevated production of RcGTA were created decades ago by chemical mutagenesis, but the nature and molecular consequences of the mutation were unknown. We show that the number of cells in a population that go on to express RcGTA genes is controlled by a stochastic process, in contrast to a genetic process. We used transposon mutagenesis along with a fluorescent protein reporter system and genome sequence data to identify a gene, rcc00280, that encodes an RTX family calcium-binding protein homologue. The Rc280 protein acts as an extracellular repressor of RcGTA gene expression by decreasing the percentage of cells that induce the production of RcGTA.IMPORTANCE GTAs catalyze horizontal gene transfer (HGT), which is important for genomic evolution because the majority of genes found in bacterial genomes have undergone HGT at some point in their evolution. Therefore, it is important to determine how the production of GTAs is regulated to understand the factors that modulate the frequency of gene transfer and thereby specify the tempo of evolution. This work describes a new type of genetic regulation in which an extracellular calcium-binding protein homologue represses the induction of the Rhodobacter capsulatus GTA, RcGTA.

Virus discovery reveals frequent infection by diverse novel members of the Flaviviridae in wild lemurs.

Lemurs are highly endangered mammals inhabiting the forests of Madagascar. In this study, we performed virus discovery on serum samples collected from 84 wild lemurs and identified viral sequence fragments from 4 novel viruses within the family Flaviviridae, including members of the genera Hepacivirus and Pegivirus. The sifaka hepacivirus (SifHV, two genotypes) and pegivirus (SifPgV, two genotypes) were discovered in the diademed sifaka (Propithecus diadema), while other pegiviral fragments were detected in samples from the indri (Indri indri, IndPgV) and the weasel sportive lemur (Lepilemur mustelinus, LepPgV). Although data are preliminary, each viral species appeared host species-specific and frequent infection was detected (18 of 84 individuals were positive for at least one virus). The complete coding sequence and partial 5' and 3' untranslated regions (UTRs) were obtained for SifHV and its genomic organization was consistent with that of other hepaciviruses, with one unique polyprotein and highly structured UTRs. Phylogenetic analyses showed the SifHV belonged to a clade that includes several viral species identified in rodents from Asia and North America, while SifPgV and IndPgV were more closely related to pegiviral species A and C, that include viruses found in humans as well as New- and Old-World monkeys. Our results support the current proposed model of virus-host co-divergence with frequent occurrence of cross-species transmission for these genera and highlight how the discovery of more members of the Flaviviridae can help clarify the ecology and evolutionary history of these viruses. Furthermore, this knowledge is important for conservation and captive management of lemurs.