Signatures of natural selection are not uniform across genes of innate immune system, but purifying selection is the dominant signature.

We tested the opposing views concerning evolution of genes of the innate immune system that (i) being evolutionary ancient, the system may have been highly optimized by natural selection and therefore should be under purifying selection, and (ii) the system may be plastic and continuing to evolve under balancing selection. We have resequenced 12 important innate-immunity genes (CAMP, DEFA4, DEFA5, DEFA6, DEFB1, MBL2, and TLRs 1, 2, 4, 5, 6, and 9) in healthy volunteers (n = 171) recruited from a region of India with high microbial load. We have compared these data with those of European-Americans (EUR) and African-Americans (AFR). We have found that most of the human haplotypes are many mutational steps away from the ancestral (chimpanzee) haplotypes, indicating that humans may have had to adapt to new pathogens. The haplotype structures in India are significantly different from those of EUR and AFR populations, indicating local adaptation to pathogens. In these genes, there is (i) generally an excess of rare variants, (ii) high, but variable, degrees of extended haplotype homozygosity, (iii) low tolerance to nonsynonymous changes, (iv) essentially one or a few high-frequency haplotypes, with star-like phylogenies of other infrequent haplotypes radiating from the modal haplotypes. Purifying selection is the most parsimonious explanation operating on these innate immunity genes. This genetic surveillance system recognizes motifs in pathogens that are perhaps conserved across a broad range of pathogens. Hence, functional constraints are imposed on mutations that diminish the ablility of these proteins to detect pathogens.

The PhenX Toolkit: get the most from your measures.

The potential for genome-wide association studies to relate phenotypes to specific genetic variation is greatly increased when data can be combined or compared across multiple studies. To facilitate replication and validation across studies, RTI International (Research Triangle Park, North Carolina) and the National Human Genome Research Institute (Bethesda, Maryland) are collaborating on the consensus measures for Phenotypes and eXposures (PhenX) project. The goal of PhenX is to identify 15 high-priority, well-established, and broadly applicable measures for each of 21 research domains. PhenX measures are selected by working groups of domain experts using a consensus process that includes input from the scientific community. The selected measures are then made freely available to the scientific community via the PhenX Toolkit. Thus, the PhenX Toolkit provides the research community with a core set of high-quality, well-established, low-burden measures intended for use in large-scale genomic studies. PhenX measures will have the most impact when included at the experimental design stage. The PhenX Toolkit also includes links to standards and resources in an effort to facilitate data harmonization to legacy data. Broad acceptance and use of PhenX measures will promote cross-study comparisons to increase statistical power for identifying and replicating variants associated with complex diseases and with gene-gene and gene-environment interactions.

Social network analysis of patient sharing among hospitals in Orange County, California.

OBJECTIVES: We applied social network analyses to determine how hospitals within Orange County, California, are interconnected by patient sharing, a system which may have numerous public health implications. METHODS: Our analyses considered 2 general patient-sharing networks: uninterrupted patient sharing (UPS; i.e., direct interhospital transfers) and total patient sharing (TPS; i.e., all interhospital patient sharing, including patients with intervening nonhospital stays). We considered these networks at 3 thresholds of patient sharing: at least 1, at least 10, and at least 100 patients shared. RESULTS: Geographically proximate hospitals were somewhat more likely to share patients, but many hospitals shared patients with distant hospitals. Number of patient admissions and percentage of cancer patients were associated with greater connectivity across the system. The TPS network revealed numerous connections not seen in the UPS network, meaning that direct transfers only accounted for a fraction of total patient sharing. CONCLUSIONS: Our analysis demonstrated that Orange County's 32 hospitals were highly and heterogeneously interconnected by patient sharing. Different hospital populations had different levels of influence over the patient-sharing network.

The role of subway travel in an influenza epidemic: a New York City simulation.

The interactions of people using public transportation in large metropolitan areas may help spread an influenza epidemic. An agent-based model computer simulation of New York City's (NYC's) five boroughs was developed that incorporated subway ridership into a Susceptible-Exposed-Infected-Recovered disease model framework. The model contains a total of 7,847,465 virtual people. Each person resides in one of the five boroughs of NYC and has a set of socio-demographic characteristics and daily behaviors that include age, sex, employment status, income, occupation, and household location and membership. The model simulates the interactions of subway riders with their workplaces, schools, households, and community activities. It was calibrated using historical data from the 1957-1958 influenza pandemics and from NYC travel surveys. The surveys were necessary to enable inclusion of subway riders into the model. The model results estimate that if influenza did occur in NYC with the characteristics of the 1957-1958 pandemic, 4% of transmissions would occur on the subway. This suggests that interventions targeted at subway riders would be relatively ineffective in containing the epidemic. A number of hypothetical examples demonstrate this feature. This information could prove useful to public health officials planning responses to epidemics.

Modeling targeted layered containment of an influenza pandemic in the United States.

Planning a response to an outbreak of a pandemic strain of influenza is a high public health priority. Three research groups using different individual-based, stochastic simulation models have examined the consequences of intervention strategies chosen in consultation with U.S. public health workers. The first goal is to simulate the effectiveness of a set of potentially feasible intervention strategies. Combinations called targeted layered containment (TLC) of influenza antiviral treatment and prophylaxis and nonpharmaceutical interventions of quarantine, isolation, school closure, community social distancing, and workplace social distancing are considered. The second goal is to examine the robustness of the results to model assumptions. The comparisons focus on a pandemic outbreak in a population similar to that of Chicago, with approximately 8.6 million people. The simulations suggest that at the expected transmissibility of a pandemic strain, timely implementation of a combination of targeted household antiviral prophylaxis, and social distancing measures could substantially lower the illness attack rate before a highly efficacious vaccine could become available. Timely initiation of measures and school closure play important roles. Because of the current lack of data on which to base such models, further field research is recommended to learn more about the sources of transmission and the effectiveness of social distancing measures in reducing influenza transmission.

Simulating school closure strategies to mitigate an influenza epidemic.

BACKGROUND: There remains substantial debate over the impact of school closure as a mitigation strategy during an influenza pandemic. The ongoing 2009 H1N1 influenza pandemic has provided an unparalleled opportunity to test interventions with the most up-to-date simulations. METHODS: To assist the Allegheny County Health Department during the 2009 H1N1 influenza pandemic, the University of Pittsburgh Models of Infectious Disease Agents Study group employed an agent-based computer simulation model (ABM) of Allegheny County, Pennsylvania, to explore the effects of various school closure strategies on mitigating influenza epidemics of different reproductive rates (R0). RESULTS: Entire school system closures were not more effective than individual school closures. Any type of school closure may need to be maintained throughout most of the epidemic (ie, at least 8 weeks) to have any significant effect on the overall serologic attack rate. In fact, relatively short school closures (ie, 2 weeks or less) may actually slightly increase the overall attack rate by returning susceptible students back into schools in the middle of the epidemic. Varying the illness threshold at which school closures are triggered did not seem to have substantial impact on the effectiveness of school closures, suggesting that short delays in closing schools should not cause concern. CONCLUSIONS: School closures alone may not be able to quell an epidemic but, when maintained for at least 8 weeks, could delay the epidemic peak for up to a week, providing additional time to implement a second more effective intervention such as vaccination.

Comprehensive defensin assay for saliva.

Defensins are highly basic cationic peptides that are important components of the innate and adaptive immune response pathways. In addition, these peptides are involved in CD8+ T cell response to HIV-1, increased pulmonary infection risk among cystic fibrosis patients, upregulated levels of HNP-5 for patients with ulcerative colitis and Crohn's disease, and monitoring HNP-3 levels as a tumor classification scheme for cutaneous T cell lymphomas, and have promise in the pharmaceutical field as a new class of antibiotics. Here we present a parallel assay for the alpha (HNP1-3) and beta (HBD1-2) classes of defensins in saliva that are naturally observed in the concentration range of 1 ng/mL to 10 microg/mL. The method utilizes solid phase extraction of saliva samples combined with liquid chromatography-tandem mass spectrometry to identify and quantitate defensin targets. The approach involves limited sample manipulation and is easily amenable to automation. The saliva samples analyzed are derived from a large cohort study focused on examining the role of polymorphisms in genes of innate and adaptive immunity in modulating the response to vaccination for two gastrointestinal tract infections: typhoid and cholera. The alpha-defensin levels observed range from 1 to 10 microg/mL and correlate well with known active concentrations against a wide variety of pathogens. The observed concentration range for beta-defensins was between the detection limit and 33 ng/mL and had a sensitivity level that was comparable to immunoassay-based detection. This method is easily adapted for use in a clinical immunology setting and can be modified for other biological matrixes. This assay will facilitate examination of the production, secretion, and regulation of defensin peptides in a direct fashion to coordinate levels of these compounds with gender, age, response to vaccination, gene copy number, and oral health.

Genetic variation and haplotype structures of innate immunity genes in eastern India.

This study reports results of an extensive and comprehensive study of genetic diversity in 12 genes of the innate immune system in a population of eastern India. Genomic variation was assayed in 171 individuals by resequencing approximately 75kb of DNA comprising these genes in each individual. Almost half of the 548 DNA variants discovered was novel. DNA sequence comparisons with human and chimpanzee reference sequences revealed evolutionary features indicative of natural selection operating among individuals, who are residents of an area with a high load of microbial and other pathogens. Significant differences in allele and haplotype frequencies of the study population were observed with the HapMap populations. Gene and haplotype diversities were observed to be high. The genetic positioning of the study population among the HapMap populations based on data of the innate immunity genes substantially differed from what has been observed for Indian populations based on data of other genes. The reported range of variation in SNP density in the human genome is one SNP per 1.19kb (chromosome 22) to one SNP per 2.18kb (chromosome 19). The SNP density in innate immunity genes observed in this study (>3SNPskb(-1)) exceeds the highest density observed for any autosomal chromosome in the human genome. The extensive genomic variation and the distinct haplotype structure of innate immunity genes observed among individuals have possibly resulted from the impact of natural selection.

Family-based detection for hereditary hemochromatosis.

The purpose of this study was to examine motivators for and barriers to family-based detection for hereditary hemochromatosis (HH). HH patients (n = 60) and HH siblings (n = 25) participated in one-on-one or group interviews. Patients and siblings understood that HH "runs in families," but not that siblings are at higher HH risk than other family members. Patient motivators included concern for siblings' health, seriousness of untreated HH, and doctor's encouragement to tell siblings that they need to seek diagnostic testing. Siblings were motivated by the seriousness of HH. Barriers included lack of symptoms, belief that HH was rare, and assumption that their doctor would have mentioned the risk of HH. Family-based detection continues to be a feasible part of an overall public health strategy to promote early detection of HH. Greater awareness of HH and its potential consequences, especially among high-risk groups, provides an additional potential avenue for public health action.

Evaluation of vaccine-induced antibody responses: impact of new technologies.

Host response to vaccination has historically been evaluated based on a change in antibody titer that compares the post-vaccination titer to the pre-vaccination titer. A four-fold or greater increase in antigen-specific antibody has been interpreted to indicate an increase in antibody production in response to vaccination. New technologies, such as the bead-based assays, provide investigators and clinicians with precise antibody levels (reported as concentration per mL) in ranges below and above those previously available through standard assays such as ELISA. Evaluations of bead assay data to determine host response to vaccination using fold change and absolute change, with a general linear model used to calculate adjusted statistics, present very different pictures of the antibody response when pre-vaccination antibody levels are low. Absolute changes in bead assay values, although not a standard computation, appears to more accurately reflect the host response to vaccination for those individuals with extremely low pre-vaccination antibody levels. Conversely, for these same individuals, fold change may be very high while post-vaccination antibodies do not achieve seroprotective levels. Absolute change provides an alternate method to characterize host response to vaccination, especially when pre-vaccination levels are very low, and may be useful in studies designed to determine associations between host genotypes and response to vaccination.

Genomic correlates of variability in immune response to an oral cholera vaccine.

Cholera is endemic to many countries. Recent major outbreaks of cholera have prompted World Health Organization to recommend oral cholera vaccination as a public-health strategy. Variation in percentage of seroconversion upon cholera vaccination has been recorded across populations. Vaccine-induced responses are influenced by host genetic differences. We have investigated association between single-nucleotide polymorphic (SNP) loci in and around 296 immunologically relevant genes and total anti-lipopolysaccharide (LPS) antibody response to a killed whole-cell vaccine, comprising LPS from multiple strains of Vibrio cholerae. Titers derived from standard vibriocidal assays were also analyzed to gain further insights on validated SNP associations. Vaccination was administered to 1000 individuals drawn from India. Data on two independent random subsets, each comprising ∼500 vaccinees, were used for discovery of genomic associations and validation, respectively. Significant associations of four SNPs and haplotypes in three genes (MARCO, TNFAIP3 and CXCL12) with AR were discovered and validated, of which two in TNFAIP3 and CXCL12 were also significantly associated with immunity (fourfold increase in vibriocidal titers). CXCL12 is a neutrophil and lymphocyte chemoattractant that is upregulated in response to V. cholerae infection. LPS in the vaccine possibly provides signals that mimic those of the live bacterium. TNFAIP3 promotes intestinal epithelial barrier integrity and provides tight junction protein regulation; possible requirements for adequate response to the vaccine. LPS is a potent activator of innate immune responses and a ligand of MARCO. Variants in this gene have been found to be associated with LPS response, but not with high vibriocidal titer level.

Synthesized Population Databases: A Geospatial Database of US Poultry Farms.

The pervasive and potentially severe economic, social, and public health consequences of infectious disease in farmed animals require that plans be in place for a rapid response. Increasingly, agent-based models are being used to analyze the spread of animal-borne infectious disease outbreaks and derive policy alternatives to control future outbreaks. Although the locations, types, and sizes of animal farms are essential model inputs, no public domain nationwide geospatial database of actual farm locations and characteristics currently exists in the United States. This report describes a novel method to develop a synthetic dataset that replicates the spatial distribution of poultry farms, as well as the type and number of birds raised on them. It combines county-aggregated poultry farm counts, land use/land cover, transportation, business, and topographic data to generate locations in the conterminous United States where poultry farms are likely to be found. Simulation approaches used to evaluate the accuracy of this method when compared to that of a random placement alternative found this method to be superior. The results suggest the viability of adapting this method to simulate other livestock farms of interest to infectious disease researchers.

Sharing Research Models: Using Software Engineering Practices for Facilitation.

Increasingly, researchers are turning to computational models to understand the interplay of important variables on systems' behaviors. Although researchers may develop models that meet the needs of their investigation, application limitations-such as nonintuitive user interface features and data input specifications-may limit the sharing of these tools with other research groups. By removing these barriers, other research groups that perform related work can leverage these work products to expedite their own investigations. The use of software engineering practices can enable managed application production and shared research artifacts among multiple research groups by promoting consistent models, reducing redundant effort, encouraging rigorous peer review, and facilitating research collaborations that are supported by a common toolset. This report discusses three established software engineering practices- the iterative software development process, object-oriented methodology, and Unified Modeling Language-and the applicability of these practices to computational model development. Our efforts to modify the MIDAS TranStat application to make it more user-friendly are presented as an example of how computational models that are based on research and developed using software engineering practices can benefit a broader audience of researchers.

Including the Group Quarters Population in the US Synthesized Population Database.

In 2005, RTI International researchers developed methods to generate synthesized population data on US households for the US Synthesized Population Database. These data are used in agent-based modeling, which simulates large-scale social networks to test how changes in the behaviors of individuals affect the overall network. Group quarters are residences where individuals live in close proximity and interact frequently. Although the Synthesized Population Database represents the population living in households, data for the nation's group quarters residents are not easily quantified because of US Census Bureau reporting methods designed to protect individuals' privacy.Including group quarters population data can be an important factor in agent-based modeling because the number of residents and the frequency of their interactions are variables that directly affect modeling results. Particularly with infectious disease modeling, the increased frequency of agent interaction may increase the probability of infectious disease transmission between individuals and the probability of disease outbreaks.This report reviews our methods to synthesize data on group quarters residents to match US Census Bureau data. Our goal in developing the Group Quarters Population Database was to enable its use with RTI's US Synthesized Population Database in the Modeling of Infectious Diseases Agent Study.

Mass commuting and influenza vaccination prevalence in new york city: protection in a mixing environment.

OBJECTIVE: Assess influenza vaccination among commuters using mass transit in New York City (NYC). METHODS: We used the 2006 NYC Community Health Survey (CHS) to analyze the prevalence of influenza immunization by commuting behaviors and to understand what socioeconomic and geographic factors may explain any differences found. RESULTS: Vaccination prevalence is significantly lower for New Yorkers who commute on public transportation compared to other New Yorkers. This difference is largely attenuated after adjusting for socio-demographic characteristics and neighborhood of residence. CONCLUSIONS: The analysis identified a low prevalence of immunization among commuters, and given the transmissibility in that setting, targeting commuters for vaccination campaigns may impede influenza spread.

A large study on immunological response to a whole-cell killed oral cholera vaccine reveals that there are significant geographical differences in response and that O blood group individuals do not elicit a higher response.

The ABO blood group system has been implicated in susceptibility to cholera or in explaining variability in the immune response to a cholera vaccine. O blood group individuals were found to be more susceptible to cholera and elicited lower vibriocidal antibody response to cholera toxin B subunit-killed oral vaccine. Based on the observations that O blood group individuals were more susceptible to cholera and that high mortality was associated with cholera, an evolutionary explanation was provided for the extremely low prevalence of the O blood group in the Gangetic Delta (West Bengal, India, and Bangladesh). However, conflicting results were reported from a later study conducted in Indonesia using a live attenuated oral cholera vaccine; O blood group individuals showed a higher vibriocidal antibody response. In a study conducted in a region of India where cholera is endemic (Kolkata, West Bengal) that comprised 992 individuals vaccinated by a killed whole-cell oral cholera vaccine, we found no statistically significant difference between O and non-O individuals either in the frequency distributions of the fold increase or in the postvaccination increase in geometric mean titer compared to the baseline. Further, in contrast to the earlier observation that the O allele frequency is extremely low in the Gangetic Delta, we have noted that the O allele frequency exceeds 0.5 in the vast majority of ethnic groups of this region. In addition, we have found large differences in response to the vaccine among residents of an area where cholera is not endemic compared to an area where cholera is endemic to The percentages of vaccinees who seroconverted in an area where cholera is not endemic (Son La province of Vietnam) was >90% compared to approximately 50% in Kolkata, India, an area where cholera is endemic.

Prevalence of high-risk indications for influenza vaccine varies by age, race, and income.

Estimates of the proportions of the population who are at high risk of influenza complications because of prior health status or who are likely to have decreased vaccination response because of immunocompromising conditions would enhance public health planning and model-based projections. We estimate these proportions and how they vary by population subgroups using national data systems for 2006-2008. The proportion of individuals at increased risk of influenza complications because of health conditions varied 10-fold by age (4.2% of children <2 years to 47% of individuals >64 years). Age-specific prevalence differed substantially by gender, by racial/ethnic groups (with African Americans highest in all age groups) and by income. Individuals living in families with less than 200% of federal poverty level (FPL) were significantly more likely to have at least one of these health conditions, compared to individuals with 400% FPL or more (3-fold greater among <2 and 30% greater among >64 years). Among children, there were significantly elevated proportions in all regions compared to the West. The estimated prevalence of immunocompromising conditions ranged from 0.02% in young children to 6.14% older adults. However, national data on race/ethnicity and income are not available for most immunocompromising conditions, nor is it possible to fully identify the degree of overlap between persons with high-risk health conditions and with immunocompromising conditions. Modifications to current national data collection systems would enhance the value of these data for public health programs and influenza modeling.

Development of a bead immunoassay to measure Vi polysaccharide-specific serum IgG after vaccination with the Salmonella enterica serovar Typhi Vi polysaccharide.

Vi polysaccharide from Salmonella enterica serotype Typhi is used as one of the available vaccines to prevent typhoid fever. Measurement of Vi-specific serum antibodies after vaccination with Vi polysaccharide by enzyme-linked immunosorbent assay (ELISA) may be complicated due to poor binding of the Vi polysaccharide to ELISA plates resulting in poor reproducibility of measured antibody responses. We chemically conjugated Vi polysaccharide to fluorescent beads and performed studies to determine if a bead-based immunoassay provided a reproducible method to measure vaccine-induced anti-Vi serum IgG antibodies. Compared to ELISA, the Vi bead immunoassay had a lower background and therefore a greater signal-to-noise ratio. The Vi bead immunoassay was used to evaluate serum anti-Vi IgG in 996 subjects from the city of Kolkata, India, before and after vaccination. Due to the location being one where Salmonella serotype Typhi is endemic, approximately 45% of the subjects had protective levels of anti-Vi serum IgG (i.e., 1 microg/ml anti-Vi IgG) before vaccination, and nearly 98% of the subjects had protective levels of anti-Vi serum IgG after vaccination. Our results demonstrate that a bead-based immunoassay provides an effective, reproducible method to measure serum anti-Vi IgG responses before and after vaccination with the Vi polysaccharide vaccine.

Protecting health care workers: a pandemic simulation based on Allegheny County.

BACKGROUND AND OBJECTIVES: The Advisory Committee on Immunization Practices has identified health care workers (HCWs) as a priority group to receive influenza vaccine. Although the importance of HCW to the health care system is well understood, the potential role of HCW in transmission during an epidemic has not been clearly established. METHODS: Using a standard SIR (Susceptible-Infected-Recovered) framework similar to previously developed pandemic models, we developed an agent-based model (ABM) of Allegheny County, PA, that incorporates the key health care system features to simulate the spread of an influenza epidemic and its effect on hospital-based HCWs. FINDINGS: Our simulation runs found the secondary attack rate among unprotected HCWs to be approximately 60% higher (54.3%) as that of all adults (34.1%), which would result in substantial absenteeism and additional risk to HCW families. Understanding how a pandemic may affect HCWs, who must be available to treat infected patients as well as patients with other medical conditions, is crucial to policy makers' and hospital administrators' preparedness planning.

A computer simulation of employee vaccination to mitigate an influenza epidemic.

BACKGROUND: Better understanding the possible effects of vaccinating employees is important and can help policymakers and businesses plan vaccine distribution and administration logistics, especially with the current H1N1 influenza vaccine in short supply. PURPOSE: This article aims to determine the effects of varying vaccine coverage, compliance, administration rates, prioritization, and timing among employees during an influenza pandemic. METHODS: As part of the H1N1 influenza planning efforts of the Models of Infectious Disease Agent Study network, an agent-based computer simulation model was developed for the Washington DC metropolitan region, encompassing five metropolitan statistical areas. Each simulation run involved introducing 100 infectious individuals to initiate a 1.3 reproductive-rate (R(0)) epidemic, consistent with H1N1 parameters to date. Another set of scenarios represented a R(0)=1.6 epidemic. RESULTS: An unmitigated epidemic resulted in substantial productivity losses (a mean of $112.6 million for a serologic 15% attack rate and $193.8 million for a serologic 25% attack rate), even with the relatively low estimated mortality impact of H1N1. Although vaccinating Advisory Committee on Immunization Practices-defined priority groups resulted in the largest savings, vaccinating all remaining workers captured additional savings and, in fact, reduced healthcare workers' and critical infrastructure workers' chances of infection. Moreover, although employee vaccination compliance affected the epidemic, once 20% compliance was achieved, additional increases in compliance provided less incremental benefit. Even though a vast majority of the workplaces in the DC metropolitan region had fewer than 100 employees, focusing on vaccinating only those in larger firms (> or =100 employees) was just as effective in mitigating the epidemic as trying to vaccinate employees in all workplaces. CONCLUSIONS: Timely vaccination of at least 20% of the large-company workforce can play an important role in epidemic mitigation.