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Researchers who use animals in science must balance statistical power with the need to satisfy the three Rs, whereby researchers are required to reduce numbers of animals, refine what they experience, and use alternatives to (replace) higher animals where possible. In repeated sampling over time-series studies, there is potential loss of power as well as ethics implications posed by repeated sampling of individual animals, where this cannot be managed or avoided. Here, we consider the mathematics of repeated sampling from three perspectives: that of the population at large, from the experience of the individual, and the conditional probability of sampled individuals being sampled again. The calculations are illustrated using four theoretical case studies across veterinary epidemiology with different practical implications and a provided R Shiny tool for researchers. Despite the availability of exact calculations, it is necessary to also consider the biological factors which may affect capture and recapture rates in sampling studies such as animal personality and response to capture. Researchers must also choose their question carefully to avoid inappropriate framing of ethical concerns around repeated sampling., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domains (TIGIT) is an inhibitory receptor on immune cells that outcompetes an activating receptor, CD226, for shared ligands. Tumor-infiltrating lymphocytes express TIGIT and CD226 on regulatory T cells (Treg) and on CD8+ T cells with tumor-reactive or exhausted phenotypes, supporting the potential of therapeutically targeting TIGIT to enhance antitumor immunity. To optimize the efficacy of therapeutic antibodies against TIGIT, it is necessary to understand IgG Fc (Fcγ) receptor binding for therapeutic benefit. In this study, we showed that combining Fc-enabled (Fce) or Fc-silent (Fcs) anti-TIGIT with antiprogrammed cell death protein 1 in mice resulted in enhanced control of tumors by differential mechanisms: Fce anti-TIGIT promoted the depletion of intratumoral Treg, whereas Fcs anti-TIGIT did not. Despite leaving Treg numbers intact, Fcs anti-TIGIT potentiated the activation of tumor-specific exhausted CD8+ populations in a lymph node-dependent manner. Fce anti-TIGIT induced antibody-dependent cell-mediated cytotoxicity against human Treg in vitro, and significant decreases in Treg were measured in the peripheral blood of patients with phase I solid tumor cancer treated with Fce anti-TIGIT. In contrast, Fcs anti-TIGIT did not deplete human Treg in vitro and was associated with anecdotal objective clinical responses in two patients with phase I solid tumor cancer whose peripheral Treg frequencies remained stable on treatment. Collectively, these data provide evidence for pharmacologic activity and antitumor efficacy of anti-TIGIT antibodies lacking the ability to engage Fcγ receptor., Significance: Fcs-silent anti-TIGIT antibodies enhance the activation of tumor-specific pre-exhausted T cells and promote antitumor efficacy without depleting T regulatory cells., (©2024 American Association for Cancer Research.)

Instructor Talk-noncontent and nonlogistical language that is focused on shaping the classroom learning environment-is a recently defined variable that may play an important role in how undergraduates experience courses. Previous research characterized Instructor Talk used by faculty teaching in biology lecture classrooms. However, graduate teaching assistants (GTAs) and laboratory classrooms represent critical factors in undergraduate education, and Instructor Talk in this context has yet to be explored. Here, we present findings analyzing Instructor Talk used by GTAs teaching in undergraduate biology laboratory classrooms. We characterized the Instructor Talk used by 22 GTA instructors across 24 undergraduate biology laboratory courses in the context of a single, urban, Hispanic-serving and Asian American and Pacific Islander-serving Institution. We found that Instructor Talk was present in every course studied, GTAs with pedagogical training and prior teaching experience used more Instructor Talk than those without, and GTAs teaching laboratory courses used more Instructor Talk than previous observations of faculty teaching lecture courses. Given the widespread use of Instructor Talk and its varying use across contexts, we predict that Instructor Talk may be a critical variable in teaching, specifically in promoting equity and inclusion, which merits continued study in undergraduate science education.

Introduction: The pace and scale of the COVID-19 pandemic, coupled with ongoing efforts by health agencies to communicate harms, have created a pressing need for data to inform messaging about smoking, vaping, and COVID-19. We examined reactions to COVID-19 and traditional health harms messages discouraging smoking and vaping., Methods: Participants were a national convenience sample of 810 US adults recruited online in May 2020. All participated in a smoking message experiment and a vaping message experiment, presented in a random order. In each experiment, participants viewed one message formatted as a Twitter post. The experiments adopted a 3 (traditional health harms of smoking or vaping: three harms, one harm, absent) × 2 (COVID-19 harms: one harm, absent) between-subjects design. Outcomes included perceived message effectiveness (primary) and constructs from the Tobacco Warnings Model (secondary: attention, negative affect, cognitive elaboration, social interactions)., Results: Smoking messages with traditional or COVID-19 harms elicited higher perceived effectiveness for discouraging smoking than control messages without these harms (all p <0.001). However, including both traditional and COVID-19 harms in smoking messages had no benefit beyond including either alone. Smoking messages affected Tobacco Warnings Model constructs and did not elicit more reactance than control messages. Smoking messages also elicited higher perceived effectiveness for discouraging vaping. Including traditional harms in messages about vaping elicited higher perceived effectiveness for discouraging vaping (p <0.05), but including COVID-19 harms did not., Conclusions: Messages linking smoking with COVID-19 may hold promise for discouraging smoking and may have the added benefit of also discouraging vaping., Competing Interests: Competing interests: None of the authors have received funding from tobacco product manufacturers. NTB, SMN, and KMR have served as paid expert consultants in litigation against e-cigarette and tobacco companies. JMS has served as a paid consultant in government litigation against tobacco companies., (© Author(s) (or their employer(s)) 2022. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Large healthcare systems provide an opportunity to disseminate evidence-based interventions to primary care. We evaluated the impact of a train-the-trainer model in two large systems to disseminate the Announcement Approach Training, which teaches providers to communicate about HPV vaccination more effectively., Methods: In collaboration with the American Cancer Society, we partnered with two midwestern healthcare systems that served over 77,000 patients ages 11 through 17. Both systems hosted a 2-hour train-the-trainer workshop. Providers from one system then conducted in-person 1-hour CME-eligible trainings, using our standard slide set and script (available at hpvIQ.org). The other system did not implement trainings, providing a natural experiment., Results: The train-the-trainer workshop included physicians, nurses and other clinical staff (n = 11/13 for intervention/comparison systems). The intervention system delivered 18 trainings to 234 physicians, nurses, and other clinic staff. From baseline to 6-month follow-up, the intervention system had an increase in HPV vaccine uptake that was larger than that of the comparison system for adolescents ages 11 through 12 (1.9%, p = .002) and ages 13 through 17 (1.5%, p = .015). Attending the training was associated with increased intentions to routinely recommend HPV vaccine when patients turn 11 or 12 (mean 4.19 (SD = 0.95) vs. 4.43 (SD = 0.83) as well as increased positive vaccine attitudes, self-efficacy, and norms (all p < .001). Participant satisfaction with the trainings was high (90%-94%)., Conclusion: The train-the-trainer model was effective in increasing provider motivation to recommend HPV vaccination and led to a small increase in vaccine uptake. Dissemination through large healthcare systems is promising but faces some challenges., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

A growing body of health determinants research recognizes that housing and health are intimately linked. This study explores the relationship between rent burden (the ratio of rent to income) and health risk behaviors among a sample of single room occupancy (SRO) building residents. Cross-sectional data were gathered from a sample of 162 residents living in privately owned, for-profit SROs in Chicago. Findings indicated that participants who had full rental subsidies and thus were designated in a no-rent-burden category were more likely to engage in risk behaviors including illicit drug use, having multiple sexual partners, and having sex without a condom, in comparison to participants with moderate or high-rent burdens. These findings suggest that interventions to increase housing stability and affordability and bolster reliable income sources (in addition to rental subsidies) may be key in reducing risk behaviors and improving health for vulnerably housed populations such as SRO residents.

Background: Fluid therapy is one of the most ubiquitous medical therapeutic interventions. There is a debate over whether colloids or crystalloids are better for fluid resuscitation. Recent large trials and meta-analyses suggest no mortality benefit and possible harm with hydroxyethyl starch (HES) use. However, these trials were conducted in critically ill and septic patients and their applicability to perioperative patients has been challenged., Objective: We aimed to evaluate the impact of HES use in scheduled and elective surgical patients., Design: Systematic review and meta-analysis of randomised controlled trials (RCTs)., Eligibility Criteria: Only RCTs comparing the use of the synthetic colloid HES with any crystalloid in adults undergoing noncardiac surgery (up to 24 h postop) were considered eligible. For each eligible trial, we extracted the outcomes of all-cause mortality within 90 days, length of hospital stay, major infectious complications, acute kidney injury (AKI) and renal replacement therapy (RRT)., Results: We identified 1555 citations, selected 90 for full-text evaluation, and identified 13 eligible RCTs. Trials were small (n = 20 to 202) with low event rates. There was a trend to increased mortality with HES within 90 days [13/373 vs. 3/368; risk ratio 2.97; 95% confidence interval (95% CI) 0.96 to 9.19; I = 0%], no difference in AKI and RRT (risk ratio 1.11; 95% CI 0.26 to 4.69; I = 34%), and no difference in major infectious complications (risk ratio 1.19; 95% CI 0.59 to 2.39; I = 0%). Patients resuscitated with HES had a shorter length of hospital stay (mean difference -1.52 days; 95% CI -2.87 to -0.18), although heterogeneity was high (I = 90%)., Conclusion: This meta-analysis, based on small studies with low event rates, suggests that there are currently insufficient data to identify a difference in outcomes associated with crystalloids and HES in scheduled or elective noncardiac surgery.

This article outlines processes followed during a two-day offsite retreat for nurse faculty planned for the purpose of curricular redesign and content mapping. Faculty identified and leveled content for all baccalaureate clinical courses and engaged in dialogue specific to promoting student connections between didactic and clinical outcomes. Faculty developed two core simulation scenarios for every clinical course and leveled learning outcomes for each. Faculty defined where within the curriculum key concepts were introduced and planned depth of knowledge, application, and synthesis from sophomore through senior levels. Evaluation revealed the process led faculty to a stronger overall understanding of the curriculum.

Background: As nursing and healthcare become more global, supported by technology, the opportunities for distance mentoring increase. Mentorship is critical to nurse educator recruitment and retention., Study Objective: The purpose of this study was to identify communication practices of nurse educators involved in mentoring at a distance., Design/settings: A qualitative design, utilizing in-person or telephone interviews was used. Participants were twenty-three protégés or mentors who were part of a yearlong distance mentoring program., Analysis Method: An iterative process of hermeneutic analysis identified three themes; this paper focuses on the theme of connectedness., Results: Participant narratives illuminate practices of connecting at a distance: meeting face-to-face, sharing personal information, experiencing reciprocity, journaling, being vulnerable, establishing one's presence, and appreciating different perspectives., Conclusion: Distance does not appear to limit the connecting potential leading to a meaningful mentoring relationship; rather, it offers possibilities that local mentoring relationships may not. Nurse educators in under-resourced countries, those in small programs without a cadre of senior faculty, and students in distance programs are among those who stand to benefit from distance mentoring relationships., (© 2013.)

Staff development departments are challenged with addressing the entry of new graduates into the work force, updating the competencies of practicing nurses, and providing nurses with educational support to reduce turnover. Knowing how to develop simulations, integrate them into teaching, and effectively assess simulated performance is becoming a core role for educators in health care. For many educators, a knowledge and skill gap exists between the demand for simulation and competence in developing and using simulation. This article shares a cost-effective, three-step train-the-trainer model to prepare nurse educators to use simulation effectively. The three steps include champion identification, champion development, and champion integration. Strategies for addressing the challenges and lessons learned in implementing this model are outlined. The model is an effective and efficient approach for developing a core of champions in any topic area. These champions can then mentor and train others throughout the organization., (Copyright 2013, SLACK Incorporated.)

Driving licence holding in Britain is increasing for older people, particularly older women. Licence holding by men aged 40-69 has saturated at about 90%, and for women aged 30-59 at about 78%. Drivers begin to surrender licences after age 70. By age 90, 38% of women and 25% of men who held a licence at age 70 have surrendered their licences. The paper shows the numbers of car drivers of different ages killed and injured in accidents since 1975 and the fatality and casualty rates per driver and per mile driven. The safety of older drivers is improving faster than that of younger age groups. The number of car driver fatalities aged 70 and over was highest between 1990 and 2004, and has subsequently reduced by almost 40%. For drivers aged 80 and over, the peak was in 2004 and the number has subsequently reduced by almost 50%. The paper uses demographic projections, forecast licence holding and the trends in fatality rates to project the numbers of fatalities for drivers of different ages in Britain. This shows that fatalities among older drivers aged are likely to continue to reduce. Fatality rates for older road users are increased by the fragility of older persons. This disguises the fact that the accident involvement rate for older drivers does not increase with age until after age 75 or 80., (Copyright © 2012. Published by Elsevier Ltd.)

Infection of domestic cats with pandemic H1N1 influenza virus has recently been documented. We conducted a seroprevalence survey and found that 17 of 78 (21.8%) cats sampled during the 2009-2010 influenza season had antibody titers ≥40 against the novel H1N1 strain by hemagglutinin-inhibition assay, compared to only 1 of 39 (2.6%) sampled in 2008 prior to emergence of the pandemic (p = 0.006). Seroprevalance of seasonal H1N1 (41.9%) and H3N2 (25.6%) viruses was similarly high. These data reflecting past infection of household cats raise the possibility that they may act as a vector of influenza transmission within households.

Objective: European countries practice a wide range of car driving license renewal procedures. These range from issuing lifelong licenses without subsequent medical checks, to issuing a license to age 70 and for 3- or 5-year periods thereafter based on self-declarations of medical fitness, to requiring medical examinations for renewal, to renewal every 5 years from the age of 45. This paper presents a case study of the different older driver licensing procedures in seven European countries and addresses the association between these procedures and older driver safety., Method: The seven countries studied consist of France, The Netherlands, the United Kingdom, Denmark, Finland, Norway, and Sweden. The first-mentioned three countries have the most relaxed license renewal procedures and least demanding medical examination requirements., Results: There is no evidence that any license renewal procedure or requirement for a medical examination has an effect on the overall road safety of drivers aged 65+, though undoubtedly there are individual drivers who should no longer be driving who might be detected by stringent renewal procedures. Considering the three countries with the most relaxed licensing procedures, The Netherlands and United Kingdom have the lowest fatality rate for car drivers aged 65+, and the rate for France is falling rapidly., Conclusions: There is also evidence that stringent renewal procedures and demanding medical examinations at renewal reduce the level of car driving licenses among older people. France, The Netherlands, and the United Kingdom have the highest level of driving license holding by people aged 65+, which has direct implications for the independent mobility of older people. Reduced mobility also has safety implications: in about half the European countries for which road accident fatality data have been analyzed, people aged 65+ are at greater risk of death as a pedestrian than as a car driver.

Various interventions have been proposed to address these ongoing needs of HIV-positive patients as they encounter challenges with medication adherence and risk reduction. This report presents the findings of a study that pilots 'DAART+', an intervention that integrates modified directly observed therapy (MDOT), and risk reduction counseling for a population of marginally housed, substance-using persons. The pilot study intended to assess the feasibility of the intervention and to obtain data to assess the intervention's potential effectiveness. The preliminary data reveal that 83% of participants who completed the intervention (n=18) had undetectable viral load (VL) (VL< or =400 copies/mL) which represents a 2.15 log(10) decrease from baseline. Risk behaviors also changed modestly with self-reported increases in condom usage.

Despite well-organized prevention efforts, HIV continues to spread in the United States. Injection drug users (IDUs) and other substance users who engage in high-risk behaviors are at particularly high risk for contracting HIV. Substance abuse(dagger) counselors therefore are in a unique position to present HIV prevention messages to their clients. This article reports the results of a study that surveyed counselors in an urban setting (n = 116) to assess their knowledge of HIV, their current HIV prevention practices, and their attitudes about integrating HIV prevention more fully into their work. The article concludes with a discussion of the survey's limitations and its implications for training.

This study examined changes in service delivery patterns of health and mental health service providers one year after a training on the fundamentals of HIV/AIDS and mental health. Paired t-tests for 424 training recipients showed significant increases in delivery of HIV-related services, and these remained significant while controlling for additional training, job changes, region (urban, rural, suburban), and provider discipline. Multiple logistic regression analysis revealed a significantly greater likelihood of providing direct services to HIV+ individuals among male providers, those with more years of HIV experience, those in counseling disciplines, and those working in a new job since the training.

As people with HIV live longer and healthier lives, ongoing prevention with positive individuals has become a new focus of care. Effective prevention with positives interventions are emerging and new interventions continue to be developed. This article discusses the development and evaluation of the prevention for positives intervention developed for a large AIDS service organization in Chicago. The intervention consists of case manager based HIV prevention education and support within residential and community settings. The article describes the intervention and presents the methods and findings of the program evaluation. The multidimensional evaluation includes formative and process evaluation elements as well as qualitative and quantitative measures (N = 94). The article concludes with a discussion of the challenges and opportunities associated with integrating prevention into care.

Background: Public insurance for testing supplies for self-monitoring of blood glucose is highly variable across Canada. We sought to determine if insured patients were more likely than uninsured patients to use self-monitoring and whether they had better glycemic control., Methods: We used baseline survey and laboratory data from patients enrolled in a randomized controlled trial examining the effect of paying for testing supplies on glycemic control. We recruited patients through community pharmacies in Alberta and Saskatchewan from Nov. 2001 to June 2003. To avoid concerns regarding differences in provincial coverage of self-monitoring and medications, we report the analysis of Alberta patients only., Results: Among our sample of 405 patients, 41% had private or public insurance coverage for self-monitoring testing supplies. Patients with insurance had significantly lower hemoglobin A(1c) concentrations than those without insurance coverage (7.1% v. 7.4%, p = 0.03). Patients with insurance were younger, had a higher income, were less likely to have a high school education and were less likely to be married or living with a partner. In multivariate analyses that controlled for these and other potential confounders, lack of insurance coverage for self-monitoring testing supplies was still significantly associated with higher hemoglobin A(1c) concentrations (adjusted difference 0.5%, p = 0.006)., Interpretation: Patients without insurance for self-monitoring test strips had poorer glycemic control.

New treatment advances have radically altered the course of HIV illness and created new challenges for HIV-affected individuals, families, and communities. This article provides a conceptual framework for understanding HIV in the multiple contexts of the client's culture, strengths, life course, and biomedical progression. The article concludes with a discussion of HIV prevention and treatment adherence as key focal points for social workers and their clients.

Epidemiological trends in HIV infection in the United States suggest existing primary and secondary prevention efforts are inadequate. Healthcare providers may be missing valuable opportunities to engage in necessary public health services, such as prevention education, risk assessment, and case finding. This study examined the HIV-related practice behaviors and training needs of physicians and nurses in the Midwest. A cross-sectional survey method was employed. A questionnaire was mailed to a probability sample of 1,500 physicians and registered nurses licensed as of August 1999 in one of six Midwestern states. A total of 534 physicians and nurses replied to the survey, and over half had received prior HIV-related continuing medical education and training. One third of nurses and 26.8 percent of physicians reported that they did not engage in any HIV-related public health role in their practice. Physicians with prior HIV continuing medical education and training were 3.1 times more likely to report HIV-related public health services in their practice than providers without prior training (p = .004). Nurses with prior experience serving HIV-infected patients were 2.0 times more likely to identify a public health role (p = .012). These findings reflect the need for greater awareness among medical providers of the importance of assuming a public health role with the HIV epidemic.

The social work profession remains an untapped resource within the community for addressing HIV/AIDS prevention and early detection. This study investigated social workers' knowledge and practices relating to HIV prevention education, risk assessment, and case finding. A six page questionnaire was mailed to a random sample of 600 licensed social workers and licensed clinical social workers in Illinois. The sample was drawn from a list provided by the state department of professional regulation, stratified by geographic location. The final response rate for the survey was 51.3% (N = 308). Social work knowledge concerning HIV and AIDS was significantly related to the variables of age, geographic location, and practice setting. Respondents scored lower on knowledge items pertaining to their ability to perform a comprehensive sexual and drug risk assessment and to appropriately advise clients on HIV antibody testing. Most of the social workers in the sample provided minimal to no HIV-related services in their clinical practice. HIV-related behaviors were also significantly related to the variables of age, geographic location, and practice setting. Multiple logistic regression was used to test whether prior HIV/AIDS training predicted the presence of HIV-related practice behaviors, controlling for potential confounding factors. Both prior HIV-related training and education and prior work experience with HIV-infected clients were statistically significant and independent predictors of current HIV-related practice behaviors reported by social workers.

Despite nearly 20 years of HIV prevention efforts, rates of new HIV infection persist at an alarming rate. As successful antiretroviral medications enable many HIV infected persons to live longer, healthier lives, interventions are necessary to support ongoing prevention and reduced risk behaviors. This article describes a survey that was used to assess the opportunities and challenges related to the integration of prevention screening into the work of HIV/AIDS case managers. The article describes the survey, reports the findings (N = 101), and concludes with a discussion of issues that must be addressed prior to incorporating prevention screening into HIV/AIDS case management.

Background: Aspergillus fumigatus is a pathogenic fungus which causes a range of diseases, particularly in the human lung. The pathological mechanism is unknown but may involve a complex mixture of biomolecules which can diffuse from the spore surface. This material is known as A fumigatus diffusate (AfD) and has previously been shown to have a range of immunosuppressive functions. It is hypothesised that AfD may influence the binding of spores to extracellular matrix (ECM) proteins and lung epithelial cells, thereby affecting the ability of the fungus to cause infection., Methods: The binding of spores to ECM proteins and to epithelial cells was carried out using a direct binding assay in microtitre plates and spores were counted by phase contrast microscopy. Rat bronchoalveolar lavage (BAL) fluid was enriched for surfactant protein D (SP-D) using maltose agarose affinity chromatography. The effects of AfD and the SP-D enriched BAL fluid were assessed by pre-incubation with ECM proteins or epithelial cells in the direct binding assay., Results: AfD enhanced the binding of spores to laminin by 137% and to A549 epithelial cells by 250%. SP-D enriched BAL fluid inhibited spore binding to ECM proteins and epithelial cells. Pre-incubation of ECM proteins and epithelial cells with SP-D enriched BAL fluid prevented the enhancement of spore binding by AfD, and pre-incubation of ECM proteins and epithelial cells with AfD prevented the inhibition of spore binding by SP-D enriched BAL fluid. This pretreatment did not prevent the enhancement of spore binding, giving an increase of 95% for collagen I, 80% for fibronectin, 75% for laminin, and 150% for A549 cells., Conclusions: The hypothesis that AfD would affect spore binding to ECM proteins and epithelial cells was confirmed. Rat BAL fluid, with SP-D as the possible bioactive agent, prevented this enhancement. The in vivo significance is unclear but the enhanced binding of spores may increase the chance of fungal infection in the lung which could be prevented by the protective effects of lung surfactant components (possibly SP-D). The results suggest that there may be competition between AfD and a BAL fluid component (possibly SP-D) for the same or similar binding sites on ECM proteins and epithelial cells. Whether this competition occurs in vivo requires further investigation.

Managed care continues to revolutionize the provision of mental health services in the United States. Long-term, open-ended therapies have been replaced by short-term, highly focused interventions. Increasingly, managed care organizations rely on standardized preferred practice guidelines to give direction and focus to social work and other therapeutic interventions. Critics argue that changes effected by managed care, particularly the use of treatment guidelines, depersonalize the client-worker relationship and significantly reduce the role of empathy in the therapeutic process. Moreover, these critics suggest that overall client satisfaction with mental health services has deteriorated. This article presents a study that examined clients' perceptions of empathy and overall satisfaction with managed behavioral health care when the clients were in unstructured individual therapy or in time-limited standardized group therapy. The results reveal no significant difference in the clients' perception of empathy or of their overall satisfaction regardless of the type of treatment they received. This article describes the rationale and design of the study, presents the results, and discusses the implications for social work practice.

This study was promoted following concern over increasing mortality on 2 farms rearing Atlantic salmon Salmo salar in the Shetland Isles, Scotland. A Mycobacterium sp. was isolated from moribund, market-sized Atlantic salmon. Biochemical tests, lipid analysis and PCR (polymerase chain reaction) techniques confirmed the bacterium to be Mycobacterium chelonae. Multiple greyish-white miliary granuloma-like nodules were observed in several tissues. Dense hard-packed nodules contained abundant acid-fast bacteria. Atlantic salmon injected with M. chelonae remained sub-clinically infected, demonstrating the chronic nature of this disease. The source of the pathogen was not identified.

Background: The fungus Aspergillus fumigatus, whose spores are present ubiquitously in the air, causes a range of diseases in the human lung. A small molecular weight (< 10 kD) heat stable toxin released from the spores of clinical and environmental isolates of A fumigatus within minutes of deposition in aqueous solution has previously been described. A key effect of the toxin was to inhibit the oxidative burst of macrophages as measured by superoxide anion release. It was hypothesised that the toxin was one of the commonly found A fumigatus hyphal toxins such as gliotoxin. This inhibitor may be an important factor which allows the fungus to colonise the lung., Methods: The spore derived inhibitor was shown to inhibit the respiratory burst of rat alveolar macrophages, as measured by the generation of superoxide anion. Samples of the spore diffusate were subject to reversed phase high performance liquid chromatography (HPLC), thin layer chromatography (TLC), high performance thin layer chromatography (HPTLC), or organic extraction followed by TLC or HPLC to identify the presence of gliotoxin, fumagillin, helvolic acid, fumigaclavine-C, and aurasperone-C. Commercially obtained preparations of the toxins gliotoxin, fumagillin and helvolic acid and extracts enriched for fumigaclavine-C and aurasperone-C were used as internal and external standards and in the respiratory burst measurements., Results: Gliotoxin, fumagillin, helvolic acid, fumigaclavine-C, and aurasperone-C were not detected in spore derived diffusate using PHLC or TLC. Using extraction procedures with solvents known to extract gliotoxin from A fumigatus culture supernatants, no gliotoxin was detected in the spore derived diffusate. Commercial gliotoxin, fumagillin, and helvolic acid or extracts enriched for fumigaclavine-C and aurasperone-C did not inhibit the oxidative burst of macrophages., Conclusions: The hypothesis that the spore derived toxin is one of the toxins derived from hyphae such as gliotoxin, helvolic acid, fumagillin, fumigaclavine-C, or aurasperone-C is not proved. The spore toxin may exert its effect through its ability to diffuse rapidly into the lung lining fluid, diminish the macrophage oxidative burst, and play a part in allowing A fumigatus to persist in the lung and manifest its well known pathogenic effects.

Background: Aspergillus fumigatus is a fungus that grows on dead and decaying organic matter in the environment and whose spores are present ubiquitously in the air. The fungus causes a range of diseases in the human lung. A study was undertaken to demonstrate and partially characterise an inhibitor of the macrophage respiratory burst from the surface of A fumigatus spores that could be an important factor in allowing the fungus to colonise the lung., Methods: The spore-derived inhibitor of the respiratory burst of rat alveolar macrophages, as measured by generation of superoxide anion, was demonstrated in Hank's balanced salt solution extracts of four clinical isolates and an environmental isolate of A fumigatus. The time course of the release of the inhibitor into aqueous solution was assessed and the cytotoxic potential of the spore-derived inhibitor towards macrophages was tested using the propidium iodide method. An oxygen electrode was used to confirm the superoxide anion measurements. Molecular weight cutoff filters were used to determine the size of the inhibitor as assessed in the respiratory burst assay and also by its ability to inhibit macrophage spreading on glass. The crude diffusate from the spore surface was fractionated by reversed phase high pressure liquid chromatography (HPLC) and the fractions analysed for inhibitory activity, protein, and carbohydrate content., Results: A small molecular weight (< 10 kD) heat stable toxin was released from the spores of clinical and environmental isolates of A fumigatus within minutes of deposition in aqueous solution. The key effect of the toxin demonstrated here was its ability to inhibit the oxidative burst of macrophages as measured by superoxide anion release. The inhibition was not due to cell death or detectable loss of membrane integrity as measured by permeability to propidium iodide. The toxin was not a scavenger of superoxide anion. Oxygen electrode studies suggested indirectly that the inhibitor acted to inhibit the assembly of the macrophage NADPH-oxidase complex. Fractions of < 10 kD also inhibited the spreading of alveolar macrophages, confirming that the toxin had an additional effect on macrophages that leads to loss of adherence or impairment of cytoskeletal function. In reversed phase HPLC fractions the inhibitory activity eluted with an associated carbohydrate, although the exact chemical nature of the toxin remains to be elucidated., Conclusions: This spore toxin may, through its ability to diffuse rapidly into lung lining fluid, diminish the macrophage respiratory burst and play a part in allowing A fumigatus to persist in the lung and manifest its well known pathogenic effects. Future research will be focused on further molecular characterisation of the toxin and elaboration of the effect of the toxin on intracellular signalling pathways involved in the activation of alveolar macrophages.

A multienzyme complex of tricarboxylic acid cycle enzymes, catalysing the consecutive reactions from fumarate to 2-oxoglutarate, has been identified in extracts of Pseudomonas aeruginosa prepared by gentle osmotic lysis of the cells. The individual enzyme activities of fumarase, malate dehydrogenase, citrate synthase, aconitase and isocitrate dehydrogenase can be used to reconstitute the complex. The citrate synthase isoenzymes, CSI and CSII, from this organism can be used either together or as the individual activities to reconstitute the complex. No complex can be reformed in the absence of CSI or CSII. Which CS isoenzyme predominates in the complex depends on the phase of growth at which the cells were harvested and the extract prepared. More CSI was found in the complex during exponential growth, whereas CSII predominated during the stationary phase. The results support the idea of a 'metabolon' in this organism, with the composition of the CS component varying during the growth cycle.

Two types of citrate synthase (CS) have been purified from Pseudomonas aeruginosa, a 'large' form (CSI) and a 'small' form (CSII). The M(r)s of the CSI and CSII isoenzymes were determined to be 240,000 +/- 16,000 (mean +/- S.E.M.) and 80,300 +/- 3800 respectively. Chemical cross-linking of the native enzymes with either dimethyl suberimidate or glutaraldehyde followed by electrophoretic analysis by SDS/PAGE showed that CSI is a hexamer and CSII is a dimer. SDS/PAGE showed that CSI and CSII each consist of a single subunit type, of M(r) 42,000 +/- 2000 and M(r) 36,500 +/- 2000 respectively. CSI and CSII were also shown to be distinct kinetically, immunologically and in terms of their regulatory properties. It is suggested that the CS isoenzymes are products of different structural genes.

Glass fibre bandages are now commonly used for definitive below-knee walking casts, in preference to plaster of Paris, since they are better able to withstand the stresses imposed upon them. This paper describes a technique for recording the cast strains in glass fibre bandages and shows typical stress levels in patient and volunteer casts. A typical map of the stress variations in walking casts during gait in a volunteer has been produced. The study shows that the high stresses recorded along the medial and lateral borders of the foot by the strain gauge technique are confirmed by clinical experience, since this is where failure is most commonly seen in below-knee walking casts. It is concluded that the current generation of polyurethane impregnated glass fibre splinting bandages are too brittle in many cases for below-knee walking casts and that a more flexible fabric would reduce the incidence of cast breakdown., (Copyright © 1990. Published by Elsevier Ltd.)

The nature of the determinants recognized by a panel of monoclonal antibodies (MAbs) raised against a cell wall antigen of Streptococcus mutans (SA I/II) was investigated. Mild periodate oxidation of SA I/II showed that MAbs Guy 1, 2, 3, and 5 recognized carbohydrate epitopes on the antigen. Glycosidases were used to identify the nature of the sugars involved in their binding. Treatment with beta-glucosidase inhibited the binding of Guy 1, 2, 3, and 5 by 90%. No competition was found for any of the MAbs between SA I/II and a series of carbohydrates, including the serotype c polysaccharide from S. mutans. The results show that MAbs Guy 1, 2, 3, and 5 recognize carbohydrate epitopes on SA I/II which are distinct from the serotype polysaccharide. The other MAbs recognized protein epitopes on SA I/II.

Cellular immunity to the hepatitis B surface antigen (HBsAg) and a liver-specific lipoprotein was studied, using the leucocyte migration test, in 38 asymptomatic blood donors found to have HBsAg in the serum. Sensitization to HBsAg was found in 26% and was related to the presence of liver damage, being detected in 47% of those with elevated serum aspartate aminotransferase but in only 13% with normal enzyme levels. The frequency of sensitization to this antigen in those with chronic persistent or chronic aggressive hepatitis on biopsy was also higher than in those with unrelated or minimal changes. The findings using the liver-specific lipoprotein as antigen were similar and there was a correlation between the results obtained with this and the hapatitis B surface antigen. This study supports the hypothesis that a T-lymphocyte response to hepatitis B virus antigen can initiate an autoimmune reaction to antigens such as liver-specific lipoprotein on the hepatocyte surface, and that this reaction may be of importance in the production of chronic liver damage. In the absence of the T-cell response, the autoimmune reaction cannot occur and the virus is able to establish a harmless symbiotic union with the host.

Citrate synthase was purified from Acinetobacter calcoaceticus and treated with the cleavable cross-linking reagent dithiobis(succinimidyl propionate). Cross-linking of the enzyme resulted in the abolition of the sigmoidal responses to inhibition by NADH and re-activation by AMP displayed by the native enzyme. Inhibition and re-activation were still observed but without any cooperativity. Cleavage of the disulphide bonds in the cross-links by treatment with dithiothreitol restored the sigmoidal characteristics of both inhibition and re-activation.

A 185,000 MW glycoprotein antigen derived from Streptococcus mutans was digested with subtilisin. Purification by reversed phase high-powered liquid chromatography (HPLC) resulted in a homogeneous 20,000 MW protein which possesses the streptococcal antigen (SA) I and II determinants. This protein was immunogenic in mice both for the 20,000 MW and the native 185,000 MW SA. Further proteolysis with subtilisin generated four peptides of 18,000, 10,000, 6000 and 4000 MW. Whereas the 20,000, 18,000 and 10,000 MW peptides contained both SAI and SAII determinants, the 4000 MW peptide possessed only the SAI and the 6000 MW peptide the SAII determinant. The 4000-SAI peptide is of special significance, as the smallest SAI material separated in the past was 150,000 MW. This was difficult to purify and proved to be protective against dental caries on immunization of rhesus monkeys.

Two forms of citrate synthase (EC 4.1.3.7) have been found in several species of Pseudomonas, a 'large' form (Mr congruent to 250,000) which is generally inhibited by NADH and reactivated by AMP, and a 'small' form (Mr congruent to 100,000) which is insensitive to these nucleotide effectors. Other species of Pseudomonas were found to contain either the 'large' or the 'small' form. Gel filtration and ion-exchange with the technique of fast protein liquid chromatography were used to resolve the enzymes. Where both citrate synthases were present, there did not appear to be an equilibrium between the two forms. The results reveal a new and complex diversity of citrate synthase within the genus Pseudomonas.

Citrate synthase from Acinetobacter calcoaceticus was subjected to proteolysis with subtilisin. Although the enzyme proved relatively resistant to inactivation by this treatment, SDS-polyacrylamide gel electrophoresis clearly revealed breakdown of the citrate synthase to smaller fragments. The regulatory responses of the native enzyme to inhibition by NADH and re-activation by AMP were retained on proteolysis, indicating that the fragments bind tightly to each other and preserve the overall cooperative molecular interactions.

The hepatitis-B surface antigen (HBsAG) may be persistently present in the serum in a few cases of active chronic hepatitis but the cause of the disease in most patients is unknown. In a study of 39 HBsAg-negative cases cell-mediated immunity to HBsAg was observed in 24 (62%), suggesting a high frequency of previous infection with the hepatitis-B virus. Hepatitis-B surface antibody was detectable by radioimmunoassay in six patients, in all of whom complexes of HBsAg were present in the serum on electron microscopy. Out of 12 patients with HBsAg-positive active chronic hepatitis who were also studied eight, including all those untreated at the time, showed a cellular response to the antigen. Evidence of sensitization to a liver-specific cell surface lipoprotein was found with similar frequency in the two groups. These results are consistent with the hypothesis that hepatitis-B virus infection is important in initiating the disease in many cases of active chronic hepatitis and that sensitization to the liver cell membrane antigen is the autoimmune process responsible for the perpetuation of the liver injury.

The effect of dissolved oxygen concentration on the metabolism of glucose in Pseudomonas aeruginosa was studied with chemostat cultures using both single-step and gradual transitions from either ammonium or glucose limitation to oxygen limitation and studying transient and steady states. The pathway of glucose metabolism was regulated by the availability of oxygen. The organism responded to oxygen limitation by adjusting its metabolism of glucose from the extracellular direct oxidative pathway, which produces gluconate and 2-oxogluconate, to the intracellular phosphorylative route. This change was a consequence of decreased activities of glucose dehydrogenase and gluconate dehydrogenase and of the transport systems for gluconate and 2-oxogluconate, and an increased activity of glucose transport, while relatively high activities of hexokinase and glucose-6-phosphate dehydrogenase were maintained. Citrate synthase, isocitrate dehydrogenase and malate dehydrogenase activities responded to changes in dissolved oxygen concentration rather than to changes in the glucose or ammonium concentrations. The effect of oxygen limitation on the oxo-acid dehydrogenases and aconitase was probably due, wholly or in part, to repression by glucose consequent upon the increase in residual glucose concentration. Succinate dehydrogenase was repressed by an increase in ammonium concentration under an oxygen limitation.

Fast protein liquid chromatography (FPLC) has been shown to be a rapid and effective method of separating isoenzymes of citrate synthase and isocitrate dehydrogenase in extracts of Pseudomonas aeruginosa and Acinetobacter calcoaceticus. The advantages of FPLC over conventional methods of fractionation are discussed and it is suggested that this may be a valuable and more general technique for isoenzyme resolution.