First Blindness Cases of Horses Infected with Setaria Digitata (Nematoda: Filarioidea) in the Republic of Korea

Ocular setariases of cattle were reported but those of equine hosts have never been reported in the Republic of Korea (Korea). We found motile worms in the aqueous humor of 15 horses (Equus spp.) from 12 localities in southern parts of Korea between January 2004 and November 2017. After the affected animals were properly restrained under sedation and local anesthesia, 10 ml disposable syringe with a 16-gauge needle was inserted into the anterior chamber of the affected eye to successfully remove the parasites. The male worm that was found in 7 of the cases showed a pair of lateral appendages near the posterior terminal end of the body. The papillar arrangement was 3 pairs of precloacal, a pair of adcloacal, and 3 pairs of postcloacal papillae, plus a central papilla just in front of the cloaca. The female worms found in the eyes of 8 horses were characterized by the tapering posterior terminal end of the body with a smooth knob. Worms were all identified as Setaria digitata (von Linstow, 1906) by the morphologic characteristics using light and electron microscopic observations. This is the first blindness cases of 15 horses infected with S. digitata (Nematoda: Filarioidea) in Korea.

Heterologous expression, chaperone mediated solubilization and purification of parasitic nematode-specific growth factor-like protein of Setaria digitata

OBJECTIVE@#To clone, express and purify a putative parasitic nematode specific protein of Setaria digitata (S. digitata), filarial nematode that infects livestock and cause significant economic losses in Far East and Asia to be used for structural and functional analyses.@*METHODS@#To characterize uncharacterized gene of S. digitata (SDUG), the herterologous expression of SDUG was carried out in the pET [cloned into pET45b(+)] expression system initially and co-expression of SDUG using chaperone plasmids pG-KJE8, pGro 7, pKJE7, pG-Tf2 and pTf16 containing chaperone proteins of dnaK-dnaJ-grpE-groES-gro-E, groES-groEL, dnaK-dnaJ-grpE, groES-groEL-tig, and tig respectively, was carried out subsequently.@*RESULTS@#Expression of SDUG was seen when Escherichia coli strain BL21(DE3) is used, while concentrating protein largely into the insoluble fraction. The co-expression of SDUG using chaperone plasmid mediated system indicated a significant increase of the protein in the soluble fraction. Of the chaperon plasmid sets, the highest amount of recombinant SDUP in the soluble fraction was seen when pGro7 was used in the presence of 2 mg/mL L-arabinose and 0.6M IPTG concentration in the culture medium and for 3 h of incubation at the temperature of 28 °C. Recombinant SDUG was purified both from soluble and insoluble fractions using Ni affinity chromatography. SDS-PAGE and western blot analyses of these proteins revealed a single band having expected size of ∼24 kDa.@*CONCLUSIONS@#SDUG seems to be more aggregate-prone and hydrophobic in nature and such protein can make soluble by correct selecting the inducer concentrations and induction temperature and its duration.

Antigenicity of filarial superoxide dismutase in human bancroftian filariasis.

Superoxide dismutase activity was measured in different stages of growth of filarial parasites (human and cattle). The activity was almost undetected or very low in microfilarial stage but in adult worms, the enzyme activity was high. The enzyme was characterized to be a Cu/Zn superoxide dismutase. Most of the enzyme activity was associated with a detergent extractable fraction of adult (Setaria) parasite. The enzyme was also detected in the in vitro released products of adult worms. The superoxide dismutase activity was completely inhibited with IgG antibody from chronic filarial patients in contrast to IgG from normal people. Filarial patients particularly have high IgG and IgM antibody levels to purified enzyme. However, individuals from non-filarial regions of Orissa are sero-negative for superoxide dismutase antibodies. Antibody response to superoxide dismutase could thus be used for filarial diagnosis.

Synthesis and release of proteins homologous to excretory-secretory antigens during embryogenesis of Setaria digitata.

Localization of antigens homologous to excretory-secretory proteins in developing embryos of Setaria digitata has been carried out by indirect fluorescent antibody test, [14C] labelling studies and Western blotting. Indirect fluorescent antibody test showed binding of excretory-secretory antibodies at perivitelline space. The fluorescent antibody binding was almost absent at small morulae stage and increasing in intensity in the successive developmental stages with maximum at coiled microfilaria stage. Hatched microfilaria did not show the presence of antigens by immunofluorescence. Immunocomplex of excretory-secretory antiserum against "amniotic fluid" collected from developing embryos of Setaria digitata labelled with [14C] amino acids showed highest radioactivity at coiled and tadpole stages and differed significantly from small morulae, big morulae and hatched microfilaria. Immunoblot analysis of amniotic fluid showed two proteins, 16·5 and 11 kDa, to be highly antigenic. The antigenic protein (11 kDa) content as seen by immuno blotting increased during embryogenesis and decreased at the stage of hatching.

Immune response to an allergenic fraction of Setaria digitata in human filariasis.

A low molecular weight antigenic fraction has been isolated from saline-soluble extracts of cattle filarial parasite Setaria digitata. This glycoprotein fraction (Fr III) which appears to have low phosphorylcholine content cross-reacted with infective larval (L3) antigens of Wuchereria bancrofti. Binding of human chronic filarial serum with L3 antigens could be inhibited partially by Fr III. The fraction elicited immediate cutaneous hypersensitivity reaction in people living in Wuchereria bancrofti endemic regions. The proportion of skin test positive cases was found to be highest in endemic normals in contrast to infected cases. IgE levels were however not different in chronic filariasis and in endemic normals (or in asymptomatic microfilaraemic carriers). On the other hand, specific IgG level was considerably enhanced only in chronic filariasis.

Phosphoenolpyruvate-succinate-glyoxylate pathway in the filarial parasite Setaria digitata.

Setaria digitata, a filarial parasite of cattle possesses certain unique characteristics like cyanide insensitivity, and lack of cytochromes. In the present study, we have shown that the parasite has an incomplete tricarboxylic acid cycle with the absence of activities of isocitrate dehydrogenase, α-ketoglutarate dehydrogenase and succinyl-CoA synthase. However the parasite showed the existence of glyoxylate cycle and phosphoenolpyruvate-succinate pathway. The widely used antifilarial drug diethylcarbamazine caused general inhibition of all enzymes of phosphoenolpyruvate-succinate pathway and glyoxylate cycle except that of fumarase and isocitrate lyase. The results may pave the way for new targets for chemotherapy in the control of filarial parasites.