Detection of pathogenic protozoa in the diagnostic laboratory: result reproducibility, specimen pooling, and competency assessment.

Stool microscopy as performed in clinical parasitology laboratories is a complex procedure with subjective interpretation. Quality assurance (QA) programs often emphasize proficiency testing as an assessment tool. We describe a result reproducibility assessment tool, which can form part of a broader QA program, and which is based on the blinded resubmission of selected clinical samples, using concordance between the reports of the initial and resubmitted specimen as an indicator. Specimens preserved in sodium acetate-acetic acid-formalin can be stored for several months for use in such a program. The presence of multiple protozoa in one specimen does not affect concordance. Some dilution of specimens occurs in this process, and this may explain poor concordance when specimens with low protozoal concentrations are resubmitted. Evaluation of this tool in a large parasitology laboratory revealed concordance rates for pathogenic protozoa (Entamoeba histolytica/Entamoeba dispar, Giardia lamblia, and Dientamoeba fragilis) of about 80%, which may be considered for use as a benchmark value. We also used this tool to demonstrate that when pairs of specimens from one patient are pooled to create a single specimen, concordance between the results of the individual and pooled specimens is high.

Wheat germ supplement reduces cyst and trophozoite passage in people with giardiasis.

The protozoan parasite Giardia lamblia is a major cause of waterborne enteric disease worldwide. Lectins are proteins that bind to carbohydrate (sugar) moieties. Potential targets for lectins are found on the surface of most single-celled organisms. Modest concentrations of wheat germ agglutinin (WGA) have been shown to inhibit G. lamblia excystation and trophozoite growth in vitro and can reduce cyst passage in mice infected with the closely related protozoan parasite, G. muris. Commercial preparations of wheat germ (WG) contain 13-53 microg of WGA per gram. We performed a double-masked, placebo-controlled study of dietary supplementation with WG in 63 subjects with giardiasis in Montreal and Lima (25 asymptomatic patients passing cysts; 38 patients with symptoms). Asymptomatic subjects received WG (2 g, 3 times a day) or placebo (cornstarch, 2 g, 3 times a day) for 10 days, followed by metronidazole (250 mg 3 times a day) for 7 days. Symptomatic subjects received metronidazole (250 mg 3 times a day) plus either WG or placebo for 7 days. Stool specimens were collected every day (Montreal) or every other day (Lima) for 10 days and on Day 35 for microscopic examination and coproantigen determination. Subjects kept a diary of symptoms for 10 days after recruitment. In asymptomatic subjects, both cyst passage and coproantigen levels were reduced by approximately 50% in those taking WG compared with the placebo group (P < 0.01 and P = 0.06, respectively). In symptomatic subjects, cyst passage and coproantigen levels fell precipitously in response to metronidazole therapy, and there were no clinically important differences between those receiving supplemental WG or placebo. However, symptoms appear to have resolved more rapidly in the subjects taking WG in addition to metronidazole. The WG supplement was well tolerated in both symptomatic and asymptomatic subjects. These data suggest that components of WG, possibly WGA, either alone or in combination with antiprotozoal agents, can influence the course of human giardiasis.

Trypanosoma cruzi infection of squirrel monkeys: comparison of blood smear examination, commercial enzyme-linked immunosorbent assay, and polymerase chain reaction analysis as screening tests for evaluation of monkey-related injuries.

BACKGROUND AND PURPOSE: Wild-caught New World monkeys (NWM) from Central or South America are often infected with Trypanosoma species, including T. cruzi. In humans, T. cruzi causes Chagas' disease. Even in closed monkey colonies, T. cruzi can be propagated by blood-to-blood exposure, sexual activity, and transplacental transmission. Animal handlers and laboratory staff who deal with blood and tissues from infected NWM are at riskfor acquiring Chagas' disease via accidental exposure. METHODS: We screened 162 blood samples from wild-caught Saimiri sp. monkeys for Trypanosoma species infections by use of blood smear examination, ELISA, and polymerase chain reaction (PCR) analysis. Blood samples from 19 employees with recent history of monkey-associated injuries also were tested. RESULTS: Six percent (10/162) of the monkey samples were T. cruzi positive on the basis of blood smear examination results, 10.4% (17/162) were positive by ELISA results, and 26.5% (43/162) were positive by PCR results. Other organisms identified by PCR analysis included T. rangeli in two animals, Plasmodium spp. in two animals (P. malariae confirmed by PCR results) and microfilariae in one animal (morphologically, Mansonella perstans). Evidence of trypanosome infection was not found in the 19 employee samples on the basis of results of any of the three aforementioned tests. CONCLUSIONS: Close attention must be paid to worker safety where wild-caught NWM are used. The PCR analysis has a clear advantage over conventional techniques (ELISA, blood smear) for screening NWM for trypanosome infections during quarantine and after employee injury.

Comparison of methods of sampling for Toxocara species and fecal coliforms in an outdoor day care environment.

OBJECTIVE: To compare three sampling methods and to pretest methods for the determination of fecal coliform (FC) counts and Toxocara species from sand in the day care outdoor environment. DESIGN: The sand samples were obtained from the play area and the sandbox of a day care centre and examined for the presence of FC and Toxocara species, the common roundworm of dogs and cats. The sampling methods included random selection and two types of judgement methods. The latter included one method where domestic animals were judged to be likely to defecate and the other where children would be likely to be playing. In addition, to obtain a global estimate of contamination, the entire areas of both the sandbox and the play area were sampled on the last day. SETTING: Outdoor day care environment. MAIN RESULTS: The most representative levels of bacterial contamination and Toxocara species originated from the combined sample of the entire surface areas rather than from any separate random or judgement method of sampling. FCs were found in all sampled areas of the sandbox (median 910 FCs/g of sand) and of the play area (median 350 FCs/g of sand). Toxocara species were recovered from a number of areas in both the sandbox and the play area. CONCLUSIONS: Research on environmental microbial contamination of outdoor day care settings would benefit from the application of standardized and validated sampling and laboratory methods.

Laboratory diagnosis for Giardia lamblia infection: A comparison of microscopy, coprodiagnosis and serology.

OBJECTIVE: To evaluate newer techniques such as coproantigen detection and serology in the diagnosis of symptomatic Giardia lamblia infection. DESIGN: Blinded comparison of copro-antigen detection (by ELISA), serology (immunoglobulin IgG and IgM anti-G lamblia by ELISA, and IgG, IgM and IgA by immunoblot) and microscopy in clinical samples. Microscopic findings for three preserved stools were considered the gold standard. SETTING: Travel medicine clinic. POPULATION STUDIED: Adults, post-travel, with gastrointestinal symptomatology. MAIN RESULTS: For 152 previously collected stools, copro-antigen detection had a sensitivity of 73 of 74 (98.6%) and a specificity of 78 of 78 (100%). In clinical samples of 62 patients, eight of the 62 patients (13%) were diagnosed with G lamblia infection on microscopy. Copro-antigen diagnosis was accurate in symptomatic patients, with sensitivity of seven of eight (87.5%) and specificity of 52 of 54 (96.8%). Serology was less accurate. IgG response to G lamblia had sensitivity of four of seven and specificity of 24 of 50 (48%), and IgM response had sensitivity of three of six and specificity 27 of 48 (56%). Western blot had a sensitivity of five of seven and a specificity of 38 of 49 (78%). CONCLUSIONS: Copro-antigen diagnosis of G lamblia is highly accurate in patients with chronic gastrointestinal complaints, while serology is less accurate and appears to be less useful diagnostically.

[Survey of parasitic prevalence in school-aged children in Guinea (1995)]. / Enquête de prévalence parasitaire chez les enfants d'âge scolaire en Guinée (1995).

UNLABELLED: A national survey was carried out to assess the prevalence of helminth infections in all four geographical zones of Guinea (the four official names should be used. Guinée forestière, Haute Guinée, Moyenne Guinée and Basse Guinée) to provide information on which a school-based intervention program could be developed. The program, financed by the World Bank, would consist of vitamin supplementation, antihelmintic treatment and hygiene education. METHODS: The survey was conducted between April and June, 1995. Two prefectures (administrative areas) were selected to represent each zone, in each prefecture, one child between the age of 10 and 14 years old from each of 100 households was included in the study. Thus 800 children were enrolled. One fresh stool and one urine specimen per child were examined for the presence of parasites using the Kato-Katz method and routine microscopy. RESULTS: The following overall prevalences were obtained: bookworm (43.9%); Schistosoma mansoni (25%); S. haematobium (19.9%) Trichuris trichiura (13.5%); Ascaris lumbricoides (9.5%); Strongyloides stercoralis (6.4%) and Taenia spp (3.8%). More than 70% of the children were infected by at least one helminth. The majority (63%) of the infections were infections with only one parasite. Only 8% had three or more helminths. More boys than girls were infected (74.3% vs. 65.2%). Helminth infection was significantly associated with region, gender and school attendance as assessed by logistic regression analysis. The prevalence of Ascaris, S. mansoni and bookworm infections were higher in Guinée forestière than any other zone, although hookworm infections were common in all zones (prevalences of between 26% and 71%). The distribution of the other helminth infections differed substantially between the regions. Trichuris was most common in Conakry, Strongyloides in Boké, Taenia in Labé, and Schistosoma infections in both Haute Guinée and Guinée forestière. CONCLUSION: The prevalence of helminth infection in Guinea is high, and there is a regional distribution. Both these factors should be taken into account when planning the school-based supplementation, treatment and education intervention program.

Common-source outbreak of acute infection due to the North American liver fluke Metorchis conjunctus.

BACKGROUND: We investigated an outbreak of acute clinical illness among 19 people who ate raw fish (sashimi) prepared from the white sucker, Catostomus commersoni, caught in a river north of Montreal, Canada. METHODS: We collected epidemiological, clinical, laboratory, and serological data on 19 individuals who ate the sashimi and six who did not. Because of the suggestive clinical picture, we set out to recover helminth parasites from uneaten fish. FINDINGS: The illness consisted of persistent upper abdominal pain, low grade fever, high blood eosinophil concentrations, and raised liver enzymes. After 10 days, opisthorchild-like eggs were found in stools. Symptoms persisted for 3 days to 4 weeks without treatment, but responded rapidly to praziquantel therapy. Necropsy of golden hamsters infected with metacercariae from uneaten fish revealed adult flukes identified as Metorchis conjunctus. INTERPRETATION: We describe an acute illness caused by the North American liver fluke M conjunctus. This is a new human disease and is the first report of a common-source outbreak of an acute illness caused by liver flukes of the family Opisthorchiidae.

Modified technique for efficient detection of microsporidia.

Changes in temperature (from room temperature to 50 degrees C) and staining time (from 90 to 10 min) were evaluated as a means of improving the detection of microsporidia from stool specimens. A blinded and independent comparison of 50 known positive matched-specimen pairs by three technologists resulted in consistently easier microscopic detection. The background is clearer, and spores stain more intensely. Staining time is reduced by 80 min.