Sleep, Immune Function, and Vaccinations in Military Personnel: Challenges and Future Directions.

The U.S. military invests substantial resources to vaccinate all personnel, including recruits, against operationally important infectious disease threats. However, research suggests that vaccine immune response and, therefore, vaccine effectiveness may be inadvertently reduced because of chronic and/or acute sleep deficiency experienced by recipients around the time of vaccination. Because sleep deficiency is expected and even necessary in deployed and training contexts, research investigations of the impacts of sleep and related physiological systems such as circadian rhythms on vaccine effectiveness in military settings are needed. Specifically, research should be aimed at understanding the effects of sleep deficiency, as well as vaccine administration schedules, on response to vaccination and clinical protection. Furthermore, knowledge gaps among military medical leadership on sleep, vaccines, and immune health should be assessed. This area of research may benefit the health and readiness of service members while also decreasing health care utilization and associated costs from illness.

Maturation and vesicle-mediated egress of primate gammaherpesvirus rhesus monkey rhadinovirus require inner tegument protein ORF52.

UNLABELLED: The tegument layer of herpesviruses comprises a collection of proteins that is unique to each viral species. In rhesus monkey rhadinovirus (RRV), a close relative of the human oncogenic pathogen Kaposi's sarcoma-associated herpesvirus, ORF52 is a highly abundant tegument protein tightly associated with the capsid. We now report that ORF52 knockdown during RRV infection of rhesus fibroblasts led to a greater than 300-fold reduction in the viral titer by 48 h but had little effect on the number of released particles and caused only modest reductions in the levels of intracellular viral genomic DNA and no appreciable change in viral DNA packaging into capsids. These data suggested that the lack of ORF52 resulted in the production and release of defective particles. In support of this interpretation, transmission electron microscopy (TEM) revealed that without ORF52, capsid-like particles accumulated in the cytoplasm and were unable to enter egress vesicles, where final tegumentation and envelopment normally occur. TEM also demonstrated defective particles in the medium that closely resembled the accumulating intracellular particles, having neither a full tegument nor an envelope. The disruption in tegument formation from ORF52 suppression, therefore, prevented the incorporation of ORF45, restricting its subcellular localization to the nucleus and appearing, by confocal microscopy, to inhibit particle transport toward the periphery. Ectopic expression of small interfering RNA (siRNA)-resistant ORF52 was able to partially rescue all of these phenotypic changes. In sum, our results indicate that efficient egress of maturing virions and, in agreement with studies on murine gammaherpesvirus 68 (MHV-68), complete tegumentation and secondary envelopment are dependent on intact ORF52. IMPORTANCE: The tegument, or middle layer, of herpesviruses comprises both viral and cellular proteins that play key roles in the viral life cycle. A subset of these proteins is present only within members of one of the three subfamilies (alphaherpesviruses, betaherpesviruses, or gammaherpesviruses) of Herpesviridae. In this report, we show that the gammaherpesvirus-specific tegument protein ORF52 is critical for maturation of RRV, the closest relative of Kaposi's sarcoma-associated herpesvirus (KSHV) (a human cancer-causing pathogen) that has undergone this type of analysis. Without ORF52, the nascent subviral particles are essentially stuck in maturation limbo, unable to acquire the tegument or outer (envelope) layers. This greatly attenuates infectivity. Our data, together with earlier work on a murine homolog, as well as a more distantly related human homolog, provide a more complete understanding of how early protein interactions involving virus-encoded tegument proteins are critical for virus assembly and are also, therefore, potentially attractive therapeutic targets.

Progressive accumulation of activated ERK2 within highly stable ORF45-containing nuclear complexes promotes lytic gammaherpesvirus infection.

De novo infection with the gammaherpesvirus Rhesus monkey rhadinovirus (RRV), a close homolog of the human oncogenic pathogen, Kaposi's sarcoma-associated herpesvirus (KSHV), led to persistent activation of the MEK/ERK pathway and increasing nuclear accumulation of pERK2 complexed with the RRV protein, ORF45 (R45) and cellular RSK. We have previously shown that both lytic gene expression and virion production are dependent on the activation of ERK [1]. Using confocal microscopy, sequential pull-down assays and FRET analyses, we have demonstrated that pERK2-R45-RSK2 complexes were restricted to the nucleus but that the activated ERK retained its ability to phosphorylate nuclear substrates throughout infection. Furthermore, even with pharmacologic inhibition of MEK beginning at 48 h p.i., pERK2 but not pERK1, remained elevated for at least 10 h, showing first order decay and a half-life of nearly 3 hours. Transfection of rhesus fibroblasts with R45 alone also led to the accumulation of nuclear pERK2 and addition of exogenous RSK augmented this effect. However, knock down of RSK during bona fide RRV infection had little to no effect on pERK2 accumulation or virion production. The cytoplasmic pools of pERK showed no co-localization with either RSK or R45 but activation of pERK downstream targets in this compartment was evident throughout infection. Together, these observations suggest a model in which R45 interacts with pERK2 to promote its nuclear accumulation, thereby promoting lytic viral gene expression while also preserving persistent and robust activation of both nuclear and cytoplasmic ERK targets.

Kind discrimination and competitive exclusion mediated by contact-dependent growth inhibition systems shape biofilm community structure.

Contact-Dependent Growth Inhibition (CDI) is a phenomenon in which bacteria use the toxic C-terminus of a large exoprotein (called BcpA in Burkholderia species) to inhibit the growth of neighboring bacteria upon cell-cell contact. CDI systems are present in a wide range of Gram-negative proteobacteria and a hallmark feature is polymorphism amongst the exoprotein C-termini (BcpA-CT in Burkholderia) and amongst the small immunity proteins (BcpI) that protect against CDI in an allele-specific manner. In addition to CDI, the BcpAIOB proteins of Burkholderia thailandensis mediate biofilm formation, and they do so independent of BcpA-mediated interbacterial competition, suggesting a cooperative role for CDI system proteins in this process. CDI has previously only been demonstrated between CDI+ and CDI- bacteria, leaving the roles of CDI system-mediated interbacterial competition and of CDI system diversity in nature unknown. We constructed B. thailandensis strains that differed only in the BcpA-CT and BcpI proteins they produced. When co-cultured on agar, these strains each participated in CDI and the outcome of the competition depended on both CDI system efficiency and relative bacterial numbers initially. Strains also participated in CDI during biofilm development, resulting in pillar structures that were composed of only a single BcpA-CT/BcpI type. Moreover, a strain producing BcpA-CT/BcpI proteins of one type was prevented from joining a pre-established biofilm community composed of bacteria producing BcpA-CT/BcpI proteins of a different type, unless it also produced the BcpI protein of the established strain. Bacteria can therefore use CDI systems for kind recognition and competitive exclusion of 'non-self' bacteria from a pre-established biofilm. Our data indicate that CDI systems function in both cooperative and competitive behaviors to build microbial communities that are composed of only bacteria that are related via their CDI system alleles.

Global research integrity in relation to the United States' research-integrity infrastructure.

Recent global attention to research integrity has led to international meetings and the development of international policies and guidelines. The United States's infrastructure for fostering research integrity (policy, instruction, oversight) has usefully supported these international initiatives. The United States cannot and should not, however, expect other national and global systems to match exactly its approach to research integrity.

Burkholderia BcpA mediates biofilm formation independently of interbacterial contact-dependent growth inhibition.

Contact-dependent growth inhibition (CDI) is a phenomenon in which Gram-negative bacteria use the toxic C-terminus of a large surface-exposed exoprotein to inhibit the growth of susceptible bacteria upon cell-cell contact. Little is known about when and where bacteria express the genes encoding CDI system proteins and how these systems contribute to the survival of bacteria in their natural niche. Here we establish that, in addition to mediating interbacterial competition, the Burkholderia thailandensis CDI system exoprotein BcpA is required for biofilm development. We also provide evidence that the catalytic activity of BcpA and extracellular DNA are required for the characteristic biofilm pillars to form. We show using a bcpA-gfp fusion that within the biofilm, expression of the CDI system-encoding genes is below the limit of detection for the majority of bacteria and only a subset of cells express the genes strongly at any given time. Analysis of a strain constitutively expressing the genes indicates that native expression is critical for biofilm architecture. Although CDI systems have so far only been demonstrated to be involved in interbacterial competition, constitutive production of the system's immunity protein in the entire bacterial population did not alter biofilm formation, indicating a CDI-independent role for BcpA in this process. We propose, therefore, that bacteria may use CDI proteins in cooperative behaviours, like building biofilm communities, and in competitive behaviours that prevent non-self bacteria from entering the community.

The Burkholderia bcpAIOB genes define unique classes of two-partner secretion and contact dependent growth inhibition systems.

Microbes have evolved many strategies to adapt to changes in environmental conditions and population structures, including cooperation and competition. One apparently competitive mechanism is contact dependent growth inhibition (CDI). Identified in Escherichia coli, CDI is mediated by Two-Partner Secretion (TPS) pathway proteins, CdiA and CdiB. Upon cell contact, the toxic C-terminus of the TpsA family member CdiA, called the CdiA-CT, inhibits the growth of CDI(-) bacteria. CDI(+) bacteria are protected from autoinhibition by an immunity protein, CdiI. Bioinformatic analyses indicate that CDI systems are widespread amongst α, ß, and γ proteobacteria and that the CdiA-CTs and CdiI proteins are highly variable. CdiI proteins protect against CDI in an allele-specific manner. Here we identify predicted CDI system-encoding loci in species of Burkholderia, Ralstonia and Cupriavidus, named bcpAIOB, that are distinguished from previously-described CDI systems by gene order and the presence of a small ORF, bcpO, located 5' to the gene encoding the TpsB family member. A requirement for bcpO in function of BcpA (the TpsA family member) was demonstrated, indicating that bcpAIOB define a novel class of TPS system. Using fluorescence microscopy and flow cytometry, we show that these genes are expressed in a probabilistic manner during culture of Burkholderia thailandensis in liquid medium. The bcpAIOB genes and extracellular DNA were required for autoaggregation and adherence to an abiotic surface, suggesting that CDI is required for biofilm formation, an activity not previously attributed to CDI. By contrast to what has been observed in E. coli, the B. thailandensis bcpAIOB genes only mediated interbacterial competition on a solid surface. Competition occurred in a defined spatiotemporal manner and was abrogated by allele-specific immunity. Our data indicate that the bcpAIOB genes encode distinct classes of CDI and TPS systems that appear to function in sociomicrobiological community development.

The problem of plagiarism.

Plagiarism is a form of research misconduct and a serious violation of the norms of science. It is the misrepresentation of another's ideas or words as one's own, without proper acknowledgement of the original source. Certain aspects of plagiarism make it less straightforward than this definition suggests. Over the past 30 years, the U.S. Federal Government has developed and refined its policies on misconduct, and Federal agencies, as well as research institutions, have established approaches to responding to allegations and instances of plagiarism. At present, efforts to avert plagiarism focus on plagiarism-detection software and instructional strategies.

Extending the Mertonian Norms: Scientists' Subscription to Norms of Research.

This analysis, based on focus groups and a national survey, assesses scientists' subscription to the Mertonian norms of science and associated counternorms. It also supports extension of these norms to governance (as opposed to administration), as a norm of decision-making, and quality (as opposed to quantity), as a evaluative norm.

The importance of organizational justice in ensuring research integrity.

The professional behavior of scientists, for good or ill, is likely associated with their perceptions of whether they are treated fairly in their work environments, including their academic department and university and by relevant regulatory bodies. These relationships may also be influenced by their own personal characteristics, such as being overcommitted to their work, and by the interactions between these factors. Theory also suggests that such associations may be mediated by negative or positive affect. We examined these issues using data from a national, mail-based survey administered in 2006 and 2007 to 5,000 randomly selected faculty from biomedical and social science departments at 50 top-tier research universities in the United States. We found that perceptions of justice in one's workplace (organizational justice) are positively associated with self-report of "ideal" behaviors and negatively associated with self-report of misbehavior and misconduct. By contrast, researchers who perceive that they are being unfairly treated are less likely to report engaging in "ideal" behaviors and more likely to report misbehavior and misconduct. Overcommitment to one's work is also associated with negative affect and interacts with perceptions of unfair treatment in ways that are associated with higher self-report of misbehavior. Thus, perceptions of fair treatment in the work environment appear to play important roles in fostering-or undermining-research integrity.

Institutions' expectations for researchers' self-funding, federal grant holding, and private industry involvement: manifold drivers of self-interest and researcher behavior.

BACKGROUND: Private industry involvement is viewed as tainting research with self-interest, whereas public funding is generally well regarded. Yet, dependence on "soft money" also triggers researcher and university self-interest. No empirical research has compared these factors' effects on academic researchers' behaviors. METHOD: In 2006-2007, a survey was mailed to 5,000 randomly selected biomedical and social science faculty at 50 top-tier research universities in the United States. Measures included a university's expectations or nonexpectations that researchers obtain external grant funding, the receipt or nonreceipt of public research funding, any relationships with private industry, and research-related behaviors ranging from the ideal, to the questionable, to misconduct. RESULTS: Being expected to obtain external funding and receiving federal research funding were both associated with significantly higher reports of 1 or more of 10 serious misbehaviors (P<.05) and neglectful or careless behaviors (P<.001). Researchers with federal funding were more likely than were those without to report having carelessly or inappropriately reviewed papers or proposals (9.6% versus 3.9%; P<.001). Those with private industry involvement were more likely than were those without to report 1 or more of 10 serious misbehaviors (28.5% versus 21.5%; P=.005) and to have engaged in misconduct (12.2% versus 7.1%; P=.004); they also were less likely to have always reported financial conflicts (96.0% versus 98.6%, P<.001). CONCLUSIONS: The free play of university and individual self-interests, combined with and contributing to the intense competition for research funding, may be undermining scientific integrity.

The perverse effects of competition on scientists' work and relationships.

Competition among scientists for funding, positions and prestige, among other things, is often seen as a salutary driving force in U.S. science. Its effects on scientists, their work and their relationships are seldom considered. Focus-group discussions with 51 mid- and early-career scientists, on which this study is based, reveal a dark side of competition in science. According to these scientists, competition contributes to strategic game-playing in science, a decline in free and open sharing of information and methods, sabotage of others' ability to use one's work, interference with peer-review processes, deformation of relationships, and careless or questionable research conduct. When competition is pervasive, such effects may jeopardize the progress, efficiency and integrity of science.

What do mentoring and training in the responsible conduct of research have to do with scientists' misbehavior? Findings from a National Survey of NIH-funded scientists.

PURPOSE: The authors examine training in the responsible conduct of research and mentoring in relation to behaviors that may compromise the integrity of science. METHOD: The analysis is based on data from the authors' 2002 national survey of 4,160 early-career and 3,600 midcareer biomedical and social science researchers who received research support from the U.S. National Institutes of Health. The authors used logistic regression analysis to examine associations between receipt of separate or integrated training in research ethics, mentoring related to ethics and in general, and eight categories of ethically problematic behavior. Analyses controlled for gender, type of doctoral degree, international degree, and disciplinary field. RESULTS: Responses were received from 1,479 early-career and 1,768 midcareer scientists, yielding adjusted response rates of 43% and 52%, respectively. Results for early-career researchers: Training in research ethics was positively associated with problematic behavior in the data category. Mentoring related to ethics and research, as well as personal mentoring, decreased the odds of researchers' engaging in problematic behaviors, but mentoring on financial issues and professional survival increased these odds. Results for midcareer researchers: Combined separate and integrated training in research ethics was associated with decreased odds of problematic behavior in the categories of policy, use of funds, and cutting corners. Ethics mentoring was associated with lowered odds of problematic behavior in the policy category. CONCLUSIONS: The effectiveness of training in obviating problematic behavior is called into question. Mentoring has the potential to influence behavior in ways that both increase and decrease the likelihood of problematic behaviors.

Normative dissonance in science: results from a national survey of u.s. Scientists.

NORMS OF BEHAVIOR IN SCIENTIFIC RESEARCH represent ideals to which most scientists subscribe. Our analysis of the extent of dissonance between these widely espoused ideals and scientists' perceptions of their own and others' behavior is based on survey responses from 3,247 mid- and early-career scientists who had research funding from the U.S. National Institutes of Health. We found substantial normative dissonance, particularly between espoused ideals and respondents' perceptions of other scientists' typical behavior. Also, respondents on average saw other scientists' behavior as more counternormative than normative. Scientists' views of their fields as cooperative or competitive were associated with their normative perspectives, with competitive fields showing more counternormative behavior. The high levels of normative dissonance documented here represent a persistent source of stress in science.

Scientists' perceptions of organizational justice and self-reported misbehaviors.

Policymakers concerned about maintaining the integrity of science have recently expanded their attention from a focus on misbehaving individuals to characteristics of the environments in which scientists work. Little empirical evidence exists about the role of organizational justice in promoting or hindering scientific integrity. Our findings indicate that when scientists believe they are being treated unfairly they are more likely to behave in ways that compromise the integrity of science. Perceived violations of distributive and procedural justice were positively associated with self-reports of misbehavior among scientists.

Normal Misbehavior: Scientists Talk about the Ethics of Research.

Those concerned with protecting the Integrity of science generally focus on the serious but rare infractions of falsification, fabrication, and plagiarism (FFP). While the violations of FFP are clear threats to the quality of scientific work and public trust in science, are they the behaviors that researchers themselves find most troubling? Noticing that scientists seldom are asked to report their perceptions of the behaviors that pose problems for the enterprise of science, we conducted six focus groups with researchers from major research universities. A total of 51 scientists participated in our focus-group discussions, which lasted from 1.5 to 2 hours each. We found that while researchers were aware of the problems of FFP, in their eyes misconduct generally is associated with more mundane, everyday problems in the work environment. These more common problems fall into four categories: the meaning of data, the rules of science, life with colleagues, and the pressures of production in science. Focus on the "normal misbehaviors" that are part of the ordinary life of researchers allows us to see the way the organization of science generates both compliance and deviance from ethical norms.