Transcriptome Sequencing Analysis and Functional Identification of Sex Differentiation Genes from the Mosquito Parasitic Nematode, Romanomermis wuchangensis.
Mosquito-transmitted diseases like malaria and dengue fever are global problem and an estimated 50-100 million of dengue or dengue hemorrhagic fever cases are reported worldwide every year. The mermithid nematode Romanomermis wuchangensis has been successfully used as an ecosystem-friendly biocontrol agent for mosquito prevention in laboratory studies. However, this nematode can not undergo sex differentiation in vitro culture, which has seriously affected their application of biocontrol in the field. In this study, based on transcriptome sequencing analysis of R. wuchangensis, Rwucmab-3, Rwuclaf-1 and Rwuctra-2 were cloned and used to investigate molecular regulatory function of sex differentiation. qRT-PCR results demonstrated that the expression level of Rwucmab-3 between male and female displayed obvious difference on the 3rd day of parasitic stage, which was earlier than Rwuclaf-1 and Rwuctra-2, highlighting sex differentiation process may start on the 3rd day of parasitic stage. Besides, FITC was used as a marker to test dsRNA uptake efficiency of R. wuchangensis, which fluorescence intensity increased with FITC concentration after 16 h incubation, indicating this nematode can successfully ingest soaking solution via its cuticle. RNAi results revealed the sex ratio of R. wuchangensis from RNAi treated groups soaked in dsRNA of Rwucmab-3 was significantly higher than gfp dsRNA treated groups and control groups, highlighting RNAi of Rwumab-3 may hinder the development of male nematodes. These results suggest that Rwucmab-3 mainly involves in the initiation of sex differentiation and the development of male sexual dimorphism. Rwuclaf-1 and Rwuctra-2 may play vital role in nematode reproductive and developmental system. In conclusion, transcript sequences presented in this study could provide more bioinformatics resources for future studies on gene cloning and other molecular regulatory mechanism in R. wuchangensis. Moreover, identification and functional analysis of sex differentiation genes may clarify the sex differentiation mechanism of R. wuchangensis, which are helpful to solve the uncompleted sex differentiation problem in vitro culture and the potential large-scale field application controlling the larvae of C. quinquefasciatus, A. aegypti and A. albopictus.
Hubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Sciences, Central China Normal University, Wuhan, China.
Mingyue
M
Duan
Hubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Sciences, Central China Normal University, Wuhan, China.
Jinfeng
J
Xiong
Hubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Sciences, Central China Normal University, Wuhan, China.
Dandan
D
Lu
Hubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Sciences, Central China Normal University, Wuhan, China.
Guoxiu
G
Wang
Hubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Sciences, Central China Normal University, Wuhan, China.
Hui
H
Ai
Am J Trop Med Hyg. 2007 Sep;77(3):467-77
17827362
Mol Biochem Parasitol. 2003 Jun;129(1):41-51
12798505
Genes Dev. 2000 Apr 15;14(8):901-6
10783162
Science. 1998 Oct 16;282(5388):430-1
9841401
PLoS One. 2012;7(1):e29811
22253787
PLoS One. 2014 May 20;9(5):e97924
24844228
Proc Natl Acad Sci U S A. 2013 Oct 1;110(40):16033-8
24046365
Nature. 1998 Feb 19;391(6669):806-11
9486653
Development. 2000 Oct;127(20):4469-80
11003845
Nature. 1998 Feb 12;391(6668):691-5
9490411
Trends Genet. 2012 Apr;28(4):175-84
22425532
Indian J Med Res. 1978 Aug;68:242-7
31332
PLoS Genet. 2011 Mar;7(3):e1002010
21408209
Development. 1995 Sep;121(9):2995-3004
7555725
Dev Biol. 2009 Jun 15;330(2):358-67
19361491
Cell. 1995 May 19;81(4):611-20
7758115
Genes Dev. 1999 Jun 1;13(11):1453-63
10364161
J Nematol. 2007 Jun;39(2):145-52
19259483
Adv Parasitol. 1985;24:307-44
3904347
Mol Biochem Parasitol. 2004 Dec;138(2):165-70
15555728
Genetics. 1988 May;119(1):43-61
3396865
Exp Parasitol. 2013 Feb;133(2):166-78
23201220
Science. 2010 Oct 1;330(6000):86-8
20929810
Nat Protoc. 2008;3(6):1101-8
18546601
Pest Manag Sci. 2014 Apr;70(4):623-7
23788440
Nat Biotechnol. 2011 May 15;29(7):644-52
21572440
PLoS Negl Trop Dis. 2015 May 28;9(5):e0003808
26020627
Genetics. 1986 Sep;114(1):53-76
3770471
J Biol Chem. 2011 Mar 25;286(12):10803-13
21262963
EMBO Rep. 2001 Oct;2(10):899-904
11600454
Parasitol Res. 2012 Apr;110(4):1435-43
21947308
Development. 1997 Feb;124(3):749-58
9043090
Mol Plant Pathol. 2009 May;10(3):389-401
19400841
Nature. 1998 Oct 29;395(6705):854
9804418
Pest Manag Sci. 2015 Nov;71(11):1471-85
26037532
Genes Dev. 2000 Dec 15;14(24):3153-65
11124807
Parasitol Res. 2013 Mar;112(3):1299-304
23306387
Parasitol Res. 2006 Sep;99(4):466-72
16642386
Am J Trop Med Hyg. 1978 Nov;27(6):1268-73
727331
J Nematol. 1992 Sep;24(3):324-9
19283004
Parasit Vectors. 2015 Feb 10;8:95
25880395
Genetics. 2000 May;155(1):105-16
10790387
J Nematol. 1981 Jul;13(3):266-74
19300761
BMC Genomics. 2013 Dec 27;14:923
24373391
Genes Dev. 2002 Sep 15;16(18):2390-402
12231628
Science. 2010 Oct 1;330(6000):88-90
20929811
Cell. 1988 Sep 23;54(7):1019-31
3046751
PLoS One. 2015 Oct 30;10(10):e0141208
26517714
Int J Parasitol. 2005 Aug;35(9):935-40
16023650