Morphological characterization ofCryptosporidium parvumlife-cycle stages in anin vitromodel system

Parasitology ◽  
2009 ◽  
Vol 137 (1) ◽  
pp. 13-26 ◽  
Author(s):  
H. BOROWSKI ◽  
R. C. A. THOMPSON ◽  
T. ARMSTRONG ◽  
P. L. CLODE
Keyword(s):  
Electron Microscopy ◽  
Life Cycle ◽  
Protozoan Parasite ◽  
Morphological Characterization ◽  
Host Cells ◽  
Epithelial Cell Line ◽  
Life Cycle Stages ◽  
Complete Life Cycle ◽  
Behavioural Characteristics

SUMMARYCryptosporidium parvumis a zoonotic protozoan parasite that mainly affects the ileum of humans and livestock, with the potential to cause severe enteric disease. We describe the complete life cycle ofC. parvumin anin vitrosystem. Infected cultures of the human ileocecal epithelial cell line (HCT-8) were observed over time using electron microscopy. Additional data are presented on the morphology, development and behavioural characteristics of the different life-cycle stages as well as determining their time of occurrence after inoculation. Numerous stages ofC. parvumand their behaviour have been visualized and morphologically characterized for the first time using scanning electron microscopy. Further, parasite-host interactions and the effect ofC. parvumon host cells were also visualized. An improved understanding of the parasite's biology, proliferation and interactions with host cells will aid in the development of treatments for the disease.

Intron-containing β-tubulin transcripts in Cryptosporidium parvum cultured in vitro

Microbiology ◽  
2004 ◽  
Vol 150 (5) ◽  
pp. 1191-1195 ◽  
Author(s):  
Xiaomin Cai ◽  
Cheryl A. Lancto ◽  
Mitchell S. Abrahamsen ◽  
Guan Zhu
Keyword(s):  
Life Cycle ◽  
Cryptosporidium Parvum ◽  
Late Life ◽  
Host Cells ◽  
Intron Splicing ◽  
Transcript Processing ◽  
Life Cycle Stages ◽  
Tubulin Mrna

The genome of Cryptosporidium parvum contains a relatively small number of introns, which includes the β-tubulin gene with only a single intron. Recently, it was observed that the intron was not removed from some of the β-tubulin transcripts in the late life cycle stages cultured in vitro. Although normally spliced β-tubulin mRNA was detected in all parasite intracellular stages by RT-PCR (e.g. HCT-8 or Caco-2 cells infected with C. parvum for 12–72 h), at 48–72 h post-infection unprocessed β-tubulin transcripts containing intact introns started to appear in parasite mRNA within infected host cells. The intron-containing transcripts could be detected by fluorescence in situ hybridization (FISH) using an intron-specific probe. The intron-containing β-tubulin transcripts appeared unique to the in vitro-cultured C. parvum, since they were not detected in parasite-infected calves at 72 h. As yet, it is unclear whether the late life cycle stages of C. parvum are partially deficient in intron-splicing or the intron-splicing processes have merely slowed, both of which would allow the detection of intron-containing transcripts. Another possible explanation is that the decay in transcript processing might simply be due to the onset of parasite death. Nonetheless, the appearance of intron-containing transcripts coincides with the arrest of C. parvum development in vitro. This unusual observation prompts speculation that the abnormal intron-splicing of β-tubulin transcripts may be one of the factors preventing complete development of this parasite in vitro. Furthermore, the presence of both processed and unprocessed introns in β-tubulin transcripts in vitro may provide a venue for studying overall mechanisms for intron-splicing in this parasite.

scholarly journals Dynamics of Toxoplasma gondii Differentiation

2004 ◽  
Vol 3 (4) ◽  
pp. 992-1003 ◽  
Author(s):  
Florence Dzierszinski ◽  
Manami Nishi ◽  
Lillian Ouko ◽  
David S. Roos
Keyword(s):  
Toxoplasma Gondii ◽  
Protozoan Parasite ◽  
Time Lapse ◽  
Host Cells ◽  
Complex Life Cycle ◽  
Fluorescent Reporter ◽  
Life Cycle Stages ◽  
Dynamic View

ABSTRACT Parasite differentiation is commonly associated with transitions between complex life cycle stages and with long-term persistence in the host, and it is therefore critical for pathogenesis. In the protozoan parasite Toxoplasma gondii, interconversion between rapidly growing tachyzoites and latent encysted bradyzoites is accompanied by numerous morphological and metabolic adaptations. In order to explore early cell biological events associated with this differentiation process, we have exploited fluorescent reporter proteins targeted to various subcellular locations. Combining these markers with efficient in vitro differentiation and time-lapse video microscopy provides a dynamic view of bradyzoite development in living cultures, demonstrating subcellular reorganization, maintenance of the mitochondrion, and missegregation of the apicoplast. Bradyzoites divide asynchronously, using both endodyogeny and endopolygeny, and are highly motile both within and between host cells. Cysts are able to proliferate without passing through an intermediate tachyzoite stage, via both the migration of free bradyzoites and the fission of bradyzoite cysts, suggesting a mechanism for dissemination during chronic infection.

scholarly journals Monoclonal Antibodies to Intracellular Stages ofCryptosporidium parvumDefine Life Cycle ProgressionIn Vitro

mSphere ◽  
2018 ◽  
Vol 3 (3) ◽  
Author(s):  
Georgia Wilke ◽  
Soumya Ravindran ◽  
Lisa Funkhouser-Jones ◽  
Jennifer Barks ◽  
Qiuling Wang ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Life Cycle ◽  
Gastrointestinal Disease ◽  
Epithelial Cell Line ◽  
Type I ◽  
Parasite Life Cycle ◽  
Major Barrier ◽  
Content Type ◽  
Life Cycle Stages

ABSTRACTAmong the obstacles hinderingCryptosporidiumresearch is the lack of anin vitroculture system that supports complete life development and propagation. This major barrier has led to a shortage of widely available anti-Cryptosporidiumantibodies and a lack of markers for staging developmental progression. Previously developed antibodies againstCryptosporidiumwere raised against extracellular stages or recombinant proteins, leading to antibodies with limited reactivity across the parasite life cycle. Here we sought to create antibodies that recognize novel epitopes that could be used to define intracellular development. We identified a mouse epithelial cell line that supportedC. parvumgrowth, enabling immunization of mice with infected cells to create a bank of monoclonal antibodies (MAbs) against intracellular parasite stages while avoiding the development of host-specific antibodies. From this bank, we identified 12 antibodies with a range of reactivities across the parasite life cycle. Importantly, we identified specific MAbs that can distinguish different life cycle stages, such as trophozoites, merozoites, type I versus II meronts, and macrogamonts. These MAbs provide valuable tools for theCryptosporidiumresearch community and will facilitate future investigation into parasite biology.IMPORTANCECryptosporidiumis a protozoan parasite that causes gastrointestinal disease in humans and animals. Currently, there is a limited array of antibodies available against the parasite, which hinders imaging studies and makes it difficult to visualize the parasite life cycle in different culture systems. In order to alleviate this reagent gap, we created a library of novel antibodies against the intracellular life cycle stages ofCryptosporidium. We identified antibodies that recognize specific life cycle stages in distinctive ways, enabling unambiguous description of the parasite life cycle. These MAbs will aid future investigation intoCryptosporidiumbiology and help illuminate growth differences between various culture platforms.

scholarly journals Lankesterella (Apicomplexa, Lankesterellidae) Blood Parasites of Passeriform Birds: Prevalence, Molecular and Morphological Characterization, with Notes on Sporozoite Persistence In Vivo and Development In Vitro

Animals ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1451
Author(s):  
Carolina Romeiro Fernandes Chagas ◽  
Josef Harl ◽  
Vytautas Preikša ◽  
Dovilė Bukauskaitė ◽  
Mikas Ilgūnas ◽  
...  
Keyword(s):  
Morphological Characterization ◽  
Life Cycles ◽  
Diagnostic Methods ◽  
Host Cells ◽  
Blood Parasites ◽  
Serinus Canaria ◽  
Long Time ◽  
Development In Vitro

Recent studies confirmed that some Hepatozoon-like blood parasites (Apicomplexa) of birds are closely related to the amphibian parasite Lankesterella minima. Little is known about the biology of these pathogens in birds, including their distribution, life cycles, specificity, vectors, and molecular characterization. Using blood samples of 641 birds from 16 species, we (i) determined the prevalence and molecular diversity of Lankesterella parasites in naturally infected birds; (ii) investigated the development of Lankesterella kabeeni in laboratory-reared mosquitoes, Culex pipiens forma molestus and Aedes aegypti; and (iii) tested experimentally the susceptibility of domestic canaries, Serinus canaria, to this parasite. This study combined molecular and morphological diagnostic methods and determined 11% prevalence of Lankesterella parasites in Acrocephalidae birds; 16 Lankesterella lineages with a certain degree of host specificity and two new species (Lankesterella vacuolata n. sp. and Lankesterella macrovacuolata n. sp.) were found and characterized. Lankesterella kabeeni (formerly Hepatozoon kabeeni) was re-described. Serinus canaria were resistant after various experimental exposures. Lankesterella sporozoites rapidly escaped from host cells in vitro. Sporozoites persisted for a long time in infected mosquitoes (up to 42 days post exposure). Our study demonstrated a high diversity of Lankesterella parasites in birds, and showed that several avian Hepatozoon-like parasites, in fact, belong to Lankesterella genus.

scholarly journals Ultrastructure of Lobosphaera reniformis (Watanabe) Komárek et Fott (Chlorellales) from King George Island, South Shetland Islands, Antarctica

2014 ◽  
Vol 63 (2) ◽  
pp. 205-210 ◽  
Author(s):  
A. Massalski ◽  
T. Mrozińska ◽  
M. Olech
Keyword(s):  
Electron Microscopy ◽  
Life Cycle ◽  
South Shetland Islands ◽  
King George Island ◽  
Vegetative Cells ◽  
Shetland Islands ◽  
Complete Life Cycle ◽  
Laboratory Cultures ◽  
First Time

<i>Lobosphaera reniformis</i> (Wat.) Kom. et Fott (=<i>Chlorella reniformis</i> Wat.) so far known only from Japan, and Papua Island, was for the first time found in Antarctica (King George Island, South Shetland Islands). In laboratory cultures a complete life cycle was obtained, and most of its stages were followed by the electron microscopy. Reproduction is by morphologically different autospores. In some large vegetative cells two Golgi apparatuses lying side by side were observed.

scholarly journals Comprehensive Assessment for Construction Materials’ Environmental Safety

2016 ◽  
Vol 5 (6) ◽  
pp. 38-47
Author(s):  
Мануйлова ◽  
Natalia Manuylova ◽  
Булычев ◽  
Sergey Bulychev ◽  
Горбачев ◽  
...  
Keyword(s):  
Life Cycle ◽  
Construction Materials ◽  
Environmental Safety ◽  
Comprehensive Assessment ◽  
Raw Material ◽  
Specific Product ◽  
Life Cycle Stages ◽  
Complete Life Cycle ◽  
Environmental Properties ◽  
Proper Material

Problems related to a comprehensive assessment of construction materials’ environmental safety, taking into account stages of products’ complete life cycle have been considered. Approaches to determination of material’s safety and environmental record as environmental characteristics of the material, regardless of its use in a specific product, and without regard to processing technology have been described. It has been proposed to consider material’s safety and environmental record as the sum of three environmental safety factors for material’s life cycle stages: production of raw material and its potential environmental hazard; processing of raw material in the material; proper material from the standpoint of its environmental safety and effects on the human body. This criterion application allows compare the environmental properties both of cognate materials and dissimilar ones.

Infection with the intracellular protozoan parasite Theileria parva induces constitutively high levels of NF-kappa B in bovine T lymphocytes

1989 ◽  
Vol 9 (11) ◽  
pp. 4677-4686
Author(s):  
V Ivanov ◽  
B Stein ◽  
I Baumann ◽  
D A Dobbelaere ◽  
P Herrlich ◽  
...  
Keyword(s):  
T Cells ◽  
Phorbol Esters ◽  
Protozoan Parasite ◽  
Lymphoproliferative Disease ◽  
Host Cells ◽  
Specific Gene ◽  
Theileria Parva ◽  
Nf Kappa B ◽  
Intracellular Protozoan Parasite

The intracellular protozoan parasite Theileria parva causes a lymphoproliferative disease of T cells in cattle and uncontrolled lymphocyte proliferation in culture. We have identified and characterized in infected cells the transcriptional activator, NF-kappa B, whose recognition motifs have been identified in several gene enhancers important for lymphocyte-specific gene expression. NF-kappa B is normally constitutively activated in nuclear extracts derived from B cells and can be induced in T cells and nonlymphoid cells by phorbol esters. Theileria-infected lymphocytes contained constitutively high levels of activated NF-kappa B in nuclear fractions and inactive NF-kappa B in cytoplasmic fractions. The inactive cytoplasmic precursor could be activated by treatment of extracts with deoxycholate, which was shown previously to dissociate NF-kappa B from an inhibitor, I kappa B. Treatment of lymphocyte extracts with 3 mM GTP stimulated NF-kappa B binding to its recognition motif in vitro, thereby distinguishing it from a related nuclear factor, H2-TF1. Selective killing of the parasite, which left the host cells intact, resulted in a rapid loss of NF-kappa B from the nuclear fractions and a slower loss from the cytoplasmic fractions. In parasitized cells, NF-kappa B could not be further stimulated by treatment with 12-O-tetradecanoylphorbol-13-acetate whereas in cells treated to remove the parasite, this compound stimulated elevated levels of NF-kappa B. We propose that high levels of activated NF-kappa B are maintained by the presence of the parasite in infected T cells. Similarly, we propose that the high levels of inactive cytoplasmic precursor are a result of increased synthesis due to the presence of the parasite.

scholarly journals Organoids and Bioengineered Intestinal Models: Potential Solutions to the Cryptosporidium Culturing Dilemma

2020 ◽  
Vol 8 (5) ◽  
pp. 715 ◽  
Author(s):  
Samantha Gunasekera ◽  
Alireza Zahedi ◽  
Mark O’Dea ◽  
Brendon King ◽  
Paul Monis ◽  
...  
Keyword(s):  
Life Cycle ◽  
Host Cells ◽  
Protozoan Parasites ◽  
Future Directions ◽  
In Vivo Models ◽  
Severe Diarrhea ◽  
Transformed Cell Lines ◽  
Gnotobiotic Piglets

Cryptosporidium is a major cause of severe diarrhea-related disease in children in developing countries, but currently no vaccine or effective treatment exists for those who are most at risk of serious illness. This is partly due to the lack of in vitro culturing methods that are able to support the entire Cryptosporidium life cycle, which has led to research in Cryptosporidium biology lagging behind other protozoan parasites. In vivo models such as gnotobiotic piglets are complex, and standard in vitro culturing methods in transformed cell lines, such as HCT-8 cells, have not been able to fully support fertilization occurring in vitro. Additionally, the Cryptosporidium life cycle has also been reported to occur in the absence of host cells. Recently developed bioengineered intestinal models, however, have shown more promising results and are able to reproduce a whole cycle of infectivity in one model system. This review evaluates the recent advances in Cryptosporidium culturing techniques and proposes future directions for research that may build upon these successes.

scholarly journals Adherence of Trichomonas vaginalis to SiHa Cells is Inhibited by Diphenyleneiodonium

2020 ◽  
Vol 8 (10) ◽  
pp. 1570
Author(s):  
Yeeun Kim ◽  
Young Ha Lee ◽  
In-Wook Choi ◽  
Bu Yeon Heo ◽  
Ju-Gyeong Kang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Trichomonas Vaginalis ◽  
Parasitic Infection ◽  
Oxidase Inhibitor ◽  
Host Cells ◽  
Epithelial Cell Line ◽  
Dependent Manner ◽  
Siha Cells ◽  
Parasite Infections

Microbial adhesion is critical for parasitic infection and colonization of host cells. To study the host–parasite interaction in vitro, we established a flow cytometry-based assay to measure the adherence of Trichomonas vaginalis to epithelial cell line SiHa. SiHa cells and T. vaginalis were detected as clearly separated, quantifiable populations by flow cytometry. We found that T. vaginalis attached to SiHa cells as early as 30 min after infection and the binding remained stable up to several hours, allowing for analysis of drug treatment efficacy. Importantly, NADPH oxidase inhibitor DPI treatment induced the detachment of T. vaginalis from SiHa cells in a dose-dependent manner without affecting host cell viability. Thus, this study may provide an understanding for the potential development of therapies against T. vaginalis and other parasite infections.

scholarly journals The ketolide antibiotics HMR 3647 and HMR 3004 are active against Toxoplasma gondii in vitro and in murine models of infection.

1997 ◽  
Vol 41 (10) ◽  
pp. 2137-2140 ◽  
Author(s):  
F G Araujo ◽  
A A Khan ◽  
T L Slifer ◽  
A Bryskier ◽  
J S Remington
Keyword(s):  
Toxoplasma Gondii ◽  
Protozoan Parasite ◽  
Host Cells ◽  
Resistant Bacteria ◽  
New Class ◽  
Human Foreskin Fibroblasts ◽  
Significant Toxicity ◽  
Different Strains

Ketolides are a new class of macrolide antibiotics that have been shown to be active against a variety of bacteria including macrolide-resistant bacteria and mycobacteria. We examined two ketolides, HMR 3647 and HMR 3004, for their in vitro and in vivo activities against the protozoan parasite Toxoplasma gondii. In vitro, both ketolides at concentrations as low as 0.05 microg/ml markedly inhibited replication of tachyzoites of the RH strain within human foreskin fibroblasts. HMR 3004 demonstrated some toxicity for host cells after they were exposed to 5 microg of the drug per ml for 72 h. In contrast, HMR 3647 did not show any significant toxicity even at concentrations as high as 25 microg/ml. In vivo, both ketolides provided remarkable protection against death in mice lethally infected intraperitoneally with tachyzoites of the RH strain or orally with tissue cysts of the C56 strain of T. gondii. A dosage of 100 mg of HMR 3647 per kg of body weight per day administered for 10 days protected 50% of mice infected with tachyzoites. The same dosage of HMR 3004 protected 100% of the mice. In mice infected with cysts, a dosage of 30 mg of HMR 3647 per kg per day protected 100% of the mice, whereas a dosage of 40 mg of HMR 3004 per kg per day protected 75% of the mice. These results demonstrate that HMR 3647 and HMR 3004 possess excellent activities against two different strains of T. gondii and may be useful for the treatment of toxoplasmosis in humans.

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