Histochemical study of the effects on abomasal mucins of Haemonchus contortus or Teladorsagia circumcincta infection in lambs.

Previously, chemical analysis of gastric fundic mucin showed that infection of sheep with Haemonchus contortus or Teladorsagia circumcincta changed the proportions of monosaccharides and decreased terminal mucin fucosylation and sialylation. To identify the effects of these parasites on the two mucin-secreting cell lineages, fundic and antral tissues were collected for histochemistry from 69 lambs aged from 3-4 to 9-10 months-of-age which had received a single infection of either H. contortus or T. circumcincta and euthanased at Day 21 or 28 post- infection respectively. All fundic tissues were stained separately with: (1) with Periodic Acid Schiff (PAS) for all mucins; (2) Alcian Blue (AB) pH 2.5 for acidic mucins (sialylated and sulphated); (3) AB pH 1 for sulphated mucins and (4) High Iron Diamine (HID) for sulphated mucins. Antral and fundic tissues from 24 lambs were also stained for acidic and neutral mucins or with specific lectins for α-1-linked fucose and for α-2,3- and α-2,6-linked sialic acids. Only mucin sulphation appeared to differ visually in uninfected lambs over this age range: there was weak staining with HID in tissues from lambs 3-6 months-of-age, but was generally more intense in those over 7 months-of-age. Sulphomucins were not apparent in surface mucous cells (SMC) or generally in the upper pits. Sialylomucins were located predominantly in the pits and glands, with small amounts of sialylated mucins in SMC and on the luminal surface, mainly in younger animals up to 6 months-of-age and less in the older animals. Parasitism markedly reduced the predominantly neutral surface mucin5AC of the SMC and pit cells, despite pit elongation in both antrum and fundus, whereas the acidic Muc6 secreted by mucus neck cells (MNC) increased along with MNC hyperplasia. Sulphated mucins were present mainly from the mid-pits downward and heavy staining was more common in older animals. In these sheep, the markedly reduced neutral mucin in the SMC and pit cells in both antrum and fundus contrasts with reported hypersecretion of mucus in the intestine, which is believed to aid in parasite expulsion. It has been proposed that intestinal goblet cell hypersecretion occurs only in resistant animals, therefore reduced mucins in the abomasum may be indicative of susceptibility to abomasal parasites.

Effects of weaning and infection with Teladorsagia circumcincta on mucin carbohydrate profiles of early weaned lambs.

Differences in mucin glycosylation in milk-fed and early weaned lambs may influence susceptibility to parasitism, particularly the greater cellular content and higher sulphation of mucins in young and unweaned lambs. Weaning also reduced the percentage of Gal (p<0.05) in fundic mucin and galactosamine (GalN) (p<0.01) in duodenal mucin, but had no noticeable effect on fucosylation or sialylation. Four experimental groups of lambs were studied (n=3): (1) 3 days old; (2) 9 weeks old milk-fed; (3) 9 weeks old weaned at 3 weeks-of-age on to lucerne chaff and cereal-based pellets (4) 9 weeks old weaned and infected with 1000 Teladorsagia circumcincta L3 twice weekly for 5 weeks. Fundic and duodenal mucin monosaccharides were analysed chemically and fundic, antral and duodenal tissues were stained with lectins, periodic acid Schiff (PAS), Alcian blue/PAS (AB/PAS) and high iron diamine. Age-related maturation of mucin glycosylation was prominent in young lambs: reduced total fundic mucins and increasing fucosylation and decreasing sialylation and sulphation of all mucins, as well as changes in the types of linkages of Fuc and sialic acids. By 9 weeks-of-age, there were no longer sialylated mucins in fundic surface mucus cells, only neutral mucins, while in Brunner's glands, there was reduced sialylation and large amounts of neutral mucins. In the neonates, both fundic and duodenal tissues contained only small amounts of mucins terminating with alpha-1,2-linked Fuc, which became the principal linkage in 9 weeks old lambs. Duodenal mucins in 3 days old lambs contained both alpha-2,6- and alpha-2,3-linked sialic acids, whereas the alpha-2,3 linkage was not present in older lambs. Parasitism increased the percentage of galactose, but reduced total and neutral fundic mucins, as well as sulphation and sialylation. There was both decreased sialylation and sulphation in duodenal mucins. Although no change in fucosylation was apparent from chemical analysis, infection reduced lectin staining for alpha-1,2-linked fucose in antral and duodenal tissues and alpha-1,6- and alpha-1,3-linked fucose in the duodenum. These changes in fundic and duodenal mucins were similar to those previously seen on Day 28 p.i. after a single infection of 4-9 months old sheep with T. circumcincta larvae.

Monosaccharide composition of fundic and duodenal mucins in sheep infected with Haemonchus contortus or Teladorsagia circumcincta.

The effects on the monosaccharide composition of fundic and duodenal mucins of parasitism by Haemonchus contortus or Teladorsagia circumcincta were investigated in sheep at 4, 6 and 9 months of age. Infected sheep were euthanased at days 21 and 28 post-infection respectively, together with uninfected controls. Fundic and duodenal mucins were purified by gel filtration and CsCl density gradient centrifugation and monosaccharides were released by heating at 95 degrees C for 6h in 2M HCl. Thin-layer chromatography identified fucose, glucosamine, galactose and galactosamine in both mucins, as well as very small amounts of sialic acids. Neither N-acetylglucosamine nor N-acetylgalactosamine was present, as these are deacetylated during acid hydrolysis to glucosamine and galactosamine, respectively. Fucose, glucosamine, galactose and galactosamine were separated and quantified by High Performance Anion Exchange Chromatography on a CarboPac PA-20 column. Sialic acids were determined by the thiobarbiturate assay. Over the age range of 4-9 months, the principal changes in the monosaccharide composition of mucins in non-infected sheep were increasing fucosylation and decreasing sialylation, as observed in other mammals. In duodenal mucins, there was a statistically significant increase in fucosylation and a decrease in sialylation (p=0.043 and 0.014, respectively), while similar trends were seen in fundic mucins. Other modifications with age in sheep mucins were decreased acetylglucosamine (N-acetylglucosamine) in the fundus and galactosamine (N-acetylgalactosamine) in the duodenum. The effects of H. contortus and T. circumcincta infection on fundic mucin monosaccharide composition were not identical, although both parasites decreased fucosylation and sialylation. Both parasites caused the same effects on duodenal mucins, however, these differed from the changes in the fundus. H. contortus infection increased the proportions of glucosamine and galactose in fundic and duodenal mucins, respectively. Mucins from the fundus of H. contortus-infected sheep had similar monosaccharide profiles at all ages, but this was not the case for T. circumcincta, in which there were lesser changes on mucins in 9 months old sheep, apart from decreased sialylation. This may indicate immunity to T. circumcincta from previous exposure in the field. The effect on duodenal mucins was similar for the two infections (decreased sialic acids, fucose and N-acetylgalactosamine and increased galactose), suggesting it may result from the immune response to the presence of worms in the abomasum. Mucin profiles from organs more accessible than the gastrointestinal tract may be useful markers for the host immune response and identify resistant, resilient or susceptible individuals.