Association between cervical dysplasia and female genital schistosomiasis diagnosed by genital PCR in Zambian women.

BACKGROUND: Female genital schistosomiasis (FGS) is a neglected tropical gynaecological disease that affects millions of women in sub-Saharan Africa (SSA). FGS is caused by Schistosoma haematobium, a parasitic carcinogen involved in the pathogenesis of squamous cell carcinoma of the bladder. Cervical cancer incidence and mortality are highest in SSA, where pre-cancerous cervical dysplasia is often detected on screening with visual inspection with acetic acid (VIA). There are no studies evaluating the association between VIA positivity and FGS diagnosed by genital PCR. METHODS: Women were recruited from the Bilharzia and HIV (BILHIV) study in Zambia a community-based study comparing genital self-sampling to provider obtained cervicovaginal-lavage for the diagnosis of FGS in women aged 18-31. FGS was defined as positive Schistosoma DNA from any genital PCR. Urogenital schistosomiasis diagnostics included urine circulating anodic antigen, urine microscopy and portable colposcopy. Participants were offered cervical cancer screening using VIA at Livingstone Central Hospital. Associations of PCR confirmed FGS and other diagnostics with VIA positivity were assessed using multivariable logistic regression. RESULTS: VIA results were available from 237 BILHIV participants. A positive Schistosoma PCR in any genital specimen was detected in 14 women (5.9%), 28.6% (4/14) of these women had positive VIA compared to 9.0% without PCR evidence of schistosome infection (20/223). Schistosoma PCR positivity in any genital specimen was strongly associated with VIA positivity (OR: 6.08, 95% CI: 1.58-23.37, P = 0.02). CONCLUSIONS: This is the first study to find an association between FGS and positive VIA, a relationship that may be causal. Further longitudinal studies are needed.

Ovarian cancer risk after salpingectomy for ectopic pregnancy or hydrosalpinx: results of the OCASE nationwide population-based database study.

STUDY QUESTION: What is the effect of salpingectomy for ectopic pregnancy or hydrosalpinx at a young age on ovarian cancer risk compared to no salpingectomy for any reason? SUMMARY ANSWER: We found no significant reduction in ovarian cancer risk after salpingectomy for ectopic pregnancy or hydrosalpinx. WHAT IS KNOWN ALREADY: Salpingectomy may reduce ovarian cancer incidence, although the lag-time between intervention and therapeutic effect remains to be elucidated. STUDY DESIGN, SIZE, DURATION: This nationwide population-based database study uses the Dutch pathology database to identify all women who underwent salpingectomy for ectopic pregnancy or hydrosalpinx between January 1990 and December 2012 and compared ovarian cancer incidence to a control group of women who had a benign dermal nevus removed, matched for age at the time and year of procedure. PARTICIPANTS/MATERIALS, SETTING, METHODS: After selection and manual control of intervention and control group, ovarian cancer incidence was recorded. Hazard ratios (HRs) with 95% CI for the development of ovarian cancer were calculated with Cox regression analyses, both unadjusted and adjusted for age. Subgroup analyses were performed to investigate lag-time between intervention and protective effect. MAIN RESULTS AND THE ROLE OF CHANCE: In all, 18 961 women were included in the intervention group; 17 106 women had a unilateral salpingectomy and 1855 had a bilateral salpingectomy. The control group consisted of 23 686 women. With 14 ovarian cancer cases in the intervention group, the incidence rate (IR) of ovarian cancer was 5.4 (95% CI 3.1-8.9) per 100 000 person-years. In the control group, there were 24 ovarian cancer cases, resulting in an IR of 7.1 (95% CI 4.7-10.5) per 100 000 person-years (P = 0.34). The age-adjusted HR for ovarian cancer was 0.76 (95% CI 0.39-1.47) after salpingectomy. Unilateral salpingectomy resulted in an age-adjusted HR of 0.81 (95% CI 0.41-1.59) and bilateral salpingectomy resulted in an age-adjusted HR of 0.43 (95% CI 0.06-3.16) based on one case. None of our subgroup analysis for lag-time resulted in a significant difference in ovarian cancer incidence between intervention and control group. The difference in ovarian cancer incidence appeared largest in women with at least 8 years of follow-up (P = 0.08). LIMITATIONS, REASONS FOR CAUTION: Due to the young population, ovarian cancer incidence is low, even at the end of follow-up. Furthermore, due to the anonymous nature of the pathology registry, we were unable to adjust for confounding factors. WIDER IMPLICATIONS OF THE FINDINGS: Although results did not reach statistical significance, they add to the available data on ovarian cancer incidence after salpingectomy. Our subgroup analysis suggests there may be no benefit in the first years following salpingectomy. STUDY FUNDING/COMPETING INTEREST(S): None. TRIAL REGISTRATION NUMBER: N/A.

Detection of Schistosoma DNA in genital specimens and urine: A comparison between five female African study populations originating from S. haematobium and/or S. mansoni endemic areas.

Female Genital Schistosomiasis (FGS) is a neglected disease affecting millions, however challenging to diagnose. This explorative descriptive study compares Schistosoma real-time PCR analysis of cervico-vaginal lavages (CVL) with corresponding urine and stool samples of 933 women from five different previously described study populations. Sampling included 310 women from an S. mansoni endemic region in Mwanza, Tanzania and 112 women from a nearby S. haematobium endemic region. Findings were compared with samples collected from S. haematobium endemic regions in South Africa from 394 women and from 117 women from Madagascar of which 79 were urine pre-selected microscopy positive cases from highly-endemic communities and 38 were urine microscopy negatives from a low-endemic community. As anticipated, urine and stool microscopy and gynecological investigations varied substantially between study populations; however, the same Schistosoma real-time PCR was performed in one reference laboratory. Schistosoma DNA was detected in 13% (120/933) of the CVL, ranging from 3% in the S. mansoni Tanzanian endemic region to 61% in the pre-selected Malagasy urine microscopy positive cases. Detectable Schistosoma DNA in CVL was associated with Schistosoma DNA in urine but not with microscopic detection of eggs in urine or by cytological examination. This study confirmed real-time PCR for the detection of Schistosoma DNA in gynecological samples to be a valuable diagnostic tool to study the distribution of FGS within schistosomiasis endemic areas.

Placenta derived factors involved in the pathogenesis of the liver in the syndrome of haemolysis, elevated liver enzymes and low platelets (HELLP): A review.

AIM: With this review we try to unravel if placenta-derived factors are able to initiate liver sinusoidal endothelial cells (LSEC) decay in HELLP syndrome and eventually cause the development of sinusoidal obstruction syndrome (SOS). BACKGROUND: Haemolysis, Elevated Liver enzymes and Low Platelets (HELLP) syndrome is a severe complication of pregnancy. It is characterized by elevated liver enzymes, low platelet count and haemolytic anaemia. The risk of developing HELLP syndrome within a pregnancy is 0.1-0.8%. The mortality rate among women with HELLP syndrome is 0-24% and the perinatal death goes up to 37%. The aetiology of HELLP syndrome is not fully understood but the pathogenesis of the liver pathology in the HELLP syndrome resembles that of a SOS with endothelial damage of the LSECs which ultimately leads to liver failure. OBJECTIVES: We hypothesize that placenta derived factors cause LSEC damage and thereby liver dysfunction. METHODS: We searched in the PubMed database for relevant articles about placenta derived factors involved in endothelial activation especially in the liver. We yielded eventually 55 relevant articles. RESULTS: Based on this literature search we associate that in HELLP syndrome there is an increase of soluble fms-like tyrosine kinase (sFlt1), vascular endothelial growth factor (VEGFR), soluble endoglin (sEng), galectin-1 (Gal-1), endothelin-1 (ET-1), Angiopoietin 2 (Angs-2), Asymmetric dimethylarginine (ADMA), activin B, inhibin A, Fas ligand (FasL) and heat shock protein 70 (Hsp70). CONCLUSION: We assume that these eleven increased placenta derived factors are responsible for LSEC damage which eventually leads to liver failure. This concept shows a possible design of the complicated pathophysiology in HELLP syndrome. However further research is required.

How can schistosome circulating antigen assays be best applied for diagnosing male genital schistosomiasis (MGS): an appraisal using exemplar MGS cases from a longitudinal cohort study among fishermen on the south shoreline of Lake Malawi.

We provide an update on diagnostic methods for the detection of urogenital schistosomiasis (UGS) in men and highlight that satisfactory urine-antigen diagnostics for UGS lag much behind that for intestinal schistosomiasis, where application of a urine-based point-of-care strip assay, the circulating cathodic antigen (CCA) test, is now advocated. Making specific reference to male genital schistosomiasis (MGS), we place greater emphasis on parasitological detection methods and clinical assessment of internal genitalia with ultrasonography. Unlike the advances made in defining a clinical standard protocol for female genital schistosomiasis, MGS remains inadequately defined. Whilst urine filtration with microscopic examination for ova of Schistosoma haematobium is a convenient but error-prone proxy of MGS, we describe a novel low-cost sampling and direct visualization method for the enumeration of ova in semen. Using exemplar clinical cases of MGS from our longitudinal cohort study among fishermen along the shoreline of Lake Malawi, the portfolio of diagnostic needs is appraised including: the use of symptomatology questionnaires, urine analysis (egg count and CCA measurement), semen analysis (egg count, circulating anodic antigen measurement and real-time polymerase chain reaction analysis) alongside clinical assessment with portable ultrasonography.

Repeated doses of Praziquantel in Schistosomiasis Treatment (RePST) - single versus multiple praziquantel treatments in school-aged children in Côte d'Ivoire: a study protocol for an open-label, randomised controlled trial.

BACKGROUND: Large scale administration of the anthelminthic drug praziquantel (PZQ) to at-risk populations is the cornerstone of schistosomiasis control, although persisting high prevalence of infections in some areas and growing concerns of PZQ resistance have revealed the limitations of this strategy. Most studies assessing PZQ efficacy have used relatively insensitive parasitological diagnostics, such as the Kato-Katz (KK) and urine-filtration methods, thereby overestimating cure rates (CRs). This study aims to determine the efficacy of repeated PZQ treatments against Schistosoma mansoni infection in school-aged children in Côte d'Ivoire using the traditional KK technique, as well as more sensitive antigen- and DNA-detection methods. METHODS: An open-label, randomised controlled trial will be conducted in school-aged children (5 to 18 years) from the region of Taabo, Côte d'Ivoire, an area endemic for S. mansoni. This 8-week trial includes four two-weekly standard doses of PZQ in the "intense treatment" intervention group and one standard dose of PZQ in the "standard treatment" control group. The efficacy of PZQ will be evaluated in stool samples using the KK technique and real-time PCR as well as in urine using the point-of-care circulating cathodic antigen test and the up-converting phosphor, lateral flow, circulating anodic antigen assay. The primary outcome of the study will be the difference in CR of intense versus standard treatment with PZQ on individuals with a confirmed S. mansoni infection measured by KK. Secondary outcomes include the difference in CR and intensity reduction rate between the intense and standard treatment groups as measured by the other diagnostic tests, as well as the accuracy of the different diagnostic tests, and the safety of PZQ. DISCUSSION: This study will provide data on the efficacy of repeated PZQ treatment on the clearance of S. mansoni as measured by several diagnostic techniques. These findings will inform future mass drug administration policy and shed light on position of novel diagnostic tools to evaluate schistosomiasis control strategies. TRIAL REGISTRATION: The study is registered at EudraCT (2016-003017-10, date of registration: 22 July 2016) and ( NCT02868385 , date of registration: 16 August 2016).

Multiplex real-time PCR for diagnosing malaria in a non-endemic setting: a prospective comparison to conventional methods.

Almost a decade ago our diagnostic laboratory implemented an in-house real-time PCR for the detection of Plasmodium DNA to diagnose malaria in parallel with conventional diagnostics, i.e., microscopy (thick and thin smears), quantitative buffy coat microscopy (QBC), and a rapid diagnostic test (RDT). Here we report our experiences and make a comparison between the different diagnostic procedures used in this non-endemic setting. All patients during the period February 2009-December 2017 suspected of malaria were prospectively tested at the moment of sample collection. Both PCR and conventional malaria diagnostics were carried out on a total of 839 specimens from 825 patients. In addition, three Plasmodium falciparum (Pf) patients were closely followed by real-time PCR and microscopy after treatment. Overall, 56 samples (55 patients) tested positive by real-time PCR, of which six were missed by microscopy and seven by QBC. RDT showed fairly good results in detecting Pf, whereas specificity was not optimal. RDT failed to detect 10 of 17 non-Pf PCR positive specimens. One Plasmodium malariae patient would have been missed if only conventional diagnostic tests had been used. The high sensitivity of the PCR was confirmed by the number of PCR positive, microscopy negative post-treatment samples. In conclusion, within our routine diagnostic setting, malaria real-time PCR not only showed a high level of agreement with the conventional methods used, but also showed higher sensitivity and better specificity. Still, for complete replacement of the conventional procedures in a non-endemic setting, the time-to-results of the real-time PCR is currently too long.

A case of delayed diagnosis of East-African trypanosomiasis in a Dutch traveller.

We present a case of East-African trypanosomiasis (EAT) in a 56-year-old Dutch woman returning from holiday in Tanzania and Kenya. The diagnosis was delayed due to the lack of suspicion and secondly because of postponed analysis of blood microscopy after negative rapid malaria antigen testing. Second stage trypanosomiasis was ruled out with liquor analysis. She was treated first with pentamidine and shortly thereafter with suramin, after which she recovered. We emphasize the use of thin/thick smear diagnostics in travellers returning from endemic countries.

Opportunistic salpingectomy in women undergoing hysterectomy: Results from the HYSTUB randomised controlled trial.

OBJECTIVE: To evaluate whether opportunistic salpingectomy in premenopausal women undergoing hysterectomy for benign indications is both hormonally and surgically safe, compared with hysterectomy without salpingectomy. STUDY DESIGN: In this multicentre randomised controlled trial, women were randomised to undergo either hysterectomy with opportunistic bilateral salpingectomy (intervention group) or standard hysterectomy with preservation of the Fallopian tubes (control group). MAIN OUTCOME MEASURES: The primary outcome was the difference in serum anti-Müllerian hormone concentration (ΔAMH), measured pre-surgery and 6 months post-surgery. Secondary outcomes were surgical outcomes and duration of hospital stay. The sample size was powered at 50 participants per group (n=100) to compare ΔAMH after hysterectomy with salpingectomy to ΔAMH after standard hysterectomy. RESULTS: Between March 2013 and December 2016, 104 women, aged 30-55 years, were randomly allocated to hysterectomy with opportunistic bilateral salpingectomy (n=52) or standard hysterectomy (n=52). The baseline characteristics did not differ between the two groups. The median ΔAMH was -0.14pmol/L (IQR -1.47-0.95) in the intervention group and 0.00pmol/L (IQR -1.05-0.80) in the control group (p=0.49). The addition of salpingectomy did not impair surgical results and it did not affect duration of hospital stay. CONCLUSION: Addition of opportunistic bilateral salpingectomy during hysterectomy did not result in a larger effect on ovarian reserve when compared with hysterectomy alone, neither did it affect surgical outcomes. Therefore, opportunistic salpingectomy seems to be a safe procedure in premenopausal women undergoing hysterectomy for benign gynaecological conditions.

Effect of anthelmintic treatment on leptin, adiponectin and leptin to adiponectin ratio: a randomized-controlled trial.

Emerging evidence suggests that helminths might confer protection against the development of type 2 diabetes. We aimed to assess the role of adipokines in mediating the effect of helminths on insulin resistance. Serum samples were obtained from a randomized-controlled trial of anthelmintic treatment in an area endemic for soil-transmitted helminths (STH), Flores Island, Indonesia. In STH-infected subjects, anthelmintic treatment significantly increased the ratio of leptin to adiponectin (treatment effect factor (95% confidence interval (CI)), P-value for interaction: 1.20 (1.06-1.35), P=0.010), which largely stemmed from a significant reduction in adiponectin (0.91 (0.85-0.98), P=0.020) and a trend for an increase in leptin level (1.10 (1.00-1.21), P=0.119). No significant effect on resistin level was observed. This increase in leptin to adiponectin ratio seemed to contribute to the observed effect of deworming on increased insulin resistance (IR) as adjustment for leptin to adiponectin ratio attenuated the effect on IR from 1.07 (1.01-1.14, P=0.023) to 1.05 (0.99-1.11, P=0.075). Anthelmintic treatment in STH-infected subjects increases leptin to adiponectin ratio which may in small part contribute to the modest increase in IR. Further studies will be needed to assess the effect of the changes in adipokine levels on the host immune response and metabolism.

The effect of three-monthly albendazole treatment on Th2 responses: Differential effects on IgE and IL-5.

Helminth parasites induce a strong Th2 response, characterized by high levels of IgE and elevated signature cytokines such as IL-5. As many global deworming programmes are underway, there is concern that this might lead to emergence of Th1-mediated pathologies when the counterbalancing helminth-induced Th2 response is absent. Therefore, we assessed the effect of deworming on Th2-mediated responses in a household-clustered randomized controlled trial in Indonesia. Total plasma IgE and whole-blood IL-5 responses to mitogen phytohaemagglutinin (PHA) were measured in 1494 and 682 subjects, respectively, at baseline, 9 and 21 months after three-monthly single-dose treatment with albendazole or placebo. Anthelmintic treatment did not result in complete removal of helminth infections in the community. However, treatment significantly decreased IgE levels in albendazole- compared to placebo-treated subjects. IL-5 responses to PHA were not significantly affected by anthelmintic treatment and tended to increase in albendazole-treated subjects, indicating that intensive treatment of helminth parasites has different outcomes on B-cell (IgE levels) and T-cell (IL-5) responses. The data shows that 2 years of deworming can have differential effects on responses typified as Th2-mediated, which needs to be taken into account when examining the impact of helminths on noncommunicable diseases.

New diagnostic tools in schistosomiasis.

Schistosomiasis is a water-based parasitic disease that affects over 250 million people. Control efforts have long been in vain, which is one reason why schistosomiasis is considered a neglected tropical disease. However, since the new millennium, interventions against schistosomiasis are escalating. The initial impetus stems from a 2001 World Health Assembly resolution, urging member states to scale-up deworming of school-aged children with the anthelminthic drug praziquantel. Because praziquantel is safe, efficacious and inexpensive when delivered through the school platform, diagnosis before drug intervention was deemed unnecessary and not cost-effective. Hence, there was little interest in research and development of novel diagnostic tools. With the recent publication of the World Health Organization (WHO) Roadmap to overcome the impact of neglected tropical diseases in 2020, we have entered a new era. Elimination of schistosomiasis has become the buzzword and this has important ramifications for diagnostic tools. Indeed, measuring progress towards the WHO Roadmap and whether local elimination has been achieved requires highly accurate diagnostic assays. Here, we introduce target product profiles for diagnostic tools that are required for different stages of a schistosomiasis control programme. We provide an update of the latest developments in schistosomiasis diagnosis, including microscopic techniques, rapid diagnostic tests for antigen detection, polymerase chain reaction (PCR) assays and proxy markers for morbidity assessments. Particular emphasis is placed on challenges and solutions for new technologies to enter clinical practice.

Clinical consequences of new diagnostic tools for intestinal parasites.

Following the success of nucleic acid-based detection in virology and bacteriology, multiplex real-time PCRs are increasingly used as first-line diagnostics in clinical parasitology, replacing microscopy. The detection and quantification of parasite-specific DNA in faeces is highly sensitive and specific and allows for cost-effective high-throughput screening. In this paper we discuss the clinical consequences of this radical change in diagnostic approach, as well as its potential drawbacks. In the Netherlands, routine diagnostic laboratories have been pioneering the implementation of multiplex real-time PCR for the detection of pathogenic intestinal protozoa and this has resulted in increased detection rates of Giardia lamblia and Cryptosporidium spp. As a consequence of this new diagnostic approach, expertise in the field of parasite morphology by conventional light microscopy seems to be disappearing in most of the high-throughput microbiological laboratories. As a result, to maintain a high standard of care, a formalized exchange of critical information between clinicians and laboratory staff is necessary to determine the most appropriate testing either in local laboratories or in reference centres, based on clinical signs and symptoms, exposure and immune status. If such a diagnostic algorithm is lacking, important infections in travellers, immigrants and immunocompromised patients may be missed.

Detection of Clonorchis sinensis in stool samples using real-time PCR.

Human clonorchiasis, caused by infection with the trematode Clonorchis sinensis, is a common health problem in East Asia. In an attempt to develop a new, sensitive method for the diagnosis of the disease, the use of a real-time PCR (targeting the internal-transcribed-spacer-2 sequence of the parasite) to detect C. sinensis-specific DNA in faecal samples has recently been evaluated. The PCR-based assay, which included an internal control to detect any inhibition of the amplification by faecal constituents in the sample, was performed on stool samples and on DNA controls representing a wide range of intestinal microorganisms. The assay appeared very specific, only showing positivity with C. sinensis and Opisthorchis felineus. The sensitivity of the assay was explored by testing 170 preselected samples of human faeces, from an endemic area of South Korea, which had known (microscopically-determined) densities of C. sinensis eggs. The sensitivity of the assay was 100% for the 74 samples that each had > 100 eggs/g and 91.4% for the other 70 samples found egg-positive by microcopy (i.e. those that had

Molecular diagnostics of intestinal parasites in returning travellers.

A new diagnostic strategy was assessed for the routine diagnosis of intestinal parasites in returning travellers and immigrants. Over a period of 13 months, unpreserved stool samples, patient characteristics and clinical data were collected from those attending a travel clinic. Stool samples were analysed on a daily basis by microscopic examination and antigen detection (i.e. care as usual), and compared with a weekly performed multiplex real-time polymerase chain reaction (PCR) analysis on Entamoeba histolytica, Giardia lamblia, Cryptosporidium and Strongyloides stercoralis. Microscopy and antigen assays of 2,591 stool samples showed E. histolytica, G. lamblia, Cryptosporidium and S. stercoralis in 0.3, 4.7, 0.5 and 0.1% of the cases, respectively. These detection rates were increased using real-time PCR to 0.5, 6.0, 1.3 and 0.8%, respectively. The prevalence of ten additional pathogenic parasite species identified with microscopy was, at most, 0.5%. A pre-selective decision tree based on travel history or gastro-intestinal complaints could not be made. With increased detection rates at a lower workload and the potential to extend with additional parasite targets combined with fully automated DNA isolation, molecular high-throughput screening could eventually replace microscopy to a large extent.

Application of a circulating-cathodic-antigen (CCA) strip test and real-time PCR, in comparison with microscopy, for the detection of Schistosoma haematobium in urine samples from Ghana.

In the detection of parasitic infection, the traditional methods based on microscopy often have low sensitivity and/or specificity compared with the newer, molecular tests. An assay based on real-time PCR and a reagent strip test for detecting circulating cathodic antigen (CCA) have both now been compared with urine filtration and microscopy, in the detection of Schistosoma haematobium infections. Urine samples, obtained from 74 'cases' in areas of Ghana with endemic S. haematobium and 79 'controls' from non-endemic areas, were each checked using the three methods. With the results of the filtration and microscopy taken as the 'gold standard', real-time PCR was found to be 100% specific and 89% sensitive whereas the CCA strips were 91% specific and 41% sensitive. With the samples found to contain > or =50 eggs/10 ml (indicating relatively intense infections), the sensitivities of the PCR and CCA were higher, at 100% and 62%, respectively. As expected, egg counts were negatively correlated with the number of amplification cycles needed, in the PCR, to give a signal that exceeded the background (r=-0.38; P<0.01). Although the real-time PCR and CCA strip tests are very different, both show promise in the detection of S. haematobium infections. The PCR has optimal specificity and high sensitivity but the specificity of the CCA strips and the sensitivity of both tools could still be improved. A more thorough re-evaluation of the sensitivity and specificity of microscopy and these newer diagnostic methods, with an estimation of the cost-effectiveness of each technique, is recommended.

Cough and alterations in semen after a tropical swim.

This case report describes a patient with cough and haematospermia shortly after visiting a Schistosoma endemic area. Numerous S. haematobium eggs were found in the ejaculate, while no eggs were seen in the urine.

Detection of diarrhoea-causing protozoa in general practice patients in The Netherlands by multiplex real-time PCR.

The diagnostic value of a multiplex real-time PCR for the detection of Entamoeba histolytica, Giardia lamblia and Cryptosporidium parvum/Cryptosporidium hominis was evaluated by comparing the PCR results obtained with those of routinely performed microscopy of faecal samples from patients consulting their general practitioner (GP) because of gastrointestinal complaints. Analysis of 722 faecal DNA samples revealed that the prevalence of G. lamblia was 9.3% according to PCR, as compared to 5.7% by microscopy. The number of infections detected was more than double in children of school age. Furthermore, G. lamblia infection was detected in 15 (6.6%) of 228 faecal samples submitted to the laboratory for bacterial culture only. C. parvum/C. hominis infections were not diagnosed by routine procedures, but DNA from these organisms was detected in 4.3% of 950 DNA samples. A strong association with age was noted, with Cryptosporidium being detected in 21.8% of 110 children aged <5 years. C. hominis was the most prevalent species. E. histolytica was not detected in this study population. Analysis of microscopy data revealed that the number of additional parasites missed by PCR was small. Overall, the study demonstrated that a multiplex real-time PCR approach is a feasible diagnostic alternative in the clinical laboratory for the detection of parasitic infections in patients consulting GPs because of gastrointestinal symptoms.