Analysis of expressed sequence tags from subtracted and unsubtracted Ctenocephalides felis hindgut and Malpighian tubule cDNA libraries.
Insect hindgut and Malpighian tubule (HMT) tissues regulate the contents of the haemolymph through the excretion of waste products and the specific reabsorption of nutrients. As such, they perform a role that is essential for survival and may contain molecular targets for insect control strategies. In order to discover genes expressed in the HMT tissues of the cat flea, Ctenocephalides felis, expressed sequence tags (ESTs) were generated from an unsubtracted HMT cDNA library and from a subtracted HMT cDNA library that had been enriched for HMT-specific cDNAs. A total of 4844 ESTs were analysed from both libraries: 3657 from the subtracted library and 1187 from the unsubtracted library. Of the 1418 distinct ESTs identified from both libraries, 953 had significant similarity to other sequences reported in the GenBank database. A comparison of the results from the two libraries confirmed that the percentages of genes likely to be involved with metabolism, cell structure, and digestion were reduced by the subtraction procedure, whereas genes likely to be involved with ion transport were enriched. Analysis of the prevalence of three individual cDNAs in each library revealed that the actin cDNA was reduced in the subtracted library whereas the cDNAs encoding allantoinase and a peritrophin-like protein were greatly enriched in the subtracted library. Northern blot analysis demonstrated that the actin cDNA was expressed in both the HMT and carcass tissues, whereas the allantoinase and peritrophin-like cDNAs were detected exclusively in the HMT tissues. In total, 97 distinct ESTs that appear to encode proteins involved with ion transport were analysed. Some of these proteins may be directly involved with diuresis or the specific reabsorption of salts and nutrients, and thus may be potential molecular targets for flea control strategies.
Gaines
P J
PJ
Heska Corporation, 1613 Prospect Parkway, Fort Collins, CO 80525, USA. gainesp@heska.com
Brandt
K S
KS
Eisele
A M
AM
Wagner
W P
WP
Bozic
C M
CM
Wisnewski
N
N
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D020224:Expressed Sequence Tags; D005260:Female; D015723:Gene Library; D007473:Ion Channels / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D005423:Siphonaptera / Q000235:genetics* / Q000378:metabolism
D018076:DNA, Complementary; D007473:Ion Channels; D012333:RNA, Messenger