Kurt P. Bottjer

CUTICULAR CARBOHYDRATES OF THREE NEMATODE SPECIES AND CHEMORECEPTION BY TRICHOSTRONGYLUS COLUBRIFORMIS

The cuticle of Haemonchus contortus, Nippostrongylus brasiliensis and Trichostrongylus colubriformis contained N-acetylgalactosamine and N-acetylglucosamine, based on the binding of fluorescein-labelled lectins. Binding of the lectins varied between the sexes and body regions of the nematodes. Treatment of male T. colubriformis with the lectin Lens culinaris agglutinin (LcA) reduced the feeding by helminths that was stimulated by histamine and the males response to their females pheromone, based on in vitro assays. Mannose residues may be involved in the helminths chemoreceptors for feeding and sexual attraction, based on the specific binding of LcA.

Trichostrongylus colubriformis: effect of anthelmintics on ingestion and oviposition.

Abstract The effects of the anthelmintics ivermectin, levaminsole, oxfendazole, piperazine citrate, pyrantel pamoate, tetramisole, and thiabendazole on ingestion and oviposition by Trichostrongylus colubriformis were determined. Six of the compounds reduced in vitro feeding at the tested doses while all drugs reduced in vivo feeding after treatment of the host. Additionally, in vitro or in vivo exposure to most anthelmintics decreased oviposition during subsequent in vitro assay. Invermectin had the most pronounced effect on in vivo and in vitro feeding and egg release. The neuromuscular activities of pharyngeal pumping and egg ejection may be suitable systems for rapid determinations of anthelmintic effects.

Effects of host serum on feeding by Trichostrongylus colubriformis (nematoda)

Summary The effect of host serum on in vitro feeding by Trichostrongylus colubriformis was studied by incubating adult helminths in goat serum containing the dye, Rhodamine B. The amount of dye ingested was determined by fluorometric analysis. Immune serum from goats infected with T. colubriformis suppressed helminth feeding, while normal serum from uninfected goats did not. Suppression of feeding by immune serum increased with the duration of the hosts infection. Heat‐inactivation (56°C) of immune serum did not affect its suppressive activity. Pre‐exposure of worms to immune serum decreased subsequent feeding activity. However, rigorous washing of helminths restored their feeding to levels that were similar to untreated worms. Indirect immunofluorescent studies with immune serum and FITC conjugated rabbit anti‐goat IgG demonstrated binding of immunoglobulin to the cuticle, stoma and excretory pore of whole worms. Feeding inhibition of immune serum was associated with IgG1 isotype. Results of the present studies indicated that IgG was responsible for in vitro suppression of T. colubriformis feeding and may be one effector of immunity to T. colubriformis in the goat.

Nippostrongylus brasiliensis: Feeding activity in the mouse

Abstract Nippostrongylus brasiliensis incorporated the fluorescent dye, Rhodamine B, while feeding in vivo. Uptake of dye in both sexes of helminth increased linearly from 30 to 120 min after the hosts were given dye per os. Feeding by female N. brasiliensis significantly exceeded that of the male at 4 and 5 days postinfection. Feeding declined in older helminths of both sexes. The density of helminths had no effect on their incorporation of dye in vivo. Feeding in male- and female-only groups of worms was similar to that seen in populations of mixed sexes. Feeding by helminths decreased during the first 36 h of food deprivation in the host, but increased during subsequent fasting of the host. Both sexes of N. brasiliensis resumed feeding within 15 min after the fasted hosts were fed. Growth of N. brasiliensis increased linearly from 4 to 7 days postinfection, based on dry weight. Seven-day-old and older females were significantly heavier than males.

Oviposition by Trichostrongylus colubriformis (Nematoda) in vitro

Summary Oviposition by females of the ruminant nematode Trichostrongylus colubriformis was influenced by in vitro conditions. Release of eggs declined linearly during incubation for 4.5 h. An average of 8.8 eggs was deposited by females during the first hour in vitro. Decreased pH, temperature, and osmolarity caused reduced production of eggs. Storage of females at 4°C had no effect on subsequent oviposition at 37°C. Oviposition was similar under light and dark conditions. Concomitant males or the females age did not influence production of eggs in vitro. The release of eggs in vitro was significantly decreased by octapamine and GABA while serotonin elevated oviposition. Thus, environmental stimuli from the host may partially modulate helminth fecundity.

Nippostrongylus brasiliensis: in vitro feeding activity.

Abstract Nippostrongylus brasiliensis incorporated the fluorescent dye, Rhodamine B, while feeding in vitro . Uptake of dye in both sexes of helminth was time-dependent with maximal incorporation occurring after 30 min. Feeding activity was elevated as the time of removal from the host increased from 1 to 8 h. Feeding was reduced in 4- or 5-day-old helminths when compared to 7-day-old or older worms. Feeding activity increased also after temperature increases, pH decreases and solute decreases. Feeding was similar under conditions of both light and dark. Feeding by males and females of N. brasiliensis decreased as the density of worms in single- or mixed-sex groups of worms was increased. Females fed significantly more than males under crowded conditions.

Characterization of and male adaptation to pheromone of femaleTrichostrongylus colubriformis (Nematoda)

Males of the ruminant nematodeTrichostrongylus colubriformis were significantly attracted to an incubate from their females during in vitro assay. Exposure of males to sera from uninfected and infected animals or selected neurotransmitters had no effect on their subsequent responsiveness to the females pheromone. In contrast, exposure to 1500 female-hours of pheromone decreased male responsiveness after moderate rinsing prior to bioassay. Rinsing of the males with larger volumes of Tyrodes solution prior to in vitro assay increased their subsequent response to the females pheromone. High-performance liquid chromatography yielded a presumptive pheromone peak with a fivefold increase in biological activity. This peak was soluble in alcohols and tetrahydrofuran, based on elution from reverse-phase Sep-Pak cartridges. Stability of the peak was increased by EDTA or ascorbic acid (10 mM). Storage for six weeks in ascorbic acid at 4 °C allowed recovery of 47.3% of the original material.

Feeding of Nippostrongylus brasiliensis in oxygen and serum

ornatum is 22 for females and 21 for males, and the sex determining mechanism is XX: XO. We thank Dr. W. J. Gladney for sending us specimens of A. inornatum from which our laboratory colony originated. Thanks are also extended to Dr. J. Mathews Pound for help with the figure and to him and Cheryl Morris for editorial and clerical assistance. This research was supported in part by Grant AI-09556 from the National Institute of Allergy and Infectious Diseases (NIH) and Contract N-00014-80-C-0546 from the Office of Naval Research.

Anthelmintic ingestion in nematodes: Enhancement by phagostimulants

Abstract Anthelmintic ingestion in nematodes: enhancement by phagostimulants. International Journal for Parasitology16: 525–527. Ingestion by intestinal nematodes is stimulated by histamine or dopamine. These biogenic amines significantly elevated the in vitro ingestion of a radiolabelled anthelmintic, morantel tartrate, by the ruminant nematode Trichostmngytus colubriformis from goats. Treatment of mice, infected by another nematode Nippostrongylus brasiliensis, with histamine and morantel tartrate reduced subsequent recoveries of worms when compared to anthelmintic treatment alone. Formulation of anthelmintic with non-pharmacological phagostimulants for nematodes may improve drug efficacy through “force feeding” of the parasites.

Depressed feeding by the nematode Trichostrongylus colubriformis in inoculated goats

loc. cit.) that centrifugation alone may initiate migration of granules and surface lamination, but no further conversion to schistosomules occurs in water. It is possible that other kinds of agitation besides centrifugation may do the same thing. Cultivation in ELAC, saline and PBS at 37 C beyond 3 hr revealed that both saline and PBS organisms were slower than ELAC organisms in onset of esophageal gland granule migration (3 hr) and development of euchromatic nuclei (4872 hr). Even ELAC organisms lagged behind in in vivo schistosomules (Stirewalt et al., 1983, loc. cit.). Glycocalyx degradation, however, occurred at the same rate in all 3 types of incubation medium, but also was much slower than in vivo (Table II). These data demonstrate that conversion of cercariae to schistosomules can be brought about by the single simple procedure of cultivation of cercariae at 37 C in a medium such as ELAC, or even in saline or PBS. The changes occur more slowly in vitro than in vivo, and in saline and PBS than ELAC. Nevertheless, with this method of conversion, the large numbers of organisms required for preparation of a vaccine consisting of irradiated schistosomules, or for other immunological investigations, are easily obtainable. This study was supported by NIH Grant RR08005.