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Infection and Immunity, January 2008, p. 221-228, Vol. 76, No. 1
0019-9567/08/$08.00+0     doi:10.1128/IAI.00701-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Essential Role of Chitinase in the Development of the Filarial Nematode Acanthocheilonema viteae{triangledown}

Babila Tachu,{dagger} Smitha Pillai,{dagger} Richard Lucius,{dagger} and Thomas Pogonka*{dagger}

Department of Molecular Parasitology, Humboldt University Berlin, Berlin, Germany

Received 23 May 2007/ Returned for modification 27 June 2007/ Accepted 8 October 2007

Chitinases of pathogens have been proposed as potential targets of vaccines or specific inhibitors. We studied the genomic organization, transcript levels, developmental expression, and biological function of chitinases in the rodent filarial nematode Acanthocheilonema viteae, a model organism for human-pathogenic filarial worms. Characterization of nine genomic clones from an A. viteae phage library and Southern blot experiments revealed the existence of three different chitinase genes, two of which could theoretically yield functional transcripts. The deduced proteins of these genes had the common modular organization of family 18 chitinases. Northern blot experiments and rapid amplification of cDNA ends-PCR with adult worms and larval stages showed that only one gene is expressed, with high variation in transcript levels, as determined by real-time PCR. Chitinase transcript levels were lowest in the late male stage 4 larva (L4) and peaked in the stage 3 larva (L3), which was corroborated by Western blotting. RNA interference (RNAi) experiments showed that treatment of L3 and adult female worms with double-stranded RNA of chitinase inhibited molting of L3 worms and hatching of microfilariae. RNAi also led to the death of 50% of female worms, revealing the essential role of chitinase in the life cycle of filarial nematodes.

* Corresponding author. Mailing address: Department of Molecular Parasitology, Humboldt University Berlin, Philippstrasse 13, 10115 Berlin, Germany. Phone: (49)-30-20936053. Fax: (49)-30-20936051. E-mail: thomas.pogonka{at}rz.hu-berlin.de

{triangledown} Published ahead of print on 15 October 2007.

Editor: J. F. Urban, Jr.

{dagger} B. Tachu, S. Pillai, and T. Pogonka performed the experiments. B. Tachu and S. Pillai contributed equally to this work. R. Lucius and T. Pogonka designed the study and supervised the experiments. All authors together analyzed the data and contributed to the preparation of the manuscript.

Infection and Immunity, January 2008, p. 221-228, Vol. 76, No. 1
0019-9567/08/$08.00+0     doi:10.1128/IAI.00701-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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