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Use of microarray hybridization to identify Brugia genes involved in mosquito infectivity

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Abstract

Brugia malayi and Brugia pahangi microfilariae (mf) require a maturation period of at least 5 days in the mammalian host to successfully infect laboratory mosquitoes. This maturation process coincides with changes in the surface composition of mf that likely are associated with changes in gene expression. To test this hypothesis, we verified the differential infectivity of immature (≤3 day) and mature (>30 day) Brugia mf for black-eyed Liverpool strain of Aedes aegypti and then assessed transcriptome changes associated with microfilarial maturation by competitively hybridizing microfilarial cDNAs to the B. malayi oligonucleotide microarray. We identified transcripts differentially abundant in immature (94 in B. pahangi and 29 in B. malayi) and mature (64 in B. pahangi and 14 in B. malayi) mf. In each case, >40% of Brugia transcripts shared no similarity to known genes or were similar to genes with unknown function; the remaining transcripts were categorized by putative function based on sequence similarity to known genes/proteins. Microfilarial maturation was not associated with demonstrable changes in the abundance of transmembrane or secreted proteins; however, immature mf expressed more transcripts associated with immune modulation, neurotransmission, transcription, and cellular cytoskeleton elements, while mature mf displayed increased transcripts potentially encoding hypodermal/muscle and surface molecules, e.g., cuticular collagens and sheath components. The results of the homologous B. malayi microarray hybridization were validated by quantitative reverse transcriptase polymerase chain reaction. These findings preliminarily lend support to the underlying hypothesis that changes in microfilarial gene expression drive maturation-associated changes that influence the parasite to develop in compatible vectors.

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Abbreviations

mf:

Microfilariae

LVP:

Black-eyed Liverpool strain of Aedes aegypti

LF:

Lymphatic filariasis

P/S:

100 μg/mL penicillin and 100 μg/mL streptomycin

PE:

Post-exposure

ESTs:

Expressed sequence tags

TIGR:

The Institute for Genomic Research

BLAST:

Basic local alignment search tool

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Acknowledgments

We are grateful for the assistance with worm transplants provided by Marty Greer and Jessica Nerenhausen. We also thank Juliet Fuhrman, Steven Williams, and Lori Saunders for helpful discussion. This project was funded by National Institutes of Health grants 1 R15 AI067295-01A1 and AI 19769. Microfilaraemic blood and infected jirds were supplied by the Filariasis Research Reagent Repository Center at the University of Georgia. K. Griffiths is a recipient of a University of Wisconsin-Oshkosh 2008 Graduate Student Collaborative Research Grant. All animal protocols were reviewed and approved by UWO and UWM Institutional Animal Care and Use Committees.

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Department of Biology and Microbiology, University of Wisconsin-Oshkosh, 800 Algoma Blvd, Oshkosh, WI, 54901, USA

    Kathryn G. Griffiths, Rebecca L. Zink, Colleen M. McDermott & Michelle L. Michalski

  2. Department of Pathobiological Sciences, University of Wisconsin Madison, Madison, WI, USA

    George F. Mayhew, Sara M. Erickson, Jeremy F. Fuchs & Bruce M. Christensen

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  1. Kathryn G. Griffiths
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  2. George F. Mayhew
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  3. Rebecca L. Zink
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  4. Sara M. Erickson
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  5. Jeremy F. Fuchs
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  6. Colleen M. McDermott
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  7. Bruce M. Christensen
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Corresponding author

Correspondence to Michelle L. Michalski.

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Supplemental Table 1

B. malayi immature mf differentially abundant transcripts (DOC 16 kb)

Supplemental Table 2

B. malayi mature mf differentially abundant transcripts (DOC 15 kb)

Supplemental Table 3

B. pahangi immature mf differentially abundant transcripts (DOC 29 kb)

Supplemental Table 4

B. pahangi mature mf differentially abundant transcripts (DOC 26 kb)

Supplemental Table 5

Abundant immature mf transcripts from B. malayi that share sequence similarity on the peptide level with C. elegans genes displaying RNAi phenotypes. Source: Wormpep (DOC 13 kb)

Supplemental Table 6

Abundant mature mf transcripts from B. malayi that share sequence similarity on the peptide level with C. elegans genes displaying RNAi phenotypes (DOC 13 kb)

Supplemental Table 7

Abundant immature mf transcripts from B. pahangi that share sequence similarity on the peptide level with C. elegans genes displaying RNAi phenotypes (DOC 17 kb)

Supplemental Table 8

Abundant mature mf transcripts from B. pahangi that share sequence similarity on the peptide level with C. elegans genes displaying RNAi phenotypes (DOC 16 kb)

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Griffiths, K.G., Mayhew, G.F., Zink, R.L. et al. Use of microarray hybridization to identify Brugia genes involved in mosquito infectivity. Parasitol Res 106, 227–235 (2009). https://doi.org/10.1007/s00436-009-1655-y

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