Impairment of larval development and a sex-ratio distortion in Litomosoides sigmodontis caused by Wolbachia-depleted microfilariae and localization of a filarial nematode phosphate permease up-regulated during Wolbachia depletion
Impairment of larval development and a sex-ratio distortion in Litomosoides sigmodontis caused by Wolbachia-depleted microfilariae and localization of a filarial nematode phosphate permease up-regulated during Wolbachia depletion
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DissertationPrüfungsdatum
01.04.2009Datum der Veröffentlichung
07.04.2009Erstgutachter
Sahl, Hans-GeorgZweitgutachter
Hoerauf, AchimMetadaten
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Inhalt
The thesis investigates the endosymbiosis between Wolbachia bacteria and their host filarial nematodes. The rodent filaria Litomosoides sigmodontis harbour Wolbachia endosymbionts and the pattern of infection and migration of these parasites mimics that of human filarial parasites such as Wuchereria bancrofti and Onchocerca volvulus. The first aim of the thesis was to study the effect of tetracycline, which depletes Wolbachia, on the development of microfilariae (L1 larvae, MF) to L3 larvae in the intermediate mite host Ornithonyssus bacoti, and to observe the development of Wolbachia-depleted L3 larvae in Mongolian gerbils (Meriones unguiculatus). Microfilaremic gerbils were treated with tetracycline for 6 weeks in drinking water (1º Tet) or left untreated (1º Con). Six weeks post-treatment, there was no difference in microfilaremia between 1º Tet and 1º Con, but there was significantly lower levels of Wolbachia in MF from 1º Tet in comparison to those in 1º Con. Naive mites were then fed on the 1º Tet and 1º Con gerbils and were used to infect uninfected gerbils (2º Tet, 2º Con). The number of MF that developed into L3 larvae in naive mites after feeding on the Wolbachia-depleted MF in 1º Tet gerbils was significantly lower than controls. Fewer female worms developed in 2º Tet gerbils but there was no significant difference in the number of male worms that developed in 2º Tet and 2º Con gerbils, resulting in a male biased sex-ratio. This suggests that Wolbachia have more influence on the development of female than male worms which is supported by the fact that male worms with significantly fewer Wolbachia could be selected by tetracycline treatment but female worms cannot. Since the worms analyzed were only exposed to tetracycline as MF, the experimental set-up rules out direct effects of tetracycline during larval development in the mites or 2º infected gerbils, suggesting that the initial loss of Wolbachia in MF was the cause of impaired larval development, not an antifilarial effect of tetracycline.
The second aim of the thesis was to study the molecular basis governing endosymbiosis between Wolbachia and their filarial host - L. sigmodontis. Previously in our research group, Heider et al., 2006 studied the gene expression of L. sigmodontis during Wolbachia depletion using differential display which revealed an up-regulation of a phosphate permease gene (Ls-ppe-1). We hypothesized that Ls-ppe-1 had an important role in nucleotide metabolism as depletion of Wolbachia induced expression of Ls-ppe-1, perhaps to compensate for lack of nucleotides in the absence of their endobacteria. To test this hypothesis, firstly, the regulation of phosphate permease during Wolbachia depletion was studied at the protein level in L. sigmodontis and O. volvulus, and secondly, the localization of phosphate permease and Wolbachia in L. sigmodontis and O. volvulus were investigated in untreated and antibiotic treated filarial worms. Results of the studies show the up-regulation of L. sigmodontis phosphate permease (Ls-PPE) both at the mRNA and protein levels and immunohistology results demonstrate that Ls-PPE is localized to areas of the worms that contain Wolbachia. Results also demonstrated the up-regulation of O. volvulus phosphate permease (Ov-PPE) at the protein level during Wolbachia depletion by doxycycline treatment of onchocerciasis. Ls-PPE and Ov-PPE are localized to areas in worms where Wolbachia are found and are up-regulated in response to Wolbachia depletion, supporting a role of phosphate permease in nematode-Wolbachia endosymbiosis. Further ultrastructural analysis using electron microscopy promises to bring insight into the molecular interaction between phosphate permeases and Wolbachia and RNAi mediated knockdown of Ls-ppe-1 in L. sigmodontis will elucidate the exact function of PPE in the Wolbachia-nematode symbiosis.
The second aim of the thesis was to study the molecular basis governing endosymbiosis between Wolbachia and their filarial host - L. sigmodontis. Previously in our research group, Heider et al., 2006 studied the gene expression of L. sigmodontis during Wolbachia depletion using differential display which revealed an up-regulation of a phosphate permease gene (Ls-ppe-1). We hypothesized that Ls-ppe-1 had an important role in nucleotide metabolism as depletion of Wolbachia induced expression of Ls-ppe-1, perhaps to compensate for lack of nucleotides in the absence of their endobacteria. To test this hypothesis, firstly, the regulation of phosphate permease during Wolbachia depletion was studied at the protein level in L. sigmodontis and O. volvulus, and secondly, the localization of phosphate permease and Wolbachia in L. sigmodontis and O. volvulus were investigated in untreated and antibiotic treated filarial worms. Results of the studies show the up-regulation of L. sigmodontis phosphate permease (Ls-PPE) both at the mRNA and protein levels and immunohistology results demonstrate that Ls-PPE is localized to areas of the worms that contain Wolbachia. Results also demonstrated the up-regulation of O. volvulus phosphate permease (Ov-PPE) at the protein level during Wolbachia depletion by doxycycline treatment of onchocerciasis. Ls-PPE and Ov-PPE are localized to areas in worms where Wolbachia are found and are up-regulated in response to Wolbachia depletion, supporting a role of phosphate permease in nematode-Wolbachia endosymbiosis. Further ultrastructural analysis using electron microscopy promises to bring insight into the molecular interaction between phosphate permeases and Wolbachia and RNAi mediated knockdown of Ls-ppe-1 in L. sigmodontis will elucidate the exact function of PPE in the Wolbachia-nematode symbiosis.
Klassifikation (DDC)
570 Biowissenschaften, BiologieArumugam, Sridhar: Impairment of larval development and a sex-ratio distortion in Litomosoides sigmodontis caused by Wolbachia-depleted microfilariae and localization of a filarial nematode phosphate permease up-regulated during Wolbachia depletion. - Bonn, 2009. - Dissertation, Rheinische Friedrich-Wilhelms-Universität Bonn.
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5N-17164
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5N-17164
@phdthesis{handle:20.500.11811/4058,
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5N-17164,
author = {{Sridhar Arumugam}},
title = {Impairment of larval development and a sex-ratio distortion in Litomosoides sigmodontis caused by Wolbachia-depleted microfilariae and localization of a filarial nematode phosphate permease up-regulated during Wolbachia depletion},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2009,
month = apr,
note = {The thesis investigates the endosymbiosis between Wolbachia bacteria and their host filarial nematodes. The rodent filaria Litomosoides sigmodontis harbour Wolbachia endosymbionts and the pattern of infection and migration of these parasites mimics that of human filarial parasites such as Wuchereria bancrofti and Onchocerca volvulus. The first aim of the thesis was to study the effect of tetracycline, which depletes Wolbachia, on the development of microfilariae (L1 larvae, MF) to L3 larvae in the intermediate mite host Ornithonyssus bacoti, and to observe the development of Wolbachia-depleted L3 larvae in Mongolian gerbils (Meriones unguiculatus). Microfilaremic gerbils were treated with tetracycline for 6 weeks in drinking water (1º Tet) or left untreated (1º Con). Six weeks post-treatment, there was no difference in microfilaremia between 1º Tet and 1º Con, but there was significantly lower levels of Wolbachia in MF from 1º Tet in comparison to those in 1º Con. Naive mites were then fed on the 1º Tet and 1º Con gerbils and were used to infect uninfected gerbils (2º Tet, 2º Con). The number of MF that developed into L3 larvae in naive mites after feeding on the Wolbachia-depleted MF in 1º Tet gerbils was significantly lower than controls. Fewer female worms developed in 2º Tet gerbils but there was no significant difference in the number of male worms that developed in 2º Tet and 2º Con gerbils, resulting in a male biased sex-ratio. This suggests that Wolbachia have more influence on the development of female than male worms which is supported by the fact that male worms with significantly fewer Wolbachia could be selected by tetracycline treatment but female worms cannot. Since the worms analyzed were only exposed to tetracycline as MF, the experimental set-up rules out direct effects of tetracycline during larval development in the mites or 2º infected gerbils, suggesting that the initial loss of Wolbachia in MF was the cause of impaired larval development, not an antifilarial effect of tetracycline.
The second aim of the thesis was to study the molecular basis governing endosymbiosis between Wolbachia and their filarial host - L. sigmodontis. Previously in our research group, Heider et al., 2006 studied the gene expression of L. sigmodontis during Wolbachia depletion using differential display which revealed an up-regulation of a phosphate permease gene (Ls-ppe-1). We hypothesized that Ls-ppe-1 had an important role in nucleotide metabolism as depletion of Wolbachia induced expression of Ls-ppe-1, perhaps to compensate for lack of nucleotides in the absence of their endobacteria. To test this hypothesis, firstly, the regulation of phosphate permease during Wolbachia depletion was studied at the protein level in L. sigmodontis and O. volvulus, and secondly, the localization of phosphate permease and Wolbachia in L. sigmodontis and O. volvulus were investigated in untreated and antibiotic treated filarial worms. Results of the studies show the up-regulation of L. sigmodontis phosphate permease (Ls-PPE) both at the mRNA and protein levels and immunohistology results demonstrate that Ls-PPE is localized to areas of the worms that contain Wolbachia. Results also demonstrated the up-regulation of O. volvulus phosphate permease (Ov-PPE) at the protein level during Wolbachia depletion by doxycycline treatment of onchocerciasis. Ls-PPE and Ov-PPE are localized to areas in worms where Wolbachia are found and are up-regulated in response to Wolbachia depletion, supporting a role of phosphate permease in nematode-Wolbachia endosymbiosis. Further ultrastructural analysis using electron microscopy promises to bring insight into the molecular interaction between phosphate permeases and Wolbachia and RNAi mediated knockdown of Ls-ppe-1 in L. sigmodontis will elucidate the exact function of PPE in the Wolbachia-nematode symbiosis.},
url = {https://hdl.handle.net/20.500.11811/4058}
}
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5N-17164,
author = {{Sridhar Arumugam}},
title = {Impairment of larval development and a sex-ratio distortion in Litomosoides sigmodontis caused by Wolbachia-depleted microfilariae and localization of a filarial nematode phosphate permease up-regulated during Wolbachia depletion},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2009,
month = apr,
note = {The thesis investigates the endosymbiosis between Wolbachia bacteria and their host filarial nematodes. The rodent filaria Litomosoides sigmodontis harbour Wolbachia endosymbionts and the pattern of infection and migration of these parasites mimics that of human filarial parasites such as Wuchereria bancrofti and Onchocerca volvulus. The first aim of the thesis was to study the effect of tetracycline, which depletes Wolbachia, on the development of microfilariae (L1 larvae, MF) to L3 larvae in the intermediate mite host Ornithonyssus bacoti, and to observe the development of Wolbachia-depleted L3 larvae in Mongolian gerbils (Meriones unguiculatus). Microfilaremic gerbils were treated with tetracycline for 6 weeks in drinking water (1º Tet) or left untreated (1º Con). Six weeks post-treatment, there was no difference in microfilaremia between 1º Tet and 1º Con, but there was significantly lower levels of Wolbachia in MF from 1º Tet in comparison to those in 1º Con. Naive mites were then fed on the 1º Tet and 1º Con gerbils and were used to infect uninfected gerbils (2º Tet, 2º Con). The number of MF that developed into L3 larvae in naive mites after feeding on the Wolbachia-depleted MF in 1º Tet gerbils was significantly lower than controls. Fewer female worms developed in 2º Tet gerbils but there was no significant difference in the number of male worms that developed in 2º Tet and 2º Con gerbils, resulting in a male biased sex-ratio. This suggests that Wolbachia have more influence on the development of female than male worms which is supported by the fact that male worms with significantly fewer Wolbachia could be selected by tetracycline treatment but female worms cannot. Since the worms analyzed were only exposed to tetracycline as MF, the experimental set-up rules out direct effects of tetracycline during larval development in the mites or 2º infected gerbils, suggesting that the initial loss of Wolbachia in MF was the cause of impaired larval development, not an antifilarial effect of tetracycline.
The second aim of the thesis was to study the molecular basis governing endosymbiosis between Wolbachia and their filarial host - L. sigmodontis. Previously in our research group, Heider et al., 2006 studied the gene expression of L. sigmodontis during Wolbachia depletion using differential display which revealed an up-regulation of a phosphate permease gene (Ls-ppe-1). We hypothesized that Ls-ppe-1 had an important role in nucleotide metabolism as depletion of Wolbachia induced expression of Ls-ppe-1, perhaps to compensate for lack of nucleotides in the absence of their endobacteria. To test this hypothesis, firstly, the regulation of phosphate permease during Wolbachia depletion was studied at the protein level in L. sigmodontis and O. volvulus, and secondly, the localization of phosphate permease and Wolbachia in L. sigmodontis and O. volvulus were investigated in untreated and antibiotic treated filarial worms. Results of the studies show the up-regulation of L. sigmodontis phosphate permease (Ls-PPE) both at the mRNA and protein levels and immunohistology results demonstrate that Ls-PPE is localized to areas of the worms that contain Wolbachia. Results also demonstrated the up-regulation of O. volvulus phosphate permease (Ov-PPE) at the protein level during Wolbachia depletion by doxycycline treatment of onchocerciasis. Ls-PPE and Ov-PPE are localized to areas in worms where Wolbachia are found and are up-regulated in response to Wolbachia depletion, supporting a role of phosphate permease in nematode-Wolbachia endosymbiosis. Further ultrastructural analysis using electron microscopy promises to bring insight into the molecular interaction between phosphate permeases and Wolbachia and RNAi mediated knockdown of Ls-ppe-1 in L. sigmodontis will elucidate the exact function of PPE in the Wolbachia-nematode symbiosis.},
url = {https://hdl.handle.net/20.500.11811/4058}
}
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