A Toll-like receptor-1 variant and its characteristic cellular phenotype is associated with severe malaria in Papua New Guinean children.

Genetic factors are likely to contribute to low severe malaria case fatality rates in Melanesian populations, but association studies can be underpowered and may not provide plausible mechanistic explanations if significant associations are detected. In preparation for a genome-wide association study, 29 candidate single-nucleotide polymorphisms (SNPs) with minor allele frequencies >5% were examined in a case-control study of 504 Papua New Guinean children with severe malaria. In parallel, an immunological substudy was performed on convalescent peripheral blood mononuclear cells (PBMCs) from cases and controls. Following stimulation with a Toll-like receptor (TLR) 1/2 agonist, effector cytokines and chemokines were assayed. The only significant genetic association observed involved a nonsynonymous SNP (TLR1rs4833095) in the TLR1 gene. A recessive (TT) genotype was associated with reduced odds of severe malaria of 0.52 (95% confidence interval (0.29-0.90), P=0.006). Concentrations of pro-inflammatory cytokines interleukin-1ß and tumour necrosis factor α were significantly higher in severe malaria cases compared with healthy controls, but lower in children with the protective recessive (TT) genotype. A genetic variant in TLR1 may contribute to the low severe malaria case fatality rates in this region through a reduced pro-inflammatory cellular phenotype.

Age-related sperm DNA methylation changes are transmitted to offspring and associated with abnormal behavior and dysregulated gene expression.

Advanced paternal age (APA) has been shown to be a significant risk factor in the offspring for neurodevelopmental psychiatric disorders, such as schizophrenia and autism spectrum disorders. During aging, de novo mutations accumulate in the male germline and are frequently transmitted to the offspring with deleterious effects. In addition, DNA methylation during spermatogenesis is an active process, which is susceptible to errors that can be propagated to subsequent generations. Here we test the hypothesis that the integrity of germline DNA methylation is compromised during the aging process. A genome-wide DNA methylation screen comparing sperm from young and old mice revealed a significant loss of methylation in the older mice in regions associated with transcriptional regulation. The offspring of older fathers had reduced exploratory and startle behaviors and exhibited similar brain DNA methylation abnormalities as observed in the paternal sperm. Offspring from old fathers also had transcriptional dysregulation of developmental genes implicated in autism and schizophrenia. Our findings demonstrate that DNA methylation abnormalities arising in the sperm of old fathers are a plausible mechanism to explain some of the risks that APA poses to resulting offspring.

Metabolism of norethisterone in the greyhound.

RATIONALE: Norethisterone has been used as a successful oral contraceptive in humans for many years. It was recently permitted for use as an oestrus suppressant in racing greyhounds. To monitor the use of norethisterone as part of a routine drug surveillance programme, knowledge of its metabolism was required to enable detection. METHODS: Gas chromatography/mass spectrometry and selective derivatisation techniques have been used to identify urinary metabolites of norethisterone following oral administration to the greyhound. Metabolites were extracted using solid-phase and liquid-liquid extraction techniques. RESULTS: Several metabolites were identified, including reduced, mono-, di- and trihydroxylated steroids. The major metabolites observed were 17α-ethynyl-5ß-estrane-3α,17ß-diol, 17α-ethynyl-5α-estrane-3ß,17ß-diol, three 17α-ethynylestranetriol stereoisomers and two 17α-ethynylestranetetrol stereoisomers. The major metabolites were predominantly excreted as glucuronic acid conjugates and detection of the administration of norethisterone was possible for up to 8 days post-dose using the methods described. The nandrolone metabolites, 19-norepiandrosterone, estranediol and 19-noretiocholanolone, were also identified in the post-administration samples collected up to 8 h after dosing the treated animals. CONCLUSIONS: The urinary metabolites identified in this study have further increased the knowledge of steroid metabolism in the greyhound, providing information to support routine drug testing programmes for greyhound racing.

Interaction of malaria with a common form of severe thalassemia in an Asian population.

In many Asian populations, the commonest form of severe thalassemia results from the coinheritance of HbE and beta thalassemia. The management of this disease is particularly difficult because of its extreme clinical diversity; although some genetic and adaptive factors have been identified as phenotypic modifiers, the reasons remain unclear. Because the role of the environment in the course of severe thalassemia has been neglected completely and because malaria due to both Plasmodium falciparum and Plasmodium vivax has been prevalent in Sri Lanka, we carried out a pilot study of patients with HbE beta thalassemia that showed high frequencies of antibodies to both parasite species and that 28.6% of the children had DNA-based evidence of current infection with P. vivax. Malarial antibodies then were assessed in patients with HbE beta thalassemia compared with those in age-matched controls. There was a significant increase in the frequency of antibodies in the thalassemic patients, particularly against P. vivax and in young children. There was also a higher frequency in those who had been splenectomized compared with those with intact spleens, although in the latter it was still higher than that in the controls. The thalassemic patients showed significant correlations between malaria antibody status and phenotype. Patients with HbE beta thalassemia may be more prone to malaria, particularly P. vivax, which is reflected in their clinical severity. Because P. vivax malaria is widespread in Asia, further studies of its interaction with HbE beta thalassemia and related diseases are required urgently as a part of ongoing thalassemia control programs.

Grip Strength, Gait Speed and Plasma Markers of Neurodegeneration in Asymptomatic Middle-aged and Older Adults.

BACKGROUND: Pragmatic biomarkers of preclinical dementia would allow for easy and large-scale screening of risk in populations. Physical function measures like grip strength and gait speed are potential predictive biomarkers but their relationship with plasma markers of Alzheimer's Disease and neurodegeneration have not been elucidated. OBJECTIVES: To examine association between physical function measures and plasma markers of Alzheimer's Disease (AD) and neurodegeneration. DESIGN: Cross-sectional and longitudinal analyses. SETTING: Community-based cohort in the city of Framingham, Massachusetts. PARTICIPANTS: 2336 participants of the Framingham Heart Study Offspring cohort with an average age of 61. MEASUREMENTS: Plasma Aß40 and Aß42 were measured in 1998-2001 (Exam-7) and plasma total tau measured 5 years later (Exam-8). Grip strength, fast walk speed and chair stand speed were measured at both exams. Quantification of Aß isoforms in plasma was performed using INNO-BIA assays and plasma total-tau was measured using Quanterix Simoa HD-1 assay. Confounder-adjusted linear regression models examined associations between physical function and plasma markers, Results: Grip strength at Exam-7 was associated with plasma Aß40 (ß -0.006, p-value 0.032) at Exam-7 and plasma total-tau (ß -0.010, p-value 0.001) at Exam-8. Grip strength and fast walk speed at Exam-8 were associated with plasma total-tau at Exam-8 (GS: ß -0.009, p 0.0005; FWS: ß -0.226, p-value <0.0001). Chair stand speed was not associated with plasma markers; Aß42 was not associated with function. CONCLUSION: Grip strength and fast walk speed are associated with plasma markers of neurodegeneration in dementia-free middle aged and older individuals. Both these measures could be used as potential screening tools for identifying individuals at a higher risk for AD and related dementias alongside other validated markers.

Oxidative stress in fungal fermentation processes: the roles of alternative respiration.

Filamentous fungi are arguably the most industrially important group of microorganisms. Production processes involving these simple eukaryotes are often highly aerobic in nature, which implies these cultures are routinely subject to oxidative stress. Despite this, little is known about how filamentous fungi cope with high levels of oxidative stress as experienced in fermenter systems. More surprisingly, much of our knowledge of oxidative stress responses in fungi comes from environmental or medical studies. Here, the current understanding of oxidative stress effects and cellular responses in filamentous fungi is critically discussed. In particular the role of alternative respiration is evaluated, and the contributions of the alternative oxidase and alternative dehydrogenases in defence against oxidative stress, and their profound influence on fungal metabolism is critically examined. Finally, the importance of further research which would underpin a less empirical approach to optimising fungal strains for the fermenter environment is emphasised.

The transmission of nosocomial pathogens in an intensive care unit: a space-time clustering and structural equation modelling approach.

We investigated the incidence of cases of nosocomial pathogens and risk factors in an intensive treatment unit ward to determine if the number of cases is dependent on location of patients and the colonization/infection history of the ward. A clustering approach method was developed to investigate the patterns of spread of cases through time for five microorganisms [methicillin-resistant Staphylococcus aureus (MRSA), Acinetobacter spp., Klebsiella spp., Candida spp., and Pseudomonas aeruginosa] using hospital microbiological monitoring data and ward records of patient-bed use. Cases of colonization/infection by MRSA, Candida and Pseudomonas were clustered in beds and through time while cases of Klebsiella and Acinetobacter were not. We used structural equation modelling to analyse interacting risk factors and the potential pathways of transmission in the ward. Prior nurse contact with colonized/infected patients, mediated by the number of patient-bed movements, were important predictors for all cases, except for those of Pseudomonas. General health and invasive surgery were significant predictors of cases of Candida and Klebsiella. We suggest that isolation and bed movement as a strategy to manage MRSA infections is likely to impact upon the incidence of cases of other opportunist pathogens.

Survey of the fatty acid composition of retail milk differing in label claims based on production management practices.

Consumers are becoming increasingly health conscious, and food product choices have expanded. Choices in the dairy case include fluid milk labeled according to production management practices. Such labeling practices may be misunderstood and perceived by consumers to reflect differences in the quality or nutritional content of milk. Our objective was to investigate nutritional differences in specialty labeled milk, specifically to compare the fatty acid (FA) composition of conventional milk with milk labeled as recombinant bST (rbST)-free or organic. The retail milk samples (n=292) obtained from the 48 contiguous states of the United States represented the consumer supply of pasteurized, homogenized milk of 3 milk types: conventionally produced milk with no specialty labeling, milk labeled rbST-free, and milk labeled organic. We found no statistical differences in the FA composition of conventional and rbST-free milk; however, these 2 groups were statistically different from organic milk for several FA. When measuring FA as a percentage of total FA, organic milk was higher in saturated FA (65.9 vs. 62.8%) and lower in monounsaturated FA (26.8 vs. 29.7%) and polyunsaturated FA (4.3 vs. 4.8%) compared with the average of conventional and rbST-free retail milk samples. Likewise, among bioactive FA compared as a percentage of total FA, organic milk was slightly lower in trans 18:1 FA (2.8 vs. 3.1%) and higher in n-3 FA (0.82 vs. 0.50%) and conjugated linoleic acid (0.70 vs. 0.57%). From a public health perspective, the direction for some of these differences would be considered desirable and for others would be considered undesirable; however, without exception, the magnitudes of the differences in milk FA composition among milk label types were minor and of no physiological importance when considering public health or dietary recommendations. Overall, when data from our analysis of FA composition of conventional milk and milk labeled rbST-free or organic were combined with previous analytical comparisons of the quality and composition of these retail milk samples, results established that there were no meaningful differences that would affect public health and that all milks were similar in nutritional quality and wholesomeness.

Hot topic: Enhancing omega-3 fatty acids in milk fat of dairy cows by using stearidonic acid-enriched soybean oil from genetically modified soybeans.

Very long chain n-3 fatty acids such as eicosapentaenoic acid (EPA; 20:5n-3) are important in human cardiac health and the prevention of chronic diseases, but food sources are limited. Stearidonic acid (SDA; 18:4n-3) is an n-3 fatty acid that humans are able to convert to EPA. In utilizing SDA-enhanced soybean oil (SBO) derived from genetically modified soybeans, our objectives were to examine the potential to increase the n-3 fatty acid content of milk fat and to determine the efficiency of SDA uptake from the digestive tract and transfer to milk fat. Three multiparous, rumen-fistulated Holstein cows were assigned randomly in a 3 x 3 Latin square design to the following treatments: 1) control (no oil infusion); 2) abomasal infusion of SDA-enhanced SBO (SDA-abo); and 3) ruminal infusion of SDA-enhanced SBO (SDA-rum). The SDA-enhanced SBO contained 27.1% SDA, 10.4% alpha-linolenic acid, and 7.2% gamma-linolenic acid. Oil infusions provided 57 g/d of SDA with equal amounts of oil infused into either the rumen or abomasum at 6-h intervals over a 7-d infusion period. Cow numbers were limited and no treatment differences were detected for DMI or milk production (22.9+/-0.5 kg/d and 32.3+/-0.9 kg/d, respectively; least squares means +/- SE), milk protein percentage and yield (3.24+/-0.04% and 1.03+/-0.02 kg/d), or lactose percentage and yield (4.88+/-0.05% and 1.55+/-0.05 kg/d). Treatment also had no effect on milk fat yield (1.36+/-0.03 kg/d), but milk fat percentage was lower for the SDA-rum treatment (4.04+/-0.04% vs. 4.30+/-0.04% for control and 4.41+/-0.05% for SDA-abo). The SDA-abo treatment increased n-3 fatty acids to 3.9% of total milk fatty acids, a value more than 5-fold greater than that for the control. Expressed as a percentage of total milk fatty acids, values (least squares means +/- SE) for the SDA-abo treatment were 1.55+/-0.03% for alpha-linolenic acid (18:3n-3), 1.86+/-0.02 for SDA, 0.23 +/- <0.01 for eicosatetraenoic acid (20:4n-3), and 0.18+/-0.01 for EPA. Transfer efficiency of SDA to milk fat represented 39.3% (range=36.8 to 41.9%) of the abomasally infused SDA and 47.3% (range=45.0 to 49.6%) when the n-3 fatty acids downstream from SDA were included. In contrast, transfer of ruminally infused SDA to milk fat averaged only 1.7% (range=1.3 to 2.1%), indicating extensive rumen biohydrogenation. Overall, results demonstrate the potential to use SDA-enhanced SBO from genetically modified soybeans combined with proper ruminal protection to achieve impressive increases in the milk fat content of SDA and other n-3 fatty acids that are beneficial for human health.

Olfaction in patients with suspected parkinsonism and scans without evidence of dopaminergic deficit (SWEDDs).

BACKGROUND: Positron emission tomography and single photon emission computed tomography scanning have 87-94% sensitivity and 80-100% specificity to differentiate patients with Parkinson's disease (PD) from control subjects and patients with essential (ET) or atypical tremor. More than 10% of patients diagnosed as early PD can have scans without evidence of dopaminergic deficiency (SWEDDs). This study investigated whether smell tests can help identify possible cases with SWEDDs. METHODS: The 40 item University of Pennsylvania Smell Test (UPSIT) was used to evaluate the sense of smell in 21 SWEDDs patients. Twenty-six ET patients, 16 patients with a diagnosis of idiopathic adult onset dystonia (D), 191 non-demented PD patients and 136 control subjects were also tested. Multiple regression analyses were used to compare the mean UPSIT score in the SWEDDs group with the other four groups (ET, D, PD and controls) after adjusting for the effects of relevant covariates. RESULTS: The mean UPSIT score for the SWEDDs group was greater than in the PD group (p<0.001) and not different from the mean UPSIT in the control (p = 0.7), ET (p = 0.4) or D (p = 0.9) groups. Smell tests indicated a high probability of PD in only 23.8% of SWEDDs as opposed to 85.3% of PD patients. CONCLUSIONS: In a patient with suspected PD, a high PD probability on smell testing favours the diagnosis of PD, and a low PD probability strengthens the indication for dopamine transporter imaging.

Sources of variation in physician adherence with clinical guidelines: results from a factorial experiment.

BACKGROUND: Health services research has documented the magnitude of health care variations. Few studies focus on provider level sources of variation in clinical decision making-for example, which primary care providers are likely to follow clinical guidelines, with which types of patient. OBJECTIVES: To estimate: (1) the extent of primary care provider adherence to practice guidelines and the unconfounded influence of (2) patient attributes and (3) physician characteristics on adherence with clinical practice guidelines. DESIGN: In a factorial experiment, primary care providers were shown clinically authentic video vignettes with actors portrayed different "patients" with identical signs of coronary heart disease (CHD). Different types of providers were asked how they would manage the different "patients" with identical CHD symptoms. Measures were taken to protect external validity. RESULTS: Adherence to some guidelines is high (over 50% of physicians would follow a third of the recommended actions), yet there is low adherence to many of them (less than 20% would follow another third). Female patients are less likely than males to receive 4 of 5 types of physical examination (p < .03); older patients are less likely to be advised to stop smoking (p < .03). Race and SES of patients had no effect on provider adherence to guidelines. A physicians' level of experience (age) appears to be important with certain patients. CONCLUSIONS: Physician adherence with guidelines varies with different types of "patient" and with the length of clinical experience. With this evidence it is possible to appropriately target interventions to reduce health care variations by improving physician adherence with clinical guidelines.

The 'Outcome Reporting in Brief Intervention Trials: Alcohol' (ORBITAL) framework: protocol to determine a core outcome set for efficacy and effectiveness trials of alcohol screening and brief intervention.

BACKGROUND: The evidence base to assess the efficacy and effectiveness of alcohol brief interventions (ABI) is weakened by variation in the outcomes measured and by inconsistent reporting. The 'Outcome Reporting in Brief Intervention Trials: Alcohol' (ORBITAL) project aims to develop a core outcome set (COS) and reporting guidance for its use in future trials of ABI in a range of settings. METHODS/DESIGN: An international Special Interest Group was convened through INEBRIA (International Network on Brief Interventions for Alcohol and Other Drugs) to inform the development of a COS for trials of ABI. ORBITAL will incorporate a systematic review to map outcomes used in efficacy and effectiveness trials of ABI and their measurement properties, using the COnsensus-based Standards for the selection of health Measurement INstruments (COSMIN) criteria. This will support a multi-round Delphi study to prioritise outcomes. Delphi panellists will be drawn from a range of settings and stakeholder groups, and the Delphi study will also be used to determine if a single COS is relevant for all settings. A consensus meeting with key stakeholder representation will determine the final COS and associated guidance for its use in trials of ABI. DISCUSSION: ORBITAL will develop a COS for alcohol screening and brief intervention trials, with outcomes stratified into domains and guidance on outcome measurement instruments. The standardisation of ABI outcomes and their measurement will support the ongoing development of ABI studies and a systematic synthesis of emerging research findings. We will track the extent to which the COS delivers on this promise through an exploration of the use of the guidance in the decade following COS publication.

Haemoglobin E beta thalassaemia in Sri Lanka.

Haemoglobin E beta thalassaemia is the commonest form of severe thalassaemia in many Asian countries, but little is known about its natural history, the reasons for clinical diversity, or its management. We studied 109 Sri Lankan patients with the disorder over 5 years. 25 patients were not receiving transfusion; transfusion was stopped with no deleterious effect in a further 37. We identified several genetic and environmental factors that might contribute to the phenotypic diversity of the disorder, including modifiers of haemoglobin F production, malaria, and age-related changes in adaptive function. Our findings suggest that haemoglobin E beta thalassaemia can be managed without transfusion in many patients, even with low haemoglobin levels. Age-related changes in the pattern of adaptation to anaemia suggest that different and more cost-effective approaches to management should be explored.

Muscle cell injury, haemolysis and dark urine in children with falciparum malaria in Papua New Guinea.

During a prospective study of red cell variants and severe malaria in children, a surprising observation was the occurrence of dark urine. Children were grouped according to urine findings: 22 had dark urine that contained a haem protein (Group I), 93 had urine of normal colour that contained a haem protein (Group II) and 236 had normal urine (Group III). To investigate the cause of dark urine, haemolysis and muscle cell injury were assessed. Intravascular haemolysis was greater in Group I than in Groups II and III. However, anaemia was more severe in Group III and is likely to have resulted mainly from extravascular haemolysis. Median plasma myoglobin concentrations were greater in Groups I and II than Group III (P = 0.00060). Plasma myoglobin was greater in children with cerebral malaria, hyperlactataemia and those who died but was not associated with acidosis. Urine myoglobin was greater in Group I than Groups II and III (P = 0.00054). It is likely that both haemoglobin and myoglobin contributed to dark urine. The association between muscle cell injury and coma suggests sequestration of parasitized red cells as a common underlying pathology. In malaria, hyperlactataemia may result directly from breakdown of muscle protein as well as tissue hypoxia.

Estrogen receptor-alpha in the inhibition of cancer growth and angiogenesis.

A high level of estrogen receptor-alpha (ER-alpha) is believed to be favorable in the prognosis and treatment of certain female cancers. ER-alpha expression in the ER-negative breast cancer cell lines inhibits their proliferation and invasive, metastatic potential in vitro. We stably overexpressed the ER-alpha in the human endometrial cancer cell line Ishikawa and showed that, unlike estradiol, high levels of ER-alpha significantly inhibit the growth of tumors xenografted from the Ishikawa cells. Subsequent to ER-alpha overexpression, in vivo down-regulation of vascular endothelial growth factor was observed in tumor xenografts. In addition, these tumors showed an inhibition of vascularization and of the angiogenic agent, integrin alphavbeta3. Involvement of a switch in the angiogenic pathways during tumorigenesis has been a recent focus of interest. Our results indicate that a high level of ER-alpha may be beneficial in the control of female cancers because of its inhibitory effect on such angiogenic pathways.

Comparing no-touch and tympanic thermometer temperature recordings.

Temperature is a vital sign which can be measured using various types of clinical thermometers. Pulmonary artery temperature is considered the 'gold standard', but this measurement is not usually clinically practical. There is currently no consensus for optimal alternative site or equipment. This research compares 178 simultaneous measurements from 5 clinical areas, using two types of thermometers: tympanic and no-touch temporal. No-touch thermometers were all set to oral equivalent. Tympanic thermometers were adjusted to either oral (n=105) or core (n=73) equivalent. Maximum acceptable difference was identified as 1oC. Two data sets (oral/core; oral/oral) were analysed using Bland-Altman method on Excel programmes, comparing all thermometers and separating oral and core-equivalent tympanics. The two thermometers were found not to be equivalent. As a simple comparison between two thermometers, this research cannot identify which thermometer is more accurate.

The Drosophila melanogaster ade5 gene encodes a bifunctional enzyme for two steps in the de novo purine synthesis pathway.

Steps 6 and 7 of de novo purine synthesis are performed by 5-aminoimidazole ribonucleotide carboxylase (AIRc) and 4-[(N-succinylamino)carbonyl]-5-aminoimidazole ribonucleotide synthetase (SAICARs), respectively. In vertebrates, a single gene encodes AIRc-SAICARs with domains homologous to Escherichia coli PurE and PurC. We have isolated an AIRc-SAICARs cDNA from Drosophila melanogaster via functional complementation with an E. coli purC purine auxotroph. This cDNA encodes AIRc yet is unable to complement an E. coli purE mutant, suggesting functional differences between Drosophila and E. coli AIRc. In vertebrates, the AIRc-SAICARs gene shares a promoter region with the gene encoding phosphoribosylamidotransferase, which performs the first step in de novo purine synthesis. In Drosophila, the AIRc-SAICARs gene maps to section 11B4-14 of the X chromosome, while the phosphoribosylamidotransferase gene (Prat) maps to chromosome 3; thus, the close linkage of these two genes is not conserved in flies. Three EMS-induced X-linked adenine auxotrophic mutations, ade4(1), ade5(1), and ade5(2), were isolated. Two gamma-radiation-induced (ade5(3) and ade5(4)) and three hybrid dysgenesis-induced (ade5(5), ade5(6), and ade5(8)) alleles were also isolated. Characterization of the auxotrophy and the finding that the hybrid dysgenesis-induced mutations all harbor P transposon sequences within the AIRc-SAICARs gene show that ade5 encodes AIRc-SAICARs.

Clinical significance of monoclonal proteins in chronic lymphocytic leukemia.

Serum monoclonal proteins were found in 36 of 111 (32%) chronic lymphocytic leukemia (CLL) patients studied using high-resolution agarose gel electrophoresis combined with immunofixation. In contrast, using immunoelectrophoresis (IEP) and serum protein electrophoresis (SPEP), monoclonal proteins were found in 20 and 4% of CLL patients, respectively. The presence of a monoclonal protein was associated with a median survival of 63 months compared to 103 months for individuals without a monoclonal spike (p < 0.012). This was independent of clinical stage. The data also suggests that the presence of a monoclonal protein may define a group of patients at greater risk for disease progression in the low and intermediate risk groups. (Rai stages, 0, I, II). Further follow-up will be required to determine if this is statistically significant.