Immune Protection Of Recombinant Trichinella Spiralis Elastase And TsE DNA Vaccine

Trichinellosis is a global distributing zoonosis caused by Trichinella spiralis(T.spiralis),which has a serious impact on livestock husbandry,social economy and national health.Veterinary vaccines against T spiralis fundamentally prevent T.spiralis infection.T.spiralis elastase gene belonging to serine protease family was obtained from T.spiralis draft genome and was highly expressed in T.spiralis intestinal infective larvae(IIL)in previous studies.Recombinant TsE(rTsE)was effective in inhibiting IIL from invading into the intestinal epithelial cell(IEC)monolayer in vitro,and might be a candidate antigen for anti-T.spiralis vaccine.1.Protective immunity in mice induced with rTsEThe rTsE was expressed in BL-21/pQE-80L-TsE.Each mouse was subcutaneously immunized by using rTsE emulsified with Montanide ISA201 adjuvantand and boosted 3 times.The results demonstrated that subcutaneous vaccination elicited a systemic humoral response(dramatically elevated levels of serum IgG and subclass IgG1/IgG2a and IgA)(P<0.0001)and a significant local enteral mucosal sIgA responses(dramatically increased levels of total sIgA and TsE-specific sIgA in intestinal washing)(P<0.001).The rTsE vaccination also produced a systemic and local mixed Th1/Th2 response with Treg function,as demonstrated by clear elevation levels of Th1 cytokines(IL-2,IFN-y),Th2 cytokine(IL-4)and Treg cytokine(IL-10)in immunized mice(P<0.0001).The immunized mice exhibited a 52.19%reduction in 7d AW and a 64.06%reduction in ML after challenge infection and the female fecundity from rTsE-immunized mice was evidently inferior to that of the control groups(P<0.01).Newborn larvae(NBLs)were cocultivated with mouse peritoneal exudate cells(PECs)and anti-rTsE serum.The antibody-dependent cell-mediated cytotoxicity(ADCC)was anti-rTsE antibody dose dependent with a decreasing trend following the increase in serum dilutions(F=44.582,P<0.0001),and an increasing trend with prolonged culture time(F=33.420,P<0.01).In conclusion,the rTsE vaccination triggered a systemic Thl/Th2 mixed immune response with Treg function and a local mucosal sIgA response,produced an effective protection against T.spiralis infection.The results indicated that TsE represented a novel target molecule for anti-T.spiralis vaccines.2.Protective immunity in mice induced with TsE DNA vaccineThe TsE gene from ML cDNA was cloned into the plasmid pcDNA3.1 as an intramuscular injection TsE DNA vaccine.After being transfected with pcDNA3.1-TsE,transcription and expression of TsE gene in hamster kidney cells(BHK-21).were detected by RT-PCR and IIFTTsE gene transcription and expression were detected in the spleen and MLN of mice immunized with the DNA vaccine by RT-PCR and IIFT experiments.It was found that the serum anti-TsE IgG antibodies in mice immunized by TsE DNA vaccine could recognize the natural TsE proteins expressed in the cuticle and stichosome of IIL and newborn larvae as well as female embryos of this parasite.Each mouse was intramuscularly immunized by pcDNA3.1-TsE and boosted 2 times.The results demonstrated that the TsE DNA vaccination elicited a systemic humoral response(dramatically increased level of serum IgG and subclass IgGl/IgG2a and IgA)(P<0.0001)and significant local enteral mucosal sIgA responses(dramatically increased level of total sIgA and TsE-specific sIgA in intestinal washing)(P<0.001).The pcDNA3.1-TsE vaccination also produced a systemic and local mixed Thl/Th2 response with Treg function,as demonstrated by clear elevation levels of Thl cytokines(IL-2,IFN-y),Th2 cytokine(IL-4)and Treg cytokine(IL-10)in immunized mice(P<0.0001).Duodenal tissue section obtained at 2 weeks after the last vaccination was dyed with AB-PAS.The amount of enteral goblet cells in pcDNA3.1-TsE vaccinated mice was obviously more than that of control group(P<0.0001).The immunized mice exhibited a 16.95%reduction in 7d AW and a 39.23%reduction in 42d ML after challenge infection,which is lower than the rTsE immunized mice.In conclusion,TsE DNA vaccine elicited a systemic concomitant Thl/Th2 response with Treg function and an enteral local sIgA response.The vaccination produced a partial protection against Trichinella challenge infection.