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Background: Malarial infections are often missed by microscopy, and most parasite carriers are asymptomatic in low-endemicity settings. Whether parasite detectability and its ability to elicit symptoms change as transmission declines remains unclear., Methods: We performed a prospective panel survey with repeated measurements on the same participants over 12 months to investigate whether Plasmodium vivax detectability by microscopy and risk of symptoms upon infection varied during a community-wide larviciding intervention in the Amazon basin of Brazil that markedly reduced vector density. We screened 1096 to 1400 residents in the intervention site for malaria by microscopy and quantitative TaqMan assays at baseline and twice during intervention., Results: We found that more P vivax infections than expected from their parasite densities measured by TaqMan assays were missed by microscopy as transmission decreased. At lower transmission, study participants appeared to tolerate higher P vivax loads without developing symptoms. We hypothesize that changes in the ratio between circulating parasites and those that accumulate in the bone marrow and spleen, by avoiding peripheral blood microscopy detection, account for decreased parasite detectability and lower risk of symptoms under low transmission., Conclusions: P vivax infections are more likely to be subpatent and remain asymptomatic as malaria transmission decreases., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Low- and middle-income countries are increasingly faced with a triple burden of malnutrition: endemic underweight, micronutrient deficiencies and rising prevalence of overweight. This study aimed to address existing knowledge gaps and to identify priority policy options in Mongolia, the Philippines and Vietnam. A landscape analysis approach was adopted using methods set out in a UNICEF global toolkit. Quantitative and qualitative data were compiled from a range of global and national sources on childhood overweight and obesity, risk factors and policy responses. Key informant interviews and validation workshops were undertaken with key food and nutrition stakeholders from government and non-government organizations to identify priority policy options for the prevention of overweight and obesity among children. Overweight and obesity among children are increasing in all three countries. Associated risk factors are related to maternal nutrition, birthweight, breastfeeding, as well as diets and physical activity shaped by increasingly obesogenic environments. Key informants identified undefined policy approaches, poor community understanding and food and beverage industry influence as barriers to addressing overweight and obesity. Key policy priorities include restricting the marketing of unhealthy food and beverages, unhealthy food and beverage taxation, introduction of front-of-pack nutrition labels and improving school nutrition environments. Mongolia, the Philippines and Vietnam are all facing an increasing burden of childhood overweight and obesity. Despite differing national contexts, similar environmental factors are driving this rise. A suite of evidence-based policies can effectively be introduced to address obesogenic environments., (© The Author(s) 2023. Published by Oxford University Press.)

Background: Trypanosoma cruzi/HIV coinfection has been described as a relevant clinical event and an emerging public health problem. Here, we describe the epidemiological patterns of deaths related to Chagas disease and HIV/AIDS coinfection in Brazil from 2000 to 2019., Methods: We performed a nationwide population-based study using mortality data obtained from the Brazilian Mortality Information System. We included all deaths recorded in Brazil from 2000 to 2019 in which Chagas disease and HIV/AIDS were mentioned on the same death certificate, either as underlying or as associated causes of death., Results: Chagas disease and HIV/AIDS were mentioned on 196/22 663 092 death certificates. HIV/AIDS was the underlying cause in 58.2% (114/196) of deaths and Chagas disease in 33.2% (65/196). The average annual mortality rate was 0.05 deaths/1 000 000 inhabitants (95% CI 0.03 to 0.09). The highest death rates were found among males, those aged 60-69 y, Afro-Brazilians, those with 1-3 y of schooling/study and residents in Chagas disease-endemic regions/states. Respiratory, infectious/parasitic and cardiovascular diseases/disorders were the associated causes of death most commonly mentioned., Conclusions: Mortality due to Chagas disease and HIV/AIDS coinfection may be largely underestimated in Brazil. Our data further reinforce the importance of screening for T. cruzi infection in HIV-infected patients from Chagas disease-endemic areas. Appropriate clinical management should be ensured for Chagas disease and HIV coinfected patients., (© The Author(s) 2021. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Background: Immunity after dengue virus (DENV) infection has been suggested to cross-protect from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and mortality., Methods: We tested whether serologically proven prior DENV infection diagnosed in September-October 2019, before the coronavirus disease 2019 (COVID-19) pandemic, reduced the risk of SARS-CoV-2 infection and clinically apparent COVID-19 over the next 13 months in a population-based cohort in Amazonian Brazil. Mixed-effects multiple logistic regression analysis was used to identify predictors of infection and disease, adjusting for potential individual and household-level confounders. Virus genomes from 14 local SARS-CoV-2 isolates were obtained using whole-genome sequencing., Results: Anti-DENV immunoglobulin G (IgG) was found in 37.0% of 1285 cohort participants (95% confidence interval [CI]: 34.3% to 39.7%) in 2019, with 10.4 (95% CI: 6.7-15.5) seroconversion events per 100 person-years during the follow-up. In 2020, 35.2% of the participants (95% CI: 32.6% to 37.8%) had anti-SARS-CoV-2 IgG and 57.1% of the 448 SARS-CoV-2 seropositives (95% CI: 52.4% to 61.8%) reported clinical manifestations at the time of infection. Participants aged >60 years were twice more likely to have symptomatic COVID-19 than children under 5 years. Locally circulating SARS-CoV-2 isolates were assigned to the B.1.1.33 lineage. Contrary to the cross-protection hypothesis, prior DENV infection was associated with twice the risk of clinically apparent COVID-19 upon SARS-CoV-2 infection, with P values between .025 and .039 after adjustment for identified confounders., Conclusions: Higher risk of clinically apparent COVID-19 among individuals with prior dengue has important public health implications for communities sequentially exposed to DENV and SARS-CoV-2 epidemics., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Background: Heterogeneity in COVID-19 morbidity and mortality is often associated with a country's health-services structure and social inequality. This study aimed to characterize social inequalities in COVID-19 mortality in São Paulo, the most populous city in Brazil and Latin America., Methods: We conducted a population-based study, including COVID-19 deaths among São Paulo residents from March to September 2020. Age-standardized mortality rates and unadjusted rate ratios (RRs) [with corresponding 95% confidence intervals (CIs)] were estimated by race, sex, age group, district of residence, household crowding, educational attainment, income level and percentage of households in subnormal areas in each district. Time trends in mortality were assessed using the Joinpoint model., Results: Males presented an 84% increase in COVID-19 mortality compared with females (RR = 1.84, 95% CI 1.79-1.90). Higher mortality rates were observed for Blacks (RR = 1.77, 95% CI 1.67-1.88) and mixed (RR = 1.42, 95% CI 1.37-1.47) compared with Whites, whereas lower mortality was noted for Asians (RR = 0.63, 95% CI 0.58-0.68). A positive gradient was found for all socio-economic indicators, i.e. increases in disparities denoted by less education, more household crowding, lower income and a higher concentration of subnormal areas were associated with higher mortality rates. A decrease in mortality over time was observed in all racial groups, but it started earlier among Whites and Asians., Conclusion: Our results reveal striking social inequalities in COVID-19 mortality in São Paulo, exposing structural inequities in Brazilian society that were not addressed by the governmental response to COVID-19. Without an equitable response, COVID-19 will further exacerbate current social inequalities in São Paulo., (© The Author(s) 2021; all rights reserved. Published by Oxford University Press on behalf of the International Epidemiological Association.)

Background: Aedes aegypti rapid larval surveys are mandatory in Brazil. Here, we retrospectively examined whether the house index estimated by larval surveys served as a useful tool in anticipating epidemics within Brazilian municipalities from 2009-2015., Methods: We used correlation indices and classification analysis stratified by year, region, population size and time after the national larval survey., Results: We found no association between the house index and the proportion of municipalities experiencing an epidemic. The sensitivity of a high score house index in predicting an epidemic was 7.20% (95% CI 6.22 to 8.33%) for all years combined. The positive predictive value of a high score house index to predict a 'true epidemic' was 38.96%, lower than the negative predictive values of a low score house index for predicting 'no epidemic' (56.96%). The highest overall sensitivity was observed in the North region (20.15%; 95% CI 17.14 to 23.53%). The sensitivity of a high score house index demonstrated a monotonic decrease with increasing time from larval collection., Conclusions: Larval surveys are surveillance tools with the potential to risk-stratify and guide dengue control programs towards judicious resource allocation. However, the national rapid larval survey performed in Brazil, in its present form, consistently underpredicts dengue epidemics., (© The Author(s) 2020. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Despite historical and contemporary evidence of its effectiveness, larval source management with insecticides remains little used by most malaria control programs worldwide. Here we show that environmentally safe biological larvicides under field conditions can significantly reduce anopheline larval density in fish farming ponds that have became major larval habitats across the Amazon Basin. Importantly, the primary local malaria vector, Anopheles darlingi Root (Diptera: Culicidae), feeds and rests predominantly outdoors, being little affected by interventions such as long-lasting insecticidal bed net distribution and indoor residual spraying. We found >95% reduction in late-instar density up to 7 d after the first application of VectoMax FG or VectoLex CG (both from Valent BioSciences), and up to 21 d after larvicide reapplication in fish ponds (n = 20) situated in the main residual malaria pocket of Brazil, irrespective of the formulation or dosage (10 or 20 kg/ha) used. These results are consistent with a substantial residual effect upon retreatment and support the use of biological larvicides to reduce the density of anopheline larvae in this and similar settings across the Amazon where larval habitats are readily identified and accessible., (© The Author(s) 2019. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Photosynthetic picoeukaryotes (PPE) are key components of primary production in marine and freshwater ecosystems. In contrast with those of marine environments, freshwater PPE groups have received little attention. In this work, we used flow cytometry cell sorting, microscopy and metabarcoding to investigate the composition of small photosynthetic eukaryote communities from six eutrophic shallow lakes in South America, Argentina. We compared the total molecular diversity obtained from PPE sorted populations as well as from filtered total plankton samples (FTP). Most reads obtained from sorted populations belonged to the classes: Trebouxiophyceae, Chlorophyceae and Bacillariophyceae. We retrieved sequences from non-photosynthetic groups, such as Chytridiomycetes and Ichthyosporea which contain a number of described parasites, indicating that these organisms were probably in association with the autotrophic cells sorted. Dominant groups among sorted PPEs were poorly represented in FTP and their richness was on average lower than in the sorted samples. A significant number of operational taxonomic units (OTUs) were exclusively found in sorting samples, emphasizing that sequences from FTP underestimate the diversity of PPE. Moreover, 22% of the OTUs found among the dominant groups had a low similarity (<95%) with reported sequences in public databases, demonstrating a high potential for novel diversity in these lakes., (© FEMS 2019.)

Fourteen fungal entomopathogenic strains were isolated from soil samples and infected field-collected fall armyworm larvae, in Guanajuato, Mexico. Isolates were identified by morphology and internal transcribed spacers sequencing. Isolates Ma22, Ma41, and Mr8 showed 99% identity with reference strains (RS) of Metarhizium anisopliae. Isolates Bb9, Bb19, Bb21, Bb40, Bb27, Bb23, and Bb39 showed identity between 99 and 100% with RS of Beauveria bassiana. Isolates Nr1, Nr2, Nr3, and Nr4 showed identity between 98 and 100% with RS of Nomuraea rileyi. Qualitative selection used one concentration (1 × 108 conidia/ml) on fall armyworm eggs and neonate larvae. Strains Ma22, Ma41, and Mr8 showed 100%, and strains Bb39, Bb23, Bb9, Bb40, Bb19, and Bb21 showed 92, 89.2, 87.6, 82.8, 58, and 38% egg mortality, respectively. Bioassays on neonate larvae showed 100% mortality with strains Ma22, Ma41, Mr8, and Bb9. Strains Bb39, Bb19, Bb27, Bb23, Bb21, and Bb40 showed 74, 60, 54, 53, 28, and 19% mortality, respectively. Bioassay estimated LC50s for strains Ma41 at 7.4 × 104, Mr8 at 8.9 × 104, and Ma22 at 10 × 104 conidia/ml, on fall armyworm eggs. LC50s on neonate larvae were estimated at 2.8 × 105, 16 × 105, 26 × 105, and 36 × 105 conidia/ml for strains Ma41, Bb9, Ma22, and Mr8, respectively. Virulence genes mad1 and mad2 were found in Mr8, Ma22, and Ma41, whereas the gen gmact was found only in the strain Ma22. Genes hyd1 and hyd2 were identified in Bb9, Bb19, Bb21, and Bb27. No correlation was observed between the virulence gene detection and the estimated LC50s. Strain Ma41 showed the highest potential to be developed as a bioinsecticide.

Objectives: To assess the radiographic appearances of the endodontic treatment in different room lighting conditions and image display devices., Methods: After approval by Research Ethical Board, 20 images were obtained from blocks with one endodontically treated tooth. Of the 20 endodontic treatments conducted in the laboratory, of those the randomly selected 10 teeth (50% of sample) received some quality defect in three parameters-apical limit, adaptation to root canal and homogeneity of the root canal filling material. Digital radiographs (DIGORA® Optime, Soredex, Milwaukee, WI) were obtained, which were evaluated (8 times) by 10 observers in two different conditions of room lighting, with bright (1000 to 1200 lux) and dim light (25 to 50 lux), using two different image display devices, smartphones and laptop computers. The observers assessed the radiographic appearances of the endodontic treatment with respect to quality of each one of three parameters individually. The data of observers' responses in terms of the quality of each parameter were statistically compared using Cronbach's α coefficient, diagnostic tests, π coefficient and ANOVA with Tukey tests, at a significant level setting of 5% (α = 0.05)., Results: The mean of reliability of observer responses ranged from 0.89 (intra-evaluator) to 0.66 (inter-evaluator). The mean values of the receiver operating characteristic (ROC) area were 0.661 (PD = 0.098), with laptop in bright lighting appraising homogeneity of the root canal filling material on the worst performance (ROC area = 0.57), which was statistically significant (p < 0.05), and 0.81 with laptop in dim lighting appraising apical limit on the best performance. Differences (p < 0.05) occurred mainly among parameters used for assessment., Conclusions: Room lighting and image display devices have very little influence in the radiographic appearances of the endodontic treatment. Thus, we suggest the use of smartphones as an acceptable image display device in daily clinical practice in routine bright lighting conditions.

Aims: To demonstrate the ability of Bacillus thuringiensis to penetrate as spore-crystal complex to the internal tissues of bean plants, and keep its insecticidal activity. To test the vertical transmission of the spore-crystal complex in Arabidopsis thaliana., Methods and Results: The experimental strain was transformed with the pMUTIN-gfp plasmid which labelled the spores of B. thuringiensis HD-73 with the GFP protein. Once the rhizosphere of the bean plants was inoculated with the labelled strain, the bacterium was recovered from leaves, stems, and petioles. Furthermore, toxicity of treated plants was significantly higher than control plants when bio-assayed on cabbage looper larvae. The labelled strain was recovered from the dead insects. When the rhizosphere of A. thaliana plants was inoculated with the labelled strain, mature seeds from these plants were surface-sterilized and grown under in vitro conditions. The labelled strain was recovered from the seedlings., Conclusions: We showed that B. thuringiensis subsp. kurstaki (HD-73) in the rhizosphere can translocate to upper tissues of bean plants, and keep its insecticidal activity. Transmission of the labelled B. thuringiensis strain passed to the next generation of A. thaliana., Significance and Impact of the Study: The role of B. thuringiensis as a potential facultative endophyte bacterium and the possible biotechnological repercussions are discussed., (© 2017 The Society for Applied Microbiology.)

Objective: To determine whether a simple quality improvement initiative consisting of a technical update and regular audit and feedback sessions will result in increased use of antenatal corticosteroids among pregnant women at risk of imminent preterm birth delivering at health facilities in the Philippines and Cambodia., Design: Non-randomized, observational study using a pre-/post-intervention design conducted between October 2013 and June 2014., Setting: A total of 12 high volume facilities providing Emergency Obstetric and Newborn Care services in Cambodia (6) and Philippines (6)., Intervention: A technical update on preterm birth and use of antenatal corticosteroids, followed by monthly audit and feedback sessions., Main Outcome Measure: The proportion of women at risk of imminent preterm birth who received at least one dose of dexamethasone., Results: Coverage of at least one dose of dexamethasone increased from 35% at baseline to 86% at endline in Cambodia (P < 0.0001) and from 34% at baseline to 56% at endline in the Philippines (P < 0.0001), among women who had births at 24-36 weeks. In both settings baseline coverage and magnitude of improvement varied notably by facility. Availability of dexamethasone, knowledge of use and cost were not major barriers to coverage., Conclusions: A simple quality improvement strategy was feasible and effective in increasing use of dexamethasone in the management of preterm birth in 12 hospitals in Cambodia and Philippines., (© The Author 2016. Published by Oxford University Press in association with the International Society for Quality in Health Care.)

The LBIT-1200 strain of Bacillus thuringiensis was recently isolated from soil, and showed a 6.4 and 9.5 increase in toxicity, against Manduca sexta and Trichoplusia ni, respectively, compared to HD-73. However, LBIT-1200 was still highly similar to HD-73, including the production of bipyramidal crystals containing only one protein of ∼130 000 kDa, its flagellin gene sequence related to the kurstaki serotype, plasmid and RepPCR patterns similar to HD-73, no production of β-exotoxin and no presence of VIP genes. Sequencing of its cry gene showed the presence of a cry1Ac-type gene with four amino acid differences, including two amino acid replacements in domain III, compared to Cry1Ac1, which may explain its higher toxicity. In conclusion, the LBIT-1200 strain is a variant of the HD-73 strain but shows a much higher toxicity, which makes this new strain an important candidate to be developed as a bioinsecticide, once it passes other tests, throughout its biotechnological development., (© FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Unlabelled: The insecticidal activity of Bacillus thuringiensis is owing to the action of Cry and Cyt proteins. In addition to the synthesis of insecticidal proteins, some strains are able to synthesize β-exotoxin, which is highly toxic to humans. In this regard, it is very important to have a simple method to detect β-exotoxin to avoid the commercial production of this type of strains. In this work, we developed a simple and fast method, using the nematode Caenorhabditis elegans to detect indirectly the synthesis of β-exotoxin by B. thuringiensis strain. Using this assay, we detected that ~60% of Mexican native strains (i.e. LBIT-471, 491, 492, 497, 507, 511, 515, 536 and 537) were toxic to the nematode (44-97% mortalities) and their β-exotoxin (βEx(+) ) production, including a positive control (NRD-12), was confirmed by HPLC. In addition, the negative controls (βEx(-) ) LBIT-436 (HD-1) and LBIT-438 and also the native strains LBIT-499, 500, 521, 522, 533 and 542, did not show a detrimental effect against nematodes larvae, neither the synthesis of β-exotoxin as determined by HPLC. Finally, we did not find a correlation between B. thuringiensis strains with similar plasmid patterns and the β-exotoxin production., Significance and Impact of the Study: In this work, we implemented a qualitative and fast bioassay using the nematode Caenorhabditis elegans to detect the production of β-exotoxin in different strains of Bacillus thuringiensis. We show that this assay is useful to detect β-exotoxin in B. thuringiensis with high reliability, helping to discriminate strains that could not be used as bioinsecticides because of their putative risk to humans. Data show that qualitative bioassay with nematodes is a potential alternative to fly larvae bioassays, and correlated with the determination of β-exotoxin by HPLC., (© 2015 The Society for Applied Microbiology.)

Aims: The objective of this study was to produce stable inclusions of chitinase ChiA74Δsp in Bacillus thuringiensis subsp. israelensis (Bti) and to assay its insecticidal activity against Aedes aegypti larvae., Methods and Results: Bti was transformed with chiA74Δsp regulated by its own promoter or by the strong chimeric cytAp/STAB-SD promoter system to generate two recombinant Bti strains. These recombinants produced their native parasporal bodies composed of Cry4Aa, Cry4Ba, Cry11Aa and Cyt1Aa and ChiA74Δsp inclusions, and showed a approx. threefold increase in both endochitinase activity and viable spore count when compared with the parental strain. Both recombinants were approximately twofold more toxic (LC50s 8·02, 9·6 ng ml(-1) ) than parental Bti (19·8 ng ml(-1) ) against 4(th) instars of A. aegypti larvae., Conclusions: ChiA74Δsp inclusions, together with the insecticidal crystals and spores of Bti increased the toxicity against A. aegypti larvae by at least twofold., Significance and Impact of the Study: We report for the first time the engineering of Bti to produce spore-parasporal body-ChiA74∆sp inclusions in the same sporangium, which are released together following autolysis. Our work lays a foundation for engineering Bti to produce more efficacious combinations of Cry4Aa, Cry4Ba, Cry11Aa, Cyt1Aa and chitinase inclusions., (© 2015 The Society for Applied Microbiology.)

Chemical analysts use analytical blanks in their analyses, but seldom is this source of uncertainty evaluated. Generally, there is great confusion. Although the numerical value of the blank, in some situations, can be negligible, its source of uncertainty cannot be. This article discusses the uncertainty contribution of the analytical blank using a numerical example of the copper content in waters by flame atomic absorption spectrometry. The results indicate that the uncertainties of the analytical blank can contribute up to 50% when the blank sample is considered in this analysis, confirming its high impact. This effect can be primarily observed where the analyte concentration approaches the lower range of the analytical curve. Even so, the blank is not always computed. Therefore, the relevance of the analytical blank can be confirmed by uncertainty evaluation.

Background: Sensitized patients (pts) may develop acute antibody-mediated rejection (AMR) due to preformed donor-specific antibodies, undetected by pre-transplant complement-dependent cytotoxicity (CDC) crossmatch (XM). We hypothesized that C4d staining in 1-h post-reperfusion biopsies (1-h Bx) could detect early complement activation in the renal allograft due to preformed donor-specific antibodies., Methods: To test this hypothesis, renal transplants (n = 229) performed between June 2005 and December 2007 were entered into a prospective study of 1-h Bx and stained for C4d by immunofluorescence. Transplants were performed against a negative T-cell CDC-XM with the exception of three cases with a positive B-cell XM., Results: All 229 1-h Bx stained negative for C4d. Fourteen pts (6%) developed AMR. None of the 14 protocol 1-h Bx stained positive for C4d in peritubular capillaries (PTC). However, all indication biopsies-that diagnosed AMR-performed at a median of 8 days after transplantation stained for C4d in PTC., Conclusions: These data show that C4d staining in 1-h Bx is, in general, not useful for the early detection of AMR when CDC-XM is negative.

Background: During haemodialysis, calcium balance can affect, or be affected by, mineral metabolism. However, when dialysate calcium concentration (d[Ca]) is chosen or kinetic models are employed to calculate calcium balance, bone remodelling is rarely considered. In this study, we examined whether bone remodelling affects calcium mass transfer during haemodialysis., Methods: We dialysed 23 patients using a d[Ca] of 1.0, 1.25, 1.5 or 1.75 mmol/L. Calcium mass transfer was measured and associated with remodelling bone factors., Results: Calcium balance varied widely depending on the d[Ca]. Calcium removal was -578 +/- 389, -468 +/- 563, +46 +/- 400 and +405 +/- 413 mg when a d[Ca] of 1.0, 1.25, 1.5 or 1.75 mmol/L was used, respectively (1.0 and 1.25 vs 1.5 and 1.75 mmol/L, P < 0.001; 1.5 vs 1.75 mmol/L, P < 0.05). Univariate analysis showed that calcium balance correlated with calcium gradient, parathyroid hormone (PTH), osteocalcin and dialysis vintage. Multivariate analysis revealed that calcium balance was dependent on calcium gradient, PTH and osteocalcin., Conclusions: These results suggest that bone remodelling could affect calcium mass transfer during haemodialysis.

Cushing's syndrome results from lengthy and inappropriate exposure to excessive concentrations of either endogenous or exogenous glucocorticoids. This case report describes a patient with a novel type of Cushing's syndrome due to the use of party drugs. A 35-year-old woman had gained 8 kg body weight in 5 months and complained of anxiety. She showed a Cushing-like appearance and mild hypertension (blood pressure, BP 150/95 mmHg). She reported daily use of increasing doses of gamma-hydroxybutyric acid (GHB), a popular party drug. ACTH plasma levels were in the upper normal range (41 ng/l), with normal plasma cortisol (0.36 micromol/l). She showed an abnormal overnight 1 mg dexamethasone suppression test (cortisol 0.38 micromol/l). The urinary excretion of free cortisol in 24 h was also increased (0.47 micromol/24 h). CT scanning of the abdomen showed normal adrenals. After stopping GHB intake she lost 7 kg body weight and her BP normalized (BP 135/80 mmHg). GHB is a popular party drug in the Netherlands, but it is also used as a narcotic and for the treatment of narcolepsy. We hypothesize that GHB may bind to the pituitary gland gamma-aminobutyric acid-B receptors leading to ACTH overproduction.

Background: In conventional haemodialysis (CHD), the morbidity and mortality rate is unacceptably high; consequently, variations in the length and frequency of the haemodialysis sessions have been studied to reduce the complications of dialysis treatment. In this sense, high-efficiency short daily haemodialysis (SDHD) has been proposed as an alternative for patients on renal replacement therapy. In this study, we have related our experience with this dialysis modality., Methods: Twenty-six patients (16 males, mean age 35.6 +/- 14.7 years) were treated by SDHD for 33.6 +/- 18.5 months (range 6-57 months). The mean time on CHD before the switch to SDHD was 25.5 +/- 31.9 months (range 1-159 months). In 23 (88.5%) patients, native arteriovenous fistulae were used for vascular access. SDHD was performed six times a week, 1.5-2 h per session, and high flux polysulfone dialysers (surface area: 1.8 m(2)) were employed. The blood flow and dialysate flow rate were 350 and 800 ml/min, respectively., Results: In this trial, the patient survival was 100%. The vascular access survival after 12, 24, 36 and 48 months on SDHD was 100, 89, 89 and 80%, respectively. There were three failures of vascular access in 72.7 patient-years (0.04 failures/patient-year). In 15 patients on SDHD during 36 consecutive months, the vascular access survival after 12, 24, 36 and 48 months was 100, 93, 93 and 84%, respectively. Also, in this group of patients, there were 0.27 hospitalizations/patient-year and 1.24 days of hospitalizations/patient-year., Conclusions: We concluded that in a long-time study of patients on SDHD the morbidity and mortality rate is very low. Furthermore, we observed that failures of vascular access are not a significant problem. Consequently, we believe that SDHD is a powerful renal replacement therapy for treatment of patients on maintenance haemodialysis.

Objective: The reactivation of human lymphotropic herpesviruses can be related to the intensity of immunosuppression. We analysed the risk of reactivation of lymphotropic herpesviruses in patients with refractory rheumatoid arthritis treated with an anti-tumour necrosis factor-alpha (TNF-alpha) agent (infliximab)., Methods: Fifteen patients were treated with infliximab (3 mg/kg) at weeks 0, 2 and 6. Samples of both plasma and peripheral blood mononuclear cells (PBMC) were obtained before treatment (week 0) and before each infusion at weeks 2 and 6. Samples were analysed using a multiplex qualitative polymerase chain reaction (PCR) for lymphotropic herpesviruses. Quantification of cytomegalovirus (CMV) viral load (copies/ml) was performed using quantitative PCR. Reactivation was defined as the presence of viral DNA in plasma. Latent infection was defined as the presence of viral DNA in PBMC samples but not in plasma., Results: On baseline, latent CMV infection was detected in eight patients (53.3%), human herpesviruses-6 (HHV-6) in two (13.3%), Epstein-Barr virus (EBV) in seven (46.6%), CMV + HHV-6 in one (6.6%), CMV + EBV in two (13.3%) and HHV-6 + EBV in one (6.6%). Viral reactivation related to infliximab treatment was not observed. There was only one patient who had HHV-6 reactivation, but this was already detected in the baseline sample., Conclusions: Infliximab treatment does not induce replication of human lymphotropic herpesviruses in patients with rheumatoid arthritis. Thus, herpesviruses prophylaxis would not be indicated in these patients.

The use of antiretroviral combination therapy in HIV has been associated with lipodystrophy and several metabolic risk factors. We postulated that patients with HIV-lipodystrophy have impaired adipose tissue free fatty acid (FFA) trapping and, consequently, increased hepatic FFA delivery. We investigated FFA, hydroxybutyric acid (HBA; reflecting hepatic FFA oxidation), and triglyceride (TG) changes after a high fat meal in HIV-infected males with (LIPO; n = 26) and without (NONLIPO; n = 12) lipodystrophy and in healthy males (n = 35). Because defective peripheral FFA trapping has been associated with impaired action of complement component 3 (C3), we also determined postprandial C3 concentrations. The LIPO group had higher homeostasis model assessment scores compared with the other groups. Areas under the curve (AUCs) for FFA, HBA, and TG were higher in the LIPO group than in the NONLIPO group or the controls. No differences in TG-AUC, FFA-AUC, and HBA-AUC were observed between the NONLIPO group and controls. In HIV-infected patients, FFA-AUC and HBA-AUC were inversely related to sc adipose tissue area. Plasma C3 showed a postprandial increase in healthy controls, but not in the HIV-infected groups. C3 was not related to body fat distribution, postprandial FFA, or HBA. The present data suggest disturbed postprandial FFA metabolism in patients with HIV-lipodystrophy, most likely due to inadequate incorporation of FFA into TG in sc adipose tissue, but do not support a major role for C3 in these patients. The higher postprandial HBA levels reflect increased hepatic FFA delivery and may aggravate insulin resistance and dyslipidemia, leading to increased cardiovascular risk.

Thyroid hormone inhibits neonatal Sertoli cell proliferation and recent results have shown that thyroid hormone upregulates cyclin-dependent kinase inhibitors (CDKIs) p27Kip1 and p21Cip1 (also known as CDKN1B and CDKN1A, respectively) in neonatal Sertoli cells. This suggests that these CDKIs, which negatively regulate the cell cycle, could be critical in Sertoli cell proliferation. Consistent with this hypothesis, mice lacking p27Kip1 develop testicular organomegaly, but Sertoli cell numbers have not been determined. Likewise, effects of loss of p21Cip1 or both p27 and p21 on Sertoli cell number and testicular development were unknown. To determine if p27 and/or p21 regulate Sertoli cell proliferation, we measured Sertoli cell proliferation at Postnatal Day 16 and testis weight, Sertoli cell number, and daily sperm production (DSP) in 4-mo-old wild-type (WT), p21 knockout (p21KO), p27 knockout (p27KO), and p27/p21 double-knockout (DBKO) mice. Testis weights were increased 27%, 42%, and 86% in adult p21KO, p27KO, and DBKO mice, respectively, compared with WT. Sertoli cell number also was increased 48%, 126%, and 126% in p21KO, p27KO, and DBKO mice, respectively, versus WT. DSP in p21KO, p27KO, and DBKO testes also showed significant increases compared with WT mice. Although DSP was increased, there were increased spermatogenic defects observed in both p27KO and DBKO mice compared with WT. These data indicate that both p27 and p21 play an inhibitory role in regulating adult Sertoli cell number such that loss of either CDKI produces primary increases in Sertoli cell number and secondary increases in DSP and testis weight. Furthermore, loss of both CDKIs causes additive effects on DSP and testis weight, suggesting a central role for these CDKIs in testis development.

Background: Fasting and postprandial triglyceride (TG) concentrations vary considerably among individuals. TG metabolism is more efficient in women than in men, which may partly explain why females are protected against atherosclerosis. Our aim was to identify gender-specific genetic influences on fasting and postprandial TG concentrations under typical living conditions in healthy, lean, normolipidemic women., Methods: We studied 40 women and 48 men. Diurnal capillary TG profiles were calculated as the integrated area under the capillary TG curve averaged over 3 days. Genotypes of the FABP-2, HL, LPL, APOE, and PPARgamma genes and the APOC-III, APOC-III/A-IV intergenic region were determined., Results: Three genes (FABP-2, APOE, and PPARgamma) had a significant additive effect only in women. Mean TG concentrations were fourfold higher in women carriers of the PPARgamma wild-type allele (P = 0.044), threefold higher in carriers of the rare FABP-2 allele (P = 0.006), and fivefold higher in carriers of the E2 allele of the APOE gene (P = 0.037) than in noncarriers. None of these effects was observed in men. The presence of two or more of these adverse alleles increased TG concentrations in a dose-dependent manner. Women carriers of three adverse alleles had postprandial TG values comparable to those for men., Conclusions: An adverse combination of common alleles of the FABP-2, APOE, and PPARgamma genes in women increases their TG concentrations to values comparable to those seen in men. Although this influence is not appreciable when studying fasting plasma TGs, it becomes apparent with use of a more sensitive index such as measurements made throughout the day.

Large triglyceride (TG)-rich lipoproteins (TRLs) circulate in the blood, but they may also be present in a marginated pool, probably attached to the endothelium. It is unknown whether statins can influence this marginated pool in vivo in humans. Intravenous fat tests were performed in familial combined hyperlipidemia (FCHL) subjects before and after atorvastatin treatment and in controls to investigate whether acute increases in apoB in TRL fractions would occur, potentially reflecting the release of this TRL from a marginated pool. After a 12-h fast, a bolus injection of 10% Intralipid was given to 12 FCHL patients before and after 16-wk treatment with atorvastatin. Twelve carefully matched controls were included. For 60 min postinjection, apoB48, apoB100, and lipids were measured in TRLs. Fasting apoB100 in all TRL fractions were 2- to 3-fold higher in untreated FCHL compared with controls. ApoB48 concentrations in chylomicron fractions increased significantly within 10 min in FCHL before and after treatment, but not in controls. ApoB100 increased significantly in the chylomicron fractions in untreated FCHL and in controls, but not in FCHL after treatment. In very low density lipoprotein 1, apoB100 increased only in untreated FCHL. In very low density lipoprotein 2, apoB100 did not change in any group. These data show that increasing the number of circulating TRLs by chylomicron-like particles, results in increased plasma apoB-TRLs, probably by acute release from a marginated pool. This is a physiological process occurring in FCHL and in healthy normolipidemic subjects, but it is more pronounced in the former. Decreased marginated TRL particles in FCHL is a novel antiatherogenic property of atorvastatin.

One of the best studied aspects of the insulin resistance syndrome in familial combined hyperlipidemia (FCHL) is impaired insulin-mediated suppression of FFA by diminished inhibition of hormone-sensitive lipase (HSL). In vitro experiments have shown that stimulation of HSL activity by catecholamines is decreased in FCHL. The aim of this study was to investigate HSL inhibition by insulin and stimulation by endogenous catecholamines in vivo in FCHL patients. Twelve FCHL subjects using lipid-lowering medication and 12 controls underwent a mental stress test after random ingestion of either 50 g glucose or placebo. After ingestion of glucose, insulin concentrations increased from 76.8 +/- 21.5 pM to a maximum of 520.2 +/- 118.4 pM (P < 0.01) in FCHL and from 38.0 +/- 5.0 to 221.7 +/- 25.1 pM (P < 0.01) in controls. The percent decreases in plasma FFA during the first hour after glucose ingestion were similar in FCHL and controls (67 +/- 5% vs. 72 +/- 3%, respectively), suggesting a comparable inhibition of HSL in both. During the placebo test, FFA increased similarly in FCHL (56 +/- 9%) and controls (57 +/- 19%). In contrast, FFA concentrations did not change during mental stress after ingestion of glucose (from 0.17 +/- 0.02 to 0.15 +/- 0.02 mmol/liter in FCHL and from 0.11 +/- 0.02 to 0.12 +/- 0.02 mmol/liter in controls). In conclusion, the present study provides in vivo evidence for intact insulin-mediated suppression of FFA in FCHL, although this inhibition of HSL was achieved by higher insulin levels, suggesting insulin resistance at the level of HSL. Secondly, the induction of HSL activity by endogenous catecholamines in vivo is not decreased in FCHL, in contrast to earlier in vitro findings. Finally, catecholamine-induced HSL activation can be inhibited by insulin in a similar manner in both FCHL and controls.

Background: The double-pool urea kinetic model requires the measurement of the blood urea concentrations 30 min after haemodialysis (C(t+30)) to calculate equilibrated Kt/V. However, it has been suggested that urea concentrations 30 min before the end of dialysis (C(t-30)) may be representative of C(t+30). The aim of this study was to validate this suggestion., Methods: Twenty-two patients underwent haemodialysis for 180, 210, and 240 min. For each patient in each dialysis session, urea exponential decay curve was calculated. Because we measured C(t+30), we calculated the time (T(c)) before the end of dialysis that blood urea concentrations would be the same as C(t+30). In an additional 33 patients, we measured blood urea concentrations at T(c) and in C(t+30)., Results: We found that C(t-30) was significantly lower than C(t+30) independent of the duration of dialysis. However, there was a significant correlation between Kt/V(t-30) and Kt/V(t+30). The T(c) was 45 min before the end of dialysis. In the additional 33 patients, C(t-45) and C(t+30) were 54+/-17 and 52+/-17 mg/dl (NS), and Kt/V(t-45) and Kt/V(t+30) were 1.27+/-0.21 and 1.29+/-0.18 (NS), respectively. There were significant correlations between C(t-45) and C(t+30) (r=0.96; P<0.001), and between Kt/V(t-45) and Kt/V(t+30) (r=0.82; P<0.001). However, when measurements were analysed individually, 48% of the data points from C(t-45) vs C(t+30), and 42% of the data points from Kt/V(t-45) vs Kt/V(t+30) fell out of the 95% confidence interval of regression line., Conclusions: Although C(t-45) is useful to estimate Kt/V when assessing mean values, it is not suitable when assessing patients individually. This study demonstrates that the best method to calculate equilibrated Kt/V was a blood sample for urea concentrations 30 min after haemodialysis.

Background: It has been reported that patients with acute renal failure (ARF) requiring haemodialysis show an improved recovery of renal function when the dialysis treatment is performed using a biocompatible membrane rather than a bioincompatible membrane. However, most recent published human trials have not been able to confirm these findings., Method: Over a 2-year period, we prospectively studied 53 patients with ARF after cadaver renal transplantation who required haemodialysis and randomized them into two treatment groups. One group underwent dialysis with a cuprophane membrane and the other group underwent haemodialysis with a more biocompatible membrane, polysulfone. All patients received an immunosuppressive regimen which included azathioprine, prednisone and cyclosporine., Results: There was no difference by patient characteristics or immunosuppressive regimen before acute tubular necrosis (ATN) recovery. In both groups the number of haemodialysis sessions required prior to the recovery of renal function (6.57+/-2.79 vs 6.05+/-2.40), the number of oliguric days (16.25+/-5.14 vs 14.40+/-4.67) and the number of hospital days (33.38+/-12.85 vs 30.10+/-11.00), were not statistically different. There was also no difference in long-term allograft outcome., Conclusion: Our data demonstrate that the use of a more biocompatible membrane had no influence on the recovery from acute renal failure after renal transplantation.

Background: Percutaneous biopsy is the method of choice for differential diagnosis of renal allograft dysfunction, although it is not risk-free. The use of less aggressive methods for diagnosis should limit the need for percutaneous biopsy to some specific situations., Methods: We analysed 42 fine-needle aspiration biopsies from 36 kidney allograft recipients immunosuppressed with quadruple sequential therapy who suffered renal allograft dysfunction. Seven cases with stable renal function were used as controls and included as non-rejection cases in the analysis. In all aspirates the Corrected Increment was calculated and an immunocytochemical analysis of renal tubular cells with the monoclonal antibodies HLA-DR and ICAM-1 was performed., Results: The Corrected Increment was increased in 13 out of 18 acute rejection cases and in one out of 31 non-rejection cases. HLA-DR expression was found in more than 30% of tubular cells from the aspirates in 16 out of 18 acute rejection cases and in eight out of 31 cases without rejection (P < 0.001). ICAM-1 expression was detected in more than 30% of tubular cells in 14 out of 18 cases with acute rejection, and in four out of 31 cases without acute rejection (P < 0.001). Interestingly, all acute vascular rejection cases (n = 6), and six out of 12 acute cellular rejection cases expressed both, HLA-DR and ICAM-1, in more than 30% of tubular cells. On the other hand, none of the non-rejection allograft dysfunctions or control samples showed more than 30% of tubular cells immunostained with both HLA-DR and ICAM-1 antibodies., Conclusions: The immunocytochemical analysis of HLA-DR and ICAM-1 on renal tubular cells taken by fine-needle aspiration biopsy, allows the diagnosis of acute cellular rejection and acute vascular rejection even when the Corrected Increment is not increased. Moreover, the risk of a core renal biopsy can be avoided when both tests are negative since an acute rejection is a remote possibility.

Unlabelled: The dietary intakes of 90 confirmed primary liver cancer cases were compared against 90 age-sex matched controls. Dietary recall, obtained by the same nutritionist, was taken for each change of residence lasting one year or more. The frequency and amounts of food items consumed were calculated into units of aflatoxin load per day using a Philippine table of aflatoxin values of these items. Of the total subjects' aflatoxin load, 51.2% came from cassava, 20.3% from corn, 6.8% from peanuts and 5.8% from sweet potato. The mean aflatoxin load per day of the cases as found to be 440% that of the controls. Upon grouping dietary aflatoxin loads (and alcohol intakes) into Heavy and Light, and upon comparing cases versus controls, the relative risk (RR) of developing primary liver cancer was found to be statistically significant in the following order of rank: cassava, peanuts, sweet potato, corn and alcohol. Boiled rice which had negligible aflatoxin content gave no difference in risk. Combining aflatoxin load and alcohol intake gave a synergistic and a statistically significant effect on RR with Light Aflatoxin, Light Alcohol as RR = 1., Results: Light Aflatoxin, Heavy Alcohol RR = 3.9; Heavy Aflatoxin, Light Alcohol RR = 17.5; Heavy Aflatoxin, Heavy Alcohol RR = 35.0.