Application of the Midi Parasep® SF technique for the detection of L1 larvae of Angiostrongylus cantonensis in faeces of infected rats.

We investigated parasitic zoonoses caused by protozoans and helminths in urban and peri-urban rat populations (Rattus norvegicus and R. rattus) in Spanish cities. Rats were trapped and then dissected to remove adult helminths, and the contents of the large intestine were retrieved for the study of parasitic forms. The Midi Parasep® solvent free (SF) technique was used to concentrate the parasites in the intestinal contents. Some of the rats studied (n = 8) were infected by the rat lungworm, Angiostongylus cantonensis, whose first stage larvae (L1) are shed in rat faeces. After the concentration technique, L1 larvae were found in the sediment of 6 of the 8 positive rats. The two negative sediment samples were due to the presence of either only adult females or, in addition to males, only young females in the lungs of the rats. In view of our results, Midi Parasep® SF turned out to be a simple, rapid, inexpensive, and sensitive method to detect nematode larvae, such as the L1 larvae of A. cantonensis (or A. costaricensis), in natural and experimentally infected rats.

Toxoplasma gondii detection and viability assays in ham legs and shoulders from experimentally infected pigs.

Epidemiological studies of toxoplasmosis show that infection in humans is mainly caused by the consumption of raw, undercooked or cured meat. Cured "Serrano" ham is a typical pork product from the Mediterranean area, highly valued for its flavour. The "Serrano" ham is prepared from pork meat and undergoes a process known as curing and a subsequent fermentation without thermal or smoking treatments. The viability of Toxoplasma gondii in hams and shoulders from experimentally infected pigs that have been subject to different curing processes has been studied in order to evaluate the best method to completely eliminate the viable protozoa. The different treatments include, i) freezing the legs and shoulders below -20 °C for 3 days before salting with marine salt, ii) salting the meat with marine salt and nitrites, iii) salting only with marine salt (traditional process) and iv) salting with marine salt and then freezing at -20 °C for 3 days after the curing period. The ham leg samples were cured for 7 months and the shoulder samples for 5 months. The presence of T. gondii in the different treatments was studied by a "magnetic-capture" method for the isolation of T. gondii DNA and a quantitative real-time PCR to estimate the T. gondii burden in the ham legs and shoulders. The infectivity capacity of T. gondii in positive samples was assayed by bioassays in mice and some physicochemical parameters, such as pH, water activity (aw) and salt content, were evaluated at the end of the curing time. In all the cases where the samples were frozen the T. gondii infectivity was eliminated. In samples in which the meat was salted in marine salt plus nitrites, the parasite viability remained for longer than in the traditional salting process. The methods described here could be useful for producers to guarantee the safety of their products.

Quantification and viability assays of Toxoplasma gondii in commercial "Serrano" ham samples using magnetic capture real-time qPCR and bioassay techniques.

"Serrano" ham is a typical pork product from the Mediterranean area, highly valued for its flavour. To make Serrano ham, pork undergoes a salting and a subsequent fermentation process known as curing. Certain pigs used for meat production are an important source of Toxoplasma gondii infection in humans. We have developed a method for quantifying and assaying the viability of the T. gondii present in commercial Serrano ham samples. A magnetic capture method for the isolation of T. gondii DNA and a qRT-PCR were used to estimate the T. gondii burden in 475 commercial samples of "Serrano" ham in two presentation formats: ham pieces and sliced ham. The infectivity capacity of T. gondii in positive samples was assayed in mice. The global prevalence of T. gondii was 8.84%, ranging from 32.35% in one of the companies to 0% prevalence in three other companies. The infectivity assays revealed that only 4.84% of the positive samples were infective. To the best of our knowledge this is the first report focussing on the prevalence of T. gondii in commercial "Serrano" ham. The method described here could be useful for producers to guarantee the safety of their products.

Influence of Er:YAG and Ti:sapphire laser irradiation on the microtensile bond strength of several adhesives to dentin.

The aim of the present study was to evaluate the influence of erbium:yttrium-aluminum-garnet (Er:YAG) and Ti:sapphire laser irradiation on the microtensile bond strength (MTBS) of three different adhesive systems to dentin. Flat dentin surfaces from 27 molars were divided into three groups according to laser irradiation: control, Er:YAG (2,940 nm, 100 µs, 2.7 W, 9 Hz) and Ti:sapphire laser (795 nm, 120 fs, 1 W, 1 kHz). Each group was divided into three subgroups according to the adhesive system used: two-step total-etching adhesive (Adper Scotchbond 1 XT, from now on XT), two-step self-etching adhesive (Clearfil SE Bond, from now on CSE), and all-in-one self-etching adhesive (Optibond All-in-One, from now on OAO). After 24 h of water storage, beams of section at 1 mm(2) were longitudinally cut from the samples. Each beam underwent traction test in an Instron machine. Fifteen polished dentin specimens were used for the surface morphology analysis by scanning electron microscopy (SEM). Failure modes of representative debonded microbars were SEM-assessed. Data were analyzed by ANOVA, chi-square test, and multiple linear regression (p < 0.05). In the control group, XT obtained higher MTBS than that of laser groups that performed equally. CSE showed higher MTBS without laser than that with laser groups, where Er:YAG attained higher MTBS than ultrashort laser. When OAO was used, MTBS values were equal in the three treatments. CSE obtained the highest MTBS regardless of the surface treatment applied. The Er:YAG and ultrashort laser irradiation reduce the bonding effectiveness when a two-step total-etching adhesive or a two-step self-etching adhesive are used and do not affect their effectiveness when an all-in-one self-etching adhesive is applied.

The helminth community of the wood mouse Apodemus sylvaticus from the Erro River valley, Navarre, Spain.

The helminth fauna of the wood mouse, Apodemus sylvaticus, in the Erro River valley (Navarre, Spain) was investigated from a total of 150 mice between February 2001 and July 2002. An overall prevalence of 90.7% was recorded and up to 14 helminth species identified. The most prevalent species was the nematode Heligmosomoides polygyrus (78.0%), whereas Syphacia stroma was the species with the highest median abundance (19.8). The detection of Calodium hepaticum, Rodentolepis straminea and the larvae of Hydatigera taeniaeformis are significant, since these helminth species could be considered potential human parasites. The helminth infracommunity comprised no more than five species. A significant predominance of monoxenous species was detected. Statistically significant differences were also found between prevalences, helminth abundance, species richness and helminth diversity of sub-populations of the wood mouse determined by host age and season of capture, which agree with most of the studies carried out on this host. This study will shed light on the helminth community of the wood mouse from a region of Spain which has not previously been documented.

The helminth community of the wood mouse Apodemus sylvaticus in a Mediterranean ecosystem in regeneration ten years after a wildfire.

This study was carried out 10 years after a wildfire in the Spanish Serra Calderona Natural Park, following a previous analysis comprising the first 5 years after the fire. Its primary aim was to elucidate the impact of this perturbation on the population biology of the wood mouse Apodemus sylvaticus, and the repercussions on its helminth community in this regenerating Mediterranean ecosystem. Second, confirmation of the ability of the parasites to tolerate environmental stressors and the effects on their transmission strategies was sought. Five hundred and sixty-four individuals of A. sylvaticus were studied in a 9-year period, from the second to the tenth post-fire year: 408 mice from the burned area and 156 from the control--non-burned--area. The helminth community for both areas and the effect of intrinsic (host age and sex) and extrinsic (site, period and year of capture) factors on helminth prevalence, abundance and diversity, and species richness were analysed. Our findings show that, after an environmental disaster, the behaviour of helminth species might be related to their pre-catastrophe presence, their biological cycles, the host's immunological condition, the change of host dynamics, the direct effects of the perturbation, and the processes related to the re-establishment of the ecological balance of a devastated ecosystem.

Geographical Information Systems risk assessment models for zoonotic fascioliasis in the South American Andes region.

The WHO recognises Fasciola hepatica to be an important human health problem. The Andean countries of Peru, Bolivia and Chile are those most severely affected by this distomatosis, though areas of Ecuador, Colombia and Venezuela are also affected. As part of a multidisciplinary project, we present results of use of a Geographical Information Systems (GIS) forecast model to conduct an epidemiological analysis of human and animal fasciolosis in the central part of the Andes mountains. The GIS approach enabled us to develop a spatial and temporal epidemiological model to map the disease in the areas studied and to classify transmission risk into low, moderate and high risk areas so that areas requiring the implementation of control activities can be identified. Current results are available on a local scale for: (1) the northern Bolivian Altiplano, (2) Puno in the Peruvian Altiplano, (3) the Cajamarca and Mantaro Peruvian valleys, and (4) the Ecuadorian provinces of Azuay, Cotopaxi and Imbabura. Analysis of results demonstrated the validity of a forecast model that combines use of climatic data to calculate of forecast indices with remote sensing data, through the classification of Normalized Difference Vegetation Index (NDVI) maps.

Prevalence of bacteria and absence of anisakid parasites in raw and prepared fish and seafood dishes in Spanish restaurants.

This study evaluated the presence of bacteria and anisakid parasites in 45 samples of raw anchovies in vinegar, a dish widely eaten in Spain, and in 227 samples of cooked fish and cephalopods served in Spanish food service establishments. Our analysis showed that, according to European and Spanish regulation, 14 to 30% of the prepared fish and cephalopod dishes exceeded the maximum allowable level for mesophilic aerobic counts, and 10 to 40% of these samples exceeded the allowable levels for Enterobacteriaceae. None of the studied samples showed evidence of anisakid parasites, Escherichia coli, Staphylococcus aureus, Salmonella, or Listeria monocytogenes. These results indicate that application of hazard analysis and critical control points, food safety training courses, and routine inspections in compliance with current European and Spanish legislation help protect consumer health.

Human Bertiella studeri in Spain, probably of African origin.

A mature tapeworm and gravid proglottids of Bertiella studeri were reported from the stools of a 33-year-old pregnant Spanish woman. The patient had spent the six months preceding this discovery in Kenya. The evidence suggests a case of parasitism imported to Spain from the African continent. The patient presented no symptoms related to the parasite and the discovery in stools of a number of active, white structures led her to collect them. A morphologic and morphometric description of the material obtained is presented. Mebendazole failed to remove the parasite but niclosamide was effective.

Validation of a mapping and prediction model for human fasciolosis transmission in Andean very high altitude endemic areas using remote sensing data.

The present paper aims to validate the usefulness of the Normalized Difference Vegetation Index (NDVI) obtained by satellite remote sensing for the development of local maps of risk and for prediction of human fasciolosis in the Northern Bolivian Altiplano. The endemic area, which is located at very high altitudes (3800-4100 m) between Lake Titicaca and the valley of the city of La Paz, presents the highest prevalences and intensities of fasciolosis known in humans. NDVI images of 1.1 km resolution from the Advanced Very High Resolution Radiometer (AVHRR) sensor on board the National Oceanic and Atmospheric Administration (NOAA) series of environmental satellites appear to provide adequate information for a study area such as that of the Northern Bolivian Altiplano. The predictive value of the remotely sensed map based on NDVI data appears to be better than that from forecast indices based only on climatic data. A close correspondence was observed between real ranges of human fasciolosis prevalence at 13 localities of known prevalence rates and the predicted ranges of fasciolosis prevalence using NDVI maps. However, results based on NDVI map data predicted zones as risk areas where, in fact, field studies have demonstrated the absence of lymnaeid populations during snail surveys, corroborated by the absence of the parasite in humans and livestock. NDVI data maps represent a useful data component in long-term efforts to develop a comprehensive geographical information system control program model that accurately fits real epidemiological and transmission situations of human fasciolosis in high altitude endemic areas in Andean countries.

A global network for the control of snail-borne disease using satellite surveillance and geographic information systems.

At a team residency sponsored by the Rockefeller Foundation in Bellagio, Italy, 10-14 April 2000 an organizational plan was conceived to create a global network of collaborating health workers and earth scientists dedicated to the development of computer-based models that can be used for improved control programs for schistosomiasis and other snail-borne diseases of medical and veterinary importance. The models will be assembled using GIS methods, global climate model data, sensor data from earth observing satellites, disease prevalence data, the distribution and abundance of snail hosts, and digital maps of key environmental factors that affect development and propagation of snail-borne disease agents. A work plan was developed for research collaboration and data sharing, recruitment of new contributing researchers, and means of access of other medical scientists and national control program managers to GIS models that may be used for more effective control of snail-borne disease. Agreement was reached on the use of compatible GIS formats, software, methods and data resources, including the definition of a 'minimum medical database' to enable seamless incorporation of results from each regional GIS project into a global model. The collaboration plan calls for linking a 'central resource group' at the World Health Organization, the Food and Agriculture Organization, Louisiana State University and the Danish Bilharziasis Laboratory with regional GIS networks to be initiated in Eastern Africa, Southern Africa, West Africa, Latin America and Southern Asia. An Internet site, www.gnosisGIS.org, (GIS Network On Snail-borne Infections with special reference to Schistosomiasis), has been initiated to allow interaction of team members as a 'virtual research group'. When completed, the site will point users to a toolbox of common resources resident on computers at member organizations, provide assistance on routine use of GIS health maps in selected national disease control programs and provide a forum for development of GIS models to predict the health impacts of water development projects and climate variation.

Morphology of Bertiella studeri (Blanchard, 1891) sensu Stunkard (1940) (Cestoda: Anoplocephalidae) of human origin and a proposal of criteria for the specific diagnosis of bertiellosis.

Human material of an African specimen of Bertiella studeri (Blanchard, 1891), a typical intestinal cestode of monkeys, is described. Mature, postmature and gravid proglottides, and eggs, previously inadequately figured, are illustrated and photographed. The description of the species agrees with that provided by Stunkard (1940). A comparative study with other descriptions of the species is made in an attempt to clarify previous findings. The morphological differences reported in various earlier descriptions of the species suggest that B. studeri should be regarded as a "B. studeri species complex" Improvements are required in the descriptions of new future findings in order to clarify the specific diagnosis of human bertiellosis. Evidence suggests that a generalised diagnosis exclusively based on egg size and geographical distribution is insufficient to differentiate B. studeiri and Bertiella mucronata (Meyner, 1895), or additional species may be affecting humans.

Increased frequency of chromosomal aberrations in railroad car painters.

The purpose of this study was to determine if exposure to paints and solvents contributes to chromosomal alterations in occupationally exposed individuals. A total of 25 male railroad and underground railroad car painters were studied. This group had a mean age of 32.7 y and a mean exposure time of 5.2 y. The results were compared with those obtained for 25 healthy (unexposed) males. The scoring of structural chromosome aberrations clearly revealed an increase in the number of all types of aberrations considered in the population of painters. This suggests that exposure to a combination of chemicals may increase genotoxicity in industrial workers.

Influence of post-cure time on the microhardness of self-adhesive resin cements inside the root canal.

PURPOSE: To compare the microhardness of several dual-cure, self-adhesive resin cements used to lute fiber posts at 24 hours and seven days after cementation. METHODS: Bovine incisors were selected to lute 15 fiber posts that were 12 mm long (FRC Postec Plus size 3, Ivoclar-Vivadent). Five resin cements were tested: Multilink Automix (Ivoclar-Vivadent), without light-curing, and the self-adhesive resin cements Maxcem Elite (Kerr), RelyX Unicem (3M ESPE), G-Cem (GC), and Smartcem 2 (Dentsply), which were light-cured for 40 seconds (LED Bluephase, Ivoclar-Vivadent). Each root was embedded in chemically cured acrylic resin and stored at 37°C for 24 hours. The roots were transversally sectioned into nine specimens that were each 1 mm thick, with three specimens corresponding to each root third. Indentations (100g, 30 seconds) were performed on each section in the resin cement, at 24 hours and seven days after cementation, using a Vickers digital microdurometer (Buehler). Data were analyzed by two-way analysis of variance, Student-Newman-Keuls test, and paired t-test (p<0.05). RESULTS: A significant influence was found (p<0.05) for the resin cement evaluated, the root third, and their interactions on microhardness values at 24 hours and seven days after post cementation. RelyX Unicem and G-Cem exhibited the highest microhardness values, whereas Multilink Automix presented the lowest. All resin cements suffered a decrease in microhardness according to root canal depth, with the exception of G-Cem and Multilink Automix at 24 hours and Smartcem 2 after seven days. After seven days, the evaluated resin cements showed a significant increase in microhardness values, with the exception of Maxcem Elite and Smartcem 2 at the coronal third. CONCLUSIONS: Microhardness of the self-adhesive resin cements when used to lute fiber posts was material-dependent and higher values were obtained in the coronal third, revealing their sensitivity to light irradiation. More information regarding the polymerization reaction of these cements is warranted. According to the current results, microhardness values were significantly higher one week after post luting.