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Axenic amastigote cultivation and in vitro development of Leishmania orientalis

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Axenic amastigote cultivation and in vitro development of Leishmania orientalis. / Chanmol, Wetpisit; Jariyapan, Narissara; Somboon, Pradya et al.
In: Parasitology Research, Vol. 118, No. 6, 01.06.2019, p. 1885-1897.

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Chanmol, W, Jariyapan, N, Somboon, P, Bates, MD & Bates, PA 2019, 'Axenic amastigote cultivation and in vitro development of Leishmania orientalis', Parasitology Research, vol. 118, no. 6, pp. 1885-1897. https://doi.org/10.1007/s00436-019-06311-z

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Chanmol W, Jariyapan N, Somboon P, Bates MD, Bates PA. Axenic amastigote cultivation and in vitro development of Leishmania orientalis. Parasitology Research. 2019 Jun 1;118(6):1885-1897. Epub 2019 Apr 10. doi: 10.1007/s00436-019-06311-z

Author

Chanmol, Wetpisit ; Jariyapan, Narissara ; Somboon, Pradya et al. / Axenic amastigote cultivation and in vitro development of Leishmania orientalis. In: Parasitology Research. 2019 ; Vol. 118, No. 6. pp. 1885-1897.

Bibtex

@article{2b6ef4897a924e59bd29443923057a27,
title = "Axenic amastigote cultivation and in vitro development of Leishmania orientalis",
abstract = "Leishmania (Mundinia) orientalis is a recently described new species that causes leishmaniasis in Thailand. To facilitate characterization of this new species, an in vitro culture system to generate L. orientalis axenic amastigotes was developed. In vitro culture conditions of the axenic culture-derived amastigotes were optimized by manipulation of temperature and pH. Four criteria were used to evaluate the resulting L. orientalis axenic amastigotes, i.e., morphology, zymographic analysis of nucleases, cyclic transformation, and infectivity to the human monocytic cell line (THP-1) cells. Results revealed that the best culture condition for L. orientalis axenic amastigotes was Grace's insect medium supplemented with FCS 20%, 2% human urine, 1% BME vitamins, and 25 μg/ml gentamicin sulfate, pH 5.5 at 35 °C. For promastigotes, the condition was M199 medium, 10% FCS supplemented with 2% human urine, 1% BME vitamins, and 25 μg/ml gentamicin sulfate, pH 6.8 at 26 °C. Morphological characterization revealed six main stages of the parasites including amastigotes, procyclic promastigotes, nectomonad promastigotes, leptomonad promastigotes, metacyclic promastigotes, and paramastigotes. Also, changes in morphology during the cycle were accompanied by changes in zymographic profiles of nucleases. The developmental cycle of L. orientalis in vitro was complete in 12 days using both culture systems. The infectivity to THP-1 macrophages and intracellular growth of the axenic amastigotes was similar to that of THP-1 derived intracellular amastigotes. These results confirmed the successful axenic cultivation of L. orientalis amastigotes. The axenic amastigotes and promastigotes can be used for further study on infection in permissive vectors and animals.",
keywords = "Leishmania, Thailand, Axenic amastigote, Promastigote, Culture, Zymography",
author = "Wetpisit Chanmol and Narissara Jariyapan and Pradya Somboon and Bates, {Michelle D.} and Bates, {Paul A.}",
note = "The final publication is available at Springer via http://dx.doi.org/10.1007/s00436-019-06311-z",
year = "2019",
month = jun,
day = "1",
doi = "10.1007/s00436-019-06311-z",
language = "English",
volume = "118",
pages = "1885--1897",
journal = "Parasitology Research",
issn = "0044-3255",
publisher = "Springer Verlag",
number = "6",

}

RIS

TY - JOUR

T1 - Axenic amastigote cultivation and in vitro development of Leishmania orientalis

AU - Chanmol, Wetpisit

AU - Jariyapan, Narissara

AU - Somboon, Pradya

AU - Bates, Michelle D.

AU - Bates, Paul A.

N1 - The final publication is available at Springer via http://dx.doi.org/10.1007/s00436-019-06311-z

PY - 2019/6/1

Y1 - 2019/6/1

N2 - Leishmania (Mundinia) orientalis is a recently described new species that causes leishmaniasis in Thailand. To facilitate characterization of this new species, an in vitro culture system to generate L. orientalis axenic amastigotes was developed. In vitro culture conditions of the axenic culture-derived amastigotes were optimized by manipulation of temperature and pH. Four criteria were used to evaluate the resulting L. orientalis axenic amastigotes, i.e., morphology, zymographic analysis of nucleases, cyclic transformation, and infectivity to the human monocytic cell line (THP-1) cells. Results revealed that the best culture condition for L. orientalis axenic amastigotes was Grace's insect medium supplemented with FCS 20%, 2% human urine, 1% BME vitamins, and 25 μg/ml gentamicin sulfate, pH 5.5 at 35 °C. For promastigotes, the condition was M199 medium, 10% FCS supplemented with 2% human urine, 1% BME vitamins, and 25 μg/ml gentamicin sulfate, pH 6.8 at 26 °C. Morphological characterization revealed six main stages of the parasites including amastigotes, procyclic promastigotes, nectomonad promastigotes, leptomonad promastigotes, metacyclic promastigotes, and paramastigotes. Also, changes in morphology during the cycle were accompanied by changes in zymographic profiles of nucleases. The developmental cycle of L. orientalis in vitro was complete in 12 days using both culture systems. The infectivity to THP-1 macrophages and intracellular growth of the axenic amastigotes was similar to that of THP-1 derived intracellular amastigotes. These results confirmed the successful axenic cultivation of L. orientalis amastigotes. The axenic amastigotes and promastigotes can be used for further study on infection in permissive vectors and animals.

AB - Leishmania (Mundinia) orientalis is a recently described new species that causes leishmaniasis in Thailand. To facilitate characterization of this new species, an in vitro culture system to generate L. orientalis axenic amastigotes was developed. In vitro culture conditions of the axenic culture-derived amastigotes were optimized by manipulation of temperature and pH. Four criteria were used to evaluate the resulting L. orientalis axenic amastigotes, i.e., morphology, zymographic analysis of nucleases, cyclic transformation, and infectivity to the human monocytic cell line (THP-1) cells. Results revealed that the best culture condition for L. orientalis axenic amastigotes was Grace's insect medium supplemented with FCS 20%, 2% human urine, 1% BME vitamins, and 25 μg/ml gentamicin sulfate, pH 5.5 at 35 °C. For promastigotes, the condition was M199 medium, 10% FCS supplemented with 2% human urine, 1% BME vitamins, and 25 μg/ml gentamicin sulfate, pH 6.8 at 26 °C. Morphological characterization revealed six main stages of the parasites including amastigotes, procyclic promastigotes, nectomonad promastigotes, leptomonad promastigotes, metacyclic promastigotes, and paramastigotes. Also, changes in morphology during the cycle were accompanied by changes in zymographic profiles of nucleases. The developmental cycle of L. orientalis in vitro was complete in 12 days using both culture systems. The infectivity to THP-1 macrophages and intracellular growth of the axenic amastigotes was similar to that of THP-1 derived intracellular amastigotes. These results confirmed the successful axenic cultivation of L. orientalis amastigotes. The axenic amastigotes and promastigotes can be used for further study on infection in permissive vectors and animals.

KW - Leishmania

KW - Thailand

KW - Axenic amastigote

KW - Promastigote

KW - Culture

KW - Zymography

U2 - 10.1007/s00436-019-06311-z

DO - 10.1007/s00436-019-06311-z

M3 - Journal article

C2 - 30972571

VL - 118

SP - 1885

EP - 1897

JO - Parasitology Research

JF - Parasitology Research

SN - 0044-3255

IS - 6

ER -