Flagellated Protozoan Parasite Research Articles

Evaluation of some micronutrients in patients infected with Giardia lamblia in Hawler city

Background and objective: The micro aerophilic flagellated protozoan parasite, Giardia lamblia, is the most common intestinal pathogen infecting human and a major diarrheal disease found throughout the world. It can cause acute or chronic diarrhea contributing to nutritional status, growth, and intellectual function. This prospective study evaluated the association between giardiasis and blood minerals disorders in Hawler city. Methods: The present study was carried out at the College of Medicine, Hawler Medical University, via collaborating study between the Department of Biochemistry and the Department of Microbiology. The study was carried out on 50 patients with giardiasis and 65 age and sex matched healthy controls. Examination of fecal samples for detection of Giardia cyst and/or trophozoite was carried out using the direct wet smear, while serum zinc, iron and copper levels were measured for both groups spectrophotometrically using commercial diagnostics kits. Results: The levels of serum zinc and iron in patients infected with Giardia lamblia were significantly lower (P

Looking through transgenic zebrafish to reveal novel mechanisms of host-parasite interaction: In vivo real-time imaging of a trypanosome infection

Trypanosomes are unicellular flagellate protozoan parasites often with a preference for living in the blood of their hosts. Trypanosoma carassii naturally infects carp, goldfish and related species. Under normal conditions fish Trypanosomes do not cause major problems in wild stocks or aquaculture fish species, so why study them? The reason is twofold: 1) they are beautiful to look at under the microscope; 2) they can help reveal detailed immune mechanisms including, in vivo macrophage polarization, immune evasion strategies, B and T cell proliferation and memory responses. Over the past years our group has used a trypanosome infection model of carp as a tool to trigger and modulate the immune system and gain insight in fundamental protective mechanisms. To further expand our toolbox and overcome some of the limitations of the carp model, we established a trypanosome infection model of zebrafish, the little twin sister of carp. Through the use of larvae, juvenile and adult transgenic fish, we are able to: 1) visualize, in real-time, the mechanisms of parasite extravasation and leukocyte-trypanosome interaction; 2) visualize the kinetics of innate immune cell activation by using Tnfa:Gfp and Il1b:Gfp expressing transgenic zebrafish lines; and 3) investigate the role of macrophages in the resolution or exacerbation of infection through ablation experiments. Where the large carp is perfect for cellular studies, the small zebrafish is an excellent tool for live imaging and genetic manipulation. Together, they form a perfect ‘twinning team’ to address fundamental questions on the evolution of the immune system and to unravel fundamental aspects of host-pathogen interaction.

RETROSPECTIVE REVIEW OF MORTALITY IN GIANT PACIFIC OCTOPUS (ENTEROCTOPUS DOFLEINI).

The giant Pacific octopus (Enteroctopus dofleini) is a popular exhibit species in public display aquaria, but information on health and disease is limited. This retrospective review evaluates time in collection and describes antemortem clinical signs and pathology of giant Pacific octopuses in an aquarium setting. Between March 2004 and December 2013, there were 19 mortalities: eight males, 10 females, and one individual whose sex was not recorded. Average time spent in collection for all octopuses was 375 ± 173 days (males 351 ± 148 days, females 410 ± 196 days). Ten (52.6%) of the octopuses were sexually mature at the time of death, six (31.6%) were not sexually mature, and reproductive status could not be determined in three octopuses (15.8%). Minimal changes were noted on gross necropsy but branchitis was histologically evident in 14 octopuses, often in conjunction with amoeboid or flagellate parasites. Senescence, parasitism, and husbandry were all important contributors to mortality and should be considered when caring for captive octopuses.

The Effect of chlorine on giardia cyst destruction isolated from different water sources in Maysan province

<p>G. lamblia was a binucleate flagellated protozoan parasite that infected the upper intestinal tract of human and many animal species. Giardiasis was the most frequently diagnosed water borne disease and the major public health concern of water utilities in the developed and developing nations. Water is an important vehicle for the transmission of Giardia to human and mammals. For identified the effect of chlorine on Giardia cyst. To detect viability of cyst in different chlorine concentration. To determine the threshold level of chlorine concentration that caused cyst destruction. Measure the pH, chlorine concentration, filtration processes and examined by zinc sulfate centrifugal flotation technique using. 50% of samples contain Giardia cysts which are untreated water sources. Cyst viability differs in different chlorine concentration in different period of time extend from few hours to more than twenty days. The extreme chlorine concentration which caused cyst destruction in hours is 1.5 mg/L.</p>

Influence of 120 kDa Pyruvate:Ferredoxin Oxidoreductase on Pathogenicity of Trichomonas vaginalis.

Trichomonas vaginalis is a flagellate protozoan parasite and commonly infected the lower genital tract in women and men. Iron is a known nutrient for growth of various pathogens, and also reported to be involved in establishment of trichomoniasis. However, the exact mechanism was not clarified. In this study, the author investigated whether the 120 kDa protein of T. vaginalis may be involved in pathogenicity of trichomonads. Antibodies against 120 kDa protein of T. vaginalis, which was identified as pyruvate:ferredoxin oxidoreductase (PFOR) by peptide analysis of MALDI-TOF-MS, were prepared in rabbits. Pretreatment of T. vaginalis with anti-120 kDa Ab decreased the proliferation and adherence to vaginal epithelial cells (MS74) of T. vaginalis. Subcutaneous tissue abscess in anti-120 kDa Ab-treated T. vaginalis-injected mice was smaller in size than that of untreated T. vaginalis-infected mice. Collectively, the 120 kDa protein expressed by iron may be involved in proliferation, adhesion to host cells, and abscess formation, thereby may influence on the pathogenicity of T. vaginalis.

Design and evaluation of a novel chitosan-based system for colon-specific drug delivery

Tritrichomonas foetus is a flagellated protozoan parasite that colonizes the feline colon causing colitis and chronic foul smelling diarrhoea. Despite the efficacy of Ronidazole in the treatment of T. foetus, Ronidazole has been reported to cause neurotoxicity in some cats due to rapid absorption in the small intestine. A novel amphoteric derivative of chitosan was synthesised and characterized. A combination of time, pH, and an enzyme controlled system was used in a study of a new compression coated tablet for delivery of Ronidazole to the colon. Axial, radial swelling and erosion of selected tablets were carried out in various media. The effect of weight ratio, enzyme and pH on in vitro drug release profile was investigated. The results show that less than 2% of the drug was released in the physiological environment of the stomach and small intestine.

Loss of the BBSome perturbs endocytic trafficking and disrupts virulence of Trypanosoma brucei

Cilia (eukaryotic flagella) are present in diverse eukaryotic lineages and have essential motility and sensory functions. The cilium's capacity to sense and transduce extracellular signals depends on dynamic trafficking of ciliary membrane proteins. This trafficking is often mediated by the Bardet-Biedl Syndrome complex (BBSome), a protein complex for which the precise subcellular distribution and mechanisms of action are unclear. In humans, BBSome defects perturb ciliary membrane protein distribution and manifest clinically as Bardet-Biedl Syndrome. Cilia are also important in several parasites that cause tremendous human suffering worldwide, yet biology of the parasite BBSome remains largely unexplored. We examined BBSome functions in Trypanosoma brucei, a flagellated protozoan parasite that causes African sleeping sickness in humans. We report that T. brucei BBS proteins assemble into a BBSome that interacts with clathrin and is localized to membranes of the flagellar pocket and adjacent cytoplasmic vesicles. Using BBS gene knockouts and a mouse infection model, we show the T. brucei BBSome is dispensable for flagellar assembly, motility, bulk endocytosis, and cell viability but required for parasite virulence. Quantitative proteomics reveal alterations in the parasite surface proteome of BBSome mutants, suggesting that virulence defects are caused by failure to maintain fidelity of the host-parasite interface. Interestingly, among proteins altered are those with ubiquitination-dependent localization, and we find that the BBSome interacts with ubiquitin. Collectively, our data indicate that the BBSome facilitates endocytic sorting of select membrane proteins at the base of the cilium, illuminating BBSome roles at a critical host-pathogen interface and offering insights into BBSome molecular mechanisms.

Preventing the transmission of American trypanosomiasis and its spread into non-endemic countries.

American trypanosomiasis, commonly known as Chagas disease, is caused by the flagellate protozoan parasite Trypanosoma cruzi. An estimated eight million people infected with T. cruzi currently reside in the endemic regions of Latin America. However, as the disease has now been imported into many non-endemic countries outside of Latin America, it has become a global health issue. We reviewed the transmission patterns and current status of disease spread pertaining to American trypanosomiasis at the global level, as well as recent advances in research. Based on an analysis of the gaps in American trypanosomiasis control, we put forward future research priorities that must be implemented to stop the global spread of the disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s40249-015-0092-7) contains supplementary material, which is available to authorized users.

Activation of MAPK Is Required for ROS Generation and Exocytosis in HMC-1 Cells Induced by Trichomonas vaginalis-Derived Secretory Products.

Trichomonas vaginalis is a flagellated protozoan parasite that causes vaginitis and cervicitis in women and asymptomatic urethritis and prostatitis in men. Mast cells have been reported to be predominant in vaginal smears and vaginal walls of patients infected with T. vaginalis. Mitogen-activated protein kinase (MAPK), activated by various stimuli, have been shown to regulate the transcriptional activity of various cytokine genes in mast cells. In this study, we investigated whether MAPK is involved in ROS generation and exocytotic degranulation in HMC-1 cells induced by T. vaginalis-derived secretory products (TvSP). We found that TvSP induces the activation of MAPK and NADPH oxidase in HMC-1 cells. Stimulation with TvSP induced phosphorylation of MAPK and p47phox in HMC-1 cells. Stimulation with TvSP also induced up-regulation of CD63, a marker for exocytosis, along the surfaces of human mast cells. Pretreatment with MAPK inhibitors strongly inhibited TvSP-induced ROS generation and exocytotic degranulation. Finally, our results suggest that TvSP induces intracellular ROS generation and exocytotic degranulation in HMC-1 via MAPK signaling.

Global Gene Expression Profiling through the Complete Life Cycle of Trypanosoma vivax.

The parasitic flagellate Trypanosoma vivax is a cause of animal trypanosomiasis across Africa and South America. The parasite has a digenetic life cycle, passing between mammalian hosts and insect vectors, and a series of developmental forms adapted to each life cycle stage. Each point in the life cycle presents radically different challenges to parasite metabolism and physiology and distinct host interactions requiring remodeling of the parasite cell surface. Transcriptomic and proteomic studies of the related parasites T. brucei and T. congolense have shown how gene expression is regulated during their development. New methods for in vitro culture of the T. vivax insect stages have allowed us to describe global gene expression throughout the complete T. vivax life cycle for the first time. We combined transcriptomic and proteomic analysis of each life stage using RNA-seq and mass spectrometry respectively, to identify genes with patterns of preferential transcription or expression. While T. vivax conforms to a pattern of highly conserved gene expression found in other African trypanosomes, (e.g. developmental regulation of energy metabolism, restricted expression of a dominant variant antigen, and expression of ‘Fam50’ proteins in the insect mouthparts), we identified significant differences in gene expression affecting metabolism in the fly and a suite of T. vivax-specific genes with predicted cell-surface expression that are preferentially expressed in the mammal (‘Fam29, 30, 42’) or the vector (‘Fam34, 35, 43’). T. vivax differs significantly from other African trypanosomes in the developmentally-regulated proteins likely to be expressed on its cell surface and thus, in the structure of the host-parasite interface. These unique features may yet explain the species differences in life cycle and could, in the form of bloodstream-stage proteins that do not undergo antigenic variation, provide targets for therapy.

Genetic diversity of Trichomonas vaginalis clinical isolates from Henan province in central China

Trichomonas vaginalis is a flagellated protozoan parasite that infects the human urogenital tract, causing the most common non-viral, sexually transmitted disease worldwide. In this study, genetic variants of T. vaginalis were identified in Henan Province, China. Fragments of the small subunit of nuclear ribosomal RNA (18S rRNA) were amplified from 32 T. vaginalis isolates obtained from seven regions of Henan Province. Overall, 18 haplotypes were determined from the 18S rRNA sequences. Each sampled population and the total population displayed high haplotype diversity (Hd), accompanied by very low nucleotide diversity (Pi). In these molecular genetic variants, 91.58% genetic variation was derived from intra-regions. Phylogenetic analysis revealed no correlation between phylogeny and geographic distribution. Demographic analysis supported population expansion of T. vaginalis isolates from central China. Our findings showing moderate-to-high genetic variations in the 32 isolates of T. vaginalis provide useful knowledge for monitoring changes in parasite populations for the development of future control strategies.

Screening natural product extracts against kinetoplastid parasites

The kinetoplastids parasites are a group of flagellated protozoan parasites characterized by the subcellular structure, kinetoplast. Trypanosoma spp and Leishmania spp are the most notable kinetoplastids with major impact on human and animal health. Human African Trypanosomiasis (HAT) caused by Trypansoma brucei and visceral leishmaniasis caused by Leishmania donovani affect mostly neglected segments of the populations. The toxicity and resistance for currently available drugs and depleted drug discovery pipeline, necessitate discovery of new drugs. Natural products are vast source of diverse drug molecules with novel chemotypes. A 384 well plate screening paradigm has been developed that covers the screening of natural products extracts against L. donovani promastigotes, L. donovani axenic amastigote, differentiated THP1 (Human Acute Monocytic Leukemia) cells infected with L. donovani amastigotes and also cytotoxicity against uninfected THP1 cells. In the initial phase 176 plant extracts, prepared from the North American plants, have been screened. Lead extracts Cupressus lawsoniana, Acer pensylvanicum, Verbesina virginica, were identified as new antitrypanosomal leads with significant selectivity. Further follow up evaluation of lead extracts is likely to identify drug leads with novel chemotypes.

Giardia - significance of zoonotic routes in human infections.

Abstract Giardia are flagellated protozoan parasites of numerous animals. Species of Giardia were originally described based, primarily, on the host from which they were isolated; later, isolates that were morphologically identical with Giardia duodenalis were synonymized with that species. Accumulating genetic evidence, however, indicates that G. duodenalis is a species complex in which members are morphologically identical, but, genetically distinct. At least eight distinct groups (=assemblages) have been identified, all of which likely constitute unique species. Although zoonotic transmission may occur in discrete circumstances, people to people transmission is likely much more important. However, the human disease burden associated with zoonotic transmission still needs to be assessed, including differences in the biology and clinical manifestations associated with the various types. Genetic targets for typing have been identified but do not work well for all assemblages, particularly assemblage B; thus, the search for appropriate targets must continue. Multilocus genotyping and standardized terminology is highly recommended as interpretations based on single markers may lead to incomplete conclusions. Combining appropriate genetic markers with well-designed epidemiological studies will be necessary to further our understanding of Giardia transmission patterns and infection sources for both people and animals.

In vitro efficacy of ethanolic extract of Artemisia absinthium (Asteraceae) against Leishmania major L. using cell sensitivity and flow cytometry assays.

Leishmaniasis is one of the most neglected human diseases with an estimated global burden ranking second in mortality and fourth in morbidity among the tropical infections. Chemotherapy involving the use of drugs like glucantime is the mainstay treatment in endemic areas of Iran. Drug resistance is increasingly prevalent, so search for alternative therapy is gathering pace. Medicinal herbs, like wormwood Artemisia, have chemical compounds effective against a number of pathogens. In this study, the efficacy of ethanol extract from Artemisia absinthium (Asteraceae) against Leishmania major L. was investigated in vitro. The outcome of different effective doses (1-40mg/ml) of ethanol extracts from this medicinal herb, A. absinthium, on a standard Iranian parasite strain of L. major was examined. The L. major promastigote cell sensitivity and mortality or viability effects due to the addition of herbal extract were measured using the MTT assay and the flow cytometry technique, respectively. There was complete agreement between the two assays. The lethal concentration (LC50) was measured as 101mg/ml. Some contrasting relationships between the medicinal herb concentrations and the viability of parasites were observed; so that there was an increased multiplication of the parasite at low concentrations of the drug, but an anti-parasitic apoptotic effect was seen at high concentrations of A. absinthium. It was concluded that there might be one or more chemical constituents within the herbal extract of wormwood which at high concentration controlled cell division and affected the relevant activity within the only one giant mitochondrion in this flagellate parasite. At low doses, however, it showed the opposite effect of leading to mitotic cell divisions.

Man-biting sand fly species and natural infection with the Leishmania promastigote in leishmaniasis-endemic areas of Ecuador

A countrywide surveillance of sand flies was performed to obtain information on their geographical distribution and natural infection by Leishmania protozoa in Ecuador. A total of 18,119 sand flies were collected by human landing collections during 32 years from 1982 to 2014, and 29 species were recognized. The most prevalent 10 species were Lutzomyia gomezi, Lu. robusta, Lu. hartmanni, Lu. shannoni, Lu. trapidoi, Lu. panamensis, Lu. maranonensis, Lu. ayacuchensis, Lu. tortura and Lu. yuilli yuilli, and their topographical and vertical distributions were identified. Among all the sand flies, only 197 (1.09%) flies of four Lutzomyia species, Lu. gomezi, Lu. trapidoi, Lu. tortura and Lu. ayacuchensis, were positive for Leishmania. Endotrypanum, a flagellate parasite not pathogenic to humans, were detected in five Lutzomyia species, Lu. robusta, Lu. hartmanni, Lu. trapidoi, Lu. panamensis and Lu. yuilli yuilli, suggesting wide vector-ranges of Endotrypanum species. These data on the genus Lutzomyia and their natural infections with Leishmania and Endotrypanum will be useful for transmission studies and surveillance of leishmaniasis.

Natural sesquiterpene lactones induce programmed cell death in Trypanosoma cruzi: A new therapeutic target?

BackgroundChagas disease or American Trypanosomiasis is caused by the flagellated protozoan parasite Trypanosoma cruzi (T. cruzi) and is recognized by the WHO as one of the world's 17 neglected tropical diseases. Only two drugs (Benznidazol, Bz and Nifurtimox, Nx) are currently accepted for treatment, however they cause severe adverse effects and their efficacy is still controversial. It is then important to explore for new drugs. PurposeProgrammed cell death (PCD) in parasites offers interesting new therapeutic targets. The aim of this work was to evaluate the induction of PCD in T. cruzi by two natural sesquiterpene lactones (STLs), dehydroleucodine (DhL) and helenalin (Hln) as compared with the two conventional drugs, Bz and Nx. Material and MethodsHln and DhL were isolated from aerial parts of Gaillardia megapotamica and Artemisia douglassiana Besser, respectively. Purity of compounds (greater than 95%) was confirmed by 13C-nuclear magnetic resonance, melting point analysis, and optical rotation. Induction of PCD in T. cruzi epimastigotes and trypomastigotes by DhL, Hln, Bz and Nx was assayed by phosphatidylserine exposure at the parasite surface and by detection of DNA fragmentation using the TUNEL assay. Trypanocidal activity of natural and synthetic compounds was assayed by measuring parasite viability using the MTT method. ResultsThe two natural STLs, DhL and Hln, induce programmed cell death in both, the replicative epimastigote form and the infective trypomastigote form of T. cruzi. Interestingly, the two conventional antichagasic drugs (Bz and Nx) do not induce programmed cell death. A combination of DhL and either Bz or Nx showed an increased effect of natural compounds and synthetic drugs on the decrease of parasite viability. ConclusionDhL and Hln induce programmed cell death in T. cruzi replicative epimastigote and infective trypomastigote forms, which is a different mechanism of action than the conventional drugs to kill the parasite. Therefore DhL and Hln may offer an interesting option for the treatment of Chagas disease, alone or in combination with conventional drugs.

Development of a combined in vitro cell culture – Quantitative PCR assay for evaluating the disinfection performance of pulsed light for treating the waterborne enteroparasite Giardia lamblia

Giardia lamblia is a flagellated protozoan parasite that is recognised as a frequent cause of water-borne disease in humans and animals. We report for the first time on the use of a combined in vitro HCT-8 cell culture-quantitative PCR assay for evaluating the efficacy of using pulsed UV light for treating G. lamblia parasites. Findings showed that current methods that are limited to using vital stains before and after cyst excystation are not appropriate for monitoring or evaluating cyst destruction post PUV-treatments. Use of the human ileocecal HCT-8 cell line was superior to that of the human colon Caco-2 cell line for in vitro culture and determining PUV sensitivity of treated cysts. G. lamblia cysts were also shown to be more resistant to PUV irradiation compared to treating similar numbers of Cryptosporidium parvum oocysts. These observations also show that the use of this HCT-8 cell culture assay may replace use of animal models for determining disinfection performances of PUV for treating both C. parvum and G. lamblia.

A critical assessment of two real-time PCR assays targeting the (SSU) rRNA and gdh genes for the molecular identification of Giardia intestinalis in a clinical laboratory

IntroductionGiardiasis is an intestinal diarrhoeal illness caused by the flagellate protozoan parasite Giardia intestinalis. Molecular techniques for the identification of G. intestinalis have generally been shown to offer a better...

Giardia and Vilém Dušan Lambl.

Giardia and Vilém Dušan Lambl.

Blackhead disease: reduced sensitivity of Histomonas meleagridis to nitarsone in vitro and in vivo.

Histomonas meleagridis, a flagellated protozoan parasite, is the causative agent of blackhead disease or histomoniasis in gallinaceous birds. Currently nitarsone (4-nitrophenylarsonic acid) is the only approved preventative drug available in the United States against blackhead disease. Initially we tested the sensitivity of three different isolates of H. meleagridis collected from outbreaks in North Carolina (strain MNC), Michigan (strain ZM), and Georgia (strain BG) to nitarsone using in vitro culture conditions. Strain ZM and strain BG at 100 and 400 ppm showed reduced growth in comparison to their respective control groups. However, there was no inhibition of growth in strain MNC treated with nitarsone at 100 ppm, while reduced growth was seen at 400 ppm. To test the resistance of strain MNC to nitarsone in vivo, turkey poults fed a nitarsone or a control diet were inoculated cloacally with H. meleagridis. The nitarsone-treated group of birds did not show any significant difference compared to that of infected control group when measuring weight gain and liver and cecal lesions scores. Histomonas meleagridis were reisolated from the nitarsone-fed turkeys and subjected to the in vitro assay. Regenerated H. meleagridis maintain their resistance to nitarsone at 100 ppm. This study demonstrates that strain MNC has acquired partial resistance to nitarsone.