In vitro growth characteristics and morphological differentiation of Leishmania martiniquensis promastigotes in different culture media

Siripattanapipong, Suradej; Boontanom, Parima; Leelayoova, Saovanee; Mungthin, Mathirut; Tan‐ariya, Peerapan

doi:10.1016/j.actatropica.2019.05.030

Cited by 10 publications

(10 citation statements)

References 23 publications

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“…Regarding the in vitro infectivity of the parasite, we found a higher agreement between the parasite load evaluated by optical microscopy and flow cytometry observing more infective promastigotes in the FCS-supplemented cultures. This result was expected taking into account most articles involving in vitro and in vivo Leishmania infection studies that describe the use of FCS supplementation (de Brito et al 2019;Santarem et al 2014;Siripattanapipong et al 2019). The removal of the FCS leads to the loss of proteins, nucleoside sources, and hemoglobin, directly affecting the growth and infectivity of the promastigotes (Carvalho et al 2009;Krishnamurthy et al 2005).…”

Section: Discussionmentioning

confidence: 88%

“…infantum promastigotes cultivated in Grace’s medium demonstrated a lower percentage of infected macrophages but high parasite load (as shown by optical microscopy and flow cytometry analysis). This can be explained by the nature of the medium which is largely used in many studies to induce infection in animal models (da Silva Jr. et al 2015 ; Silva et al 2018 ; Siripattanapipong et al 2019 ). Based on our data, we can suggest that Grace’s medium, instead of inducing procyclic promastigotes, probably induces the formation of metacyclic promastigotes that explains the high parasite load in the infected macrophages in vitro.…”

Section: Discussionmentioning

confidence: 99%

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Liver infusion tryptose (LIT): the best choice for growth, viability, and infectivity of Leishmania infantum parasites

et al. 2020

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Leishmania spp. parasites have a complex biological cycle presenting basically two different morphological stages, the amastigote and promastigote forms. In vitro cultivation allows a more complete study of the biological aspects of these parasites, indicating better conditions for infection, immunoassay tests, drug evaluations, and vaccines. Thus, we evaluated the three most used culture media for Leishmania spp., Grace's insect cell culture medium (Grace's), liver infusion tryptose (LIT), and Schneider's insect medium (Schneider's), without supplementation or supplemented with fetal calf serum (FCS) and bovine serum albumin (Albumin) to evaluate the growth, viability, and infectivity of the L. infantum promastigotes. It was observed that promastigote forms have a better growth in LIT and Schneider's with or without FCS when compared to that in Grace's. The supplementation with albumin promoted greater viability of the parasites independent of the medium. For in vitro infection of J774.A1 macrophages using light microscopy and flow cytometry analyses, FCS-supplemented LIT and Grace's promoted higher percentage of infected macrophages and parasite load compared with Schneider's media. Taken together, our results demonstrated that the supplementation of LIT culture medium with FCS is the most suitable strategy to cultivate Leishmania infantum parasites enabling the maintenance of growth and infective parasites for research uses.

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Section: Discussionmentioning

confidence: 88%

Section: Discussionmentioning

confidence: 99%

Liver infusion tryptose (LIT): the best choice for growth, viability, and infectivity of Leishmania infantum parasites

et al. 2020

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“…These results are consistent with a study in L. martiniquensis , another member of the subgenus L . ( Mundinia ), in which SIM promoted metacyclogenesis and provided the highest metacyclic ratio ( Siripattanapipong et al., 2019 ). M199 has been used broadly and suggested for mass culture of Leishmania parasites ( Siripattanapipong et al., 2019 ).…”

Section: Discussionmentioning

confidence: 99%

Stimulation of metacyclogenesis in Leishmania (Mundinia) orientalis for mass production of metacyclic promastigotes

Chanmol

Jariyapan

Preativatanyou

et al. 2022

Front. Cell. Infect. Microbiol.

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Leishmania (Mundinia) orientalis is a human pathogen causing leishmaniasis and studies on the properties of metacyclic promastigotes, the parasite’s infective stage, are required for a better understanding of its transmission and infection. However, information on cultivation for mass production of L. orientalis metacyclic promastigotes and factors that stimulate their metacyclogenesis is limited. Therefore, the objective of this study was to develop a suitable methodology for generating promastigote cultures containing a high proportion and number of L. orientalis metacyclic promastigotes. Various media, i.e., Schneider’s insect medium, Medium 199 and Grace’s insect medium, supplemented with various quantities of dithiothreitol, Basal Medium Eagle vitamins, pooled human urine, and fetal bovine serum, were optimized for metacyclogenesis. The results revealed that the optimum culture medium and conditions of those tested were Schneider’s insect medium supplemented with 100 μM dithiothreitol, 1% (v/v) Basal Medium Eagle vitamins, 2% (v/v) pooled human urine, and 10% (v/v) fetal bovine serum, pH 5.0 at 26°C. We also demonstrated that L. orientalis metacyclic promastigotes could be purified and enriched by negative selection using peanut lectin. Under these culture conditions, the highest yield of metacyclic promastigotes was obtained with a significantly higher percentage of parasite survival, resistance to complement-mediated lysis, and infection index in THP-1 macrophage cells compared to parasites cultured without media supplements at neutral pH. This is the first report providing a reliable method for mass production of L. orientalis metacyclic promastigotes for in vivo infections and other experimental studies of this emerging parasite in the future.

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“…Promastigotes in the exponential phase of growth were conducted with the same conditions as previously studied. 33 Cells were harvested by centrifugation at 3000 g for 10 min at room temperature and washed three times in phosphate-buffered saline (PBS) buffer pH 7.4. Approximately 10 9 parasites were estimated by counting with a hemocytometer prior to extracting proteins.…”

Section: Methodsmentioning

confidence: 99%

Utilizing Quantitative Proteomics to Identify Species-Specific Protein Therapeutic Targets for the Treatment of Leishmaniasis

et al. 2022

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Leishmaniasis is a tropical disease caused by Leishmania parasites, which are transmitted through the bites of infected sandflies. We focused on the emergence of leishmaniasis in Thailand caused by a species ( Leishmania orientalis ). Treatment by chemotherapy is not effective against L. orientalis . Hence, we intended to solve this issue using a proteomics approach to investigate protein profiles and in silico analysis for the identification of antigenic proteins from L. orientalis , Leishmania martiniquensis , and Leishmania donovani . Using principal component analysis (PCA), protein profile comparisons indicated that different species of Leishmania are different at the protein level. Proteomics analysis identified 6099 proteins. Among these proteins, 1065 proteins were used for further analysis. There were 16 proteins that were promising candidates for therapeutic aspects as they were abundantly expressed and common to all species. In silico analysis of protein’s antigenicity revealed that eight proteins had the potential for the development of antigenic molecules. Protein profile information and these antigenic proteins may play key roles in the pathogeny of leishmaniasis and can be used as novel therapeutic targets against leishmaniasis in the future.

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In vitro growth characteristics and morphological differentiation of Leishmania martiniquensis promastigotes in different culture media

Cited by 10 publications

References 23 publications

Liver infusion tryptose (LIT): the best choice for growth, viability, and infectivity of Leishmania infantum parasites

Liver infusion tryptose (LIT): the best choice for growth, viability, and infectivity of Leishmania infantum parasites

Stimulation of metacyclogenesis in Leishmania (Mundinia) orientalis for mass production of metacyclic promastigotes

Utilizing Quantitative Proteomics to Identify Species-Specific Protein Therapeutic Targets for the Treatment of Leishmaniasis

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