An AC-5 cathepsin B-like protease purified fromHaemonchus contortusexcretory secretory products shows protective antigen potential for lambs

Abstract

The immunogenic properties of cysteine proteases obtained from excretory/secretory products (ES) of Haemonchus contortus were investigated with a fraction purified with a recombinant H. contortus cystatin affinity column. The enrichment of H. contortus ES for cysteine protease was confirmed with substrate SDS-PAGE gels since the cystatin-binding fraction activity was three times higher than total ES, despite representing only 3% of total ES. This activity was inhibited by a specific cysteine protease inhibitor (E64) and by recombinant cystatin. The one-dimensional profile of the cystatin-binding fraction displayed a single band with a molecular mass of 43 kDa. Mass spectrometry showed this to be AC-5, a cathepsin B-like cysteine protease which had not been identified in ES products of H. contortus before. The cystatin binding fraction was tested as an immunogen in lambs which were vaccinated three times (week 0, 2.5 and 5), challenged with 10 000 L3 H. contortus (week 6) before necropsy and compared to unvaccinated challenge controls and another group given total ES (n = 10 per group). The group vaccinated with cystatin-binding proteins showed 36% and 32% mean worm burden and eggs per gram of faeces (EPG) reductions, respectively, compared to the controls but total ES was almost without effect. After challenge the cystatin-binding proteins induced significantly higher local and systemic ES specific IgA and IgG responses.

Secretion of Protective Antigens by Tissue-Stage Nematode Larvae Revealed by Proteomic Analysis and Vaccination-Induced Sterile Immunity

“…ACE levels in L4 ES were comparable to those in HES, as determined by emPAI, an expression pattern confirmed by RPKM transcript analysis, which revealed low or absent expression in L3 and eggs, and peak expression by day 5 L4 parasites, maintained in adults ( Tables S1, S2, S3 ). In contrast, significant products which are preferentially secreted by the luminal adults, and which have previously been reported to be secreted by parasitic nematodes, include proteases such as astacins and zinc metalloproteases [40]–[42], cathepsins [43], [44] and trypsin-like and other serine proteases [45]–[47], serpin [48] and Kunitz protease inhibitors [49], and predicted chitinases and chitin-binding proteins [50].…”

Immune response in goats vaccinated with thiol‐binding proteins from Haemonchus contortus

“…The specific humoural response included increased levels of peripheral and mucosal IgG and IgA. Although in some vaccine trials, it has not been possible to establish correlations between serum‐specific antibody levels and protection, and in many other cases, a clear association between serum humoural response and protection has been described after immunization with different H. contortus antigens . Similarly, the role of antibodies at the local level on the immunoprotection against H. contortus and other gastric nematodes has also been demonstrated .…”

Relative expression analysis of IL‐5 and IL‐6 genes in tropical sheep breed Pelibuey infected with Haemonchus contortus

“…Abomasal infection of H. contortus is a mechanical process because the third (L 3 ) and fourth (L 4 ) stages develop in the abomasal tissue, where they may be vulnerable to immune attack . Endoparasitic larvae use enzymatic mechanisms such as cathepsin B‐like cysteine protease and zinc metalloproteinase to digest the abomasal tissue for nutrients and survival, although these enzymes have demonstrated antigenic activation . The nature of the antigen to which a T H 0 responds largely determines whether it will differentiate into a collection of T H 1 or T H 2 cells.…”