SUMMARY The genus Entamoeba contains many species, six of which (Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, Entamoeba polecki, Entamoeba coli, and Entamoeba hartmanni) reside in the human intestinal lumen. Entamoeba histolytica is the causative agent of amebiasis and is considered a leading parasitic cause of death worldwide in humans. Although recent studies highlight the recovery of E. dispar and E. moshkovskii from patients with gastrointestinal symptoms, there is still no convincing evidence of a causal link between the presence of these two species and the symptoms of the host. New approaches to the identification of E. histolytica are based on detection of E. histolytica-specific antigen and DNA in stool and other clinical samples. Several molecular diagnostic tests, including conventional and real-time PCR, have been developed for the detection and differentiation of E. histolytica, E. dispar, and E. moshkovskii in clinical samples. The purpose of this review is to discuss different methods that exist for the identification of E. histolytica, E. dispar, and E. moshkovskii which are available to the clinical diagnostic laboratory. To address the need for a specific diagnostic test for amebiasis, a substantial amount of work has been carried out over the last decade in different parts of the world. The molecular diagnostic tests are increasingly being used for both clinical and research purposes. In order to minimize undue treatment of individuals infected with other species of Entamoeba such as E. dispar and E. moshkovskii, efforts have been made for specific diagnosis of E. histolytica infection and not to treat based simply on the microscopic examination of Entamoeba species in the stool. The incorporation of many new technologies into the diagnostic laboratory will lead to a better understanding of the public health problem and measures to control the disease.

Aspergillus section Circumdati or the Aspergillus ochraceus group, includes species with rough walled stipes, biseriate conidial heads, yellow to ochre conidia and sclerotia that do not turn black. Several species are able to produce mycotoxins including ochratoxins, penicillic acids, and xanthomegnins. Some species also produce drug lead candidates such as the notoamides. A polyphasic approach was applied using morphological characters, extrolite data and partial calmodulin, β-tubulin and ITS sequences to examine the evolutionary relationships within this section. Based on this approach the section Circumdati is revised and 27 species are accepted, introducing seven new species: A. occultus, A. pallidofulvus, A. pulvericola, A. salwaensis, A. sesamicola, A. subramanianii and A. westlandensis. In addition we correctly apply the name A. fresenii (≡ A. sulphureus (nom. illeg.)). A guide for the identification of these 27 species is provided. These new species can be distinguished from others based on morphological characters, sequence data and extrolite profiles. The previously described A. onikii and A. petrakii were found to be conspecific with A. ochraceus, whilst A. flocculosus is tentatively synonymised with A. ochraceopetaliformis, despite extrolite differences between the two species. Based on the extrolite data, 13 species of section Circumdati produce large amounts of ochratoxin A: A. affinis, A. cretensis, A. fresenii, A. muricatus, A. occultus, A. ochraceopetaliformis (A. flocculosus), A. ochraceus, A. pseudoelegans, A. pulvericola, A. roseoglobulosus, A. sclerotiorum, A. steynii and A. westerdijkiae. Seven additional species produce ochratoxin A inconsistently and/or in trace amounts: A. melleus, A. ostianus, A. persii, A. salwaensis, A. sesamicola, A. subramanianii and A. westlandensis. The most important species regarding potential ochratoxin A contamination in agricultural products are A. ochraceus, A. steynii and A. westerdijkiae.

This study investigated the presence of Entamoeba histolytica, Entamoeba dispar, and Entamoeba moshkovskii in stool samples from a patient population in Sydney, Australia. Stool samples were tested by microscopy and PCR. Five patients were found with E. histolytica infections, while E. dispar and E. moshkovskii were observed in 63 (70.8%) and 55 (61.8%) patients, respectively, by PCR. This is the first study in Australia using molecular techniques to determine the presence of E. histolytica, E. dispar, and E. moshkovskii.The genus Entamoeba comprises six species (Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, Entamoeba poleki, Entamoeba coli, and Entamoeba hartmanni) that live in the human intestinal lumen. E. histolytica, E. dispar, and E. moshkovskii are morphologically identical but are different biochemically and genetically (1,2,3,5,6). Although E. histolytica is recognized as a pathogen, the ability of the other two species to cause disease is unclear. E. moshkovskii, for example, is considered primarily a free-living ubiquitous amoeba found in anoxic sediments (2), and E. dispar is considered primarily a commensal of the human gut (3,5).Early studies of amebiasis in Australia have reported that the incidence of Entamoeba species varies from 1 to 4% in urban and rural communities, respectively (14). In another study, Law et al. (8) reported a 37% prevalence of Entamoeba in men who have sex with men. However, these studies did not differentiate E. histolytica from E. dispar or E. moshkovskii. The prevalence of E. histolytica, E. dispar, and E. moshkovskii (hereafter called the E complex) in the Australian population therefore remains unknown. The present study investigated the presence of the E complex in clinical samples by microscopy and PCR directly in stool samples collected from patients presenting with gastrointestinal symptoms.All of the stool specimens (from a diverse patient population) submitted to the Department of Microbiology at St. Vincent's Hospital, Sydney, Australia, during January 2003 to June 2006 for investigation of diarrhea were included in this study. Specimens from outpatients were collected by the patient and submitted to the laboratory as fresh specimens along with a portion mixed with sodium acetate-acetic acid-formalin (SAF). Specimens from inpatients or received without a portion fixed in SAF were immediately preserved in SAF upon arrival at the laboratory. The SAF-fixed specimens underwent permanent staining with a modified iron-hematoxylin stain according to the manufacturer's recommendations (Fronine, Australia). Patients diagnosed with members of the E complex by microscopy of stained smears underwent further investigations.DNA was extracted from fecal specimens (without fixatives) from patients either fresh or after storage at Ϫ20°C (immediately frozen upon arrival at the laboratory). Briefly, the stool sample (200 mg) was washed twice with 1 ml of sterile phosphate-buffered saline (pH 7.2), centrifuged for 5 min at 14,000 ϫ g, and DNA extracted according t...

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.

Entamoeba moshkovskii is considered to be a free-living ameba, which is morphologically similar, but biochemically and genetically different, to Entamoeba histolytica and Entamoeba dispar. However, recent studies have suggested that E. moshkovskii may be a "potential" pathogen, with infections giving rise to diarrhea and other intestinal disorders. Microscopy and polymerase chain reaction (PCR) targeting the 18S ribosomal (r) DNA was performed on fecal samples collected from patients presenting with diarrhea and other gastrointestinal disorders (test group), as well as fecal samples collected from healthy controls (control group). Of the 110 patients microscopically positive for the Entamoeba species, 55/110 (50%) samples were positive for E. moshkovskii in the test group of patients presenting with diarrhea. E. moshkovskii was the only pathogen detected (including bacteria or viruses) in 3/55 (5.5%) of the test group of patients presenting with diarrhea and abdominal pain. The DNA of E. moshkovskii was not detected in the fecal samples collected from the healthy controls. These results suggest that E. moshkovskii may not simply be a commensal of the human gastrointestinal tract and provides evidence for E. moshkovskii as a "potential" pathogen in the case of diarrhea and other gastrointestinal disorders. Further studies are needed to determine the role of E. moshkovskii in causing diarrheal diseases in our population.

The present study was conducted to compare two stool antigen detection kits with PCR for the diagnosis of Entamoeba histolytica infections by using fecal specimens submitted to the Department of Microbiology at St. Vincent's Hospital, Sydney, and the Institute of Medical and Veterinary Science, Adelaide, Australia. A total of 279 stool samples containing the E complex (E. histolytica, Entamoeba dispar, and Entamoeba moshkovskii) were included in this study. The stool specimens were tested by using two commercially produced enzyme immunoassays (the Entamoeba CELISA PATH and TechLab E. histolytica II kits) to detect antigens of E. histolytica. DNA was extracted from all of the samples with a Qiagen DNA stool mini kit (Qiagen, Hilden, Germany), and a PCR targeting the small-subunit ribosomal DNA was performed on all of the samples. When PCR was used as a reference standard, the CELISA PATH kit showed 28% sensitivity and 100% specificity. The TechLab ELISA (enzyme-linked immunosorbent assay) kit did not prove to be useful in detecting E. histolytica, as it failed to identify any of the E. histolytica samples which were positive by PCR. With the TechLab kit, cross-reactivity was observed for three specimens, one of which was positive for both E. dispar and E. moshkovskii while the other two samples contained E. moshkovskii. Quantitative assessment of the PCR and ELISA results obtained showed that the ELISA kits were 1,000 to 10,000 times less sensitive, and our results show that the CELISA PATH kit and the TechLab ELISA are not useful for the detection of E. histolytica in stool samples from patients in geographical regions where this parasite is not endemic.

Two yeast strains (INY29 and INY13) representing a novel yeast species were isolated from the hypersaline marine environment of the Inland Sea, Qatar. Phylogenetic analysis based on the D1/D2 domains of the large subunit (LSU) regions and internal transcribed spacer (ITS1 and ITS2) regions showed that the two strains represent a single species in the genus Naganishia that is distinct from other species. These two strains were classified as members of the genus Naganishia and clustered in a strongly supported clade represented by Naganishia albidus in the Filobasidiales order in the phylogenetic tree drawn from ITS and D1/D2 sequences. The novel species was most closely related to the type strain of Naganishia cerealis but the two species differed by 1 % sequence divergence (four substitutions and one gap) in the D1/D2 domains and (five substitutions and one gap) in the ITS regions. In contrast to the closest relative, N. cerealis, the novel yeast species assimilated melibiose, glycerol, meso-erythritol, dl-lactate, methanol, propane 1-2-diol, butane 2-3-diol, and grew at 35 °C. The name Naganishia qatarensis sp. nov. is proposed to accommodate these strains, with INY29 as the holotype.

The morphology, ontogenesis, and phylogenetic relationships of a halophile euplotid ciliates, Euplotes qatarensis nov. spec., isolated from the Khor Al-Adaid Lagoon in Qatar were investigated based on live observation as well as protargol- and silver nitrate-impregnated methods. The new species is characterised by a combination of features: the halophile habitat, a cell size of 50-65 × 33-40 μm, seven dorsal ridges, 10 commonly sized frontoventral cirri, two widely spaced marginal cirri, 10 dorsolateral kineties, and a double silverline pattern. The morphogenesis is similar to that of its congeners: (i) the oral primordium develops hypoapokinetally and the parental oral apparatus is retained; (ii) the frontoventral-transverse field of five streaks gives rise to the frontal, ventral, and transverse cirri, but not to the cirri I/1 and the marginal cirri; (iii) the dorsal somatic ciliature develops by intrakinetal proliferation of basal bodies in two anlagen per kinety that are just anterior and posterior to the future division furrow; (iv) the caudal cirri are formed by the two rightmost dorsolateral kineties. The SSU rDNA sequence of E. qatarensis branches with full support in the Euplotopsis elegans-Euplotes nobilii-Euplotopsis raikovi clade. The closest related publicly available SSU rDNA sequence is the one of E. nobilii, with which E. qatarensis has 93.4% sequence similarity. Euplotes parawoodruffi Song & Bradbury, 1997 is transferred to the genus Euplotoides based on the absence of frontoventral cirrus VI/3.