[Low dose naltrexone in the treatment of dissociative symptoms]. / Niedrig dosiertes Naltrexon in der Behandlung dissoziativer Symptome.

BACKGROUND: Following the hypothesis that blocking opioid receptors leads to a decline in opiate-modulated dissociative phenomena, experiences with naltrexone as medication for dissociative symptoms have been gained since 1999 (mainly in doses of 25-100 mg/day). PATIENTS AND METHODS: In this study patients with severe trauma-related and dissociative disorders were treated with naltrexone in doses of 2-6 mg/day (0.06 mg/kg body weight). RESULTS: The low dose treatment with naltrexone proved to be effective whereby 11 out of 15 patients reported immediate positive effects and 7 described a lasting helpful effect. The majority of patients who felt positive effects reported a clearer perception of both their surroundings and their inner life. Assessment of reality and dealing with it improved as did the perception of their own body and affects as well as self-regulation. The treatment was very low in side effects. CONCLUSION: Treatment with low-dose naltrexone may be a helpful element in the treatment of patients with complex posttraumatic stress disorder. However, it has to be realized that the decrease of dissociation may lead patients to a not yet resolvable challenge, in as much as dissociation had previously been a necessary mechanism of self-protection.

Predicting human West Nile virus infections with mosquito surveillance data.

West Nile virus (WNV) has become established across the Americas with recent heightened activity causing significant human illness. Surveillance methods to predict the risk of human infection are urgently needed to initiate timely preventative measures and justify the expense of implementing costly or unpopular control measures, such as aerial spraying or curfews. We quantified the links between mosquito surveillance data and the spatiotemporal patterns of 3,827 human WNV cases reported over 5 years in Colorado from 2003 to 2007. Mosquito data were strongly predictive of variation in the number of human WNV infections several weeks in advance in both a spatiotemporal statewide analysis and temporal variation within counties with substantial numbers of human cases. We outline several ways to further improve the predictive power of these data and we quantify the loss of information if no funds are available for testing mosquitoes for WNV. These results demonstrate that mosquito surveillance provides a valuable public health tool for assessing the risk of human arboviral infections, allocating limited public health resources, and justifying emergency control actions.

Irrigated agriculture is an important risk factor for West Nile virus disease in the hyperendemic Larimer-Boulder-Weld area of north central Colorado.

This study focused on two West Nile virus (WNV) disease outbreak years, 2003 and 2007, and included a three-county area (Larimer, Boulder, and Weld) in North Central Colorado that is hyperendemic for WNV disease. We used epidemiological data for reported WNV disease cases at the census tract scale to: (1) elucidate whether WNV disease incidence differs between census tracts classified as having high versus lower human population density (based on a threshold value of 580 persons/km2) and (2) determine associations between WNV disease incidence and habitat types suitable as development sites for the larval stage of Culex mosquito vectors. WNV disease incidence was significantly elevated in census tracts with lower human population density, compared with those with high density of human population, in both 2003 (median per census tract of 223 and 143 cases per 100,000 population, respectively) and 2007 (median per census tract of 46 and 19 cases per 100,000 population). This is most likely related, in large part, to greater percentages of coverage in less densely populated census tracts by habitats suitable as development sites for Culex larvae (open water, developed open space, pasture/hay, cultivated crops, woody wetlands, and emergent herbaceous wetlands) and, especially, for the subset of these habitats made up by irrigated agricultural land (pasture/hay and cultivated crops) that presumably serve as major producers of the locally most important vector of WNV to humans: Culex tarsalis. A series of analyses produced significant positive associations between greater coverage of or shorter distance to irrigated agricultural land and elevated WNV disease incidence. As an exercise to produce data with potential to inform spatial implementation schemes for prevention and control measures within the study area, we mapped the spatial patterns, by census tract, of WNV disease incidence in 2003 and 2007 as well as the locations of census tracts that had either low (<25th percentile) or high (>75th percentile) WNV disease incidence in both outbreak years (relative to the incidence for each year). This revealed substantial changes from 2003 to 2007 in the spatial pattern for census tracts within the study area with high WNV disease incidence and suggests a dynamic and evolving scenario of WNV transmission to humans that needs to be taken into account for prevention and control measures to stay current and represent the most effective use of available resources.

Seasonal patterns for entomological measures of risk for exposure to Culex vectors and West Nile virus in relation to human disease cases in northeastern Colorado.

We examined seasonal patterns for entomological measures of risk for exposure to Culex vectors and West Nile virus (family Flaviviridae, genus Flavivirus, WNV) in relation to human WNV disease cases in a five-county area of northeastern Colorado during 2006-2007. Studies along habitat/elevation gradients in 2006 showed that the seasonal activity period is shortened and peak numbers occur later in the summer for Culex tarsalis Coquillett females in foothills-montane areas >1600 m compared with plains areas <1600 m in Colorado's Front Range. Studies in the plains of northeastern Colorado in 2007 showed that seasonal patterns of abundance for Cx. tarsalis and Culex pipiens L. females differed in that Cx. tarsalis reached peak abundance in early July (mean of 328.9 females per trap night for 18 plains sites), whereas the peak for Cx. pipiens did not occur until late August (mean of 16.4 females per trap night). During June-September in 2007, which was a year of intense WNV activity in Colorado with 578 reported WNV disease cases, we recorded WNV-infected Cx. tarsalis females from 16 of 18 sites in the plains. WNV infection rates in Cx. tarsalis females increased gradually from late June to peak in mid-August (overall maximum likelihood estimate for WNV infection rate of 8.29 per 1000 females for the plains sites in mid-August). No WNV-infected Culex mosquitoes were recorded from sites >1600 m. The vector index for abundance of WNV-infected Cx. tarsalis females for the plains sites combined exceeded 0.50 from mid-July to mid-August, with at least one site exceeding 1.00 from early July to late August. Finally, we found that abundance of Cx. tarsalis females and the vector index for infected females were strongly associated with weekly numbers of WNV disease cases with onset 4-7 wk later (female abundance) or 1-2 wk later (vector index).

Colorado surveillance program for chronic wasting disease transmission to humans: lessons from 2 highly suspicious but negative cases.

OBJECTIVE: To describe 2 patients with rapidly progressive dementia and risk factors for exposure to chronic wasting disease (CWD) in whom extensive testing negated the possible transmission of CWD. Design/ METHODS: We describe the evaluation of 2 young adults with initial exposure histories and clinical presentations that suggested the possibility of CWD transmission to humans. Patients A 52-year-old woman with possible laboratory exposure to CWD and a 25-year-old man who had consumed meat from a CWD endemic area. INTERVENTIONS: Clinical evaluation, neuropathological examination, and genetic testing. RESULTS: Neuropathological and genetic assessment in the 2 patients proved the diagnoses of early-onset Alzheimer disease and a rare genetic prion disease. CONCLUSION: No convincing cases of CWD transmission to humans have been detected in our surveillance program.

West Nile viremic blood donors and risk factors for subsequent West Nile fever.

BACKGROUND: While increasing age is a known risk factor for neuroinvasive West Nile virus (WNV) disease, little is known about risk factors for West Nile fever (WNF). In 2003, United States blood centers identified WN (West Nile) viremic donors using nucleic acid-amplification tests (NATs), making it possible to prospectively determine risk factors for WNF. We report the characteristics of WN viremia among donors at Colorado's largest blood center and risk factors for WNF in viremic donors. METHODS: Prospective public health surveillance was conducted in WN viremic donors. NAT-reactive donors who developed WNV-specific IgM antibody were considered viremic donors. Demographic data were abstracted from blood center records for all viremic donors aged>or=18 years. Standardized telephone questionnaires were administered a median of 39 days following donation. Donors reporting fever and headache within seven days following donation were considered West Nile fever (WNF) cases. RESULTS: Of 66,771 donations screened from July 1-October 31, 146 (0.22%) were from viremic donors (1:457 donations). Of 135 surveyed donors, 81 (60%) were male. The median age was 49 years (range: 18-78). Forty-one (30%) donors developed WNF; of these, 12 (29%) visited a physician. Other reported symptoms included body aches (98%), eye pain (63%), and skin rash (61%). The risk of WNF was inversely correlated with age (odds ratio: 0.95 for every 1-year increase in age; 95% CI 0.91, 0.99; p=0.008). CONCLUSIONS: WN viremia was frequently identified in Colorado blood donors during the 2003 WNV epidemic. The high frequency of WNF and subsequent physician visits among healthy blood donors suggest substantial morbidity from WNF in the general population. The inverse correlation between age and WNF is a new finding and its pathophysiology is unknown.

Outbreak of Q fever associated with a horse-boarding ranch, Colorado, 2005.

Coxiella burnetii is a bacterium located worldwide that can cause Q fever when inhaled. We describe an outbreak of Q fever associated with a horse-boarding ranch that had acquired two herds of goats. We conducted case finding and cohort studies among persons who boarded horses on the ranch and ranchers and among residents in the surrounding community, and conducted sampling of the goats and environment, to determine risk factors for infection and guide public health interventions. Sixty-six ranchers and persons who boarded horses on the ranch were interviewed; 62 (94%) were not professional ranchers. Twenty persons (53%) of 38 persons tested had evidence of infection with C. burnetii. Contact with goats was associated with seropositivity, including having helped birth goats (relative risk [RR] 2.4, 95% confidence interval [CI] 1.6-3.6), having had contact with newborn goats (RR 2.3, CI 1.2-4.3), having vaccinated goats (RR 2.1, CI 1.3-3.5), having had contact with stillbirths or newborns that died (RR 2.1, CI 1.2-3.7), and having fed goats (RR 2.1, CI 1.0-4.3). Among 138 tested persons living within 1 mile of the ranch, 11 (8%) demonstrated evidence of C. burnetii infection; eight seropositive persons (73%) had no direct contact with the ranch. Testing of the soil and goats with an IS1111 polymerase chain reaction (PCR) assay confirmed the presence of C. burnetii among the herd and in the environment. This outbreak of Q fever was caused by exposure to infected goats, but exposure to the environment likely played a secondary role. Laypersons should not participate in the birthing process of goats; professionals who come into contact with birthing goats should be educated on reducing their infection risk. This is the first time an IS1111 PCR assay has been used in an outbreak investigation in the United States.

West Nile virus disease: a descriptive study of 228 patients hospitalized in a 4-county region of Colorado in 2003.

BACKGROUND: Risk factors for complications of West Nile virus disease and prognosis in hospitalized patients are incompletely understood. METHODS: Demographic characteristics and data regarding potential risk factors, hospitalization, and dispositions were abstracted from medical records for residents of 4 Colorado counties who were hospitalized in 2003 with West Nile virus disease. Univariate and multivariate analyses were used to identify factors associated with West Nile encephalitis (WNE), limb weakness, or death by comparing factors among persons with the outcome of interest with factors among those without the outcome of interest. RESULTS: Medical records of 221 patients were reviewed; 103 had West Nile meningitis, 65 had WNE, and 53 had West Nile fever. Respiratory failure, limb weakness, and cardiac arrhythmia occurred in all groups, with significantly more cases of each in the WNE group. Age, alcohol abuse, and diabetes were associated with WNE. Age and WNE were associated with limb weakness. The mortality rate in the WNE group was 18%; age, immunosuppression, requirement of mechanical ventilation, and history of stroke were associated with death. Only 21% of patients with WNE who survived returned to a prehospitalization level of function. The estimated incidence of West Nile fever cases that required hospitalization was 6.0 cases per 100,000 persons; West Nile fever was associated with arrhythmia, limb weakness, and respiratory failure. CONCLUSIONS: Persons with diabetes and a reported history of alcohol abuse and older persons appear to be at increased risk of developing WNE. Patients with WNE who have a history of stroke, who require mechanical ventilation, or who are immunosuppressed appear to be more likely to die. Respiratory failure, limb weakness, and arrhythmia occurred in all 3 categories, but there were significantly more cases of all in the WNE group.

Genetic divergence of rabies viruses from bat species of Colorado, USA.

Molecular epidemiological studies have linked many cryptic human rabies cases in the United States with exposure to rabies virus (RV) variants associated with insectivorous bats. In Colorado, bats accounted for 98% of all reported animal rabies cases between 1977 and 1996. The genetic divergence of RV was investigated in bat and terrestrial animal specimens that were submitted for rabies diagnosis to the Colorado Department of Public Health and Environment (CDPHE), Colorado, USA. RV isolates from animal specimens across the United States were also included in the analysis. Phylogenetic analyses were performed on partial nucleoprotein (N) gene sequences, which revealed seven principal clades. RV associated with the colonial big brown bat, Eptesicus fuscus, an bats of the genus Myotis were found to segregate into two distinct clades (I and IV). Clade I was harbored by E. fuscus and Myotis species, but was also identified in terrestrial animals such as domestic cats and striped skunks (Mephitis mephitis). Clade IV was divided into subclades IVA, IVB, and IVC; IVA was identified in E. fuscus, and Myotis species bats, and also in a fox; subclades IVB and IVC circulated predominantly in E. fuscus. Clade II was formed by big free-tailed bat (Nyctinomops macrotis) and striped skunk (Mephitis mephitis) samples. Clade III included RVs that are maintained by generally solitary, migratory bats such as the silver-haired bat (Lasionycteris noctivagans) and bats of the genus Lasiurus. Big brown bats were found to harbor this RV variant. None of the Colorado specimens segregated with clades V and VII that harbor RVs associated with terrestrial animals. Different species of bats had the same RV variant, indicating active inter-species rabies transmission. In Colorado, animal rabies occurs principally in bats, and the identification of bat RVs in cat, gray fox Urocyon cinereoargenteus), and striped skunks demonstrated the importance of rabies spillover from bats to domestic and terrestrial wildlife species.

[Intestinal helminthiasis in school children in Haiti in 2002]. / Helminthoses intestinales en milieu scolaire en Haïti en 2002.

A survey on intestinal helminths in school children was conducted in Haiti in 2002. This first nationwide study involving the entire country was stratified by department according to urban and rural zones using the cluster method. Focusing on elementary school children (n=5792; age range 3 to 20 years), it involved 26 urban and 49 rural schools randomly selected. Stools were preserved in formalin and examined by the Ritchie technique. Thirty-four per cent of stools (1981/5792) tested positive for intestinal helminths with the following parasites identified: Ascaris lumbricoides (27.3%), Trichuris trichiura (7.3%), Necator americanus (3.8%), Hymenolepsis nana (2%), Taenia sp. (0.3%) and Strongyloides stercoralis (0.2%). The helminth prevalence was higher in rural (38.4%) compared to urban areas (30%). There was no significant difference in prevalence by sex and age. The importance of geohelminths changed from one department to another with the highest prevalence found in the Southern department of Grande Anse (73.7%) and the lowest prevalence in the Center department (20.6%). Five out of the country's nine departments had a similar prevalence varying from 25.5% to 28.2%. Intestinal helminthic polyparasitism was observed in a percentage of infested school children comprise between 3.4% and 28.6% according in relation to the geographical area. A program to fight against geohelminths in school children should be initiated as a public health priority. Albendazole is the drug of choice. Frequency of drug distribution should be based on the prevalence of geohelminths in each department.

An interstate outbreak of tick-borne relapsing fever among vacationers at a Rocky Mountain cabin.

In July 1995, an outbreak of acute febrile illness affected 11 (48%) of 23 family members from Nebraska and Kansas who had vacationed at a Colorado cabin in June. Similar symptoms were identified among five (17%) of 30 additional persons from Nebraska, Kansas, Florida, and Texas who had vacationed at the same cabin. Symptoms suggested tick-borne relapsing fever (TBRF). Although no spirochetes were detected in available blood smears from five case-patients, Borrelia hermsii was cultured from the blood of one case-patient and two chipmunks trapped near the cabin. Case-patients were more likely than non-ill cabin visitors to have slept on the floor (odds ratio [OR] = 28.0, 95% confidence interval [CI] = 3.0-258) or in the top bunk bed (OR = 5.2, 95% CI = 1.1-25.1). Tick-borne relapsing fever should considered in the differential diagnosis of fever in patients who have stayed overnight in mountain cabins in the western United States.

In vitro analysis of immunoprotective effects of topical sunscreens.

The aim of this study was to evaluate procedures for the assessment of the immunoprotective capacities of topical sunscreens, using cell cultures. The exposure of humans to solar or ultraviolet radiation has been shown to induce numerous changes at the level of the immunoresponsiveness of the skin, which could be described as immune suppression. In this study, a sunscreen containing the commercially available UVB filters octyl triazone, phenylbenzimidazole sulfonic acid and methylbenzylidene camphor, was tested because of its capacity to protect from the immunosuppressive effects of UVB radiation. In contrast to control suncreens with a comparable sun protector factor (SPF), but that did not contain the UVB-filter octyl triazone, the immunoprotective sunscreen protected from two forms of immune suppression as assessed in vitro. Expression of the immune-relevant cellular communication structure intercellular adhesion molecule-1 is decreased on the cell surface of epidermal cells under the influence of UVB. This effect of immune suppression is totally abolished by the immunoprotective capacity of the tested sunscreen, as monitored cytofluorometrically. The mixed-lymphocyte reaction (MLR) was also used to assess the protective capacities of sunscreens against UVB-radiation-induced suppression of the immune response. Again, this UVB-induced suppression of the MLR was prevented by the sunscreen containing octyl triazone. In summary, the results of this study demonstrate the usefulness of the above-mentioned methods for the in vitro evaluation of the immunoprotective properties of topical sunscreens.

Differentiation of monocytes in vitro: Influence of different types of substances.

The monocyte is one of the cell types activated in inflammatory processes to leave the blood and to enter the site of tissue inflammation through the endothelium as part of the cellular infiltrate. In tissue, monocytes differentiate to macrophages. This process is characterized by the expression of differentiation antigens, which can be monitored by monoclonal antibodies. A set of monoclonal antibodies has been used to design a test to analyse the ability of substances to influence the pattern of antigen expression during macrophage differentiation. Irritant substances stimulate the enhanced expression of an 'acute marker', which indicates the infiltration of macrophages into acutely inflamed tissues. Anti-inflammatory substances stimulate the enhanced expression of a 'middle marker' associated with the down-regulatory phase of macrophages. The stimulation of the 'acute marker' is positively correlated with the potential of substances and products to induce or elicit adverse skin reactions.

Combined in vitro assay for photohaemolysis and haemoglobin oxidation as part of a phototoxicity test system assessed with various phototoxic substances.

12 well-known phototoxicants used in the EC/COLIPA (The European Cosmetic, Toiletry and Perfumery Association) interlaboratory validation trial on 'Photoirritation in vitro' have been analysed in view of their ability to induce photohaemolysis. As well as, or instead of, this effect, oxidative reactions involving oxyhaemoglobin have been monitored, by assessing intra- and extracellular methaemoglobin formation, as an additional parameter in the screening of deleterious photoeffects on the cellular protein of red blood cells. A combination of both photohaemolysis and photo-oxidation of haemoglobin is proposed for the investigation of light-induced toxicity in an in vitro test system. Piroxicam and 8-methoxypsoralen did not produce any marked phototoxic alterations in this system. Rose bengal and amiodarone demonstrated predominantly membrane effects, whereas the antibiotics tetra- and doxycycline damaged exclusively the intracellular haemoglobin. All other phototoxins caused both photohaemolysis and photo-oxidation of haemoglobin.

Immunocompetent cells for in vitro screening of skin irritation.

The present studies were aimed at evaluating procedures for assessing the immunmodulatory effects of chemicals and preparations on macrophage differentiation and lymphocyte proliferation in cell cultures. The effects of 10 drugs and anti-inflammatory agents were monitored by determining thymidine incorporation into phytohaemagglutinin (PHA)-stimulated T cells in the lymphocyte transformation test (LTT) and the expression of two surface antigens on macrophages in the macrophage differentiation assay (MDA). One antigen was found on macrophages in acute inflamed tissue. The other was detected on those found in recovering tissue. These parameters were compared with mean skin irritation scores for 12 known cosmetic products from epicutaneous patch testing. Finally, these parameters were also used to study six cosmetic test formulae with unknown irritation potentials subjected to blind testing during phase 2 of the "CTFA Evaluation of Alternatives Program". Immunosuppressive agents were detected in both systems. Agents, thought to be pro-inflammatory, were monitored in the MDA by the acute inflammation marker. Skin irritation scores of known preparations correlated well with those of expressed acute inflammation markers in the MDA (r(s) = 0.714), but no clear relationship was detectable in the LTT. In contrast one of the CTFA samples tested blind revealed a strong response in both tests. The roll-on antiperspirant stimulated T-cell proliferation and induce a strong expression of the acute inflammation marker on macrophages. Based on these findings further studies are in progress to evaluate the usefulness of these in vitro tests for predicting dermal irritation.

Performance of the Pollen Tube Growth Test in the COLIPA Validation Study on Alternatives to the Rabbit Eye Irritation Test.

In the present paper, we describe and analyse the performance and the results of the pollen tube growth (PTG) test applied to the COLIPA international validation study of in vitro alternatives to the Draize eye irritation test. The PTG test, based on photometric quantification of in vitro pollen tube mass production, was used by three independent laboratories to estimate the acute eye irritation potentials of 23 ingredients and 32 cosmetic formulations. Basing on historical Draize test data and on IC(50) values of previously tested cosmetic formulations, a mathematical formula was generated to predict rabbit eye irritation potentials from PTG test results. Statistical evaluation of the calculated modified maximal average scores (MMAS) revealed a high prediction capability of the PTG test in regard to the finished formulations but a relatively low one for alcohols, higher concentrated cationic surfactants, and acidic and alkaline materials. Furthermore, our results indicated that the PTG test was able to produce precise IC(50) values without any limitations from all of the 55 test substances with good intra- and interlaboratory reproducibility. From these findings we suggest that the PTG test is not a validated test at present but is considered to be a potent candidate for further validation processes. For this purpose an additional prediction model for ingredient classes as mentioned above must be generated.

Validation study of alternatives to the Draize eye irritation test in Germany: Cytotoxicity testing and HET-CAM test with 136 industrial chemicals.

According to OECD guideline 405 revised in 1987 Draize eye tests need not be performed for severely irritating and corrosive chemicals if results from 'well-validated alternative studies' are presented. In 1988 a validation study on alternatives to the Draize eye test was started in Germany to establish 'well-validated alternative methods' for this purpose. During database development, the last stage of the validation programme, 136 chemicals from the German chemical industry were classified in a blind trial with the 3T3 cell neutral red/kenacid blue cytotoxicity assay and the hen's egg chorioallantoic membrane (HET-CAM) test using fertile chicken eggs. The major goal of this stage of validation was to demonstrate the feasibility and limitations of the two alternative methods. Chemicals were, therefore, selected as representatives of chemical structural groups as well as of physicochemical and toxicological properties. In addition, some of the chemicals were chosen because they were of interest to the cosmetic and detergent industries. Draize eye testing data in vivo were provided by industry. In contrast to data from a previous interlaboratory assessment trial, it was impossible to correlate cytotoxicity data to the EEC classification for in vivo eye irritation. However, seven of 10 severely irritating chemicals (EEC labelling R-41) could be identified correctly in the HET-CAM assay, whereas test conditions of the study described here did not allow identification of irritating chemicals (EEC labelling R-36). The HET-CAM test is, therefore, fulfilling the criteria of a 'well-validated alternative method' according to OECD guideline 405 and should be incorporated into eye irritation testing at the earliest possible stage to reduce effectively the suffering of rabbits in the Draize eye test. Although an 80% correct prediction of 'non-labelled' chemicals in the HET-CAM test is encouraging, for safety assessment of non-irritant chemicals, for use as cosmetic formulations, for example, both government and industry will accept an in vitro assay only if its prediction of the absence of irritant properties is 100% correct.

COLIPA validation project on in vitro eye irritation tests for cosmetic ingredients and finished products (phase I): the red blood cell test for the estimation of acute eye irritation potentials. Present status.

The red blood cell test (RBC test) is part of the COLIPA Validation Project on Alternatives to Draize Eye Irritation. It shows good intra- and interlaboratory reproducibility (reliability) and represents one of the promising in vitro alternatives of this project with a good fit to prediction models (relevance) for the assessment of acute ocular irritancy caused by certain classes of chemicals (mainly surfactants) and formulations. Results obtained during the period of test development, prevalidation, and validation are summarized. The method is based on that of Pape et al. (1987), Pape and Hoppe (1990) and Lewis et al. (1993). The protocol has two endpoints: cellular lysis and changes in protein conformation which can be correlated with initial events in tissue injury inducing inflammatory responses as assessed by Draize eye irritation scoring. Both endpoints are detected by spectrophotometric changes in the haemoglobin absorption at 541nm. The protocol also includes a set of prediction models (PM). One PM is designed to predict three classes of irritancy (classification model) based on both endpoints and the three other PMs are designed to predict modified maximum average scores (MMAS) by algorithms based on data from cellular lysis only. These three PMs [with prediction intervals (PIs)] are: (i) for surfactant ingredients, (ii) for surfactant containing finished products, and (iii) for both groups together. The three PMs are based on a common algorithm derived from historic data. It is shown that PMs derived from historic data from several laboratories, by the same procedure, also produce a good fit with the presented data. Therefore, participating laboratories concluded that the protocol as used in this formal validation study can be considered to be validated for the estimation of acute eye irritation potential of surfactant-containing finished products.

EEC/COLIPA project on in vitro phototoxicity testing: First results obtained with a Balb/c 3T3 cell phototoxicity assay.

In a joint validation project eight laboratories from the European Cosmetic Industry Association (COLIPA) as well as FRAME (England) and ZEBET (Germany) are trying to develop validated in vitro methods to be incorporated into new international guidelines for acute phototoxicity testing. The first stage of the study involved selection of the most promising in vitro phototoxicity tests for further validation. 20 chemicals with known phototoxic properties (12 phototoxins, four UV-absorbing non-phototoxins and four non-UV absorbing non-phototoxins) were tested under identical conditions of UV exposure conditions (sun simulator, UVA 5 J/cm(2)) in a standardized cytotoxicity assay with Balb/c 3T3 fibroblasts (endpoint: neutral red uptake, NRU). 19 of the 20 chemicals were correctly classified by the 3T3 NRU phototoxicity test, and therefore, this simple assay for phototoxicity seems very promising and should be validated further.

Interlaboratory assessment of alternatives to the Draize eye irritation test in Germany.

A national interlaboratory study to validate two alternative methods to the Draize rabbit's eye test, co-ordinated by ZEBET at the German Federal Health Office (BGA), is described. The aim of the study is to classify chemicals according to their irritation potential using the neutral red/kenacid blue (NR/KB) cytotoxicity assay and the hen's egg chorioallantoic membrane (HET-CAM) test. During the last two years 12 toxicology laboratories from industry, universities and other research institutions have tested 32 substances from a variety of chemical classes, characterized by a broad spectrum of locally irritating properties, using the NR/KB cytotoxicity test and the HET-CAM assay. Intra- and interlaboratory reproducibility of the two methods was investigated under standardized conditions. The so-far limited evaluation of the interlaboratory assessment phase of validation indicates that the results of the Draize rabbit's eye test correlate better with the results of the HET-CAM test than with those of the cytotoxicity test as far as false negative results are concerned. However, the intra- and interlaboratory reproducibility of the cytoxicity test is better than that of the HET-CAM test. The validation project has recently entered the stage of database development during which 150 chemicals will be tested in seven laboratories to provide information on whether and to what extent the NR/KB test and the HET-CAM test can replace the Draize rabbit's eye test for the classification and labelling of chemicals with regard to their eye irritation potential.