Results suggest that hematophagous transfer of Mhm and Mhf into fleas occurred and that C felis is a possible vector for Mhf via hematophagous activity.

The larval stage of the intestinal nematode, Trichinella spiralis, secretes and displays on its cuticle a number of antigenically cross-reactive glycoproteins. These so-called TSL-1 antigens induce a powerful antibody response in parasitized animals. In rats, anti-TSL-1 antibodies mediate a protective immunity that expels invading larvae from the intestine. The vast majority of anti-TSL-1 antibodies are specific for glycans. Although the biological functions of TSL-1 antigens are not known, the powerful effect of glycan-specific antibodies on the intestinal survival of T. spiralis suggests that they play an important role in parasite establishment. Little is known about the structures of the glycans present on the TSL-1 glycoproteins. Recent studies have suggested, however, that the antigens contain very unusual glycans (Wisnewski, N., McNeil, M., Grieve, R.B. and Wassom, D.L., Mol. Biochem. Parasitol., 61, 25-36, 1993). Sugar and linkage analysis of the combined secreted products unexpectedly showed that a major terminal sugar is tyvelose (3,6-dideoxy-D-arabino-hexose; Tyv) which has previously been found only in certain gram-negative bacterial lipopolysaccharides. In this paper, we report the first rigorous structural study of oligosaccharides released from TSL-1 antigens by peptide N-glycosidase F digestion. Using strategies based on fast atom bombardment mass spectrometry (FAB-MS), we have discovered a novel family of tri- and tetra-antennary N-glycans whose antennae are comprised of the tyvelose-capped structure: Tyv1,3GalNAc beta 1,4(Fuc alpha 1,3)GlcNAc beta 1-. Thus a major population of TSL-1 glycans contains clusters of hydrophobic terminal structures which are likely to be highly immunogenic.

No abstract

No abstract

cDNAs encoding two different epoxide hydrolases (nCfEH1 and nCfEH2) were cloned from a cDNA library prepared from the wandering larval stage of the cat flea, Ctenocephalides felis. Predicted translations of the open reading frames indicated the clones encoded proteins of 464 (CfEH1) and 465 (CfEH2) amino acids. These proteins have a predicted molecular weight of 53 kDa and a putative 22 amino acid N-terminal hydrophobic membrane anchor. The amino acid sequences are 77% identical, and both are homologous to previously isolated epoxide hydrolases from Manduca sexta, Trichoplusia ni, and Rattus norvegicus. Purification of native juvenile hormone epoxide hydrolase (JHEH) from unfed adult cat fleas generated a partially pure protein that hydrolyzed juvenile hormone III to juvenile hormone III-diol. The amino terminal sequence of this;50-kDa protein is identical to the deduced amino terminus of the protein encoded by the nCfEH1 clone. Affinity-purified rabbit polyclonal antibodies raised against Escherichia coli-expressed HisCfEH1 recognized a approximately 50-kDa protein present in the partially purified fraction containing JHEH activity. Immunohistochemistry experiments using the same affinity-purified rabbit polyclonal antibodies localized the epoxide hydrolase in developing oocytes, fat body, and midgut epithelium of the adult flea. The presence of JHEH in various flea life stages and tissues was assessed by Northern blot and enzymatic activity assays. JHEH mRNA expression remained relatively constant throughout the different flea larval stages and was slightly elevated in the unfed adult flea. JHEH enzymatic activity was highest in the late larval, pupal, and adult stages. In all stages and tissues examined, JHEH activity was significantly lower than juvenile hormone esterase (JHE) activity, the other enzyme responsible for JH catalysis.

No abstract

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.

No abstract