SUMMARY Several enteric protozoa cause severe morbidity and mortality in both humans and animals worldwide. In developed settings, enteric protozoa are often ignored as a cause of diarrheal illness due to better hygiene conditions, and as such, very little effort is used toward laboratory diagnosis. Although these protozoa contribute to the high burden of infectious diseases, estimates of their true prevalence are sometimes affected by the lack of sensitive diagnostic techniques to detect them in clinical and environmental specimens. Despite recent advances in the epidemiology, molecular biology, and treatment of protozoan illnesses, gaps in knowledge still exist, requiring further research. There is evidence that climate-related changes will contribute to their burden due to displacement of ecosystems and human and animal populations, increases in atmospheric temperature, flooding and other environmental conditions suitable for transmission, and the need for the reuse of alternative water sources to meet growing population needs. This review discusses the common enteric protozoa from a public health perspective, highlighting their epidemiology, modes of transmission, prevention, and control. It also discusses the potential impact of climate changes on their epidemiology and the issues surrounding waterborne transmission and suggests a multidisciplinary approach to their prevention and control.

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Dientamoeba fragilis is a protozoan that inhabits the human gut. It is approximately 100 years since Dientamoeba's discovery and first description when it was described as a rare and harmless commensal. Since then it has struggled to gain recognition as a pathogen despite the evidence supporting its pathogenicity. Dientamoeba remains neglected, probably due to the misconceptions that it is uncommon and non-pathogenic. Usually, carriage of Dientamoeba is associated with symptoms such as abdominal pain and diarrhea. Moreover, antimicrobial therapy followed by resolution of symptoms coincides with the eradication of Dientamoeba. This manuscript reviews the scientific literature relating to Dientamoeba's prevalence and pathogenicity. While much of the evidence supporting its pathogenicity is only circumstantial, it is apparent that most researchers agree that Dientamoeba is pathogenic. Therefore, in symptomatic patients who harbor Dientamoeba and no other pathogen, Dientamoeba should be considered as the etiological agent and treated as such.

The aim of this study was to describe the first development and evaluation of a multiplex tandem PCR (MT-PCR) assay for the detection and identification of 4 common pathogenic protozoan parasites, Cryptosporidium spp., Dientamoeba fragilis, Entamoeba histolytica, and Giardia intestinalis, from human clinical samples. A total of 472 fecal samples submitted to the Department of Microbiology at St. Vincent's Hospital were included in the study. The MT-PCR assay was compared to four real-time PCR (RT-PCR) assays and microscopy by a traditional modified iron hematoxylin stain. The MT-PCR detected 28 G. intestinalis, 26 D. fragilis, 11 E. histolytica, and 9 Cryptosporidium sp. isolates. Detection and identification of the fecal protozoa by MT-PCR demonstrated 100% correlation with the RT-PCR results, and compared to RT-PCR, MT-PCR exhibited 100% sensitivity and specificity, while traditional microscopy of stained fixed fecal smears exhibited sensitivities and specificities of 56% and 100% for Cryptosporidium spp., 38% and 99% for D. fragilis, 47% and 97% for E. histolytica, and 50% and 100% for G. intestinalis. No cross-reactivity was detected in 100 stool samples containing various other bacterial, viral, and protozoan species. The MT-PCR assay was able to provide rapid, sensitive, and specific simultaneous detection and identification of the four most important diarrhea-causing protozoan parasites that infect humans. This study also highlights the lack of sensitivity demonstrated by microscopy, and thus, molecular methods such as MT-PCR must be considered the diagnostic methods of choice for enteric protozoan parasites.Enteric protozoa continue to be the most commonly encountered parasitic diseases and to cause significant morbidity and mortality throughout both developed and developing regions of the world, affecting millions of people each year (22). While Blastocystis hominis is the most common protozoan parasite detected in stool samples, the major etiological agents of parasitic diarrhea are considered to be Cryptosporidium spp., Dientamoeba fragilis, Entamoeba histolytica, and Giardia intestinalis (22).E. histolytica is a pathogenic amoeboid protozoan parasite for which humans are the primary reservoir (27). It is a potentially invasive pathogen and the causative agent of amebiasis, with approximately 50 million cases and 100,000 deaths annually (28). The clinical presentation can range from asymptomatic carriage to gastrointestinal disease and invasive disease. E. histolytica is morphologically identical to the nonpathogenic species Entamoeba dispar and Entamoeba moshkovskii, though genetic differences have confirmed their separation into independent species (4, 20). Due to this conserved morphology, stained smears of stool specimens are insufficient for differentiation of the species. Currently, the only way to differentiate the pathogenic E. histolytica from the nonpathogenic strains via microscopy is to visualize ingested red blood cells within the E. histolytica trophozoite; however, this is rarely seen i...

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A prospective study was conducted over a 30-month period, in which fecal specimens from 6,750 patients were submitted to the Department of Microbiology at St. Vincent's Hospital, Sydney, Australia. Trophozoites of Dientamoeba fragilis were detected in 60 (0.9%) patients by permanent staining, and confirmation was performed by PCR. Gastrointestinal symptoms were present in all patients, with diarrhea and abdominal pain the most common symptoms. Thirty-two percent of patients presented with chronic symptoms. The average age of infected patients was 39.8 years. No correlation was found between D. fragilis and Enterobius vermicularis, a proposed vector of transmission for D. fragilis. The genetic diversity of 50 D. fragilis isolates was examined by PCR, and the PCR products were analyzed for the presence of restriction fragment length polymorphisms. These results showed no variation in the small-subunit rRNA gene and demonstrated a single genotype for all Australian isolates. This study shows the potential pathogenic properties of D. fragilis and the need for all laboratories to routinely test for this organism.Dientamoeba fragilis is a trichomonad parasite found in the gastrointestinal tract of humans and implicated as a cause of gastrointestinal disease. Dientamoeba fragilis has been found in most parts of the world in both rural and cosmopolitan areas (10). The prevalence of this organism in Australia varies greatly, from 0.4% to 16.8%, in patients presenting with gastrointestinal complaints (1,22).No cyst stage has been observed, and only the trophozoites are detected in stool samples. Definitive diagnosis is based on prompt fixation and permanent staining, as the trophozoites degenerate rapidly, within hours of been passed, and demonstration of their characteristic nuclear structure cannot be achieved in unstained preparations (24). Daily shedding of D. fragilis trophozoites has been shown to be highly variable, with intermittent shedding occurring regularly, necessitating multiple sampling for maximum chances of detection (20).Molecular techniques for the diagnosis of D. fragilis show much promise, with PCR demonstrating excellent sensitivity and specificity (18). Such techniques have been used successfully for the diagnosis of other pathogenic protozoa (11,19).Molecular genotyping and sequence analysis have demonstrated that D. fragilis exists as two genetically distinct forms (9,15,18,26). Stark et al. (18) sequenced the SSU rRNA gene of seven Australian D. fragilis isolates, and the data generated from the seven showed no variation among them. These observations support the notion that D. fragilis is a clonal species. The sequences from the Australian isolates, however, differed from the sequence of the D. fragilis strain Bi/PA (ATCC 30948; GenBank accession no. U37461) and were found to be similar to those found in a recent study in the Netherlands (15). The true incidence of the wild-type and variant forms in Australia needs to be established and to determine if such variation has any influence on the pa...

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A prospective, comparative study of the prevalence of enteric protozoa was determined among human immunodeficiency virus (HIV)- positive and HIV-negative men who have sex with men (MSM) in Sydney, Australia. A total of 1,868 patients submitted stool specimens; 1,246 were from MSM (628 HIV positive and 618 HIV positive) and 622 from non-MSM were examined over a 36-month period. A total of 651 (52.2%) stool specimens from MSM were positive for protozoa compared with 85 (13%) from non-MSM. There was a significant difference in the prevalence of Blastocystis hominis, Endolimax nana, Entamoeba histolytica/dispar complex, Entamoeba hartmanni, Iodamoeba butschlii, and Enteromonas hominis detected between MSM and non-MSM (P<0.001). The only notable difference between HIV-negative and HIV-positive MSM was that HIV-infected MSM were found to more likely have a Cryptosporidium parvum infection. Entamoeba histolytica was found in 3 patients, E. dispar in 25, and E. moshkovskii in 17, all of whom were MSM. When compared with a control group, MSM were significantly more likely to harbor intestinal protozoa and have multiple parasites present. The results of this study show high rates of enteric parasites persist in MSM and highlight the importance of testing for intestinal parasites in MSM. This is the first report of E. moshkovskii from MSM.