Active surveillance documents rates of clinical care seeking due to respiratory illness.

BACKGROUND: Respiratory viral infections are a leading cause of disease worldwide. However, the overall community prevalence of infections has not been properly assessed, as standard surveillance is typically acquired passively among individuals seeking clinical care. METHODS: We conducted a prospective cohort study in which participants provided daily diaries and weekly nasopharyngeal specimens that were tested for respiratory viruses. These data were used to analyze healthcare seeking behavior, compared with cross-sectional ED data and NYC surveillance reports, and used to evaluate biases of medically attended ILI as signal for population respiratory disease and infection. RESULTS: The likelihood of seeking medical attention was virus-dependent: higher for influenza and metapneumovirus (19%-20%), lower for coronavirus and RSV (4%), and 71% of individuals with self-reported ILI did not seek care and half of medically attended symptomatic manifestations did not meet the criteria for ILI. Only 5% of cohort respiratory virus infections and 21% of influenza infections were medically attended and classifiable as ILI. We estimated 1 ILI event per person/year but multiple respiratory infections per year. CONCLUSION: Standard, healthcare-based respiratory surveillance has multiple limitations. Specifically, ILI is an incomplete metric for quantifying respiratory disease, viral respiratory infection, and influenza infection. The prevalence of respiratory viruses, as reported by standard, healthcare-based surveillance, is skewed toward viruses producing more severe symptoms. Active, longitudinal studies are a helpful supplement to standard surveillance, can improve understanding of the overall circulation and burden of respiratory viruses, and can aid development of more robust measures for controlling the spread of these pathogens.

Longitudinal active sampling for respiratory viral infections across age groups.

BACKGROUND: Respiratory viral infections are a major cause of morbidity and mortality worldwide. However, their characterization is incomplete because prevalence estimates are based on syndromic surveillance data. Here, we address this shortcoming through the analysis of infection rates among individuals tested regularly for respiratory viral infections, irrespective of their symptoms. METHODS: We carried out longitudinal sampling and analysis among 214 individuals enrolled at multiple New York City locations from fall 2016 to spring 2018. We combined personal information with weekly nasal swab collection to investigate the prevalence of 18 respiratory viruses among different age groups and to assess risk factors associated with infection susceptibility. RESULTS: 17.5% of samples were positive for respiratory viruses. Some viruses circulated predominantly during winter, whereas others were found year round. Rhinovirus and coronavirus were most frequently detected. Children registered the highest positivity rates, and adults with daily contacts with children experienced significantly more infections than their counterparts without children. CONCLUSION: Respiratory viral infections are widespread among the general population with the majority of individuals presenting multiple infections per year. The observations identify children as the principal source of respiratory infections. These findings motivate further active surveillance and analysis of differences in pathogenicity among respiratory viruses.

Advection of surface-derived organic carbon fuels microbial reduction in Bangladesh groundwater.

Chronic exposure to arsenic (As) by drinking shallow groundwater causes widespread disease in Bangladesh and neighboring countries. The release of As naturally present in sediment to groundwater has been linked to the reductive dissolution of iron oxides coupled to the microbial respiration of organic carbon (OC). The source of OC driving this microbial reduction--carbon deposited with the sediments or exogenous carbon transported by groundwater--is still debated despite its importance in regulating aquifer redox status and groundwater As levels. Here, we used the radiocarbon ((14)C) signature of microbial DNA isolated from groundwater samples to determine the relative importance of surface and sediment-derived OC. Three DNA samples collected from the shallow, high-As aquifer and one sample from the underlying, low-As aquifer were consistently younger than the total sediment carbon, by as much as several thousand years. This difference and the dominance of heterotrophic microorganisms implies that younger, surface-derived OC is advected within the aquifer, albeit more slowly than groundwater, and represents a critical pool of OC for aquifer microbial communities. The vertical profile shows that downward transport of dissolved OC is occurring on anthropogenic timescales, but bomb (14)C-labeled dissolved OC has not yet accumulated in DNA and is not fueling reduction. These results indicate that advected OC controls aquifer redox status and confirm that As release is a natural process that predates human perturbations to groundwater flow. Anthropogenic perturbations, however, could affect groundwater redox conditions and As levels in the future.

XRCC1 polymorphisms and breast cancer risk from the New York Site of the Breast Cancer Family Registry: A family-based case-control study.

BACKGROUND: XRCC1 is a scaffold protein involved in the early and late stages of Base Excision Repair (BER). Three DNA polymorphisms occur in XRCC1, resulting in non-synonymous amino acid changes, which could alter the binding or regulatory activities of XRCC1. MATERIALS AND METHODS: We used a family-based case-control study design to evaluate the association between XRCC1 polymorphisms and breast cancer risk. Participants were breast cancer cases and their unaffected sisters enrolled in the New York Site of the Breast Cancer Family Registry. Conditional logistic regression was used to assess associations between genotype and breast cancer. XRCC1 mRNA levels and DNA nicking activity were measured in lymphoblastoid cell lines from unaffected sisters to determine whether the XRCC1 R399Q polymorphism has a functional effect on expression or protein activity. RESULTS: XRCC1 194W was associated with a non-significant increase in breast cancer, while XRCC1 280H and XRCC1 399Q were associated with a non-significant decrease in breast cancer. We found a significant increase in XRCC1 expression in 399Q/Q lymphoblastoid cell lines from unaffected sisters (n=28, P=0.03). An increase in median nicking activity was not statistically significant. CONCLUSIONS: Our results suggest that XRCC1 399Q may alter mRNA expression and DNA repair phenotype, although the main effects of the genotype were not significantly associated with familial cancer risk. Additional research on the regulation of XRCC1 expression will contribute to an understanding of how this polymorphism may impact disease risk.

Microbial mineral weathering for nutrient acquisition releases arsenic.

Tens of millions of people in Southeast Asia drink groundwater contaminated with naturally occurring arsenic. How arsenic is released from the sediment into the water remains poorly understood. Here, we show in laboratory experiments that phosphate-limited cells of Burkholderia fungorum mobilize ancillary arsenic from apatite. We hypothesize that arsenic mobilization is a by-product of mineral weathering for nutrient acquisition. The released arsenic does not undergo a redox transformation but appears to be solubilized from the apatite mineral lattice during weathering. Analysis of apatite from the source area in the Himalayan basin indicates the presence of elevated levels of arsenic, with an average concentration of 210 mg/kg. The rate of arsenic release is independent of the initial dissolved arsenic concentration and occurs at phosphate levels observed in Bangladesh aquifers. We also demonstrate the presence of the microbial phenotype that releases arsenic from apatite in Bangladesh aquifer sediments and groundwater. These results suggest that microbial mineral weathering for nutrient acquisition could be an important mechanism for arsenic mobilization.

Importance of a C-terminal conserved region of Chk1 for checkpoint function.

BACKGROUND: The protein kinase Chk1 is an essential component of the DNA damage checkpoint pathway. Chk1 is phosphorylated and activated in the fission yeast Schizosaccharomyces pombe when cells are exposed to agents that damage DNA. Phosphorylation, kinase activation, and nuclear accumulation are events critical to the ability of Chk1 to induce a transient delay in cell cycle progression. The catalytic domain of Chk1 is well-conserved amongst all species, while there are only a few regions of homology within the C-terminus. A potential pseudosubstrate domain exists in the C-terminus of S. pombe Chk1, raising the possibility that the C-terminus acts to inhibit the catalytic domain through interaction of this domain with the substrate binding site. METHODOLOGY/PRINCIPAL FINDINGS: To evaluate this hypothesis, we characterized mutations in the pseudosubstrate region. Mutation of a conserved aspartic acid at position 469 to alanine or glycine compromises Chk1 function when the mutants are integrated as single copies, demonstrating that this domain of Chk1 is critical for function. Our data does not support, however, the hypothesis that the domain acts to inhibit Chk1 function as other mutations in the amino acids predicted to comprise the pseudosubstrate do not result in constitutive activation of the protein. When expressed in multi-copy, Chk1D469A remains non-functional. In contrast, multi-copy Chk1D469G confers cell survival and imposes a checkpoint delay in response to some, though not all forms of DNA damage. CONCLUSIONS/SIGNIFICANCE: Thus, we conclude that this C-terminal region of Chk1 is important for checkpoint function and predict that a limiting factor capable of associating with Chk1D469G, but not Chk1D469A, interacts with Chk1 to elicit checkpoint activation in response to a subset of DNA lesions.

Mus81-Eme1-dependent and -independent crossovers form in mitotic cells during double-strand break repair in Schizosaccharomyces pombe.

During meiosis, double-strand breaks (DSBs) lead to crossovers, thought to arise from the resolution of double Holliday junctions (HJs) by an HJ resolvase. In Schizosaccharomyces pombe, meiotic crossovers are produced primarily through a mechanism requiring the Mus81-Eme1 endonuclease complex. Less is known about the processes that produces crossovers during the repair of DSBs in mitotic cells. We employed an inducible DSB system to determine the role of Rqh1-Top3 and Mus81-Eme1 in mitotic DSB repair and crossover formation in S. pombe. In agreement with the meiotic data, crossovers are suppressed in cells lacking Mus81-Eme1. And relative to the wild type, rqh1Delta cells show a fourfold increase in crossover frequency. This suppression of crossover formation by Rqh1 is dependent on its helicase activity. We found that the synthetic lethality of cells lacking both Rqh1 and Eme1 is suppressed by loss of swi5(+), which allowed us to show that the excess crossovers formed in an rqh1Delta background are independent of Mus81-Eme1. This result suggests that a second process for crossover formation exists in S. pombe and is consistent with our finding that deletion of swi5(+) restored meiotic crossovers in eme1Delta cells. Evidence suggesting that Rqh1 also acts downstream of Swi5 in crossover formation was uncovered in these studies. Our results suggest that during Rhp51-dependent repair of DSBs, Rqh1-Top3 suppresses crossovers in the Rhp57-dependent pathway while Mus81-Eme1 and possibly Rqh1 promote crossovers in the Swi5-dependent pathway.

Rqh1 blocks recombination between sister chromatids during double strand break repair, independent of its helicase activity.

Many questions remain about the process of DNA double strand break (DSB) repair by homologous recombination (HR), particularly concerning the exact function played by individual proteins and the details of specific steps in this process. Some recent studies have shown that RecQ DNA helicases have a function in HR. We studied the role of the RecQ helicase Rqh1 with HR proteins in the repair of a DSB created at a unique site within the Schizosaccharomyces pombe genome. We found that DSBs in rqh1(+) cells, are predominantly repaired by interchromosomal gene conversion, with HR between sister chromatids [sister-chromatid conversion (SCC)], occurring less frequently. In Deltarqh1 cells, repair by SCC is favored, and gene conversion rates slow significantly. When we limited the potential for SCC in Deltarqh1 cells by reducing the length of the G2 phase of the cell cycle, DSB repair continued to be predominated by SCC, whereas it was essentially eliminated in wild-type cells. These data indicate that Rqh1 acts to regulate DSB repair by blocking SCC. Interestingly, we found that this role for Rqh1 is independent of its helicase activity. In the course of these studies, we also found nonhomologous end joining to be largely faithful in S. pombe, contrary to current belief. These findings provide insight into the regulation of DSB repair by RecQ helicases.

RMI1/NCE4, a suppressor of genome instability, encodes a member of the RecQ helicase/Topo III complex.

SGS1 encodes a DNA helicase whose homologues in human cells include the BLM, WRN, and RECQ4 genes, mutations in which lead to cancer-predisposition syndromes. Clustering of synthetic genetic interactions identified by large-scale genetic network analysis revealed that the genetic interaction profile of the gene RMI1 (RecQ-mediated genome instability, also known as NCE4 and YPL024W) was highly similar to that of SGS1 and TOP3, suggesting a functional relationship between Rmi1 and the Sgs1/Top3 complex. We show that Rmi1 physically interacts with Sgs1 and Top3 and is a third member of this complex. Cells lacking RMI1 activate the Rad53 checkpoint kinase, undergo a mitotic delay, and display increased relocalization of the recombination repair protein Rad52, indicating the presence of spontaneous DNA damage. Consistent with a role for RMI1 in maintaining genome integrity, rmi1Delta cells exhibit increased recombination frequency and increased frequency of gross chromosomal rearrangements. In addition, rmi1Delta strains fail to fully activate Rad53 upon exposure to DNA-damaging agents, suggesting that Rmi1 is also an important part of the Rad53-dependent DNA damage response.

A postsynaptic role for Rhp55/57 that is responsible for cell death in Deltarqh1 mutants following replication arrest in Schizosaccharomyces pombe.

Following replication arrest, multiple cellular responses are triggered to maintain genomic integrity. In fission yeast, the RecQ helicase, Rqh1, plays a critical role in this process. This is demonstrated in Deltarqh1 cells that, following treatment with hydroxyurea (HU), undergo an aberrant mitosis leading to cell death. Previous data suggest that Rqh1 functions with homologous recombination (HR) in recovery from replication arrest. We have found that loss of the HR genes rhp55(+) or rhp57(+), but not rhp51(+) or rhp54(+), suppresses the HU sensitivity of Deltarqh1 cells. Much of this suppression requires Rhp51 and Rhp54. In addition, this suppression is partially dependent on swi5(+). In budding yeast, overexpressing Rad51 (the Rhp51 homolog) minimized the need for Rad55/57 (Rhp55/57) in nucleoprotein filament formation. We overexpressed Rhp51 in Schizosaccharomyces pombe and found that it greatly reduced the requirement for Rhp55/57 in recovery from DNA damage. However, overexpressing Rhp51 did not change the Deltarhp55 suppression of the HU sensitivity of Deltarqh1, supporting an Rhp55/57 function during HR independent of nucleoprotein filament formation. These results are consistent with Rqh1 playing a role late in HR following replication arrest and provide evidence for a postsynaptic function for Rhp55/57.

The severe slow growth of Deltasrs2 Deltarqh1 in Schizosaccharomyces pombe is suppressed by loss of recombination and checkpoint genes.

Our interest in the Schizosaccharomyces pombe RecQ helicase, rqh1+, led us to investigate the function of a related putative DNA helicase, srs2+. We identified the srs2+ homolog in S.pombe, and found that srs2+ is not essential for cell viability. A Deltasrs2 Deltarqh1 double mutant grows extremely slowly with aberrant shaped cells and low viability. This slow growth does not appear to be related to stalled replication, as Deltasrs2 Deltarqh1 cells showed higher survival rates, compared with Deltarqh1, when stalled forks were increased by UV irradiation or hydroxy urea treatment. Consistent with this result, we found that Deltasrs2 Deltarqh1 cells progress through S-phase with a slight delay, but undergo a checkpoint-dependent arrest presumably at G2/M. Further, we found that Deltasrs2 Deltarqh1 slow growth is related to recombination, as loss of either the rhp51+ or rhp57+ recombination genes improves cell growth in the double mutant. Deltasrs2 is also synthetic lethal with Deltarhp54, another homologous recombination gene. This lethality is suppressed in a Deltarhp51 background. Together, these results demonstrate a clear genetic interaction between rqh1+, srs2+ and the genes of the homologous recombination pathway.

A single unbranched S-phase DNA damage and replication fork blockage checkpoint pathway.

The eukaryotic intra-S-phase checkpoint, which slows DNA synthesis in response to DNA damage, is poorly understood. Is DNA damage recognized directly, or indirectly through its effects on replication forks? Is the slowing of S phase in part because of competition between DNA synthesis and recombination/repair processes? The results of our genetic analyses of the intra-S-phase checkpoint in the fission yeast, Schizosaccharomyces pombe, suggest that the slowing of S phase depends weakly on the helicases Rqh1 and Srs2 but not on other recombination/repair pathways. The slowing of S phase depends strongly on the six checkpoint-Rad proteins, on Cds1, and on Rad4/Cut5 (similar to budding yeast Dpb11, which interacts with DNA polymerase epsilon) but not on Rhp9 (similar to budding yeast Rad9, necessary for direct damage recognition). These results suggest that, in fission yeast, the signal activating the intra-S-phase checkpoint is generated only when replication forks encounter DNA damage.