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Fluorescence In Situ Hybridization (FISH) Assays for Diagnosing Malaria in Endemic Areas.

Shah J, Mark O, Weltman H, Barcelo N, Lo W, Wronska D, Kakkilaya S, Rao A, Bhat ST, Sinha R, Omar S, O'bare P, Moro M, Gilman RH, Harris N - PLoS ONE (2015)

Bottom Line: The P. falciparum (PF) FISH assay and P. vivax (PV) FISH assay detect and differentiate between P. falciparum and P. vivax respectively from other Plasmodium species.The FISH assays are more sensitive than Giemsa.The sensitivities of P-Genus, PF and PV FISH assays were found to be 98.2%, 94.5% and 98.3%, respectively compared to 89.9%, 83.3% and 87.9% for the detection of Plasmodium, P. falciparum and P. vivax by Giemsa staining respectively.

View Article: PubMed Central - PubMed

Affiliation: ID-FISH Technology Inc., Palo Alto, California, United States of America; IGeneX Inc. Palo Alto, California, United States of America.

ABSTRACT
Malaria is a responsible for approximately 600 thousand deaths worldwide every year. Appropriate and timely treatment of malaria can prevent deaths but is dependent on accurate and rapid diagnosis of the infection. Currently, microscopic examination of the Giemsa stained blood smears is the method of choice for diagnosing malaria. Although it has limited sensitivity and specificity in field conditions, it still remains the gold standard for the diagnosis of malaria. Here, we report the development of a fluorescence in situ hybridization (FISH) based method for detecting malaria infection in blood smears and describe the use of an LED light source that makes the method suitable for use in resource-limited malaria endemic countries. The Plasmodium Genus (P-Genus) FISH assay has a Plasmodium genus specific probe that detects all five species of Plasmodium known to cause the disease in humans. The P. falciparum (PF) FISH assay and P. vivax (PV) FISH assay detect and differentiate between P. falciparum and P. vivax respectively from other Plasmodium species. The FISH assays are more sensitive than Giemsa. The sensitivities of P-Genus, PF and PV FISH assays were found to be 98.2%, 94.5% and 98.3%, respectively compared to 89.9%, 83.3% and 87.9% for the detection of Plasmodium, P. falciparum and P. vivax by Giemsa staining respectively.

No MeSH data available.


Related in: MedlinePlus

Blood smears tested with Plasmodium genus FISH assay.Blood smears from different Plasmodium species P. falciparum, P. vivax, P. knowlesi, P. ovale and P. malariae were analyzed using Plasmodium genus FISH assay. (1) All the processed smears were read with a 100X objective in a fluorescence microscope. (2) P. falciparum. P. vivax, P. knowlesi and negative control smears were read with a 100X objective on a regular microscope with a LED unit. Green fluorescence indicates the presence of Plasmodium ribosomal RNA (rRNA). (A) P. falciparum including crescent shaped gametocytes (B) P. vivax; (C) P. knowlesi; (D) P. ovale; (E) P. malariae; and (N) Negative Control; (A1) P. falciparum;. (B1) P. vivax; (C1) P. knowlesi; and (N1) Negative Control.
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pone.0136726.g001: Blood smears tested with Plasmodium genus FISH assay.Blood smears from different Plasmodium species P. falciparum, P. vivax, P. knowlesi, P. ovale and P. malariae were analyzed using Plasmodium genus FISH assay. (1) All the processed smears were read with a 100X objective in a fluorescence microscope. (2) P. falciparum. P. vivax, P. knowlesi and negative control smears were read with a 100X objective on a regular microscope with a LED unit. Green fluorescence indicates the presence of Plasmodium ribosomal RNA (rRNA). (A) P. falciparum including crescent shaped gametocytes (B) P. vivax; (C) P. knowlesi; (D) P. ovale; (E) P. malariae; and (N) Negative Control; (A1) P. falciparum;. (B1) P. vivax; (C1) P. knowlesi; and (N1) Negative Control.

Mentions: In the P-Genus FISH assay, the Plasmodium specific probe is labeled with an Alexa 488 green fluorescent dye. Therefore, all the Plasmodium parasites would appear green under the blue filter (Excitation 492 nm; emission 530 nm) when viewed in a fluorescence microscope (Fig 1). Normal controls and non-malaria parasites will not be visible under the blue filter (Fig 1).


Fluorescence In Situ Hybridization (FISH) Assays for Diagnosing Malaria in Endemic Areas.

Shah J, Mark O, Weltman H, Barcelo N, Lo W, Wronska D, Kakkilaya S, Rao A, Bhat ST, Sinha R, Omar S, O'bare P, Moro M, Gilman RH, Harris N - PLoS ONE (2015)

Blood smears tested with Plasmodium genus FISH assay.Blood smears from different Plasmodium species P. falciparum, P. vivax, P. knowlesi, P. ovale and P. malariae were analyzed using Plasmodium genus FISH assay. (1) All the processed smears were read with a 100X objective in a fluorescence microscope. (2) P. falciparum. P. vivax, P. knowlesi and negative control smears were read with a 100X objective on a regular microscope with a LED unit. Green fluorescence indicates the presence of Plasmodium ribosomal RNA (rRNA). (A) P. falciparum including crescent shaped gametocytes (B) P. vivax; (C) P. knowlesi; (D) P. ovale; (E) P. malariae; and (N) Negative Control; (A1) P. falciparum;. (B1) P. vivax; (C1) P. knowlesi; and (N1) Negative Control.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4558036&req=5

pone.0136726.g001: Blood smears tested with Plasmodium genus FISH assay.Blood smears from different Plasmodium species P. falciparum, P. vivax, P. knowlesi, P. ovale and P. malariae were analyzed using Plasmodium genus FISH assay. (1) All the processed smears were read with a 100X objective in a fluorescence microscope. (2) P. falciparum. P. vivax, P. knowlesi and negative control smears were read with a 100X objective on a regular microscope with a LED unit. Green fluorescence indicates the presence of Plasmodium ribosomal RNA (rRNA). (A) P. falciparum including crescent shaped gametocytes (B) P. vivax; (C) P. knowlesi; (D) P. ovale; (E) P. malariae; and (N) Negative Control; (A1) P. falciparum;. (B1) P. vivax; (C1) P. knowlesi; and (N1) Negative Control.
Mentions: In the P-Genus FISH assay, the Plasmodium specific probe is labeled with an Alexa 488 green fluorescent dye. Therefore, all the Plasmodium parasites would appear green under the blue filter (Excitation 492 nm; emission 530 nm) when viewed in a fluorescence microscope (Fig 1). Normal controls and non-malaria parasites will not be visible under the blue filter (Fig 1).

Bottom Line: The P. falciparum (PF) FISH assay and P. vivax (PV) FISH assay detect and differentiate between P. falciparum and P. vivax respectively from other Plasmodium species.The FISH assays are more sensitive than Giemsa.The sensitivities of P-Genus, PF and PV FISH assays were found to be 98.2%, 94.5% and 98.3%, respectively compared to 89.9%, 83.3% and 87.9% for the detection of Plasmodium, P. falciparum and P. vivax by Giemsa staining respectively.

View Article: PubMed Central - PubMed

Affiliation: ID-FISH Technology Inc., Palo Alto, California, United States of America; IGeneX Inc. Palo Alto, California, United States of America.

ABSTRACT
Malaria is a responsible for approximately 600 thousand deaths worldwide every year. Appropriate and timely treatment of malaria can prevent deaths but is dependent on accurate and rapid diagnosis of the infection. Currently, microscopic examination of the Giemsa stained blood smears is the method of choice for diagnosing malaria. Although it has limited sensitivity and specificity in field conditions, it still remains the gold standard for the diagnosis of malaria. Here, we report the development of a fluorescence in situ hybridization (FISH) based method for detecting malaria infection in blood smears and describe the use of an LED light source that makes the method suitable for use in resource-limited malaria endemic countries. The Plasmodium Genus (P-Genus) FISH assay has a Plasmodium genus specific probe that detects all five species of Plasmodium known to cause the disease in humans. The P. falciparum (PF) FISH assay and P. vivax (PV) FISH assay detect and differentiate between P. falciparum and P. vivax respectively from other Plasmodium species. The FISH assays are more sensitive than Giemsa. The sensitivities of P-Genus, PF and PV FISH assays were found to be 98.2%, 94.5% and 98.3%, respectively compared to 89.9%, 83.3% and 87.9% for the detection of Plasmodium, P. falciparum and P. vivax by Giemsa staining respectively.

No MeSH data available.


Related in: MedlinePlus