Limits...
A new methodology for evaluation of nematode viability.

Ferreira SR, Mendes TA, Bueno LL, de Araújo JV, Bartholomeu DC, Fujiwara RT - Biomed Res Int (2015)

Bottom Line: Furthermore, data on motility test presented an inverse correlation with fluorimetric data when ivermectin was used.Our results showed that lower concentrations of drugs were effective to interfere in the processes of cellular transport while higher drugs concentrations were necessary in order to result in any damage to cell integrity.The methodology described in this work might be useful for studies that aim to evaluate the viability of nematodes, particularly for testing of new anthelminthic compounds using an easy, economic, reproducible, and no time-consuming technique.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Parasitologia, Universidade Federal de Minas Gerais, Avenida Antônio Carlos 6627, Pampulha, 31270-901 Belo Horizonte, MG, Brazil.

ABSTRACT
Nematodes infections are responsible for debilitating conditions and economic losses in domestic animals as well as livestock and are considered an important public health problem due to the high prevalence in humans. The nematode resistance for drugs has been reported for livestock, highlighting the importance for development of new anthelmintic compounds. The aim of the current study was to apply and compare fluorimetric techniques using Sytox and propidium iodide for evaluating the viability of C. elegans larvae after treatment with anthelmintic drugs. These fluorescent markers were efficient to stain larvae treated with ivermectin and albendazole sulfoxide. We observed that densitometric values were proportional to the concentration of dead larvae stained with both markers. Furthermore, data on motility test presented an inverse correlation with fluorimetric data when ivermectin was used. Our results showed that lower concentrations of drugs were effective to interfere in the processes of cellular transport while higher drugs concentrations were necessary in order to result in any damage to cell integrity. The methodology described in this work might be useful for studies that aim to evaluate the viability of nematodes, particularly for testing of new anthelminthic compounds using an easy, economic, reproducible, and no time-consuming technique.

Show MeSH

Related in: MedlinePlus

Correlation of paralysis and number of pixels from fluorimetric assays. (a) Propidium iodide and (b) Sytox. Statistical significance was determined by Spearman rank correlation.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4383492&req=5

fig6: Correlation of paralysis and number of pixels from fluorimetric assays. (a) Propidium iodide and (b) Sytox. Statistical significance was determined by Spearman rank correlation.

Mentions: Figure 3 shows the densitometric data of the larvae treated with albendazole sulfoxide and ivermectin and stained with propidium iodide. Our data suggest that the higher concentrations of drugs might affect the mechanisms used to expel the marker, once propidium iodide labels the nucleic acids when it remains within the cell. The densitometric data of larvae treated with the same drugs and stained with Sytox suggests that higher concentrations of drugs were able to induce damage in the cellular integrity of the larvae, because this marker is not able to overcome intact membranes (Figure 4). Concerning the ivermectin treatment, we observed that at the highest concentrations all larvae were practically inert (Figure 5). For instance, at concentration of 1000 uM, 100% of paralysis was observed when compared to control group (P < 0.05). A negative correlation (Spearman rank correlation, P < 0.05) was observed when paralysis and densitometry data were compared (Figure 6). The albendazole sulfoxide treatment induced a slightly reduction of the movement; however, body straight shapes and absence movement were not observed; thus, motility assay could not be properly performed (data not shown). The IC50 dates are demonstrated at Table 1, where different IC50 were observed. IC50 data for albendazole sulfoxide motility test were not included once the larvae did not fit the criteria used for impairment of motility. Larvae were considered paralyzed when presenting straight body and absence of any motility; the treatment with albendazole sulfoxide induced reduction of the larval movement but body straight shapes and absence of movement were not observed.


A new methodology for evaluation of nematode viability.

Ferreira SR, Mendes TA, Bueno LL, de Araújo JV, Bartholomeu DC, Fujiwara RT - Biomed Res Int (2015)

Correlation of paralysis and number of pixels from fluorimetric assays. (a) Propidium iodide and (b) Sytox. Statistical significance was determined by Spearman rank correlation.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4383492&req=5

fig6: Correlation of paralysis and number of pixels from fluorimetric assays. (a) Propidium iodide and (b) Sytox. Statistical significance was determined by Spearman rank correlation.
Mentions: Figure 3 shows the densitometric data of the larvae treated with albendazole sulfoxide and ivermectin and stained with propidium iodide. Our data suggest that the higher concentrations of drugs might affect the mechanisms used to expel the marker, once propidium iodide labels the nucleic acids when it remains within the cell. The densitometric data of larvae treated with the same drugs and stained with Sytox suggests that higher concentrations of drugs were able to induce damage in the cellular integrity of the larvae, because this marker is not able to overcome intact membranes (Figure 4). Concerning the ivermectin treatment, we observed that at the highest concentrations all larvae were practically inert (Figure 5). For instance, at concentration of 1000 uM, 100% of paralysis was observed when compared to control group (P < 0.05). A negative correlation (Spearman rank correlation, P < 0.05) was observed when paralysis and densitometry data were compared (Figure 6). The albendazole sulfoxide treatment induced a slightly reduction of the movement; however, body straight shapes and absence movement were not observed; thus, motility assay could not be properly performed (data not shown). The IC50 dates are demonstrated at Table 1, where different IC50 were observed. IC50 data for albendazole sulfoxide motility test were not included once the larvae did not fit the criteria used for impairment of motility. Larvae were considered paralyzed when presenting straight body and absence of any motility; the treatment with albendazole sulfoxide induced reduction of the larval movement but body straight shapes and absence of movement were not observed.

Bottom Line: Furthermore, data on motility test presented an inverse correlation with fluorimetric data when ivermectin was used.Our results showed that lower concentrations of drugs were effective to interfere in the processes of cellular transport while higher drugs concentrations were necessary in order to result in any damage to cell integrity.The methodology described in this work might be useful for studies that aim to evaluate the viability of nematodes, particularly for testing of new anthelminthic compounds using an easy, economic, reproducible, and no time-consuming technique.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Parasitologia, Universidade Federal de Minas Gerais, Avenida Antônio Carlos 6627, Pampulha, 31270-901 Belo Horizonte, MG, Brazil.

ABSTRACT
Nematodes infections are responsible for debilitating conditions and economic losses in domestic animals as well as livestock and are considered an important public health problem due to the high prevalence in humans. The nematode resistance for drugs has been reported for livestock, highlighting the importance for development of new anthelmintic compounds. The aim of the current study was to apply and compare fluorimetric techniques using Sytox and propidium iodide for evaluating the viability of C. elegans larvae after treatment with anthelmintic drugs. These fluorescent markers were efficient to stain larvae treated with ivermectin and albendazole sulfoxide. We observed that densitometric values were proportional to the concentration of dead larvae stained with both markers. Furthermore, data on motility test presented an inverse correlation with fluorimetric data when ivermectin was used. Our results showed that lower concentrations of drugs were effective to interfere in the processes of cellular transport while higher drugs concentrations were necessary in order to result in any damage to cell integrity. The methodology described in this work might be useful for studies that aim to evaluate the viability of nematodes, particularly for testing of new anthelminthic compounds using an easy, economic, reproducible, and no time-consuming technique.

Show MeSH
Related in: MedlinePlus