First reported foodborne outbreak associated with microsporidia, Sweden, October 2009.

Microsporidia are spore-forming intracellular parasites that infrequently cause disease in immunocompetent persons. This study describes the first report of a foodborne microsporidiosis outbreak which affected persons visiting a hotel in Sweden. Enterocytozoon bieneusi was identified in stool samples from 7/11 case-patients, all six sequenced samples were genotype C. To confirm that this was not a chance finding, 19 stool samples submitted by healthy persons from a comparable group who did not visit the hotel on that day were tested; all were negative for microsporidia. A retrospective cohort study identified 135 case-patients (attack rate 30%). The median incubation period was 9 days. Consumption of cheese sandwiches [relative risk (RR) 4·1, 95% confidence interval (CI) 1·4-12·2] and salad (RR 2·1, 95% CI 1·1-4) were associated with illness. Both items contained pre-washed, ready-to-eat cucumber slices. Microsporidia may be an under-reported cause of gastrointestinal outbreaks; we recommend that microsporidia be explored as potential causative agents in food- and waterborne outbreaks, especially when no other organisms are identified.

Microscopic diagnosis of sodium acetate-acetic acid-formalin-fixed stool samples for helminths and intestinal protozoa: a comparison among European reference laboratories.

The present study aimed to compare the diagnostic performance of different European reference laboratories in diagnosing helminths and intestinal protozoa, using an ether-concentration method applied to sodium acetate-acetic acid-formalin (SAF)-preserved faecal samples. In total, 102 stool specimens were analysed during a cross-sectional parasitological survey in urban farming communities in Côte d'Ivoire. Five SAF-preserved faecal samples were prepared from each specimen and forwarded to the participating reference laboratories, processed and examined under a microscope adhering to a standard operating procedure (SOP). Schistosoma mansoni (cumulative prevalence: 51.0%) and hookworm (cumulative prevalence: 39.2%) were the predominant helminths. There was excellent agreement (kappa > 0.8; p < 0.001) among the reference laboratories for the diagnosis of S. mansoni, hookworm, Trichuris trichiura and Ascaris lumbricoides. Moderate agreement (kappa = 0.54) was found for Hymenolepis nana, and lesser agreement was observed for other, less prevalent helminths. The predominant intestinal protozoa were Entamoeba coli (median prevalence: 67.6%), Blastocystis hominis (median prevalence: 55.9%) and Entamoeba histolytica/Entamoeba dispar (median prevalence: 47.1%). Substantial agreement among reference laboratories was found for E. coli (kappa = 0.69), but only fair or moderate agreement was found for other Entamoeba species, Giardia intestinalis and Chilomastix mesnili. There was only poor agreement for B. hominis, Isospora belli and Trichomonas intestinalis. In conclusion, although common helminths were reliably diagnosed by European reference laboratories, there was only moderate agreement between centres for pathogenic intestinal protozoa. Continued external quality assessment and the establishment of a formal network of reference laboratories is necessary to further enhance both accuracy and uniformity in parasite diagnosis.

Receptor-mediated endocytosis in an apicomplexan parasite (Toxoplasma gondii).

Endocytosis mechanisms are poorly known in apicomplexan parasites. Here, we show that extracellular tachyzoites of Toxoplasma gondii bind and internalize heparin-like sulfated glycans in a specific, saturable manner. Discrete binding of the glycan occurs at the anterior third of the tachyzoite, where it is rapidly concentrated inside single tubulo vesicular compartments that become multiple with time. The compound is held for several hours intracellularly with no apparent exocytosis or acidification. Incubation in the continuous presence of fluorescein isothiocyanate-conjugated heparin enhances the binding and internalization of this ligand by live tachyzoites. Two tachyzoite surface polypeptides exhibit strong binding and specificity for heparin, making them candidate receptors. Uptake of fluid-phase endocytic tracers occurs via nonspecific pinocytosis in the same region of the parasite cell, but with much lower efficiency. These observations show that extracellular tachyzoites can acquire molecules through both receptor-specific and fluid-phase endocytic mechanisms. Understanding the physiological relevance of these processes for the extracellular and intracellular stages of T. gondii may bring about direct targeting of the parasite by drug delivery into the tachyzoites.

A protease inhibitor associated with the surface of Toxoplasma gondii.

Toxoplasma gondii has a broad host-range including man and a variety of warm-blooded animals. The ability to infect and survive in this wide spectrum of hosts suggests highly evolved mechanisms to handle the harsh environments encountered. Here we show that extracellular tachyzoites are resistant to milligram levels of trypsin and describe the presence of an inhibitor of trypsin associated with the surface of T. gondii, TgTI. TgTI has an estimated molecular mass of 37000 dalton and is encoded by the TgTI-gene which is found at low abundance as an expressed sequence tag (EST) in both the bradyzoite and tachyzoite stages. The inhibitory binding region was found to be in the N-terminus of TgTI where aminoacid-alignment to earlier described protease inhibitors demonstrates 75% similarity. In functional analysis, recombinant TgTI-protein inhibits the activity of trypsin approximately 10 times more efficiently than an inhibitor isolated from soybean. In contrast to other known trypsin inhibitors, TgTI also possesses a predicted membrane-binding region. Polyclonal antibodies raised against recombinant TgTI bind to the surface of the tachyzoite stage as seen both by immunofluorescence and immunoprecipitation of surface labelled parasite proteins. The high survival rate of the parasite in the upper gastrointestinal tract may be enhanced by the presence of the TgTI-molecule.