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KCNE3 is a single-pass integral membrane protein that regulates numerous voltage-gated potassium channel functions such as KCNQ1. Previous solution NMR studies suggested a moderate degree of curved α-helical structure in the transmembrane domain (TMD) of KCNE3 in lyso-myristoylphosphatidylcholine (LMPC) micelles and isotropic bicelles with the residues T71, S74 and G78 situated along the concave face of the curved helix. During the interaction of KCNE3 and KCNQ1, KCNE3 pushes its transmembrane domain against KCNQ1 to lock the voltage sensor in its depolarized conformation. A cryo-EM study of KCNE3 complexed with KCNQ1 in nanodiscs suggested a deviation of the KCNE3 structure from its independent structure in isotropic bicelles. Despite the biological significance of KCNE3 TMD, the conformational properties of KCNE3 are poorly understood. Here, all atom molecular dynamics (MD) simulations were utilized to investigate the conformational dynamics of the transmembrane domain of KCNE3 in a lipid bilayer containing a mixture of POPC and POPG lipids (3:1). Further, the effect of the interaction impairing mutations (V72A, I76A and F68A) on the conformational properties of the KCNE3 TMD in lipid bilayers was investigated. Our MD simulation results suggest that the KCNE3 TMD adopts a nearly linear α helical structural conformation in POPC-POPG lipid bilayers. Additionally, the results showed no significant change in the nearly linear α-helical conformation of KCNE3 TMD in the presence of interaction impairing mutations within the sampled time frame. The KCNE3 TMD is more stable with lower flexibility in comparison to the N-terminal and C-terminal of KCNE3 in lipid bilayers. The overall conformational flexibility of KCNE3 also varies in the presence of the interaction-impairing mutations. The MD simulation data further suggest that the membrane bilayer width is similar for wild-type KCNE3 and KCNE3 containing mutations. The Z-distance measurement data revealed that the TMD residue site A69 is close to the lipid bilayer center, and residue sites S57 and S82 are close to the surfaces of the lipid bilayer membrane for wild-type KCNE3 and KCNE3 containing interaction-impairing mutations. These results agree with earlier KCNE3 biophysical studies. The results of these MD simulations will provide complementary data to the experimental outcomes of KCNE3 to help understand its conformational dynamic properties in a more native lipid bilayer environment.

Background: The purpose of the current study was to determine the utility of early follicular phase follicle-stimulating hormone (FSH) testing in patients undergoing in vitro fertilization (IVF)., Methods: This was a retrospective review of patients from 2012 to 2015 at Mayo Clinic in Rochester, Minnesota, USA. Included subjects had a normal anti-Müllerian hormone (AMH) of 1 to 9 ng/ml and antral follicle count (AFC) of 10 to 29. Patients were stratified by FSH level when associated estradiol was less than 50 ng/ml . In total, 225 patients were categorized into three groups: high FSH (FSH ≥10 IU/L ; n= 36), normal FSH (>5 IU/L and <10 IU/L ; n=170), and low FSH (FSH ≤5 IU/L ; n= 19). ANOVA and multiple logistic regression were used for statistical comparisons and for evaluation of the relationships between variables; significance level was set at <0.05., Results: There were no significant differences in demographics, IVF cycle type, or peak estradiol level between the groups. Patients with a high basal FSH level had a similar clinical pregnancy rate and live birth rate compared to controls and patients with low FSH. High FSH level was associated with decreased follicular development (17 versus 22; p<0.01), oocyte yield (15 versus 18; p=0.02), and embryo yield (8 versus 10; p=0.04) despite higher total doses of gonadotropins., Conclusion: Patients with normal AMH and AFC levels could be further stratified into lower responders and starting doses of medications can be adjusted based on high basal FSH levels. Therefore, it is suggested to counsel patients on pregnancy outcomes which seem to be quite similar regardless of the FSH level., Competing Interests: Conflict of Interest The authors have no relevant financial or non-financial interests to disclose., (Copyright© 2023, Avicenna Research Institute.)

Background: Brugada syndrome (BrS) is an inherited arrhythmia syndrome caused by loss-of-function variants in the cardiac sodium channel gene SCN5A (sodium voltage-gated channel alpha subunit 5) in ≈20% of subjects. We identified a family with 4 individuals diagnosed with BrS harboring the rare G145R missense variant in the cardiac transcription factor TBX5 (T-box transcription factor 5) and no SCN5A variant., Methods: We generated induced pluripotent stem cells (iPSCs) from 2 members of a family carrying TBX5-G145R and diagnosed with Brugada syndrome. After differentiation to iPSC-derived cardiomyocytes (iPSC-CMs), electrophysiologic characteristics were assessed by voltage- and current-clamp experiments (n=9 to 21 cells per group) and transcriptional differences by RNA sequencing (n=3 samples per group), and compared with iPSC-CMs in which G145R was corrected by CRISPR/Cas9 approaches. The role of platelet-derived growth factor (PDGF)/phosphoinositide 3-kinase (PI3K) pathway was elucidated by small molecule perturbation. The rate-corrected QT (QTc) interval association with serum PDGF was tested in the Framingham Heart Study cohort (n=1893 individuals)., Results: TBX5-G145R reduced transcriptional activity and caused multiple electrophysiologic abnormalities, including decreased peak and enhanced "late" cardiac sodium current (I Na ), which were entirely corrected by editing G145R to wild-type. Transcriptional profiling and functional assays in genome-unedited and -edited iPSC-CMs showed direct SCN5A down-regulation caused decreased peak I Na , and that reduced PDGF receptor ( PDGFRA [platelet-derived growth factor receptor α]) expression and blunted signal transduction to PI3K was implicated in enhanced late I Na . Tbx5 regulation of the PDGF axis increased arrhythmia risk due to disruption of PDGF signaling and was conserved in murine model systems. PDGF receptor blockade markedly prolonged normal iPSC-CM action potentials and plasma levels of PDGF in the Framingham Heart Study were inversely correlated with the QTc interval ( P <0.001)., Conclusions: These results not only establish decreased SCN5A transcription by the TBX5 variant as a cause of BrS, but also reveal a new general transcriptional mechanism of arrhythmogenesis of enhanced late sodium current caused by reduced PDGF receptor-mediated PI3K signaling.

Introduction: In the United States, 19 states permit recreational use of cannabis, with 16 more permitting medical use (Marijuana Policy Project, 2021). Concerns remain about whether liberalized policies result in increased adolescent cannabis use. To date, limited evidence exists that the statewide prevalence of adolescent cannabis use increased in states with liberalized policies. However, analyses at local levels show some negative impacts. Thus, we analyzed if living in a ZIP code with a dispensary (ZCWD) was associated with adolescent cannabis use., Methods: Dispensary ZIP codes from public records were matched to self- reported ZIP codes on the Illinois Youth Survey (IYS). We compared past 30-day and past-year cannabis use among youth living in a ZCWD and not living in a ZCWD., Results: About one in eight adolescents (12.8%, n = 1,348) in the weighted sample (n=10,569) resided in a ZCWD. Overall, past 30-day use was lower among youth who lived in ZIP codes with dispensaries (OR = .69, p < .05), with variation by grade. For example, only 10 th (OR = .62, p < .05) and 12 th graders (OR = .59, p < .05) living in a ZCWD had lower odds of past 30-day cannabis use. Additionally, only 12 th graders in a ZCWD had lower odds of past-year use (OR = .70, p < .05). Finally, suburban youth living in a ZCWD also had lower odds of cannabis use (OR = .54, p < .01)., Conclusion/discussion: Cannabis use was significantly lower among 10 th and 12 th graders living in a ZCWD. Additional research should continue to monitor evolving state policies and whether they are associated with adolescent cannabis use., (© 2022 Authors et al.)

KCNE3 is a potassium channel accessory transmembrane protein that regulates the function of various voltage-gated potassium channels such as KCNQ1. KCNE3 plays an important role in the recycling of potassium ion by binding with KCNQ1. KCNE3 can be found in the small intestine, colon, and in the human heart. Despite its biological significance, there is little information on the structural dynamics of KCNE3 in native-like membrane environments. Molecular dynamics (MD) simulations are a widely used as a tool to study the conformational dynamics and interactions of proteins with lipid membranes. In this study, we have utilized all-atom molecular dynamics simulations to characterize the molecular motions and the interactions of KCNE3 in a bilayer composed of: a mixture of POPC and POPG lipids (3:1), POPC alone, and DMPC alone. Our MD simulation results suggested that the transmembrane domain (TMD) of KCNE3 is less flexible and more stable when compared to the N- and C-termini of KCNE3 in all three membrane environments. The conformational flexibility of N- and C-termini varies across these three lipid environments. The MD simulation results further suggested that the TMD of KCNE3 spans the membrane width, having residue A69 close to the center of the lipid bilayers and residues S57 and S82 close to the lipid bilayer membrane surfaces. These results are consistent with previous biophysical studies of KCNE3. The outcomes of these MD simulations will help design biophysical experiments and complement the experimental data obtained on KCNE3 to obtain a more detailed understanding of its structural dynamics in the native membrane environment.

Single-cell RNA-seq and single-cell assay for transposase-accessible chromatin (ATAC-seq) technologies are used extensively to create cell type atlases for a wide range of organisms, tissues, and disease processes. To increase the scale of these atlases, lower the cost and pave the way for more specialized multiome assays, custom droplet microfluidics may provide solutions complementary to commercial setups. We developed HyDrop, a flexible and open-source droplet microfluidic platform encompassing three protocols. The first protocol involves creating dissolvable hydrogel beads with custom oligos that can be released in the droplets. In the second protocol, we demonstrate the use of these beads for HyDrop-ATAC, a low-cost noncommercial scATAC-seq protocol in droplets. After validating HyDrop-ATAC, we applied it to flash-frozen mouse cortex and generated 7996 high-quality single-cell chromatin accessibility profiles in a single run. In the third protocol, we adapt both the reaction chemistry and the capture sequence of the barcoded hydrogel bead to capture mRNA, and demonstrate a significant improvement in throughput and sensitivity compared to previous open-source droplet-based scRNA-seq assays (Drop-seq and inDrop). Similarly, we applied HyDrop-RNA to flash-frozen mouse cortex and generated 9508 single-cell transcriptomes closely matching reference single-cell gene expression data. Finally, we leveraged HyDrop-RNA's high capture rate to analyze a small population of fluorescence-activated cell sorted neurons from the Drosophila brain, confirming the protocol's applicability to low input samples and small cells. HyDrop is currently capable of generating single-cell data in high throughput and at a reduced cost compared to commercial methods, and we envision that HyDrop can be further developed to be compatible with novel (multi) omics protocols., Competing Interests: FD, JI, CB, GH, CF, JJ, KT, VC, JW, GM, Jd, SP, SA No competing interests declared, (© 2022, De Rop et al.)

Introduction: Orthodontic residents face challenges unparalleled to their predecessors, including competitive marketplaces, rising debt burdens, and changing demographics that have contributed to the shift in initial career choice from owner to employee. We aim to understand factors important to orthodontists at different career stages and the impact on job satisfaction through a sequential mixed methodology study., Methods: Semistructured interviews were conducted with 25 orthodontic residents and practitioners. A survey was developed from qualitative findings that explored career decision-making of orthodontic residents and practicing orthodontists, with descriptive and bivariate statistical analyses (n = 343 orthodontists and 185 residents)., Results: Graduating orthodontic residents are choosing employment over ownership as their initial job, prioritizing high income to offset the educational debt. The majority of residents report thinking about their debt very often to all of the time and find it very to extremely stressful. Current residents have long-term goals of ownership, whereas practicing doctors of both genders became owners at equal frequencies. However, on average, women earn $119,000 less per year and report being the primary family caretaker more often than men., Conclusions: Excellent patient care, work-life balance, and sufficient income are the most important drivers for career choices and job satisfaction for orthodontists of all ages. Despite debt, most residents and working orthodontists report a high level of job satisfaction and would recommend the field of orthodontics to others., (Copyright © 2021 American Association of Orthodontists. Published by Elsevier Inc. All rights reserved.)

Background: Although rates of nonmedical opioid use are highest in late adolescence and emerging adulthood, efforts to understand the extent of the heterogeneity in opioid misuse during this time have been limited. The current study aimed to derive and define typologies of opioid use in high school students at the onset of emerging adulthood., Methods: Survey responses from a statewide sample of high school students aged 18 and 19 (N = 26,223) were analyzed. Group-based comparisons between participants reporting opioid use and those not reporting opioid use were conducted. Among those reporting opioid use (n = 1,636), we conducted a latent class analysis (LCA) to identify heterogeneous subgroups of opioid users on the basis of non-medical use of prescription opioids (NMUPO) and heroin use. The resulting classes were then compared across various risk and protective factors using multinominal logistic regression., Results: Consistent differences were observed between participants using opioids and participants not using opioids, with moderate to large effect sizes. Results from LCA revealed three subclasses: NMUPO-Any Use, NMUPO To Get High, and Heroin Use. Subclass differences were observed for non-opioid substance use, mental health, and demographics., Conclusions: Findings from this study underscore the variability of youth who engage in opioid use in late adolescence. Results also indicate that opioid use during adolescence is likely indicative of a broader set of substance use and mental health issues., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Research using mHealth apps has the potential to positively impact health care management and outcomes. However, choosing an appropriate mHealth app may be challenging for the health researcher. The author team used existing evaluation tools, checklists, and guidelines to assess selected mHealth apps to identify strengths, challenges, and potential gaps within existing evaluation tools. They identified specific evaluation tool components, questions, and items most effective in examining app content, usability, and features, including literacy demand and cultural appropriateness; technical information; practical aspects of app functionality; and evolving capabilities of mobile medical apps. Challenges included the subjective nature of the results, time required to complete the evaluation, lack of emphasis on evidence-based content, and inadequate tool flexibility. Health researchers considering the integration of mobile apps into research will benefit from evaluation tools that assess both evidence-based content and the ability of the mobile app to securely integrate with other digital technologies involved in patient care. Next steps will include the involvement of health care providers and professionals, including nurses a wide range of expertise, to develop an mHealth evaluation tool that focuses on identifying quality, evidence-based mobile apps into patient outcomes research., (© 2019 John Wiley & Sons Ltd.)

Surgical treatment of wrist arthritis in the younger patient population remains a challenging issue, and various surgical options need to be carefully considered for each patient. Proximal row carpectomy and 4-corner arthrodesis have proven to be reliable options for reducing pain and restoring adequate function in most young high-demand patients if the lunate facet is spared. Selective neurectomy has proven to be a promising stand-alone or complementary procedure for the treatment of pain that is associated with wrist arthritis in patients of all ages, with spared motion and the opportunity to perform additional procedures if pain continues. Alternative procedures, including capitolunate arthrodesis, total wrist arthrodesis, radial styloidectomy, total wrist arthroplasty, and wrist hemiarthroplasty, have considerable strengths and weaknesses and need to be studied further in younger patients.

Background: Dementia is the leading cause of disability worldwide, and interventions aimed at reducing the prevalence and burden of the disease are urgently needed. Maintain Your Brain (MYB) is a randomized controlled trial of a multimodal digital health intervention targeting modifiable dementia risk factors to combat cognitive decline and potentially prevent dementia. In addition to behavioral modules targeting mood, nutrition, and physical exercise, a new Brain Training System (BTS) will deliver computerized cognitive training (CCT) throughout the trial to provide systematic, challenging, and personally adaptive cognitive activity., Objective: This paper aimed to describe the design and development of BTS., Methods: BTS has been designed with a central focus on the end user. Raw training content is provided by our partner NeuroNation and delivered in several innovative ways. A baseline cognitive profile directs selection and sequencing of exercises within and between sessions and is updated during the 10-week 30-session module. Online trainers are available to provide supervision at different levels of engagement, including face-to-face share-screen coaching, a key implementation resource that is triaged by a "red flag" system for automatic tracking of user adherence and engagement, or through user-initiated help requests. Individualized and comparative feedback is provided to aid motivation and, for the first time, establish a social support network for the user based on their real-world circle of friends and family., Results: The MYB pilot was performed from November 2017 to March 2018. We are currently analyzing data from this pilot trial (n=100), which will make up a separate research paper. The main trial was launched in June 2018. Process and implementation data from the first training module (September to November 2018) are expected to be reported in 2019 and final trial outcomes are anticipated in 2022., Conclusions: The BTS implemented in MYB is focused on maximizing adherence and engagement with CCT over the short and long term in the setting of a fully digital trial, which, if successful, could be delivered economically at scale., Trial Registration: Australian New Zealand Clinical Trials Registry ACTRN12618000851268; https://www.anzctr.org.au /Trial/Registration/TrialReview.aspx?id=370631&isReview=true., (©Courtney Campbell Walton, Amit Lampit, Christos Boulamatsis, Harry Hallock, Polly Barr, Jeewani Anupama Ginige, Henry Brodaty, Tiffany Chau, Megan Heffernan, Perminder Singh Sachdev, Maria A Fiatarone Singh, Michael Valenzuela. Originally published in JMIR Aging (http://aging.jmir.org), 27.02.2019.)

Many women of reproductive age in the United States are marginally iodine deficient, perhaps because the salt in processed foods is not iodized. Iodine deficiency, per se, can interfere with normal brain development in their offspring; in addition, it increases vulnerability to the effects of certain environmental pollutants, such as nitrate, thiocyanate, and perchlorate. Although pregnant and lactating women should take a supplement containing adequate iodide, only about 15% do so. Such supplements, however, may not contain enough iodide and may not be labeled accurately. The American Thyroid Association recommends that pregnant and lactating women take a supplement with adequate iodide. The American Academy of Pediatrics recommends that pregnant and lactating women also avoid exposure to excess nitrate, which would usually occur from contaminated well water, and thiocyanate, which is in cigarette smoke. Perchlorate is currently a candidate for regulation as a water pollutant. The Environmental Protection Agency should proceed with appropriate regulation, and the Food and Drug Administration should address the mislabeling of the iodine content of prenatal/lactation supplements.

A concerted effort to control malaria in Africa has produced dramatic reductions in childhood death in the past decade. This early success has prompted the global community to commit to eradication of malaria deaths and eventually all transmission. Evidence suggests that this is a feasible goal using currently available interventions, augmented with newer tools such as vaccines, which are in development. Malaria deaths are entirely preventable now, and our sustained political and financial commitment to continue to prevent these deaths hangs in the balance.

Purpose: To report a case of peripheral ulcerative keratitis and necrotizing scleritis precipitated by trauma in a patient with mixed cryoglobulinemia due to hepatitis C viral infection., Methods: Case report and literature review., Results: A 62-year-old man with a history of mixed cryoglobulinemia developed an episode of necrotizing scleritis and peripheral ulcerative keratitis one month after repair of a traumatic scleral defect with patch grafting. This episode resolved following treatment with high-dose corticosteroids and the patient underwent successful repeat patch grafting along with a free conjunctival autograft. This is the second reported case of necrotizing scleritis and peripheral ulcerative keratitis associated with mixed cryoglobulinemia., Conclusion: Ophthalmologists should be aware of the association between mixed cryoglobulinemia and necrotizing scleritis/peripheral ulcerative keratitis. Patients with this condition experiencing ocular trauma or undergoing ocular surgery should be monitored closely.

Purpose: To report a case of corneal melting in a patient with fornix foreshortening being treated with reformulated generic diclofenac., Methods: Case report., Results: An asymptomatic 76-year old man presented with central corneal melting while being treated with reformulated generic diclofenac. This formulation did not contain the vitamin E derivative, tocophersolan, which has been felt to contribute to corneal melting associated with generic diclofenac in the past. Other factors for corneal melting included multiple medication use and altered forniceal architecture, but did not include previous corneal or cataract surgery. Following discontinuation of the topical medications and temporary tarsorrhapy, the defect reepithelialized., Conclusion: In patients with a compromised ocular surface, reformulated generic diclofenac has the potential to cause corneal melting without prior cataract or refractive surgery. Use should not be indiscriminate or without close supervision.

Background: Since 2005, malaria control scale-up has progressed in many African countries. Controlled studies of insecticide-treated mosquito nets (ITNs), indoor residual spraying (IRS), intermittent preventive treatment during pregnancy (IPTp) and malaria case management suggested that when incorporated into national programmes a dramatic health impact, likely more than a 20% decrease in all-cause childhood mortality, was possible. To assess the extent to which national malaria programmes are achieving impact the authors reviewed African country programme data available through 2009., Methods: National survey data, published literature, and organization or country reports produced during 2000-2009 were reviewed to assess available malaria financing, intervention delivery, household or target population coverage, and reported health benefits including infection, illness, severe anaemia, and death., Results: By the end of 2009, reports were available for ITN household ownership (n = 34) and IPTp use (n = 27) in malaria-endemic countries in Africa, with at least two estimates (pre-2005 and post-2005 intervals). Information linking IRS and case management coverage to impact were more limited. There was generally at least a three-fold increase in household ITN ownership across these countries between pre-2005 (median of 2.4% of households with at least one ITN) and post-2005 (median of 32.5% of households with at least one ITN). Ten countries had temporal data to assess programme impact, and all reported progress on at least one impact indicator (typically on mortality); in under-five year mortality rates most observed a decline of more than 20%. The causal relationship between malaria programme scale-up and reduced child illness and mortality rates is supported by biologic plausibility including mortality declines consistent with experience from intervention efficacy trials, consistency of findings across multiple countries and different epidemiologic settings, and temporal congruity where morbidity and mortality declines have been documented in the 18 to 36 months following intervention scale-up., Conclusions: Several factors potentially have contributed to recent health improvement in African countries, but there is substantial evidence that achieving high malaria control intervention coverage, especially with ITNs and targeted IRS, has been the leading contributor to reduced child mortality. The documented impact provides the evidence required to support a global commitment to the expansion and long-term investment in malaria control to sustain and increase the health impact that malaria control is producing in Africa.

Zambia national survey, administrative, health facility, and special study data were used to assess progress and impact in national malaria control between 2000 and 2008. Zambia malaria financial support expanded from US$9 million in 2003 to US$ approximately 40 million in 2008. High malaria prevention coverage was achieved and extended to poor and rural areas. Increasing coverage was consistent in time and location with reductions in child (age 6-59 months) parasitemia and severe anemia (53% and 68% reductions, respectively, from 2006 to 2008) and with lower post-neonatal infant and 1-4 years of age child mortality (38% and 36% reductions between 2001/2 and 2007 survey estimates). Zambia has dramatically reduced malaria transmission, disease, and child mortality burden through rapid national scale-up of effective interventions. Sustained progress toward malaria elimination will require maintaining high prevention coverage and further reducing transmission by actively searching for and treating infected people who harbor malaria parasites.

Objective: To characterize the epidemiology and microbiology of ventilator-associated pneumonia (VAP) in a long-term acute care hospital (LTACH)., Design: Retrospective study of prospectively identified cases of VAP., Setting: Single-center, 207-bed LTACH with the capacity to house 42 patients requiring mechanical ventilation, evaluated from April 1, 2006, through January 31, 2008., Methods: Data on the occurrence of VAP were collected prospectively as part of routine infection surveillance at Radius Specialty Hospital. After March 2006, Radius Specialty Hospital implemented a bundle of interventions for the prevention of VAP (hereafter referred to as the VAP-bundle approach). A case of VAP was defined as a patient who required mechanical ventilation at Radius Specialty Hospital for at least 48 hours before any symptoms of pneumonia appeared and who met the Centers for Disease Control and Prevention criteria for VAP. Sputum samples were collected from a tracheal aspirate if there was clinical suspicion of VAP, and these samples were semiquantitatively cultured., Results: During the 22-month study period, 23 cases of VAP involving 19 patients were associated with 157 LTACH admissions (infection rate, 14.6%), corresponding to a rate of 1.67 cases per 1,000 ventilator-days, which is a 56% reduction from the VAP rate of 3.8 cases per 1,000 ventilator-days reported before the implementation of the VAP-bundle approach (P< .001). Microbiological data were available for 21 (91%) of 23 cases of VAP. Cases of VAP in the LTACH were frequently polymicrobial (mean number +/- SD, 1.78+/-1.0 pathogens per case of VAP), and 20 (95%) of 21 cases of VAP had at least 1 pathogen (Pseudomonas species, Acinetobacter species, gram-negative bacilli resistant to more than 3 antibiotics, or methicillin-resistant Staphylococcus aureus) cultured from a sputum sample. LTACH patients with VAP were more likely to have a neurological reason for ventilator dependence, compared with LTACH patients without VAP (69.6% of cases of VAP vs 39% of cases of respiratory failure; P= .014). In addition, patients with VAP had a longer length of LTACH stay, compared with patients without VAP (median length of stay, 131 days vs 39 days; P= .002). In 6 (26%) of 23 cases of VAP, the patient was eventually weaned from use of mechanical ventilation. Of the 19 patients with VAP, 1 (5%) did not survive the LTACH stay., Conclusions: The VAP rate in the LTACH is lower than the VAP rate reported in acute care hospitals. Cases of VAP in the LTACH were frequently polymicrobial and were associated with multidrug-resistant pathogens and increased length of stay. The guidelines from the Centers for Disease Control and Prevention that are aimed at reducing cases of VAP appear to be effective if applied in the LTACH setting.

We sought to evaluate this "response-to-injury" hypothesis of atherosclerosis by studying the interaction between systolic blood pressure (SBP) and LDL- cholesterol (LDL-C) in predicting the presence of coronary artery calcification (CAC) in asymptomatic men. We studied 526 men (46+/-7 years of age) referred for electron-beam tomography (EBT) exam. The prevalence of CAC was determined across LDL-C tertiles (low: <115 mg/dl; middle: 115-139 mg/dl; high: >or=140 mg/dl) within tertiles of SBP (low: <121 mmHg; middle: 121-130 mmHg; high: >or=131 mmHg). CAC was found in 220 (42%) men. There was no linear trend in the presence of CAC across LDL-C tertiles in the low (p=0.6 for trend) and middle (p=0.3 for trend) SBP tertile groups, respectively. In contrast, there was a significant trend for increasing CAC with increasing LDL-C (1st: 44%; 2nd: 49%; 3rd: 83%; p<0.0001 for trend) in the high SBP tertile group. In multivariate logistic analyses (adjusting for age, smoking, triglyceride levels, HDL-cholesterol levels, body mass index, and fasting glucose levels), the odds ratio for any CAC associated with increasing LDL-C was significantly higher in those with highest SBP levels, whereas no such relationship was observed among men with SBP in the lower two tertiles. An interaction term (LDL-C x SBP) incorporated in the multivariate analyses was statistically significant (p=0.038). The finding of an interaction between SBP and LDL-C relation to CAC in asymptomatic men support the response-to-injury model of atherogenesis.

With its 2006-2011 National Malaria Strategic Plan, Zambia committed to control malaria at a national scale. This scale-up for impact approach was facilitated by sound business planning and financing in 2006 of approximately US$35 million. Compared with surveys in 2001 and 2004, a 2006 national survey of 14,681 persons in 2,999 households at the end of the transmission season showed substantial coverage increases for preventive interventions. Ownership and use rates of insecticide-treated mosquito nets (ITNs) among vulnerable groups doubled, with 44% of households owning ITNs and 23% of children less than five years of age and 24% of pregnant women using them. Roll Back Malaria Abuja targets for intermittent preventive treatment in pregnancy (IPTp) were exceeded, with 62% of pregnant women receiving at least two doses of IPTp. As of 2006, Zambia is demonstrating substantial progress toward the national targets (80% population coverage rates for the interventions) and aspires to show that malaria need not be its leading health problem, and that malaria control is a sound national investment.

A renaissance in commitment to malaria control is transforming the perspectives and aspirations of the global community, prompting a consideration of goals for confronting a disease that is responsible for legendary death and suffering in Africa. The results in several countries are producing confidence that current control interventions can result in a dramatic reduction in the burden that malaria causes. However, the complexities of the challenges that must be addressed for comprehensive Africa programming are formidable in terms of the time required and the resources that will have to be mobilized. Progress toward elimination of the malaria burden in the African region in the next 5 years will be the critical benchmark for the feasibility of a comprehensive global campaign to eliminate and potentially eradicate malaria.

Background: Pulmonary infections can be life threatening for children with neuromuscular diseases who have impaired ability to clear secretions. Intrapulmonary percussive ventilation (IPV) is a pneumatic device that delivers air and aerosol to the lungs at frequencies of 200 to 300 cycles per minute at peak pressures from 20 to 40 cm H(2)O. Anecdotal reports and pilot studies show its safety and effectiveness in mobilizing secretions in patients with cystic fibrosis., Objective: To test the hypothesis that IPV used in a pulmonary program for adolescents with neuromuscular disease would reduce the number of days of antibiotic use for pulmonary infection., Methods: A randomized, controlled study was conducted to compare efficacy of IPV with incentive spirometry (IS) in reducing number of days of antibiotic use in adolescents with neuromuscular disease. The secondary endpoints were the number of respiratory infections, hospitalizations, and school days missed., Results: A total of 18 patients were enrolled (9 IPV, 9 IS). Antibiotic use was significantly higher with IS (24/1000 patient-days) compared with IPV (0/1000 patient-days), (incidence rate ratio, 43; 95% confidence interval, 6-333). The IS group spent more days hospitalized (4.4/1000 patient-days vs 0/1000 patient-days) than the IPV group (incidence rate ratio, 8.5; 95% confidence interval, 1.1-67). The IPV group had 0 episodes of pneumonia or bacterial bronchitis compared with 3 events in the IS group, although this did not meet statistical significance., Conclusion: Intrapulmonary percussive ventilation as part of a preventive pulmonary regimen reduced days of antibiotic use and hospitalization for respiratory illness in adolescents with neuromuscular disease.

Systemic Staphylococcus aureus infection is associated with significant morbidity and mortality arising from both bacterial and host immune factors. IL-18 is a pro-inflammatory cytokine of the IL-1 superfamily that exhibits broad functional effects in innate and acquired immune responses and which has been found in high levels in several chronic inflammatory and autoimmune diseases. Over-expression of IL-18 may promote early resolution of infection or could promote a detrimental exaggerated immune response. This was explored in a model of S. aureus infection. We report increased mortality in Swiss mice that were given recombinant IL-18 prior to inoculation with S. aureus LS-1. IL-18 administration prior to infection induced preferentially enhanced IFN-gamma mRNA expression in peripheral blood leukocytes and spleen, especially splenic NK cells. This correlated with increased IFN-gamma protein detection in serum, and leukocyte and spleen cultures at subsequent discrete time points. These data suggest that increased mortality following gram-positive infection in autoimmune diseases could in part reflect the impact of high levels of pleiotropic pro-inflammatory cytokines such as IL-18 present prior to the onset of infection.

Physiological stimuli using reactive oxygen species (ROS) as second messengers caused nucleotide-specific base modifications in the hypoxic response element of the VEGF gene in lung vascular cells, with the 3' guanine of the HIF-1 DNA recognition sequence uniformly targeted. Modeling this effect by replacing the targeted guanine with an abasic site increased incorporation of HIF-1 and the bi-functional DNA repair enzyme and transcriptional coactivator, Ref-1/Ape1, into the transcriptional complex and engendered more robust reporter gene expression. Oxidants generated in the context of physiological signaling thus affect nuclear DNA integrity and may facilitate gene expression by optimizing DNA-protein interactions.

This study examined the development of deictic and representational gestures in 12 typically developing children from 6 to 24 months of age. Gestures were categorized into J. Bruner's (1981) 3 broad (and 8 specific) communicative functions: behavior regulation (i.e., requesting objects, requesting actions, protesting), joint attention (i.e., commenting, requesting information), and social interaction (i.e., representational gestures, attention seeking, social games). Ongoing parental completion of researcher-created gesture recording forms and monthly researcher observational confirmation were used to capture the emergence and consistent use of targeted gestures. Within each specific functional category, a hierarchy of development was documented for the gestures and behaviors used to signal that intent. This study provides rich detail as to the order of emergence of common deictic and representational gestures and their relationship to other preceding and concomitant behaviors that children use to signal their intentions. Furthermore, the results document younger ages of emergence, in comparison with previous studies, for most of the targeted gestures and provide insight into the controversy in the literature regarding the relative emergence of declarative and imperative gestures.

Background: IL-18 is a cytokine which is known to have an important role in the development of a Th1 lymphocyte response. As such, it may have a regulatory role in asthma by modifying Th2 lymphocyte responses. Cigarette smoking may amplify the airway inflammation associated with asthma., Objective: This study investigated if IL-18 could be detected in induced sputum from asthmatics and normal subjects and if smoking altered IL-18 levels., Methods: Induced sputum was obtained from asthmatic (31 smokers, 35 non-smokers) and normal (20 smokers, 20 non-smokers) subjects. All smokers had a smoking history of > or =15 pack years. IL-18 levels in sputum supernatant were measured by ELISA. IL-18 mRNA expression and cellular localization were assessed by quantitative PCR and immunocytochemistry, respectively., Results: Smoking was associated with a significant reduction in IL-18 levels (median (interquartile range) - smokers 20 (0-102) pg/mL vs. non-smokers 358 (50-876) pg/mL, P<0.001). This was more pronounced in asthmatics (smokers, 47 (40-64) pg/mL vs. non-smokers, 530 (30-1484) pg/mL; P<0.001) than in normal subjects (smokers, 25 (0-78) pg/mL vs. non-smokers, 247 (50-656) pg/mL; P<0.01). Within each of the smoking and non-smoking groups there was no significant difference in IL-18 levels between asthmatic and normal subjects. There was no correlation between sputum IL-18 levels and any specific cell type in the sputum samples nor serum IgE levels. IL-18 mRNA expression was reduced in asthmatic smokers compared with non-smokers. IL-18 production was localized to sputum macrophages by immunocytochemistry., Conclusions: IL-18 is detectable in induced sputum samples from both asthmatic and normal subjects. Cigarette smoking significantly reduces sputum IL-18 levels. This effect is more pronounced in asthmatics than in normal subjects.

The co-transcription factor and DNA repair enzyme, Redox effector factor-1/apurinic/apyrimidinic endonuclease (Ref-1/Ape), facilitates DNA binding and transcriptional activity of a number of transactivating factors, including those governing hypoxia-induced gene expression HIF-1. It is not known, however, whether Ref-1/Ape is a component of the hypoxic transcriptional complex. Electrophoretic mobility shift assays failed to detect direct DNA binding of Ref-1/Ape to either the HIF-1 or AP1 DNA recognition sequences present in the hypoxic response element of the VEGF gene. However, immunodepletion of Ref-1/Ape from nuclear extract prevented DNA binding of ATF/CREB and HIF-1 to the HIF-1 DNA recognition sequence. DNA affinity-precipitation analyses showed that Ref-1/Ape was part of the multiprotein transcriptional complex forming on a 64-mer sequence encompassing a minimal hypoxic response element. Immunodepletion of Ref-1/Ape prevented probe association with HIF-1, p300, ATF, and CREB. Co-immunoprecipitation experiments indicated that Ref-1/Ape present in nuclear extract interacted with HIF-1 and p300 but not ATF/CREB. However, when Ref-1/Ape was immunoprecipitated from the oligonucleotide probe, both HIF-1 and p300 remained probe-associated while ATF/CREB co-immunoprecipitated. These findings suggest that Ref-1/Ape is a critical component of the hypoxia-inducible transcriptional complex forming on the VEGF gene's hypoxic response element and that the presence of Ref-1/Ape in the complex is required for the apparent high affinity association between HIF-1 and its DNA recognition sequence.

Insecticide-treated nets (ITNs) have been shown to reduce the burden of malaria in African villages by providing personal protection and, if coverage of a community is comprehensive, by reducing the infective mosquito population. We do not accept the view that scaling-up this method should be by making villagers pay for nets and insecticide, with subsidies limited so as not to discourage the private sector. We consider that ITNs should be viewed as a public good, like vaccines, and should be provided via the public sector with generous assistance from donors. Our experience is that teams distributing free ITNs, replacing them after about 4 years when they are torn and retreating them annually, have high productivity and provide more comprehensive and equitable coverage than has been reported for marketing systems. Very few of the free nets are misused or sold. The estimated cost would be an annual expenditure of about US$295 million to provide for all of rural tropical Africa where most of the world's malaria exists. This expenditure is affordable by the world community as a whole, but not by its poorest members. Recently, funding of this order of magnitude has been committed by donor agencies for malaria control.

In spite of increasing resistance, chloroquine remains the primary drug for treatment of malaria in most sub-Saharan African countries. We evaluated the effect of drug treatment policy on the case-fatality rates of children, adjusting for differing distributions of malaria and severe anemia. In 1991, 63% of children were treated with chloroquine while the remaining 37% were treated with a regimen that would eliminate and clear parasitemia. Case-fatality rates were 13% and 4.1%, respectively; the proportion of deaths attributable to chloroquine treatment was 69%. The trend in case-fatality rates for malaria decreased as an increasing proportion of children received an effective treatment regimen; adjusted malaria case-fatality rates were 5.1%, 3.6%, and 3.3% in 1992, 1993, and 1994, respectively, when 85% of children in 1992 and 97% of children in 1993-1994 received effective therapy. These 4 years of data provide strong evidence that continued use of chloroquine in areas with resistance is contributing to excess Plasmodium falciparum-related deaths.

During bacterial infections, the balance between resolution of infection and development of sepsis is dependent upon the macrophage response to bacterial products. We show that priming of murine bone marrow-derived macrophages (BMMs) with CSF-1 differentially regulates the response to two such stimuli, LPS and immunostimulatory (CpG) DNA. CSF-1 pretreatment enhanced IL-6, IL-12, and TNF-alpha production in response to LPS but suppressed the same response to CpG DNA. CSF-1 also regulated cytokine gene expression in response to CpG DNA and LPS; CpG DNA-induced IL-12 p40, IL-12 p35, and TNF-alpha mRNAs were all suppressed by CSF-1 pretreatment. CSF-1 pretreatment enhanced LPS-induced IL-12 p40 mRNA but not TNF-alpha and IL-12 p35 mRNAs, suggesting that part of the priming effect is posttranscriptional. CSF-1 pretreatment also suppressed CpG DNA-induced nuclear translocation of NF-kappaB and phosphorylation of the mitogen-activated protein kinases p38 and extracellular signal-related kinases-1/2 in BMMs, indicating that early events in CpG DNA signaling were regulated by CSF-1. Expression of Toll-like receptor (TLR)9, which is necessary for responses to CpG DNA, was markedly suppressed by CSF-1 in both BMMs and thioglycolate-elicited peritoneal macrophages. CSF-1 also down-regulated expression of TLR1, TLR2, and TLR6, but not the LPS receptor, TLR4, or TLR5. Hence, CSF-1 may regulate host responses to pathogens through modulation of TLR expression. Furthermore, these results suggest that CSF-1 and CSF-1R antagonists may enhance the efficacy of CpG DNA in vivo.

Parasite survival and host health may depend on the ability of the parasite to modulate the host immune response by the release of immunomodulatory molecules. Excretory-secretory (ES)-62, one such well-defined molecule, is a major secreted protein of the rodent filarial nematode Acanthocheilonema viteae, and has homologues in human filarial nematodes. Previously we have shown that ES-62 is exclusively associated with a Th2 Ab response in mice. Here we provide a rationale for this polarized immune response by showing that the parasite molecule suppresses the IFN-gamma/LPS-induced production, by macrophages, of bioactive IL-12 (p70), a key cytokine in the development of Th1 responses. This suppression of the induction of a component of the host immune response extends to the production of the proinflammatory cytokines IL-6 and TNF-alpha, but not NO. The molecular mechanism underlying these findings awaits elucidation but, intriguingly, the initial response of macrophages to ES-62 is to demonstrate a low and transient release of these cytokines before becoming refractory to further release induced by IFN-gamma/LPS. The relevance of our observations is underscored by the finding that macrophages recovered from mice exposed to "physiological" levels of ES-62 by the novel approach of continuous release from implanted osmotic pumps in vivo were similarly refractory to release of IL-12, TNF-alpha, IL-6, but not NO, ex vivo. Therefore, our results suggest that exposure to ES-62 renders macrophages subsequently unable to produce Th1/proinflammatory cytokines. This likely contributes to the generation of immune responses with an anti-inflammatory Th2 phenotype, a well-documented feature of filarial nematode infection.

ST2/ST2L, a member of the IL-1R gene family, is expressed by fibroblasts, mast cells, and Th2, but not Th1, cells. It exists in both membrane-bound (ST2L) and soluble forms (ST2). Although ST2L has immunoregulatory properties, its ligand, cellular targets, and mode of action remain unclear. Using a soluble ST2-human IgG fusion protein, we demonstrated that ST2 bound to primary bone marrow-derived macrophages (BMM) and that this binding was enhanced by treatment with LPS. The sST2 treatment of BMMs inhibited production of the LPS-induced proinflammatory cytokines IL-6, IL-12, and TNF-alpha but did not alter IL-10 or NO production. Treatment of BMMs with sST2 down-regulated expression of Toll-like receptors-4 and -1 but induced nuclear translocation of NF-kappaB. Administration of sST2 in vivo after LPS challenge significantly reduced LPS-mediated mortality and serum levels of IL-6, IL-12, and TNF-alpha. Conversely, blockade of endogenous ST2 through administration of anti-ST2 Ab exacerbated the toxic effects of LPS. Thus, ST2 has anti-inflammatory properties that act directly on macrophages. We demonstrate here a novel regulatory pathway for LPS-induced shock via the ST2-Toll-like receptor 4 route. This may be of considerable therapeutic potential for reducing the severity and pathology of inflammatory diseases.

Agmatine, a product of arginine decarboxylation in mammalian cells, is believed to govern cell polyamines by inducing antizyme, which in turn suppresses ornithine decarboxylase (ODC) activity and polyamine uptake. However, since agmatine is structurally similar to the polyamines, it is possible that it exerts antizyme-independent actions on polyamine regulatory pathways. The present study determined whether agmatine inhibited ODC activity and polyamine transport in rat pulmonary artery endothelial cells (PAECs) by an antizyme-dependent mechanism. Agmatine caused time-dependent reductions in ODC activity, which occurred before increases in antizyme. Interventions that suppressed proteasome function caused large increases in ODC activity but failed to attenuate inhibitory effects of agmatine. When agmatine was present in the culture medium, 14C-polyamine uptake was competitively inhibited as evidenced by substantial elevations in K(m) values. If PAECs were incubated with agmatine for periods sufficient to increase antizyme, there were modest decreases in V(max) for putrescine and spermidine but not for spermine. These effects of agmatine on polyamine transport were insensitive to protein synthesis inhibition. Collectively, our findings show that agmatine decreases ODC activity and polyamine transport in PAECs, but a causal role for antizyme in these actions of agmatine is difficult to establish. Nevertheless, these observations are consistent with a model in which PAECs express both antizyme-1 and -2, but only the latter contributes to agmatine-mediated suppression of ODC activity.

Objective: The antidepressant action of ECT may be related to its anticonvulsant properties. Positron emission tomography (PET) studies of regional cerebral metabolic rate for glucose were used to test this hypothesis., Method: Ten patients with major depression were studied with PET before and approximately 5 days after a course of bilateral ECT. Statistical parametric mapping was used to identify regions of decreased cerebral glucose metabolism., Results: Widespread regions of decreased regional cerebral glucose metabolism were identified after ECT, especially in the frontal and parietal cortex, anterior and posterior cingulate gyrus, and left temporal cortex. A region-of-interest analysis similarly indicated post-ECT reductions in regional cerebral glucose metabolism., Conclusions: ECT reduces neuronal activity in selected cortical regions, a potential anticonvulsant and antidepressant effect.

A large-scale longitudinal cohort project was initiated in western Kenya in June 1992. The primary purpose of the project was to study Plasmodium falciparum malaria in a highly endemic area using a comprehensive and multidisciplinary approach, which included epidemiology, entomology, and immunology. Between June 1992 and July 1994, pregnant women living in 15 rural villages were identified during a monthly census and 1,164 were enrolled. The women were followed-up throughout their pregnancy and they, along with their newborn infants and direct siblings of the infants' less than 15 years of age, were monitored over time. As of May 1995, 1,017 infants had been born to these women. This paper presents the design and general methodology used in this study and describes the initial experience with intense monitoring of a large population over a prolonged period.

A large-scale longitudinal cohort project was initiated in western Kenya in June 1992. Between June 1992 and July 1994, 1,848 children less than 15 years of age were monitored prospectively for a mean of 236 days. During this period, 12,035 blood smears were examined for malaria and only 34% were found to be negative. Parasite prevalence (all species) decreased with age (from a high of 83% among children 1-4 years old to 60% among children 10-14 years old). Even more dramatic decreases were noted in the prevalence of high density falciparum infection (from 37% among children 12-23 months old to < 1% among 10-14-year-old children) and in clinical malaria (20% to 0.3% in the same age groups). Children < 1 year of age accounted for 55% of all cases of anemia detected. Anemia was consistently associated with high density infection in children < 10 years of age (20% to 210% increased risk relative to aparasitemic children). These results demonstrate the relationship between high-density malaria infection and two clinical manifestations of malarial illness.

Objective: To determine the effect of transfusion on hematologic recovery and mortality among severely anemic children during and after hospitalization in rural Kenya., Design: Prospective cohort., Methods: We collected clinical and laboratory information on all severely anemic children (hemoglobin < 5.0 g/dl) and a 33% sample of children with hemoglobin < or = 5.0 g/dl who were admitted to the pediatric ward of a rural Kenyan hospital during a 6 month study period. Children were followed during hospitalization and at 4 and 8 weeks after admission., Results: Overall, 303 (25%) of the 1223 hospitalized children had hemoglobin < 5.0 g/dl, 30% of whom died during the study period. Severely anemic children who were transfused had a higher mean hemoglobin level at discharge (9.0 g/dl) than non-transfused children (5.8 g/dl, P < 0.001) and maintained a higher mean hemoglobin during the 8-week follow-up period. However, the presence of malaria parasitemia on follow-up negated the benefit of transfusion on hematologic recovery at both 4- and 8-week visits (longitudinal linear model, least square means, P > 0.05). Transfusion was associated with improved survival among children with respiratory distress who received transfusions within the first 2 days of hospitalization., Conclusions: The use of transfusion can be improved by targeting use of blood to severely anemic children with cardiorespiratory compromise, improving immediate availability of blood, and treating severely anemic children with effective antimalarial therapy.

Problem/condition: Malaria is caused by infection with one of four species of Plasmodium (P. falciparum, P. vivax, P. ovale, and P. malariae), which are transmitted by the bite of an infective female Anopheles sp. mosquito. Most malaria cases in the United States occur among persons who have traveled to areas (i.e., other countries) in which disease transmission is ongoing. However, cases are transmitted occasionally through exposure to infected blood products, by congenital transmission, or by local mosquito-borne transmission. Malaria surveillance is conducted to identify episodes of local transmission and to guide prevention recommendations., Reporting Period Covered: Cases with onset of illness during 1993., Description of System: Malaria cases confirmed by blood smear are reported to local and/or state health departments by health-care providers and/or laboratories. Case investigations are conducted by local and/or state health departments, and the reports are transmitted to CDC., Results: CDC received reports of 1,275 cases of malaria in persons in the United States and its territories who had onset of symptoms during 1993; this number represented a 40% increase over the 910 malaria cases reported for 1992. P. vivax, P. falciparum, P. ovale, and P. malariae were identified in 52%, 36%, 4%, and 3% of cases, respectively. The species was not determined in the remaining 5% of cases. The 278 malaria cases in U.S. military personnel represented the largest number of such cases since 1972; 234 of these cases were diagnosed in persons returning from deployment in Somalia during Operation Restore Hope. In New York City, the number of reported cases increased from one in 1992 to 130 in 1993. The number of malaria cases acquired in Africa by U.S. civilians increased by 45% from 1992; of these, 34% had been acquired in Nigeria. The 45% increase primarily reflected cases reported by New York City. Of U.S. civilians who acquired malaria during travel, 75% had not used a chemoprophylactic regimen recommended by CDC for the area in which they had traveled. Eleven cases of malaria had been acquired in the United States: of these cases, five were congenital; three were induced; and three were cryptic, including two cases that were probably locally acquired mosquito-borne infections. Eight deaths were associated with malarial infection., Interpretation: The increase in the reported number of malaria cases was attributed to a) the number of infections acquired during military deployment in Somalia and b) complete reporting for the first time of cases from New York City., Actions Taken: Investigations were conducted to collect detailed information concerning the eight fatal cases and the 11 cases acquired in the United States. Malaria prevention guidelines were updated and disseminated to health-care providers. Persons who have a fever or influenza-like illness after returning from a malarious area should seek medical care, regardless of whether they took antimalarial chemoprophylaxis during their stay. The medical evaluation should include a blood smear examination for malaria. Malaria can be fatal if not diagnosed and treated rapidly. Recommendations concerning prevention and treatment of malaria can be obtained from CDC.

Optimal treatment of Plasmodium falciparum-related paediatric anaemia can result in improved haematological recovery and survival. Clinical predictors are needed to identify children with anaemia in settings where laboratory measurements are not available. The use of conjunctival (eyelid), palmar, nailbed, and tongue pallor to detect children with moderate anaemia (haemoglobin, 5.0-7.9 g/dl) or severe anaemia (haemoglobin, < 5.0 g/dl) was evaluated among children seen at an outpatient and inpatient setting in a hospital in western Kenya. Severe nailbed or severe palmar pallor had the highest sensitivity (62% and 60%, resp.), compared with severe conjunctival pallor (sensitivity = 31%), to detect children with severe anaemia in the outpatient setting. Children with moderate anaemia were best identified by the presence of nailbed or palmar pallor (sensitivity = 90% for both signs), compared with conjunctival pallor (sensitivity = 81%). Clinical signs of respiratory distress, in addition to the presence of severe pallor, did not increase the recognition of children requiring hospitalization for severe anaemia. Among inpatients, the sensitivity of severe nailbed pallor (59%) was highest for detecting children with severe anaemia, although the sensitivity of severe conjunctival pallor and severe palmar pallor was the same (53% for both signs). Presence of conjunctival pallor (sensitivity = 74%) was similar in sensitivity to both nailbed and palmar pallor (70% for both signs) among children with moderate anaemia. The sensitivity of tongue pallor was low among all children evaluated. Low haemoglobin levels were significantly associated with the likelihood of being smear-positive for P. falciparum. This study demonstrates that clinical criteria can be used to identify children with moderate and severe anaemia, thus enabling implementation of treatment algorithms. Children aged < 36 months who live in an area with P. falciparum malaria should receive treatment with an effective antimalarial drug if they have pallor.

In 1993, the World Health Organization completed the development of a draft algorithm for the integrated management of childhood illness (IMCI), which deals with acute respiratory infections, diarrhoea, malaria, measles, ear infections, malnutrition, and immunization status. The present study compares the performance of a minimally trained health worker to make a correct diagnosis using the draft IMCI algorithm with that of a fully trained paediatrician who had laboratory and radiological support. During the 14-month study period, 1795 children aged between 2 months and 5 years were enrolled from the outpatient paediatric clinic of Siaya District Hospital in western Kenya; 48% were female and the median age was 13 months. Fever, cough and diarrhoea were the most common chief complaints presented by 907 (51%), 395 (22%), and 199 (11%) of the children, respectively; 86% of the chief complaints were directly addressed by the IMCI algorithm. A total of 1210 children (67%) had Plasmodium falciparum infection and 1432 (80%) met the WHO definition for anaemia (haemoglobin < 11 g/dl). The sensitivities and specificities for classification of illness by the health worker using the IMCI algorithm compared to diagnosis by the physician were: pneumonia (97% sensitivity, 49% specificity); dehydration in children with diarrhoea (51%, 98%); malaria (100%, 0%); ear problem (98%, 2%); nutritional status (96%, 66%); and need for referral (42%, 94%). Detection of fever by laying a hand on the forehead was both sensitive and specific (91%, 77%). There was substantial clinical overlap between pneumonia and malaria (n = 895), and between malaria and malnutrition (n = 811). Based on the initial analysis of these data, some changes were made in the IMCI algorithm. This study provides important technical validation of the IMCI algorithm, but the performance of health workers should be monitored during the early part of their IMCI training.