Nancy Wisnewski

Characterization of novel fucosyl- and tyvelosyl-containing glycoconjugates from Trichinella spiralis muscle stage larvae

The monosaccharide composition of an affinity-purified family of antigenically-related Trichinella spiralis larval glycoproteins was determined by gas chromatography/mass spectrometry. This group of 6 major glycoproteins, designated TSL-1, originates in the muscle stage (L1) larval stichosome. They are present on the L1 surface and in excretory/secretory products of L1 larvae, are stage-specific, and are highly immunodominant. The glycosyl composition of the TSL-1 antigens was remarkable in 2 respects: (1) fucose accounted for 36 molar percent of the glycosyl residues; and (2) a 3,6-dideoxyhexose was identified, which accounted for at least 24 molar percent of the glycosyl residues. Previously, 3,6-dideoxyhexoses have been found only in certain Gram-negative bacterial lipopolysaccharides and in ascaroside alcohols (ascarylose) of Ascaris eggs. The 3,6-dideoxyhexose found in the TSL-1 antigens also was found in ES. This Trichinella sugar has been chemically identified as a 3,6-dideoxyarabinohexose, the same as found in Ascaris eggs. However, the absolute configuration of the TSL-1 sugar is D-(tyvelose), not L-(ascarylose) as is found in Ascaris eggs. Methylation analysis indicated that the TSL-1 3,6-dideoxy-D-arabinohexose was present entirely as non-reducing terminal residues. Approximately 83% of the fucose was also present as non-reducing terminal residues, with the remaining fucose found as 3,4-linked branched residues.

Antibody to the Dirofilaria immitis aspartyl protease inhibitor homologue is a diagnostic marker for feline heartworm infections.

Feline heartworm disease, caused by the filarial nematode Dirofilaria immitis, has been diagnosed with increased frequency in areas endemic for canine heartworm infection. The routine methods for determining the infection status of dogs, such as identification of circulating microfilariae in blood or identification of circulating antigen in serum, plasma or blood, have proven inadequate for screening cats. The inadequacies are due to the likelihood of single-sex infections and clinical disease during prepatent infections. Current antibody detection methodologies rely on crude or partially purified worm antigen preparations that may result in poor specificity. This report describes the cloning, expression, and diagnostic utility of the D. immitis homologue (PDi33) of the Onchocerca volvulus aspartyl protease inhibitor (Ov33). PDi33 is present in all stages that occur in the mammalian host (microfilariae, L3, L4, adult males, and females) and is released by adults cultured in vitro. An indirect enzyme-linked immunosorbent assay (ELISA) using antibody to recombinant PDi33 as a diagnostic marker for infection in cats was very sensitive and was useful for identifying prepatent infections. Testing of sera from cats infected with common gastrointestinal parasites also indicated excellent specificity. The same ELISA in dogs, although demonstrating reasonable sensitivity and specificity, appeared to be of less value as compared with the currently accepted antigen detection methodologies.

Vaccine Research and Development for the Prevention of Filarial Nematode Infections

The development of vaccines for the prevention of filarial nematode infections is in a state of relative infancy in comparison to vaccines for other parasitic diseases, such as schistosomiasis and malaria. There are many reasons for this slow start. Some of the principal problems are: (1) the lengthy and complex life cycle of these organisms with attendant complex immune responses, (2) the unique characteristics associated with a relatively large number of different pathogens, (3) the lack of suitable model systems for study of medically important infections, (4) the paucity of parasite material for antigen discovery and recombinant library construction, (5) the lack of substantial evidence suggesting the natural occurrence of protective immune responses, and (6) the limited data on mechanisms responsible for protective immunity. As technical hurdles are considered, it is also critical to focus on the characteristics of a vaccine necessary for its eventual utility. In the case of a vaccine for D. immitis a completely successful product will need to approach a 99+% efficacy. This is because of the 99+% efficacy of competitive chemotherapeutic products and the fact that microfilaremia observed on blood examination, resulting from as few as two worms, would present as a vaccine failure. Although very low worm burdens in large dogs could be perceived as success in the context of protection from clinical disease, because of the option of virtually complete chemoprophylactic protection, the typical veterinary practitioner would probably fail to appreciate less than complete vaccine protection. In contrast, a vaccine that produced a reduction in adult worm burdens without complete protection in either lymphatic filariasis or onchocerciasis would be very important. Highly effective chemoprophylactic agents are not widely available for prevention of the human filariases, and dramatically reduced clinical disease provided by less than a completely effective vaccine could occur as the result of fewer adult worms. The importance of developing these vaccines has outweighed the obstacles to this research. There has been a great deal of epidemiological and experimental evidence to suggest a vaccine is feasible and antigen discovery has progressed relatively rapidly within just the past few years. Efforts to generate appropriate larval cDNA libraries are beginning to yield dividends and a variety of fascinating vaccine candidates have been cloned. Additional antigen discovery, research on appropriate modalities for overexpression of genes from these parasites, and the complex tasks associated with vaccinology remain as significant research and development obstacles.(ABSTRACT TRUNCATED AT 400 WORDS)