Previous studies have shown that the secreted phosphorylcholine-containing glycoprotein of filarial nematodes, ES-62, is only present in the post-infective life-cycle stages, but that the mRNA is transcribed throughout the worm's life-cycle. The aim of this current study was to investigate whether the presence or absence of protein expression simply reflects differences in mRNA abundance. To this end, we investigated the relative abundance of ES-62 using TaqMan real time RT-PCR, in different life-cycle stages of 2 model filarial nematode parasites, Acanthocheilonema viteae and Brugia pahangi. For B. pahangi, microfilariae, infective larvae and adult worms were each found to have approximately similar levels of ES-62 mRNA. However, the corresponding stages of A. viteae differed greatly from each other with a pattern of increased mRNA production with maturation. As a rule A. viteae had higher levels of ES-62 mRNA than B. pahangi, and this was particularly noticeable in the adult stage where the difference was approximately 3500-fold higher. However, this significant difference in mRNA abundance was not reflected in the quantity of ES-62 protein secreted by the adult worms of each species, as A. viteae only secreted ~3 times as much ES-62 as B. pahangi. Thus, overall, the results obtained from this study indicate that ES-62 protein production does not solely reflect mRNA levels, and also suggest that the 2 nematodes may employ different mechanisms for regulating protein production.

Infection with the endosymbiotic bacteria Wolbachia is widespread in filarial nematodes. Previous studies have suggested concordance between the phylogeny of Wolbachia with that of their nematode hosts. However, there is only one published molecular phylogenetic study of filarial species, based on the 5S rRNA gene spacer. The phylogeny proposed by this study is partially incongruent with previous classifications of filarial nematodes, based on morphological characters. Furthermore, both traditional classifications and molecular phylogenies are, in part, inconsistent with the phylogeny of Wolbachia. Here we report mitochondrial cytochrome oxidase I (COI) gene sequences for 11 species of filaria and for another spirurid nematode which was included as an outgroup. In addition, 16S rRNA, wsp and ftsZ gene sequences were generated for the Wolbachia of several filarial species, in order to complete the available data sets and further resolve the phylogeny of Wolbachia in nematodes. We used these data to evaluate whether nematode and Wolbachia phylogenies are concordant. Some of the possible phylogenetic reconstructions based on COI gene were congruent with the phylogeny of Wolbachia and supported the grouping of the rodent filaria Litomosoides sigmodontis with the lymphatic filariae (i.e. Brugia spp. and Wuchereria spp.) and the sister group relationship of Dirofilaria spp. and Onchocerca spp. However, the placement of the Wolbachia-free filaria Acanthocheilonema viteae is ambiguous and dependent on the phylogenetic methods used.

The filarial nematodes Wuchereria bancrofti, Brugia malayi and Onchocerca volvulus represent major public health problems in the Tropics. Effective diagnosis of infection with these parasites is required both for administration of drugs to infected individuals and for monitoring of control programs. However parasitological diagnosis is associated with a number of problems including frequently inadequate sensitivity, long pre-patency of infection and inconvenience for patients. For these reasons there has been considerable effort expended in developing other forms of diagnosis, in particular immunoassays for measuring antibody and circulating parasite antigen as well as molecular-biology-based assays for detecting parasite DNA. This article reviews the progress and achievements obtained to date. The latter include the development of ELISAs employing recombinant antigen for detection of antibody to O. volvulus which have both high sensitivity and specificity, the commercial availability of immunoassays to measure circulating antigen in W. bancrofti infection and the generation of specific DNA-based detection systems for all three parasites.

The mechanism of linkage of phosphorylcholine (PC) to excretory–secretory products (ES) of adult Brugia pahangi has been investigated. Biosynthetic radio-isotope labelling of ES with [3H]choline followed by SDS–PAGE/fluorography revealed a smear of molecular weight approximately 40–100 kDa which loses its radiolabel following exposure to N-glycosidase F, but not mild alkali. PC is thus attached to this smear of molecules via N-type glycans, a mechanism of linkage previously observed with respect to PC–ES of Acanthocheilonema viteae. Western blotting analysis of non-radiolabelled ES demonstrated the existence of additional PC–ES which were insensitive to N-glycosidase F, but not to alkali. This second group of molecules is therefore likely to contain PC linked to O-glycans. Filarial nematodes may thus utilize 2 classes of glycan for attachment of PC. Examination of B. pahangi and A. viteae whole worm extracts by Western blotting indicated that their PC content could not be cleaved by N-glycosidase F and hence the use of N-type glycans may be restricted to a subset of ES products. The implications of these findings with respect to developing inhibitors of PC attachment for use as anti-filarial drugs are discussed.