Limited transmission of SARS-CoV-2 in schools in Ireland during the 2020-2021 school year.

BackgroundThe role of schools in SARS-CoV-2 transmission has been a debated topic since the beginning of the COVID-19 pandemic.AimTo examine SARS-CoV-2 transmission in all schools in Ireland during the 2020-21 school year.MethodsIn a national descriptive cross-sectional study, we investigated PCR-confirmed cases of COVID-19 among students (aged < 20 years) and staff (aged ≥ 20 years) who attended school during their infectious period to identify school close contacts. SARS-CoV-2 PCR test results of all school close contacts were pooled to obtain an overall positivity rate and to stratify positivity rate by school setting and role (i.e. student or staff).ResultsIn total, 100,474 individuals were tested as close contacts in 1,771 schools during the 2020-21 school year. An overall close contact positivity rate of 2.4% was observed across all schools (n = 2,373 secondary cases). The highest positivity rate was seen in special schools (3.4%), followed by primary (2.5%) and post-primary schools (1.8%) (p < 0.001). Of the close contacts identified, 90.5% (n = 90,953) were students and 9.5% (n = 9,521) were staff. Overall, students had a significantly higher positivity rate than staff (2.4% vs 1.8%, p < 0.001).ConclusionThis study demonstrated that a low level of SARS-CoV-2 transmission occurred in Irish schools during the 2020-21 academic year. In the event of future pandemics, and as the COVID-19 pandemic continues, there is a need to carefully weigh up the harms and benefits associated with disrupted education to mitigate infectious disease transmission before reflexively closing classes or schools.

Multisystem inflammatory syndrome in the context of paediatric COVID-19 infection in the Republic of Ireland April 2020 to April 2021.

AIM: Our aim was to describe the epidemiology of multisystem inflammatory syndrome in children (MIS-C) in the Republic of Ireland, in the context of all cases of COVID-19 in children, during the first year of the SARS-CoV-2 pandemic. METHODS: Cases of MIS-C were identified by prospective surveillance in Irish hospitals from April 2020 to April 2021. Paediatric COVID-19 cases and outbreaks in schools or childcare facilities were notified to and routinely investigated by Public Health. Univariate and bivariate analyses were carried out in Excel, Stata and JMP statistical package. RESULTS: Fifty-four MIS-C cases (median age 7.58 years; males 57%) were identified over the study period. MIS-C incidence was higher in certain ethnicities ('black' 21.3/100,000 [95% CI 4.3-38.4]; and 'Irish Traveller' 14.7/100,000 [95% CI -5.7-35.1]) than those of 'white' ethnicity (3.4 /100,000). MIS-C cases occurred in three temporal clusters, which followed three distinct waves of community COVID-19 infection, irrespective of school closures. Formal contact tracing identified an epidemiological link with a COVID-19-infected family member in the majority of MIS-C cases (77%). In contrast, investigation of COVID-19 school outbreaks demonstrated no epidemiological link with MIS-C cases during the study period. CONCLUSION: Efforts at controlling SARS-CoV-2 transmission in the community may be a more effective means to reduce MIS-C incidence than school closures. Establishing a mandatory reporting structure for MIS-C will help delineate the role of risk factors such as ethnicity and obesity and the effect of vaccination on MIS-C incidence.

The Study to Investigate COVID-19 Infection in People Living in Ireland (SCOPI): A seroprevalence study, June to July 2020.

BackgroundRobust data on SARS-CoV-2 population seroprevalence supplement surveillance data in providing evidence for public health action.AimTo conduct a SARS-CoV-2 population-based seroprevalence survey in Ireland.MethodsUsing a cross-sectional study design, we selected population samples from individuals aged 12-69 years in counties Dublin and Sligo using the Health Service Executive Primary Care Reimbursement Service database as a sampling frame. Samples were selected with probability proportional to the general population age-sex distribution, and by simple random sampling within age-sex strata. Antibodies to SARS-CoV-2 were detected using the Abbott Architect SARS-CoV-2 IgG Assay and confirmed using the Wantai Assay. We estimated the population SARS-CoV-2 seroprevalence weighted for age, sex and geographic area.ResultsParticipation rates were 30% (913/3,043) and 44% (820/1,863) in Dublin and Sligo. Thirty-three specimens had detectable SARS-CoV-2 antibodies (1.9%). We estimated weighted seroprevalences of 3.12% (95% confidence interval (CI): 2.05-4.53) and 0.58% (95% CI: 0.18-1.38) for Dublin and Sligo, and 1.69% (95% CI: 1.13-2.41) nationally. This equates to an estimated 59,482 (95% CI: 39,772-85,176) people aged 12-69 years nationally having had infection with SARS-CoV-2, 3.0 (95% CI: 2.0-4.3) times higher than confirmed notifications. Ten participants reported a previous laboratory-confirmed SARS-CoV-2 -infection; eight of these were antibody-positive. Twenty-five antibody-positive participants had not reported previous laboratory-confirmed infection.ConclusionThe majority of people in Ireland are unlikely to have been infected with SARS-CoV-2 by June-July 2020. Non-pharmaceutical public health measures remained key pending widespread availability of vaccination, and effective treatments.

Prevalence estimates of dementia in older adults in rural Kilimanjaro 2009-2010 and 2018-2019: is there evidence of changing prevalence?

INTRODUCTION: Although limited, existing epidemiological data on dementia in sub-Saharan Africa indicate that prevalence may be increasing; contrasting with recent decreases observed in high-income countries. We have previously reported the age-adjusted prevalence of dementia in rural Tanzania in 2009-2010 as 6.4% (95% confidence interval [CI] 4.9-7.9) in individuals aged ≥70 years. We aimed to repeat a community-based dementia prevalence study in the same setting to assess whether prevalence has changed. METHODS: This was a two-phase door-to-door community-based cross-sectional survey in Kilimanjaro, Tanzania. In Phase I, trained primary health workers screened all consenting individuals aged ≥60 years from 12 villages using previously validated, locally developed, tools (IDEA cognitive screen and IDEA-Instrumental Activities of Daily Living questionnaire). Screening was conducted using a mobile digital application (app) on a hand-held tablet. In Phase II, a stratified sample of those identified in Phase I were clinically assessed using the DSM-5 criteria and diagnoses subsequently confirmed by consensus panel. RESULTS: Of 3011 people who consented, 424 screened positive for probable dementia and 227 for possible dementia. During clinical assessment in Phase II, 105 individuals met DSM-5 dementia criteria. The age-adjusted prevalence of dementia was 4.6% (95% CI 2.9-6.4) in those aged ≥60 years and 8.9% (95% CI 6.1-11.8) in those aged ≥70 years. Prevalence rates increased significantly with age. CONCLUSIONS: The prevalence of dementia in this rural Tanzanian population appears to have increased since 2010, although not significantly. Dementia is likely to become a significant health burden in this population as demographic transition continues.

Negative supercoiling of DNA by gyrase is inhibited in Salmonella enterica serovar Typhimurium during adaptation to acid stress.

DNA in intracellular Salmonella enterica serovar Typhimurium relaxes during growth in the acidified (pH 4-5) macrophage vacuole and DNA relaxation correlates with the upregulation of Salmonella genes involved in adaptation to the macrophage environment. Bacterial ATP levels did not increase during adaptation to acid pH unless the bacterium was deficient in MgtC, a cytoplasmic-membrane-located inhibitor of proton-driven F1 F0 ATP synthase activity. Inhibiting ATP binding by DNA gyrase and topo IV with novobiocin enhanced the effect of low pH on DNA relaxation. Bacteria expressing novobiocin-resistant (NovR ) derivatives of gyrase or topo IV also exhibited DNA relaxation at acid pH, although further relaxation with novobiocin was not seen in the strain with NovR gyrase. Thus, inhibition of the negative supercoiling activity of gyrase was the primary cause of enhanced DNA relaxation in drug-treated bacteria. The Salmonella cytosol reaches pH 5-6 in response to an external pH of 4-5: the ATP-dependent DNA supercoiling activity of purified gyrase was progressively inhibited by lowering the pH in this range, as was the ATP-dependent DNA relaxation activity of topo IV. We propose that DNA relaxation in Salmonella within macrophage is due to acid-mediated impairment of the negative supercoiling activity of gyrase.

If it transcribes, we can sequence it: mining the complexities of host-pathogen-environment interactions using RNA-seq.

Host-pathogen interactions are exceedingly complex because they involve multiple host tissues, often occur in the context of normal microflora, and can span diverse microenvironments. Although decades of gene expression studies have provided detailed insights into infection processes, technical challenges have restricted experiments to single pathogenic species or host tissues. RNA-sequencing (RNA-seq) has revolutionized the study of gene expression because in addition to quantifying transcriptional output, it allows detection and characterization of all transcripts in a genome. Here, we review how refined approaches to RNA-seq are used to map the transcriptional networks that control host-pathogen interactions. These enhanced techniques include dRNA-seq and term-seq for the fine-scale mapping of transcriptional start and termination sites, and dual RNA-seq for simultaneous sequencing of host and bacterial pathogen transcriptomes. Dual RNA-seq experiments are currently limited to in vitro infection systems that do not fully reflect the complexities of the in vivo environment, thus a challenge is to develop in vivo model systems and experimental approaches that address the biological heterogeneity of host environments, followed by the integration of RNA-seq with other genome-scale datasets to identify the transcriptional networks that mediate host-pathogen interactions.

The Impact of 18 Ancestral and Horizontally-Acquired Regulatory Proteins upon the Transcriptome and sRNA Landscape of Salmonella enterica serovar Typhimurium.

We know a great deal about the genes used by the model pathogen Salmonella enterica serovar Typhimurium to cause disease, but less about global gene regulation. New tools for studying transcripts at the single nucleotide level now offer an unparalleled opportunity to understand the bacterial transcriptome, and expression of the small RNAs (sRNA) and coding genes responsible for the establishment of infection. Here, we define the transcriptomes of 18 mutants lacking virulence-related global regulatory systems that modulate the expression of the SPI1 and SPI2 Type 3 secretion systems of S. Typhimurium strain 4/74. Using infection-relevant growth conditions, we identified a total of 1257 coding genes that are controlled by one or more regulatory system, including a sub-class of genes that reflect a new level of cross-talk between SPI1 and SPI2. We directly compared the roles played by the major transcriptional regulators in the expression of sRNAs, and discovered that the RpoS (σ38) sigma factor modulates the expression of 23% of sRNAs, many more than other regulatory systems. The impact of the RNA chaperone Hfq upon the steady state levels of 280 sRNA transcripts is described, and we found 13 sRNAs that are co-regulated with SPI1 and SPI2 virulence genes. We report the first example of an sRNA, STnc1480, that is subject to silencing by H-NS and subsequent counter-silencing by PhoP and SlyA. The data for these 18 regulatory systems is now available to the bacterial research community in a user-friendly online resource, SalComRegulon.

Bacterial pathogen gene regulation: a DNA-structure-centred view of a protein-dominated domain.

The mechanisms used by bacterial pathogens to regulate the expression of their genes, especially their virulence genes, have been the subject of intense investigation for several decades. Whole genome sequencing projects, together with more targeted studies, have identified hundreds of DNA-binding proteins that contribute to the patterns of gene expression observed during infection as well as providing important insights into the nature of the gene products whose expression is being controlled by these proteins. Themes that have emerged include the importance of horizontal gene transfer to the evolution of pathogens, the need to impose regulatory discipline upon these imported genes and the important roles played by factors normally associated with the organization of genome architecture as regulatory principles in the control of virulence gene expression. Among these architectural elements is the structure of DNA itself, its variable nature at a topological rather than just at a base-sequence level and its ability to play an active (as well as a passive) part in the gene regulation process.

RNA-seq Brings New Insights to the Intra-Macrophage Transcriptome of Salmonella Typhimurium.

Salmonella enterica serovar Typhimurium is arguably the world's best-understood bacterial pathogen. However, crucial details about the genetic programs used by the bacterium to survive and replicate in macrophages have remained obscure because of the challenge of studying gene expression of intracellular pathogens during infection. Here, we report the use of deep sequencing (RNA-seq) to reveal the transcriptional architecture and gene activity of Salmonella during infection of murine macrophages, providing new insights into the strategies used by the pathogen to survive in a bactericidal immune cell. We characterized 3583 transcriptional start sites that are active within macrophages, and highlight 11 of these as candidates for the delivery of heterologous antigens from Salmonella vaccine strains. A majority (88%) of the 280 S. Typhimurium sRNAs were expressed inside macrophages, and SPI13 and SPI2 were the most highly expressed pathogenicity islands. We identified 31 S. Typhimurium genes that were strongly up-regulated inside macrophages but expressed at very low levels during in vitro growth. The SalComMac online resource allows the visualisation of every transcript expressed during bacterial replication within mammalian cells. This primary transcriptome of intra-macrophage S.-Typhimurium describes the transcriptional start sites and the transcripts responsible for virulence traits, and catalogues the sRNAs that may play a role in the regulation of gene expression during infection.

An infection-relevant transcriptomic compendium for Salmonella enterica Serovar Typhimurium.

Bacterial transcriptional networks consist of hundreds of transcription factors and thousands of promoters. However, the true complexity of transcription in a bacterial pathogen and the effect of the environments encountered during infection remain to be established. We present a simplified approach for global promoter identification in bacteria using RNA-seq-based transcriptomic analyses of 22 distinct infection-relevant environmental conditions. Individual RNA samples were combined to identify most of the 3,838 Salmonella enterica serovar Typhimurium promoters in just two RNA-seq runs. Individual in vitro conditions stimulated characteristic transcriptional signatures, and the suite of 22 conditions induced transcription of 86% of all S. Typhimurium genes. We highlight the environmental conditions that induce the Salmonella pathogenicity islands and present a small RNA expression landscape of 280 sRNAs. This publicly available compendium of environmentally controlled expression of every transcriptional feature of S. Typhimurium constitutes a useful resource for the bacterial research community.

ProP is required for the survival of desiccated Salmonella enterica serovar typhimurium cells on a stainless steel surface.

Consumers trust commercial food production to be safe, and it is important to strive to improve food safety at every level. Several outbreaks of food-borne disease have been caused by Salmonella strains associated with dried food. Currently we do not know the mechanisms used by Salmonella enterica serovar Typhimurium to survive in desiccated environments. The aim of this study was to discover the responses of S. Typhimurium ST4/74 at the transcriptional level to desiccation on a stainless steel surface and to subsequent rehydration. Bacterial cells were dried onto the same steel surfaces used during the production of dry foods, and RNA was recovered for transcriptomic analysis. Subsequently, dried cells were rehydrated and were again used for transcriptomic analysis. A total of 266 genes were differentially expressed under desiccation stress compared with a static broth culture. The osmoprotectant transporters proP, proU, and osmU (STM1491 to STM1494) were highly upregulated by drying. Deletion of any one of these transport systems resulted in a reduction in the long-term viability of S. Typhimurium on a stainless steel food contact surface. The proP gene was critical for survival; proP deletion mutants could not survive desiccation for long periods and were undetectable after 4 weeks. Following rehydration, 138 genes were differentially expressed, with upregulation observed for genes such as proP, proU, and the phosphate transport genes (pstACS). In time, this knowledge should prove valuable for understanding the underlying mechanisms involved in pathogen survival and should lead to improved methods for control to ensure the safety of intermediate- and low-moisture foods.

sRNAs and the virulence of Salmonella enterica serovar Typhimurium.

The combination of genomics and high-throughput cDNA sequencing technologies has facilitated the identification of many small RNAs (sRNAs) that play a central role in the post-transcriptional gene regulation of Salmonella enterica serovar Typhimurium. To date, most of the functionally characterized sRNAs have been involved in the regulation of processes which are not directly linked to virulence. Just five sRNAs have been found to affect the ability of Salmonella to replicate within mammalian cells, but the precise regulatory mechanisms that are used by sRNAs to control Salmonella pathogenicity at the post-transcriptional level remain to be identified. It is anticipated that an improved understanding of sRNA biology will shed new light on the virulence of Salmonella.

The transcriptional landscape and small RNAs of Salmonella enterica serovar Typhimurium.

More than 50 y of research have provided great insight into the physiology, metabolism, and molecular biology of Salmonella enterica serovar Typhimurium (S. Typhimurium), but important gaps in our knowledge remain. It is clear that a precise choreography of gene expression is required for Salmonella infection, but basic genetic information such as the global locations of transcription start sites (TSSs) has been lacking. We combined three RNA-sequencing techniques and two sequencing platforms to generate a robust picture of transcription in S. Typhimurium. Differential RNA sequencing identified 1,873 TSSs on the chromosome of S. Typhimurium SL1344 and 13% of these TSSs initiated antisense transcripts. Unique findings include the TSSs of the virulence regulators phoP, slyA, and invF. Chromatin immunoprecipitation revealed that RNA polymerase was bound to 70% of the TSSs, and two-thirds of these TSSs were associated with σ(70) (including phoP, slyA, and invF) from which we identified the -10 and -35 motifs of σ(70)-dependent S. Typhimurium gene promoters. Overall, we corrected the location of important genes and discovered 18 times more promoters than identified previously. S. Typhimurium expresses 140 small regulatory RNAs (sRNAs) at early stationary phase, including 60 newly identified sRNAs. Almost half of the experimentally verified sRNAs were found to be unique to the Salmonella genus, and <20% were found throughout the Enterobacteriaceae. This description of the transcriptional map of SL1344 advances our understanding of S. Typhimurium, arguably the most important bacterial infection model.