Endotoxin Measurement Research Articles

Influence of sample preservation on endotoxin measurement in water

Appropriate preservation of a range of water types prior to analysis for endotoxin was investigated, including sample storage and addition of sodium thiosulphate. Biologically active endotoxin in water samples was assayed using a chromogenic Limulus Amoebocyte Lysate (LAL) assay. Statistical analysis of measured mean endotoxin levels obtained for samples with and without sodium thiosulphate showed no significant difference in results. There was a 44% mean decline in the concentration of detectable endotoxin in water samples stored at -80 degrees C for 4 weeks compared with samples stored at 4 degrees C and analysed within 24 hours. Freezing of water samples at -80 degrees C in pyrogen-free containers for 4 weeks or longer, then thawing may lead to considerable endotoxin loss; however the addition of sodium thiosulphate to water samples interferes minimally with the LAL assay. These results provide methodological information that can be used to assist researchers in future water endotoxin monitoring studies. The validation and standardisation of water sample preservation protocols are necessary, given the likely increase in the quantification of endotoxin levels in a variety of water sources and the use of such results for health effect determinations.

Bacterial translocation and change in intestinal permeability in patients after abdominal surgery

The purpose of this study was to investigate bacterial translocation and change in intestinal permeability in patients after abdominal surgery. Sixty-three patients undergoing elective abdominal surgery were enrolled in the study. Blood samples were collected prior to operation and 2, 24, 48 h after surgery for bacterial culture, microbial DNA extraction, plasma D-lactate and endotoxin measurement. PCR analysis was performed after DNA extraction, with beta-lactosidase gene of E. coli and 16S rRNA gene as target genes. All patients were observed for a period of 30 days for infectious complications. Our results showed that no bacterial DNA was detected before surgery, but after operation it was found in 12 patients (19.0%). Bacterial DNA was detected in 41.7% (10/24) of SIRS patients and 5.1% (2/39) of non-SIRS patients (P<0.01). About 83.3% of PCR-positive patients developed systemic inflammatory response syndrome (SIRS), but only 27.5% of PCR-negative patients did so (P<0.01). Two thirds of PCR-positive patients developed infectious complications, while none of PCR-negative patients did (P<0.01). The blood culture was positive only in 3 patients (4.8%), who were all PCR-positive. E. coli DNA was found in 66.7% of the PCR-positive patients. The plasma levels of D-lactate and endotoxin were elevated significantly 2, 24 and 48 h after operation in PCR-positive patients, with a significant positive correlation found between them (r=0.91, P<0.01). It is concluded that increased intestinal permeability was closely related with bacterial translocation. Intestinal bacterial translocation (most commonly E. coli) might occur at early stage (2 h) after abdominal surgery. Postoperative SIRS and infection might bear a close relationship with bacterial translocation.

Comparison of Endotoxin and Particle Bounce in Marple Cascade Samplers With and Without Impaction Grease

ABSTRACT The health of persons engaged in agricultural activities are often related or associated with environmental exposures in their workplace. Accurately measuring, analyzing, and reporting these exposures is paramount to outcomes interpretation. This paper describes issues related to sampling air in poultry barns with a cascade impactor. Specifically, the authors describe how particle bounce can affect measurement outcomes and how the use of impaction grease can impact particle bounce and laboratory analyses such as endotoxin measurements. This project was designed to (1) study the effect of particle bounce in Marple cascade impactors that use polyvinyl chloride (PVC) filters; (2) to determine the effect of impaction grease on endotoxin assays when sampling poultry barn dust. A pilot study was undertaken utilizing six-stage Marple cascade impactors with PVC filters. Distortion of particulate size distributions and the effects of impaction grease on endotoxin analysis in samples of poultry dust distributed into a wind tunnel were studied. Although there was no significant difference in the overall dust concentration between utilizing impaction grease and not, there was a greater than 50% decrease in the mass median aerodynamic diameter (MMAD) values when impaction grease was not utilized. There was no difference in airborne endotoxin concentration or endotoxin MMAD between filters treated with impaction grease and those not treated. The results indicate that particle bounce should be a consideration when sampling poultry barn dust with Marple samplers containing PVC filters with no impaction grease. Careful consideration should be given to the utilization of impaction grease on PVC filters, which will undergo endotoxin analysis, as there is potential for interference, particularly if high or low levels of endotoxin are anticipated.

Acute effects after occupational endotoxin exposure at a spa

Two spa workers reported such symptoms as fever, shivering, palpitation, arthralgia, and diarrhea after performing seaweed massages on clients at a spa center. This study was carried out to determine whether the symptoms were related to exposure to endotoxin. Personal and stationary air sampling for the measurement of airborne endotoxin was carried out at the spa during the preparation of a bath and the following seaweed massage. In addition, the impact of storage time on the concentration of endotoxin in the seaweed was investigated. The measurements confirmed exposure to aerosolized endotoxin at the spa (11 ng/m (2)and 22 ng/m (3)). The endotoxin concentration in the stored seaweed increased as the storage time increased, from 360 ng/g seaweed for fresh seaweed to 33100 ng/g seaweed for seaweed stored for >20 weeks. Organic dust toxic syndrome was diagnosed for two workers who performed seaweed massages at a spa center at which aerosolized endotoxin was measured. In order to minimize entotoxin exposure during massages, it is important to use fresh seaweed or seaweed kept well cooled for no more than 2-3 weeks.

A microbiological survey of bicarbonate-based replacement circuits in continuous veno-venous hemofiltration*

The potential for clinically significant transfer of pyrogen-inducing material in dialysate and substitution fluids is well recognized in the setting of chronic hemodialysis and hemodiafiltration and has led to the establishment of strict standards for microbiological purity. Preliminary evidence has indicated the potential for fluid contamination in continuous renal replacement therapy, and although the scale of the problem in contemporary, industry-standard equipment is unclear. We aimed to define the microbial integrity of modern continuous veno-venous hemofiltration (CVVH) replacement fluid circuitry. Twenty-four CVVH replacement fluid circuits (mean lifespan, 34.2 hours; range, 4-86) were studied at completion of therapy. The integrated critical care unit and cardiothoracic intensive care unit of the Freeman Hospital, Newcastle upon Tyne, United Kingdom, between January and August 2007. Patients with renal failure receiving treatment with CVVH. Nil. Culture and endotoxin assays of replacement fluid, culture of endoluminal swabs, and electron microscopy of harvested tubing. Of the 24 replacement fluid cultures, nine (mean lifespan 32.8 hours, range 5-79) breached European Pharmacopoeia standards for ultrapure water (<0.1 colony-forming units/mL). One of 24 endotoxin measurements breached European Pharmacopoeia standards (<0.03 endotoxin units/mL). Internal tubing cultures were negative, but electron microscopy revealed 13 of the 24 collected tubing samples to be contaminated with biofilm. Only seven of the 24 studied circuits proved to be free from microbial contamination. We have confirmed frequent breaches of microbial integrity in industry-standard, bicarbonate-based CVVH, indicating the potential for added risk to the vulnerable, critically ill patient. These findings are of particular concern given the need for systemic infusion of replacement fluid. Measures to reduce the levels of contamination and their impact are discussed.

Translocation of bacterial NOD2 agonist and its link with inflammation

IntroductionThe gut is often considered as the motor of critical illness through bacterial translocation, which amplifies the inflammatory response and alters the immune status. However, systemic bacterial translocation was rarely proven and endotoxin measurement only reflects translocation of Gram-negative-derived products. The process could be more frequently identified if peptidoglycan, derived from both Gram-negative and Gram-positive bacteria, was measured.MethodsWe developed a new tool to detect circulating peptidoglycan-like structure using a NOD2-transfected cell line. We also measured plasma and cell-associated endotoxin and different plasma markers of inflammation. We studied 21 patients undergoing abdominal aortic surgery (AAS), and 21 patients undergoing carotid artery surgery (CAS) were included as negative controls. Patients were sampled during surgery until two days post-surgery.ResultsIn 90.5% of the AAS patients, a NOD2 agonist peak was detected in plasma before aortic clamping, but after gut manipulation by the surgeon, and persisted after blood reperfusion. As expected, no peak was detected in plasma from CAS patients (P = 0.003). Leukocyte-bound endotoxin appeared after blood reperfusion in 71% of the AAS patients, and circulating endotoxin was detected for 57% of them. The levels of interleukin (IL)-6, IL-10 and of inflammatory markers (C-reactive protein, procalcitonin) were maximal at postoperative day 1 or 2 in AAS patients. The levels of circulating NOD2 agonist positively correlated with those of cortisol and IL-10.ConclusionsThe measurement of circulating NOD2 agonist gives a higher informative tool than that of circulating endotoxin for early and sensitive detection of the translocation of bacterial products. The data suggest that circulating NOD2 agonist contributes to further enhance the stress response following surgery.

Newly developed endotoxin measurement method (endotoxin activity assay) may reflect the severity of sepsis

Endotoxin (ET) is a structural molecule of the Gram-negative bacilli extracellular membrane, which activates target cells including macrophages and neutrophils, and causes septic shock. But it is known that the conventional ET measurement method has many problems; for example, a discrepancy between plasma ET concentration and clinical manifestation in the septic patient. We therefore evaluate the usefulness of a new developed method to measure the plasma ET activity (endotoxin activity assay (EAA)) [1] in patients under sepsis compared with the ordinary method of the limulus amebocyte lysate (LAL) assay.

Measurement of Airborne Bacteria and Endotoxin Generated During Dental Cleaning

Dynamic dental instruments generate abundant aerosols in the work environment. Dental unit waterlines (DUWL) support a large microbial population and can be a significant source of bioaerosols generated during dental treatments. This study was conducted to characterize bioaerosol generation during dental treatments performed in standardized conditions. Culture-based method (R2A, and blood agar with and without O2) and fluorescence microscopy were used. Dental cleaning procedures were performed in an isolated treatment room with controlled ventilation rate. Andersen microbial samplers were used to collect culturable bioaerosols generated before (baseline), during, and after 2 hr of dental treatments. Inhalable dust samplers were used to measure total bioaerosols content in dental hygienist's and patients' breathing zones. AGI-30 were used for the collection of the endotoxin. The use of fluorescence microscopy in combination with culture demonstrated that dental staff and patients were exposed to up to 1.86 E+05 bacteria/m3 generated during treatments. Fortunately, bioaerosols returned to baseline within 2 hr after the dental procedures. The small diameter of the aerosols generated (<1 μm) suggests that the risk of contact between the aerosolized bacteria and the respiratory system of exposed individuals is likely to occur.

Variability in Endotoxin Exposure Levels and Consequences for Exposure Assessment

Workers in many industries are exposed to endotoxins, which may cause adverse health effects. In exposure assessment, information about exposure variability is essential. However, variability in exposure has rarely been investigated for biological agents and more specifically for endotoxin. Therefore, variance components and determinants of exposure were studied in a large database with >2000 endotoxin measurements. Data from 10 individual studies were combined to create a database with 2010 personal inhalable dust and endotoxin measurements, of which 1650 were repeated measurements. Exposure groups were defined based on job codes. Between- and within-worker variance components were estimated for different grouping strategies, and determinants of exposure were studied using mixed effects models. Inhalable dust and endotoxin exposure levels are summarized for 46 industries and 4 broadly defined sectors. The between-worker variability exceeded the within-worker variability overall and within sectors and subsectors, and variance components were larger for endotoxin than for dust. Between-worker variability also exceeded within-worker variability in nearly half of the exposure groups based upon industries or job code within industries for endotoxin exposure and in 10% of the groups for dust exposure. Among other things, dustiness of the process, contact with animals, bulk production, presence of plant material or a cyclic process appeared as determinants of exposure, which largely explained the between-worker variability. Exposure groups were much less homogeneous for endotoxin exposure than for dust exposure. This is distinctly different than for chemical exposure. Large variability in measured exposure levels is inherent to endotoxin exposure, which is caused in part by determinants that influence growth of microorganisms. These findings have major consequences for the design of future occupational intervention and epidemiological studies. The measurement effort needs to be greater than exposure assessment for chemical agents which demonstrate lower exposure variability, especially when evaluating endotoxin exposure for compliance testing. The established determinants of exposure give direction for potential exposure control, although more information about determinants of day-to-day variability in exposure is still needed to be able to effectively control endotoxin exposure.

Effect of Extraction and Assay Media on Analysis of Airborne Endotoxin

The measurement of airborne endotoxins is thus far not standardized. Earlier studies reported higher endotoxin yields when Tween 20 was added to the media used for filter extraction and in the Limulus amebocyte lysate (LAL) assay. This study compared four common media and assessed the effects of Tween during extraction and analysis separately. Parallel airborne dust samples from five work environments (n = 250) were used to compare the four media (pyrogen-free water [PFW], PFW-Tween 20, PFW-Tris, and PFW-triethylamine-phosphate [TAP]) and an extraction time of 10 or 60 min. A subset of the extracts in PFW or PFW-Tween (n = 40) were analyzed in parallel LAL assays with PFW or PFW-Tween as the assay medium. The results produced by a shorter extraction time or the presence of Tris were similar to the results for the reference procedure (PFW and 60 min of shaking). The use of PFW-TAP showed overall lower yields and a deviant calibration curve. The presence of Tween in the extraction medium resulted in significantly (P < 0.05) higher endotoxin yields from all dust types, independent of the effect of Tween in the assay. Tween in the LAL assay, however, also strongly inhibited the reactivity of the lipopolysaccharide (LPS) standard, thus shifting the calibration curve to higher values. The inhibition of LPS in test samples was less pronounced and varied between dust sources, resulting in enhanced calculated concentrations. This assay effect could be circumvented by diluting extracts at least 50-fold before the LAL assay. In conclusion, of the media tested, only Tween enhances the efficiency of endotoxin extraction from airborne dust samples in a consistent manner. We recommend extraction in PFW-Tween combined with dilution and LAL analysis in PFW.

Work Area Measurements as Predictors of Personal Exposure to Endotoxin and Cotton Dust in the Cotton Textile Industry

To determine if work area measurements of endotoxin and/or cotton dust obtained from the vertical elutriator (VE) can be used to predict levels of personal endotoxin exposure as measured by the Institute of Occupational Medicine (IOM) inhalable dust sampler in the cotton textile industry. Fifty-six work area cotton dust samples were collected from 14 areas and 82 personal cotton dust samples were collected from 41 workers in three textile mills (Mills A, B and C) in Shanghai, China. Cotton dust concentrations were determined gravimetrically from sample filters, of which endotoxin concentrations were determined using a kinetic chromogenic modification of the limulus amoebocyte lysate assay. Linear regression models were used to determine the association between log IOM personal endotoxin concentration and log VE area endotoxin concentration. Median cotton dust and endotoxin concentrations measured from VE area samples in the three mills were 0.36 mg m(-3) and 1280.76 endotoxin units per cubic meter (EU m(-3)), respectively, compared to 1.74 mg m(-3) and 2226.83 EU m(-3) from IOM personal samples. Excluding samples from weaving processes, we observed linear associations between VE area measures of endotoxin and IOM personal endotoxin concentrations; VE area concentration of endotoxin explained 83 and 89% of the total variation in IOM personal endotoxin concentration for Mills A and B, respectively (Mill A: R2 = 0.83, P < 0.0001; Mill B: R2 = 0.89, P < 0.0001). Although area measures of cotton dust was also a significant predictor of person endotoxin, the model explained less of the variance in personal endotoxin measurements. Specific to the conditions of the textile mills investigated in this study, work area measurements of endotoxin, but not cotton dust, may be reasonable proxies for personal levels, at least for rank-ordering exposures.

Environmental determinants of atopic eczema phenotypes in relation to asthma and atopic sensitization

There is still uncertainty about the determinants of atopic eczema (AE). To explain the heterogeneity of the disease, different phenotypes of AE have been suggested. The cross-sectional PARSIFAL study included 14 893 school-age children of farmers or children attending Steiner schools and their respective reference groups. A detailed questionnaire was completed, and house dust was collected for the measurement of endotoxin and glucans. Atopic sensitization was defined by allergen-specific IgE levels in the serum. In multivariate analyses, helping with haying was the only variable related to a farming environment having a consistent inverse association with both current symptoms and a doctor's diagnosis of AE [aOR = 0.65 (95% CI: 0.46-0.93) and 0.73 (0.51-1.05)], respectively. Severe lower respiratory tract infections (LRTI) in the first 2 years of life and usage of antibiotics ever were found to be positively related only to asthma-associated AE, whereas the effect of LRTI on AE without asthma had an opposite effect. Levels of beta(1-->3)-glucans in mattress dust were inversely related to a doctor's diagnosis of asthma-associated AE [aOR = 0.75 (0.57-0.98)], and endotoxin levels to current symptoms of asthma-associated AE [aOR = 0.73 (0.57-0.94)]. The analyses of the PARSIFAL study revealed two different phenotypes of AE, depending on the association with asthma and wheezing ever. With regard to the hygiene hypothesis, help with haying, exposure to beta(1-->3)-glucans and endotoxin were found to be inversely associated with the AE phenotype associated with asthma and wheezing.

Lymph invasion route of bacteria and endotoxin and CD4+/CD8+ T cells ratio changes of draining lymph node of burn infection wound

The aim of this study was to determine the lymphatic invasion route of bacteria and endotoxin of burn wounds infected by Pseudomonas aeruginosa and moreover, the effect of P. aeruginosa infection of the burn wound on the draining lymph node and lymph fluid. Male Wistar rats were subjected to unilateral hind limb burn + wound infected by P. aeruginosa (infection limbs group) and contralateral hind limb burn alone (burn limbs group). On hours 6, 24 and 72 after infection, rats were killed, the common iliac lymph nodes (CILN) was collected for the culture of P. aeruginosa. Lymph fluid in the efferent lymph trunk of CILN was collected for the measurement of endotoxin by the Limulus Amebocyte Lysate test. Lymph fluid tumor necrosis factor-alpha (TNF-α) concentration was measured by enzyme-linked immunosorbent assay (ELISA). The CD4+/CD8+ T cells ratio of CILN was subjected to flow cytometry analysis. The results showed bacteria invasion incidence, endotoxin and TNF-α concentrations were significantly higher in the infection limb group when compared to the burn limb group ( P < 0.01). The CD4+/CD8+ T cell ratio was significantly lower on post-burn wound infection hours 72 ( P < 0.05). This study provides evidence that bacteria and endotoxin of burn wound infected by P. aeruginosa invade draining lymph node and lymph fluid. P. aeruginosa infection of the burn wound can increase TNF-α level of the draining lymph fluid and decrease CD4+/CD8+ T cells ratio of the draining lymph node.

Natural History and Risk Factors of Early Respiratory Responses to Exposure to Cotton Dust in Newly Exposed Workers

A prospective study of newly exposed cotton workers was performed to investigate the natural history of respiratory symptoms and lung function changes. A total of 157 workers naive to cotton dust exposure were investigated by questionnaire, spirometry, and skin tests. They were examined before employment (baseline) and at the end of the first week, and the first, third, sixth, and 12th month after starting work. Acute airway response was defined as either a cross-first-shift or a cross-week fall in forced expiratory volume in one second (FEV1). The longitudinal change of lung function over the year was also calculated. Five hundred seventy-two personal dust sampling and 191 endotoxin measurements were performed to assess the exposure. Forty percent of workers reported work-related symptoms in the first week of the study. Smoking, endotoxin, and dust concentrations were risk factors for all work-related symptoms. Acute airway responses were witnessed after immediate exposure. Female status was the only factor found to be predictive of acute airway response. The mean longitudinal fall in FEV1 at 1 year was 65.5 mL (standard error = 37.2). Age, early respiratory symptoms, and early fall in cross-week FEV1 were found to predict the 12-month fall in FEV1. Cross-first-shift and cross-week falls in FEV1 reduced in magnitude during the course of the study. This study of workers naive to cotton dust exposure has demonstrated that respiratory symptoms and acute airway responses develop early following first exposure, and a tolerance effect develops in those workers with the continued exposure. Current smoking and increasing exposure predicts the development of work-related lower respiratory tract symptoms, while early symptoms and acute airway changes across the working week predict the longitudinal loss of lung function at 1 year.

Ultrapure Dialysate for Home Hemodialysis?

Use of ultrapure dialysate (bacteria < 0.1 CFU/mL and endotoxin < 0.03 EU/mL) is associated with a reduction in inflammation and morbidity in patients treated with conventional thrice-weekly dialysis. The improved outcomes obtained with more frequent dialysis schedules have reawakened interest in home hemodialysis. More frequent dialysis also appears to reduce inflammation, and whether combining more frequent dialysis with use of ultrapure dialysate will have an additive effect on inflammation and its consequences remains unclear. Routinely producing ultrapure dialysate in a home environment with a conventional hemodialysis machine poses technical challenges related to the design of the equipment and the intermittent nature of hemodialysis. Solutions to these problems include use of a system in which the water-treatment equipment is fully integrated with the dialysis machine, use of dry-powder cartridges or sterile prepackaged liquids for bicarbonate concentrate, and use of a bacteria-retentive and endotoxin-retentive filter for final purification of the dialysate immediately before it enters the dialyzer. Alternatively, ultrapure dialysate may be achieved with newer machines designed specifically for home hemodialysis that use a new batch of dialysate for each treatment. The volume of dialysate available with these machines, however, currently limits their use to short-daily dialysis.

Size Distribution of Airborne Mist and Endotoxin-Containing Particles in Metalworking Fluid Environments

The objective of the study was to investigate size-selective concentrations of airborne particles and endotoxin in metalworking fluid (MWF) environments. The experiments were conducted under two conditions: (1) MWF collected in the field was aerosolized with a laboratory-scale simulator (MWF simulator) in the laboratory; and (2) MWFs were aerosolized during routine field operations. All experiments included size-selective measurement of airborne concentrations of particle numbers and endotoxin mass using an electrical low-pressure impactor. During field sampling, the total microbial and endotoxin concentrations in the air were also measured with a BioSampler, and the mass concentration of MWF mists was measured with a photometer. Airborne particle concentrations were highest in the fine particle size ranges in the areas affected by MWFs. Relatively high concentrations of endotoxin were detected at particle size below 0.39 μm, which is smaller than the size of intact bacterial cells. The total microbial and endotoxin analysis revealed high microbial contamination in one sampling site although the total particle mass was not elevated. It was concluded that MWF sites can be contaminated with high concentrations of fine particles, and these fine particles may contain microbial components, such as endotoxin. The results call for the size-selective measurement of particles and endotoxin for more comprehensive exposure assessment in MWF facilities.

Evaluation of Endotoxaemia in the Prognosis and Treatment of Scouring Merino Lambs

SummaryThis study looked at measurement of endotoxaemia as a tool in determining prognosis and probable response to treatment in scouring lambs. One hundred eighty‐three lambs in the first 15–20 days of life, from eight Merino sheep farms located in the region of La Serena, south‐west Spain, were used in this experiment. Scouring and normal/control lambs were selected following a clinical examination, the scouring group was further divided into subgroups, specifically those that did or did not survive 72 h following treatment. At the time of the clinical examination, faecal and blood samples were taken. Faecal culture and commercial faecal antigen tests for detection of enteropathogens in faeces and serum endotoxin measurement using chromogenic lymulus amoebocyte lysate (LAL) were carried out. Scouring lambs received 0.07 mg/kg liveweight halofuginone once a day for 3 days, a single oral dose of 0.20 mg/kg liveweight of spectinomycin and oral rehydration fluid. The pathogens isolated were Cryptosporidium spp. and Escherichia coli. The case fatality rate was 51% in the scouring lambs. Postmortem findings were consistent with enterotoxigenic E. coli infection. The concentration of endotoxin was 0.18 ± 0.12 ng/ml in the control group, 0.35 ± 0.17 ng/ml in the surviving lambs and 0.46 ± 0.14 ng/ml in the non‐surviving lambs. Significant differences between groups were found. Case fatality rate of the scouring lambs with endotoxaemia below 0.30 ng/ml was 0%, while it was 100% above 0.50 ng/ml. These results may be utilized as a prognostic indicator in lambs affected by E. coli and Cryptosporidium that will help aid in decision‐making as to whether to treat a lamb or not based on its chances of survival.

Measurements of endotoxin on ambient loaded PM filters after long-term storage

At present there exist unsolved questions, whether endotoxin measurements on PM filters show valid measurements after long-term storage of loaded PM filters. As part of the TRAPCA study we collected particles with a 50% aerodynamic cut-off diameter of 2.5 μm (PM2.5) at 40 outdoor sites and particles less than 10 μm in size (PM10) at a subset of these sites (N=12) in Munich. The PM filters were cut in halves and endotoxin was measured in the extract of one filter half 2–3 years after PM collection. Here we present the results of repeated endotoxin measurements, where the association between previously measured endotoxin concentrations, measurements from stored eluates, and measurements in extracts of the second filter halves were determined. No statistically significant changes were seen between our old measurements and those from the stored eluates. Additionally, no change was observed between previously measured endotoxin concentrations and those from the stored set of filter halves. We found an average increase of 5% between our old measurements and those from the stored eluates and of 19% between the old measurements and those from the stored set of filter halves. The correlations between the previously measured endotoxin concentrations and the repeated measurements from the stored eluates were high (r=0.789) for EU/m3 PM2.5. Since endotoxin was evaluated from the same set of filter halves, it is obvious that storage over three years did not substantially effect the correlation of the endotoxin content. Regarding the outliers from previous measurements unsymmetric endotoxin spots on one out of 48 filter halves could be determined. Apart from this we conclude that the endotoxin is homogenously distributed on the filter halves and that endotoxin levels, which were repeatedly measured from filters stored over three years, could be determined validly.

Risk factors for asthma and allergy associated with urban migration: background and methodology of a cross-sectional study in Afro-Ecuadorian school children in Northeastern Ecuador (Esmeraldas-SCAALA Study).

BackgroundAsthma and allergic diseases are becoming increasingly frequent in children in urban centres of Latin America although the prevalence of allergic disease is still low in rural areas. Understanding better why the prevalence of asthma is greater in urban migrant populations and the role of risk factors such as life style and environmental exposures, may be key to understand what is behind this trend.Methods/designThe Esmeraldas-SCAALA (Social Changes, Asthma and Allergy in Latin America) study consists of cross-sectional and nested case-control studies of school children in rural and urban areas of Esmeraldas Province in Ecuador. The cross-sectional study will investigate risk factors for atopy and allergic disease in rural and migrant urban Afro-Ecuadorian school children and the nested case-control study will examine environmental, biologic and social risk factors for asthma among asthma cases and non-asthmatic controls from the cross-sectional study. Data will be collected through standardised questionnaires, skin prick testing to relevant aeroallergen extracts, stool examinations for parasites, blood sampling (for measurement of IgE, interleukins and other immunological parameters), anthropometric measurements for assessment of nutritional status, exercise testing for assessment of exercise-induced bronchospasm and dust sampling for measurement of household endotoxin and allergen levels.DiscussionThe information will be used to identify the factors associated with an increased risk of asthma and allergies in migrant and urbanizing populations, to improve the understanding of the causes of the increase in asthma prevalence and to identify potentially modifiable factors to inform the design of prevention programmes to reduce the risk of allergy in urban populations in Latin America.

Inflammatory potency of dust from the indoor environment and correlation to content of NAGase and fungi

The aim of this study was to analyse the contribution of microbial factors to the inflammatory potency of dust (PD). Floor dust was sampled three times from 12 rooms in two schools. The potency of floor dust was measured as interleukin-8 secretion from the lung epithelial cell line A549 after exposure to dust. Measurements of endotoxin, NAGase activity and cultivable fungi in the dust were made. For endotoxin a difference was found between sampling days and for NAGase a difference was found between the schools. The carpeted staff rooms of the two schools had a significantly higher amount of dust/m 2 and endotoxin/m 2, and the PD/m 2 and NAGase activity/m 2 were also higher than in the classrooms with smooth flooring. The PD/m 2 correlated with all fungal parameters except total cultivable fungi. NAGase/m 2 correlated with PD, endotoxin and cultivable Cladosporium sp. per rarea, and was nearly significantly correlated to total cultivable fungi and Penicillium spp. Therefore; microbiological activity especially from fungi may contribute to the inflammatory potency of floor dust. Carpet flooring may act as a “sink” for microorganisms resulting in a higher inflammatory potency of floor dust, which may reflect building-related symptoms in occupants.