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A histochemical study of the Nras/let-60 activity in filarial nematodes.

Geary JF, Lovato R, Wanji S, Guderian R, O'Neill M, Specht S, Madrill N, Geary TG, Mackenzie CD - Parasit Vectors (2015)

Bottom Line: An immunocytochemical approach using an antibody against human Nras was used to identify and detect changes in the nematode homolog let-60 that is associated with cell growth and maintenance.A statistically significant decrease in the general staining intensity of Nras/let-60 was observed in adult female O. volvulus treated with ivermectin when compared with parasites from untreated patients.Nras/let-60, as detected by immunocytochemical staining, is decreased in ivermectin-treated adult female O. volvulus relative to untreated control specimens, suggesting a suppressive effect of ivermectin on the overall biochemical activity of these parasites.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathobiology and Diagnostic Investigation, Michigan State University, East Lansing, MI, 48824, USA. gearyjam@msu.edu.

ABSTRACT

Background: Control and elimination of filarial pathogens is a central focus of major global health efforts directed at parasitic diseases of developing countries. Accomplishment of these goals would be markedly enhanced by the enhanced destruction of the adult stage of filariae. The identification of new, more quantitative biomarkers that correlate with mortality or chemotherapeutic damage to adult filariae, would greatly facilitate, for example, the development of new macrofilaricides.

Methods: An immunocytochemical approach using an antibody against human Nras was used to identify and detect changes in the nematode homolog let-60 that is associated with cell growth and maintenance. Single Onchocerca volvulus nodules were removed from each of 13 patients treated with ivermectin (as part of a community-wide mass drug administration programme), and from each of 13 untreated individuals; these 26 nodules were stained with the anti-Nras antibody. The localization and degree of positivity of Nras/let-60 staining were assessed subjectively and compared between the two groups; the positivity of staining was also quantified, using image analysis, in a subgroup of these nodules. In addition, the specific morphological association between Nras/let-60 and the Wolbachia endosymbiont present in these parasites was also observed in 4 additional filarial species using an anti-Wolbachia surface protein (WSP) antibody under light and confocal microscopy.

Results: Nras/let-60 is present in many structures within the adult female worms. A statistically significant decrease in the general staining intensity of Nras/let-60 was observed in adult female O. volvulus treated with ivermectin when compared with parasites from untreated patients. Nras/let-60 staining was frequently observed to be co-localized with WSP in O.volvulus, Brugia malayi, Litomosoides sigmodontis and Dirofilaria immitis. Nras/let60 is also present in Onchocerca ochengi.

Conclusion: Nras/let-60, as detected by immunocytochemical staining, is decreased in ivermectin-treated adult female O. volvulus relative to untreated control specimens, suggesting a suppressive effect of ivermectin on the overall biochemical activity of these parasites. Co-localization of Nras/let-60 and WSP suggests the possibility that the endosymbiont utilizes this nematode protein as part of a mutualistic relationship. Nras/let60 appears to be a useful biomarker for assessing the health of filariae.

No MeSH data available.


Related in: MedlinePlus

a Female O.volvulus stained with anti-WSP demonstrating the presence of Wolbachia. b A serial section stained the anti-Nras stain showing an intense punctate pattern associated with Wolbachia
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Fig5: a Female O.volvulus stained with anti-WSP demonstrating the presence of Wolbachia. b A serial section stained the anti-Nras stain showing an intense punctate pattern associated with Wolbachia

Mentions: Anti-WSP staining clearly identified individual organisms in various tissues (Fig. 5a); staining intensity and distribution were independent of the ivermectin treatment status. The number of bacteria varied within a tissue, such as the hypodermis, depending on the location within the worm, with some areas being free of organisms, whereas large numbers were present in others. Variation in presence and number of Wolbachia in tissues (asymmetrical distribution) was also evident in developing forms. As Nras/let-60 staining also exhibited a punctate nature in certain areas of the worm, namely those reported to contain Wolbachia, such as the hypodermis, a BlastP search was undertaken to investigate the possibility that an Nras/let-60 homolog was present in Wolbachia. This search compared human Nras and B. malayi let-60 to the Wolbachia endosymbiont of B. malayi. In both cases, the closest bacterial protein returned was Elongation Factor 4 Wolbachia (EF-4), previously annotated as GTP-binding protein Lep-A [GenBank YP_198497.1]. A global amino acid alignment showed very low sequence homology between this protein and human Nras or B. malayi let-60 (Additional file 2: Figure S2), suggesting that the anti-human Nras antibody is unlikely to recognize EF-4 in Wolbachia.Fig. 5


A histochemical study of the Nras/let-60 activity in filarial nematodes.

Geary JF, Lovato R, Wanji S, Guderian R, O'Neill M, Specht S, Madrill N, Geary TG, Mackenzie CD - Parasit Vectors (2015)

a Female O.volvulus stained with anti-WSP demonstrating the presence of Wolbachia. b A serial section stained the anti-Nras stain showing an intense punctate pattern associated with Wolbachia
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4493820&req=5

Fig5: a Female O.volvulus stained with anti-WSP demonstrating the presence of Wolbachia. b A serial section stained the anti-Nras stain showing an intense punctate pattern associated with Wolbachia
Mentions: Anti-WSP staining clearly identified individual organisms in various tissues (Fig. 5a); staining intensity and distribution were independent of the ivermectin treatment status. The number of bacteria varied within a tissue, such as the hypodermis, depending on the location within the worm, with some areas being free of organisms, whereas large numbers were present in others. Variation in presence and number of Wolbachia in tissues (asymmetrical distribution) was also evident in developing forms. As Nras/let-60 staining also exhibited a punctate nature in certain areas of the worm, namely those reported to contain Wolbachia, such as the hypodermis, a BlastP search was undertaken to investigate the possibility that an Nras/let-60 homolog was present in Wolbachia. This search compared human Nras and B. malayi let-60 to the Wolbachia endosymbiont of B. malayi. In both cases, the closest bacterial protein returned was Elongation Factor 4 Wolbachia (EF-4), previously annotated as GTP-binding protein Lep-A [GenBank YP_198497.1]. A global amino acid alignment showed very low sequence homology between this protein and human Nras or B. malayi let-60 (Additional file 2: Figure S2), suggesting that the anti-human Nras antibody is unlikely to recognize EF-4 in Wolbachia.Fig. 5

Bottom Line: An immunocytochemical approach using an antibody against human Nras was used to identify and detect changes in the nematode homolog let-60 that is associated with cell growth and maintenance.A statistically significant decrease in the general staining intensity of Nras/let-60 was observed in adult female O. volvulus treated with ivermectin when compared with parasites from untreated patients.Nras/let-60, as detected by immunocytochemical staining, is decreased in ivermectin-treated adult female O. volvulus relative to untreated control specimens, suggesting a suppressive effect of ivermectin on the overall biochemical activity of these parasites.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathobiology and Diagnostic Investigation, Michigan State University, East Lansing, MI, 48824, USA. gearyjam@msu.edu.

ABSTRACT

Background: Control and elimination of filarial pathogens is a central focus of major global health efforts directed at parasitic diseases of developing countries. Accomplishment of these goals would be markedly enhanced by the enhanced destruction of the adult stage of filariae. The identification of new, more quantitative biomarkers that correlate with mortality or chemotherapeutic damage to adult filariae, would greatly facilitate, for example, the development of new macrofilaricides.

Methods: An immunocytochemical approach using an antibody against human Nras was used to identify and detect changes in the nematode homolog let-60 that is associated with cell growth and maintenance. Single Onchocerca volvulus nodules were removed from each of 13 patients treated with ivermectin (as part of a community-wide mass drug administration programme), and from each of 13 untreated individuals; these 26 nodules were stained with the anti-Nras antibody. The localization and degree of positivity of Nras/let-60 staining were assessed subjectively and compared between the two groups; the positivity of staining was also quantified, using image analysis, in a subgroup of these nodules. In addition, the specific morphological association between Nras/let-60 and the Wolbachia endosymbiont present in these parasites was also observed in 4 additional filarial species using an anti-Wolbachia surface protein (WSP) antibody under light and confocal microscopy.

Results: Nras/let-60 is present in many structures within the adult female worms. A statistically significant decrease in the general staining intensity of Nras/let-60 was observed in adult female O. volvulus treated with ivermectin when compared with parasites from untreated patients. Nras/let-60 staining was frequently observed to be co-localized with WSP in O.volvulus, Brugia malayi, Litomosoides sigmodontis and Dirofilaria immitis. Nras/let60 is also present in Onchocerca ochengi.

Conclusion: Nras/let-60, as detected by immunocytochemical staining, is decreased in ivermectin-treated adult female O. volvulus relative to untreated control specimens, suggesting a suppressive effect of ivermectin on the overall biochemical activity of these parasites. Co-localization of Nras/let-60 and WSP suggests the possibility that the endosymbiont utilizes this nematode protein as part of a mutualistic relationship. Nras/let60 appears to be a useful biomarker for assessing the health of filariae.

No MeSH data available.


Related in: MedlinePlus