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Field testing of Schistosoma japonicum DNA vaccines in cattle in China
Vaccine 20 (2002) 3629–3631
Short communication
Field testing of Schistosoma japonicum DNA vaccines in cattle in China Fuhui Shi a , Yaobi Zhang b , Jiaojiao Lin a , Xin Zuo c , Wei Shen a , Yiumin Cai a , Ping Ye a , Quentin D. Bickle b , Martin G. Taylor b,∗ a
Shanghai Institute of Animal Parasitology, Chinese Academy of Agricultural Sciences, 3 Lane 345 Shi-long Road, Shanghai 200232, PR China b Immunology Unit, Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WCIE 7HT, UK c Da Li Zhou Animal Husbandry and Veterinary Work Station, 36 Xing Fu Road, Xia Guan 671000, Yunnan Province, PR China Received 25 February 2002; received in revised form 22 July 2002; accepted 9 August 2002
Abstract Vaccines are needed to reduce the zoonotic reservoir of Schistosoma japonicum infection in bovines in China. We have developed two experimental DNA vaccines and have already shown these to be capable of inducing partial protection in water buffalo naturally exposed to the risk of S. japonicum infection in the field. We now report a similar field trial in cattle, the other major bovine reservoir host species in China. Groups of cattle were vaccinated with the VRSj28 vaccine or the VRSj23 vaccine, or, to test whether protection could be enhanced by combination vaccination, with both these DNA vaccines together. After vaccination, the cattle were exposed to natural infection in the field for a period of 54 days. Worm and egg counts carried out at the end of the experiment showed that each of the vaccine groups showed partial resistance, and that combined vaccination was not more effective than vaccination with the individual plasmids. © 2002 Elsevier Science Ltd. All rights reserved. Keywords: Schistosoma japonicum; DNA vaccines; Cattle
1. Introduction There are currently 0.9 million people in China infected with Schistosoma japonicum with a further 40 million at risk of infection [1,2], but this infection is a zoonosis infecting not only man but also a wide range of wild and domestic animals. In China, the most important animal reservoirs of schistosome infection are cattle and water buffalo: in a nation-wide schistosomiasis survey in 1995, the overall prevalence of S. japonicum in water buffalo was found to be 9.6% and in cattle 7.2% [3]. In many endemic areas, these bovines are responsible for a very large proportion of the total contamination of the environment with schistosome eggs and are thus instrumental in maintaining transmission of the infection [4]. China has made major investments in schistosomiasis control over a period of more than four decades which have yielded great public health benefits but schistosomiasis remains endemic in many areas because the major control strategy—annual chemotherapy of all diagnosed human and bovine cases—does not stop transmission of the infection. A ∗ Corresponding author. Tel.: +44-20-7927-2463; fax: +44-20-7323-5687. E-mail address: [email protected] (M.G. Taylor).
particular difficulty is that in bovines only partial cures are achievable using the maximum safe drug dose. For example, we recently found that when 15 naturally-infected water buffalo were treated with the standard dose of praziquantel used in the National Schistosomiasis Control Programme (a single oral dose of 25 mg/kg), only one-third of them were completely cured, i.e. became negative in the faecal egg hatching test (Ye Ping et al., unpublished observations). The development of vaccines to reduce schistosome infection in bovines is thus an attractive goal [5]; we envisage that such vaccines would be used in conjunction with selective population chemotherapy of both the human and the bovine populations. Our early experiments with live, irradiation-attenuated vaccines demonstrated the proof-of-principle of animal vaccination: vaccinated water buffalo and cattle were partially resistant when subsequently exposed to infection under field conditions [6]. However, this type of vaccine was impractical for large-scale field use and we therefore developed a panel of recombinant-derived defined antigen vaccines and showed that several of these could induce partial protection in sheep when administered as recombinant proteins in Freund’s adjuvants or BCG [7]. Since these adjuvants were unacceptable for widespread field application we then developed DNA vaccine formulations of the most promising antigens, a S. japonicum
0264-410X/02/$ – see front matter © 2002 Elsevier Science Ltd. All rights reserved. PII: S 0 2 6 4 - 4 1 0 X ( 0 2 ) 0 0 3 9 8 - 5
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F. Shi et al. / Vaccine 20 (2002) 3629–3631
Table 1 Field trial of DNA vaccines in cattle: worm recoveries, faecal egg hatching tests, and tissue egg counts in liver Group (n)
Mean worm recovery (S.D.)
R (%)a
P
Mean miracidial countb (S.D.)
R (%)a
P
Tissue egg count, eggs/g (S.D.)
R (%)a
P
VRSj28 (13) VRSj23 (13) VRSj28 + VRSj23 (12) Control (12)
35.9 42.3 39.2 63.6
44 33 38 –
0.008 0.03 0.04 –
18.2 25.2 26.7 78.6
77 66 68 –
0.04 0.07 0.10 –
183.3 147.9 115.9 225.0
19 34 48 –
0.58 0.18 0.03 –
a b
(22.3) (21.0) (29.8) (25.1)
(14.2) (21.3) (28.8) (91.1)
(216.7) (140.4) (89.6) (124.0)
Percentage reduction compared to control group. Miracidia/50 g faeces 54 days after first field exposure to infection.
28 kDa glutathione S-transferase (GST) and a S. japonicum 23 kDa transmembrane protein and showed that both of these could induce partial protection in sheep (tested in the laboratory) and water buffalo (tested in both laboratory and field trials) [8]. We have now backed up these experiments by carrying out a field trial in cattle, the other major species of reservoir host in China. In our buffalo experiments, we had only administered the DNA vaccines singly: therefore, another aim of the present experiment was to test whether protection levels could be raised by co-administering the two DNA vaccines.
2. Materials and methods Our field trial was carried out using 26 male and 24 female yearling Chinese yellow cattle at San-Ying Xiang in Er-Yuan County in Yunnan Province. This is a mountainous area, at 1700 m, and the site chosen was an irrigated grassland plateau that is used by local farmers for grazing their approximately 5000 dairy cattle. According to local veterinary records, the prevalence of schistosomiasis in these herds was 10, 5, 7 and 9% in the years 1998–2001, respectively. The experimental cattle were purchased and maintained during vaccination in the non-endemic Yang-Bi County, 100 km from San-Ying Xiang. They were randomly allocated into four groups by sex and body weight and immunised intramuscularly three times at monthly intervals with 100 g of either the VRSj28 or the VRSj23 DNA vaccine alone, or were injected with 100 g of the VRSj28 and VRSj23 DNA vaccines simultaneously, or were unvaccinated controls. One week after the last vaccination the cattle were transported to San-Ying Xiang, where they were exposed for 8 weeks to the risk of infection by being pastured in the field. The cattle were bled weekly for antibody studies. Production of the DNA vaccines and recombinant proteins, the IgG ELISA methods, and the parasitological methods used were all as described in our previous publications [7,8]. Briefly, full-length genes for Sj28 GST and Sj23 were amplified from cDNA by PCR, cloned into the VR1020 vector, and plasmids purified using the Qiagen Plasmid DNA purification kit. For antibody detection by ELISA, the full-length Sj28 GST gene was cloned into the pET-15b vector and the untagged protein was expressed in E. coli (BL21(DE3)pLysS)
and purified by glutathione-sepharose chromatography. The large hydrophilic domain of Sj23 was expressed in pGEX and purified over glutathione agarose. Recovery of adult schistosomes from the cattle by perfusion, egg hatching tests on faecal samples and tissue egg counts on liver samples were all carried out as described previously [8].
3. Results No significant levels of anti-rSj28GST IgG were detected by ELISA in cattle vaccinated with VRSj28 alone, but after the third vaccination significant levels were detected in the cattle immunised with the combined vaccine (data not shown). No significant levels of anti-rSj23 IgG were detected in any group. In faecal egg hatching tests reductions of 66–77% were seen in the vaccinated groups, compared with the non-immunised controls, and the reduction in the VRSj28-vaccinated group was statistically significant. Significant reductions of 33–44% in worm recovery were seen in the vaccinated groups, and tissue egg counts in the liver were also reduced, especially in the group given both vaccines (Table 1).
4. Discussion We previously found that two DNA vaccines (VRSj28 and VRSj23) were able to induce partial protection against experimental challenge in sheep and against both experimental and natural field challenge in water buffalo [8]. The protective effects were manifested as reductions in the mean worm and/or egg counts in vaccinated compared to control animals. It was particularly encouraging to find that in the buffalo field trial, each of the DNA vaccines induced significant reductions in worm recovery (39 and 38% in the VRSj28 and VRSj23 groups, respectively) and that there were substantial reductions in the numbers of viable eggs excreted by both vaccinated groups (62 and 50% for the VRSj28 and VRSj23 vaccine groups, respectively). Likewise, in the present trial in cattle VRSj28 induced a 44% reduction in worm recovery and a 77% reduction in the numbers of miracidia hatching from faeces, and VRSj23 induced 33 and 68% worm and miracidium reductions, respectively. Although both we [7]
F. Shi et al. / Vaccine 20 (2002) 3629–3631
and others [9–12] have developed recombinant protein-based S. japonicum vaccines and shown that these can protect livestock against experimental challenges, this and our previous trials [8] with VRSj28 and VRSj23 represent the first reported trials of S. japonicum DNA vaccines in farm animals under either laboratory or field conditions. The combined vaccines did induce a somewhat greater reduction in tissue egg density than either of the single vaccines, but overall co-administration of the VRSj23 and VRSj28 DNA vaccines was not significantly more protective than vaccination with the individual vaccines. The reason for this is not apparent, since we have not as yet identified immunological correlates of protection in DNA-vaccinated bovines. As in our previous experiments in sheep and water buffalo [8] levels of specific IgG were not indicative of protection status, and further work is needed to determine which specific antibody isotype responses and/or cytokine responses induced by these vaccines are associated with protection against infection. We have recently shown that in mouse experiments, vaccination with a ‘cocktail’ of four different DNA plasmids, encoding Sj28GST, Sj23, Sj62 and Sj14-3-3 induced significant protection to S. japonicum in CBA/Ca mice [13], whereas apart from a DNA vaccine construct of one S. japonicum vaccine candidate (paramyosin) [14], DNA vaccine versions of several other S. japonicum candidate antigens did not induce significant protection in mice when injected individually [15–19]. Therefore, further trials in bovines would be of interest to test whether vaccination with multiple DNA plasmids can significantly improve on the protection levels conferred by vaccination with individual DNA vaccines administered singly.
Acknowledgements This work was supported by the UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases (TDR) Grant #970950 and a 863 High Technology Project Grant #102-07-04-03 from the Ministry of Science and Technology, PR China. We are grateful to Dr. R.H. Zaugg of Vical Inc. for kindly supplying the VR1020 vector. References [1] WHO. Report of the WHO Informal Consultation on Schistosomiasis Control. WHO/CDS/CPC/SIP/99.2. Geneva: WHO; 1998. p. 1–45. [2] WHO. Report of the World Health Organisation Informal Consultation on Schistosomiasis in Low Transmission Areas: Control Strategies and Criteria for Elimination. WHO/CDS/CPC/SIP/2001.1. Geneva: WHO; 2001. p. 1–51.
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[3] Anon. Epidemic status of schistosomiasis in China, a nation-wide sampling survey in 1995. Office of Endemic Diseases Control, Ministry of Health, PR China. Beijing: Press of China. Schistosomiasis Control Journal; 1997. [4] Wang TP, Ge JH, Wu WD, et al. Infection sources of schistosomiasis and their role in the transmission in lake and marshland regions in Anhui Province. Chin J Parasitic Dis Control 1998;11:196–9. [5] Hsu SY, Hsu HF, Xu ST, et al. Vaccination against bovine schistosomiasis japonica with highly X-irradiated schistosomula. Am J Trop Med Hyg 1983;32:367–70. [6] Hsu SY, Xu ST, He YX, et al. Vaccination of bovines against schistosomiasis japonica with highly irradiated schistosomula in China. Am J Trop Med Hyg 1984;33:891–8. [7] Taylor MG, Huggins MC, Shi F, et al. Production and testing of Schistosoma japonicum candidate vaccine antigens in the natural ovine host. Vaccine 1998;16:1290–8. [8] Shi F, Zhang Y, Ye P, et al. Laboratory and field evaluation of Schistosoma japonicum DNA vaccines in sheep and water buffalo in China. Vaccine 2002;20:462–7. [9] McManus DP, Wong JY, Zhou J, et al. Recombinant paramyosin (rec-Sj-97) tested for immunogenicity and vaccine efficacy against Schistosoma japonicum in mice and water buffaloes. Vaccine 2002;20:870–8. [10] Liu SX, Song GC, Xu YX, Yang W, McManus DP. Anti-fecundity immunity induced in pigs vaccinated with recombinant Schistosoma japonicum 26 kDa glutathione S-transferase. Parasite Immunol 1995;17:335–40. [11] Shuxian L, Yongkang H, Guangchen S, Xing-Song L, Yuxin X, McManus DP. Anti-fecundity immunity to Schistosoma japonicum induced in Chinese water buffaloes (Bos buffelus) after vaccination with recombinant 26 kDa glutathione S-transferase (reSjc26GST). Vet Parasitol 1997;69:39–47. [12] Chen H, Nara T, Zeng X, et al. Vaccination of domestic pig with recombinant paramyosin against Schistosoma japonicum in China. Vaccine 2000;18:2142–6. [13] Zhang Y, Taylor MG, Johansen MV, Bickle QD. Vaccination of mice with a cocktail DNA vaccine induces a Th-1 type immune response and partial protection against Schistosoma japonicum infection. Vaccine 2002;20:724–30. [14] Zhou S, Liu S, Song GC, Xu Y, Sun W. Protective immunity induced by the full-length cDNA encoding paramyosin of Chinese Schistosoma japonicum. Vaccine 2000;18:3196–204. [15] Waine GJ, Yang W, Scott JC, McManus DP, Kalinna BH. DNA-based vaccination using Schistosoma japonicum (Asian blood-fluke) genes. Vaccine 1997;15:846–8. [16] Waine GJ, Mazzer DR, McManus DP. DNA immunization by intramuscular injection or gene gun induces specific IgG antibodies against a Schistosoma japonicum 22 kDa antigen, Sj22, when fused to the murine Ig K-chain secretory leader sequence. Parasite Immunol 1999;21:53–6. [17] Waine GJ, Alarcon JB, Qiu C, McManus DP. Genetic immunization of mice with DNA encoding the 23 kDa transmembrane surface protein of Schistosoma japonicum (Sj23) induces antigen-specific immunoglobulin G antibodies. Parasite Immunol 1999;21:377–81. [18] Scott JC, McManus DP. Molecular cloning and enzymatic expression of the 28-kDa glutathione S-transferase of Schistosoma japonicum: evidence for sequence variation but lack of consistent vaccine efficacy in the murine host. Parasitol Int 2000;49:289–300. [19] Zhang Y, Taylor MG, Gregoriadis G, McCrossan MV, Bickle QD. Immunogenicity of plasmid DNA encoding the 62 kDa fragment of Schistosoma japonicum myosin. Vaccine 2000;18:2102–9.
Short communication
Field testing of Schistosoma japonicum DNA vaccines in cattle in China Fuhui Shi a , Yaobi Zhang b , Jiaojiao Lin a , Xin Zuo c , Wei Shen a , Yiumin Cai a , Ping Ye a , Quentin D. Bickle b , Martin G. Taylor b,∗ a
Shanghai Institute of Animal Parasitology, Chinese Academy of Agricultural Sciences, 3 Lane 345 Shi-long Road, Shanghai 200232, PR China b Immunology Unit, Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WCIE 7HT, UK c Da Li Zhou Animal Husbandry and Veterinary Work Station, 36 Xing Fu Road, Xia Guan 671000, Yunnan Province, PR China Received 25 February 2002; received in revised form 22 July 2002; accepted 9 August 2002
Abstract Vaccines are needed to reduce the zoonotic reservoir of Schistosoma japonicum infection in bovines in China. We have developed two experimental DNA vaccines and have already shown these to be capable of inducing partial protection in water buffalo naturally exposed to the risk of S. japonicum infection in the field. We now report a similar field trial in cattle, the other major bovine reservoir host species in China. Groups of cattle were vaccinated with the VRSj28 vaccine or the VRSj23 vaccine, or, to test whether protection could be enhanced by combination vaccination, with both these DNA vaccines together. After vaccination, the cattle were exposed to natural infection in the field for a period of 54 days. Worm and egg counts carried out at the end of the experiment showed that each of the vaccine groups showed partial resistance, and that combined vaccination was not more effective than vaccination with the individual plasmids. © 2002 Elsevier Science Ltd. All rights reserved. Keywords: Schistosoma japonicum; DNA vaccines; Cattle
1. Introduction There are currently 0.9 million people in China infected with Schistosoma japonicum with a further 40 million at risk of infection [1,2], but this infection is a zoonosis infecting not only man but also a wide range of wild and domestic animals. In China, the most important animal reservoirs of schistosome infection are cattle and water buffalo: in a nation-wide schistosomiasis survey in 1995, the overall prevalence of S. japonicum in water buffalo was found to be 9.6% and in cattle 7.2% [3]. In many endemic areas, these bovines are responsible for a very large proportion of the total contamination of the environment with schistosome eggs and are thus instrumental in maintaining transmission of the infection [4]. China has made major investments in schistosomiasis control over a period of more than four decades which have yielded great public health benefits but schistosomiasis remains endemic in many areas because the major control strategy—annual chemotherapy of all diagnosed human and bovine cases—does not stop transmission of the infection. A ∗ Corresponding author. Tel.: +44-20-7927-2463; fax: +44-20-7323-5687. E-mail address: [email protected] (M.G. Taylor).
particular difficulty is that in bovines only partial cures are achievable using the maximum safe drug dose. For example, we recently found that when 15 naturally-infected water buffalo were treated with the standard dose of praziquantel used in the National Schistosomiasis Control Programme (a single oral dose of 25 mg/kg), only one-third of them were completely cured, i.e. became negative in the faecal egg hatching test (Ye Ping et al., unpublished observations). The development of vaccines to reduce schistosome infection in bovines is thus an attractive goal [5]; we envisage that such vaccines would be used in conjunction with selective population chemotherapy of both the human and the bovine populations. Our early experiments with live, irradiation-attenuated vaccines demonstrated the proof-of-principle of animal vaccination: vaccinated water buffalo and cattle were partially resistant when subsequently exposed to infection under field conditions [6]. However, this type of vaccine was impractical for large-scale field use and we therefore developed a panel of recombinant-derived defined antigen vaccines and showed that several of these could induce partial protection in sheep when administered as recombinant proteins in Freund’s adjuvants or BCG [7]. Since these adjuvants were unacceptable for widespread field application we then developed DNA vaccine formulations of the most promising antigens, a S. japonicum
0264-410X/02/$ – see front matter © 2002 Elsevier Science Ltd. All rights reserved. PII: S 0 2 6 4 - 4 1 0 X ( 0 2 ) 0 0 3 9 8 - 5
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F. Shi et al. / Vaccine 20 (2002) 3629–3631
Table 1 Field trial of DNA vaccines in cattle: worm recoveries, faecal egg hatching tests, and tissue egg counts in liver Group (n)
Mean worm recovery (S.D.)
R (%)a
P
Mean miracidial countb (S.D.)
R (%)a
P
Tissue egg count, eggs/g (S.D.)
R (%)a
P
VRSj28 (13) VRSj23 (13) VRSj28 + VRSj23 (12) Control (12)
35.9 42.3 39.2 63.6
44 33 38 –
0.008 0.03 0.04 –
18.2 25.2 26.7 78.6
77 66 68 –
0.04 0.07 0.10 –
183.3 147.9 115.9 225.0
19 34 48 –
0.58 0.18 0.03 –
a b
(22.3) (21.0) (29.8) (25.1)
(14.2) (21.3) (28.8) (91.1)
(216.7) (140.4) (89.6) (124.0)
Percentage reduction compared to control group. Miracidia/50 g faeces 54 days after first field exposure to infection.
28 kDa glutathione S-transferase (GST) and a S. japonicum 23 kDa transmembrane protein and showed that both of these could induce partial protection in sheep (tested in the laboratory) and water buffalo (tested in both laboratory and field trials) [8]. We have now backed up these experiments by carrying out a field trial in cattle, the other major species of reservoir host in China. In our buffalo experiments, we had only administered the DNA vaccines singly: therefore, another aim of the present experiment was to test whether protection levels could be raised by co-administering the two DNA vaccines.
2. Materials and methods Our field trial was carried out using 26 male and 24 female yearling Chinese yellow cattle at San-Ying Xiang in Er-Yuan County in Yunnan Province. This is a mountainous area, at 1700 m, and the site chosen was an irrigated grassland plateau that is used by local farmers for grazing their approximately 5000 dairy cattle. According to local veterinary records, the prevalence of schistosomiasis in these herds was 10, 5, 7 and 9% in the years 1998–2001, respectively. The experimental cattle were purchased and maintained during vaccination in the non-endemic Yang-Bi County, 100 km from San-Ying Xiang. They were randomly allocated into four groups by sex and body weight and immunised intramuscularly three times at monthly intervals with 100 g of either the VRSj28 or the VRSj23 DNA vaccine alone, or were injected with 100 g of the VRSj28 and VRSj23 DNA vaccines simultaneously, or were unvaccinated controls. One week after the last vaccination the cattle were transported to San-Ying Xiang, where they were exposed for 8 weeks to the risk of infection by being pastured in the field. The cattle were bled weekly for antibody studies. Production of the DNA vaccines and recombinant proteins, the IgG ELISA methods, and the parasitological methods used were all as described in our previous publications [7,8]. Briefly, full-length genes for Sj28 GST and Sj23 were amplified from cDNA by PCR, cloned into the VR1020 vector, and plasmids purified using the Qiagen Plasmid DNA purification kit. For antibody detection by ELISA, the full-length Sj28 GST gene was cloned into the pET-15b vector and the untagged protein was expressed in E. coli (BL21(DE3)pLysS)
and purified by glutathione-sepharose chromatography. The large hydrophilic domain of Sj23 was expressed in pGEX and purified over glutathione agarose. Recovery of adult schistosomes from the cattle by perfusion, egg hatching tests on faecal samples and tissue egg counts on liver samples were all carried out as described previously [8].
3. Results No significant levels of anti-rSj28GST IgG were detected by ELISA in cattle vaccinated with VRSj28 alone, but after the third vaccination significant levels were detected in the cattle immunised with the combined vaccine (data not shown). No significant levels of anti-rSj23 IgG were detected in any group. In faecal egg hatching tests reductions of 66–77% were seen in the vaccinated groups, compared with the non-immunised controls, and the reduction in the VRSj28-vaccinated group was statistically significant. Significant reductions of 33–44% in worm recovery were seen in the vaccinated groups, and tissue egg counts in the liver were also reduced, especially in the group given both vaccines (Table 1).
4. Discussion We previously found that two DNA vaccines (VRSj28 and VRSj23) were able to induce partial protection against experimental challenge in sheep and against both experimental and natural field challenge in water buffalo [8]. The protective effects were manifested as reductions in the mean worm and/or egg counts in vaccinated compared to control animals. It was particularly encouraging to find that in the buffalo field trial, each of the DNA vaccines induced significant reductions in worm recovery (39 and 38% in the VRSj28 and VRSj23 groups, respectively) and that there were substantial reductions in the numbers of viable eggs excreted by both vaccinated groups (62 and 50% for the VRSj28 and VRSj23 vaccine groups, respectively). Likewise, in the present trial in cattle VRSj28 induced a 44% reduction in worm recovery and a 77% reduction in the numbers of miracidia hatching from faeces, and VRSj23 induced 33 and 68% worm and miracidium reductions, respectively. Although both we [7]
F. Shi et al. / Vaccine 20 (2002) 3629–3631
and others [9–12] have developed recombinant protein-based S. japonicum vaccines and shown that these can protect livestock against experimental challenges, this and our previous trials [8] with VRSj28 and VRSj23 represent the first reported trials of S. japonicum DNA vaccines in farm animals under either laboratory or field conditions. The combined vaccines did induce a somewhat greater reduction in tissue egg density than either of the single vaccines, but overall co-administration of the VRSj23 and VRSj28 DNA vaccines was not significantly more protective than vaccination with the individual vaccines. The reason for this is not apparent, since we have not as yet identified immunological correlates of protection in DNA-vaccinated bovines. As in our previous experiments in sheep and water buffalo [8] levels of specific IgG were not indicative of protection status, and further work is needed to determine which specific antibody isotype responses and/or cytokine responses induced by these vaccines are associated with protection against infection. We have recently shown that in mouse experiments, vaccination with a ‘cocktail’ of four different DNA plasmids, encoding Sj28GST, Sj23, Sj62 and Sj14-3-3 induced significant protection to S. japonicum in CBA/Ca mice [13], whereas apart from a DNA vaccine construct of one S. japonicum vaccine candidate (paramyosin) [14], DNA vaccine versions of several other S. japonicum candidate antigens did not induce significant protection in mice when injected individually [15–19]. Therefore, further trials in bovines would be of interest to test whether vaccination with multiple DNA plasmids can significantly improve on the protection levels conferred by vaccination with individual DNA vaccines administered singly.
Acknowledgements This work was supported by the UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases (TDR) Grant #970950 and a 863 High Technology Project Grant #102-07-04-03 from the Ministry of Science and Technology, PR China. We are grateful to Dr. R.H. Zaugg of Vical Inc. for kindly supplying the VR1020 vector. References [1] WHO. Report of the WHO Informal Consultation on Schistosomiasis Control. WHO/CDS/CPC/SIP/99.2. Geneva: WHO; 1998. p. 1–45. [2] WHO. Report of the World Health Organisation Informal Consultation on Schistosomiasis in Low Transmission Areas: Control Strategies and Criteria for Elimination. WHO/CDS/CPC/SIP/2001.1. Geneva: WHO; 2001. p. 1–51.
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[3] Anon. Epidemic status of schistosomiasis in China, a nation-wide sampling survey in 1995. Office of Endemic Diseases Control, Ministry of Health, PR China. Beijing: Press of China. Schistosomiasis Control Journal; 1997. [4] Wang TP, Ge JH, Wu WD, et al. Infection sources of schistosomiasis and their role in the transmission in lake and marshland regions in Anhui Province. Chin J Parasitic Dis Control 1998;11:196–9. [5] Hsu SY, Hsu HF, Xu ST, et al. Vaccination against bovine schistosomiasis japonica with highly X-irradiated schistosomula. Am J Trop Med Hyg 1983;32:367–70. [6] Hsu SY, Xu ST, He YX, et al. Vaccination of bovines against schistosomiasis japonica with highly irradiated schistosomula in China. Am J Trop Med Hyg 1984;33:891–8. [7] Taylor MG, Huggins MC, Shi F, et al. Production and testing of Schistosoma japonicum candidate vaccine antigens in the natural ovine host. Vaccine 1998;16:1290–8. [8] Shi F, Zhang Y, Ye P, et al. Laboratory and field evaluation of Schistosoma japonicum DNA vaccines in sheep and water buffalo in China. Vaccine 2002;20:462–7. [9] McManus DP, Wong JY, Zhou J, et al. Recombinant paramyosin (rec-Sj-97) tested for immunogenicity and vaccine efficacy against Schistosoma japonicum in mice and water buffaloes. Vaccine 2002;20:870–8. [10] Liu SX, Song GC, Xu YX, Yang W, McManus DP. Anti-fecundity immunity induced in pigs vaccinated with recombinant Schistosoma japonicum 26 kDa glutathione S-transferase. Parasite Immunol 1995;17:335–40. [11] Shuxian L, Yongkang H, Guangchen S, Xing-Song L, Yuxin X, McManus DP. Anti-fecundity immunity to Schistosoma japonicum induced in Chinese water buffaloes (Bos buffelus) after vaccination with recombinant 26 kDa glutathione S-transferase (reSjc26GST). Vet Parasitol 1997;69:39–47. [12] Chen H, Nara T, Zeng X, et al. Vaccination of domestic pig with recombinant paramyosin against Schistosoma japonicum in China. Vaccine 2000;18:2142–6. [13] Zhang Y, Taylor MG, Johansen MV, Bickle QD. Vaccination of mice with a cocktail DNA vaccine induces a Th-1 type immune response and partial protection against Schistosoma japonicum infection. Vaccine 2002;20:724–30. [14] Zhou S, Liu S, Song GC, Xu Y, Sun W. Protective immunity induced by the full-length cDNA encoding paramyosin of Chinese Schistosoma japonicum. Vaccine 2000;18:3196–204. [15] Waine GJ, Yang W, Scott JC, McManus DP, Kalinna BH. DNA-based vaccination using Schistosoma japonicum (Asian blood-fluke) genes. Vaccine 1997;15:846–8. [16] Waine GJ, Mazzer DR, McManus DP. DNA immunization by intramuscular injection or gene gun induces specific IgG antibodies against a Schistosoma japonicum 22 kDa antigen, Sj22, when fused to the murine Ig K-chain secretory leader sequence. Parasite Immunol 1999;21:53–6. [17] Waine GJ, Alarcon JB, Qiu C, McManus DP. Genetic immunization of mice with DNA encoding the 23 kDa transmembrane surface protein of Schistosoma japonicum (Sj23) induces antigen-specific immunoglobulin G antibodies. Parasite Immunol 1999;21:377–81. [18] Scott JC, McManus DP. Molecular cloning and enzymatic expression of the 28-kDa glutathione S-transferase of Schistosoma japonicum: evidence for sequence variation but lack of consistent vaccine efficacy in the murine host. Parasitol Int 2000;49:289–300. [19] Zhang Y, Taylor MG, Gregoriadis G, McCrossan MV, Bickle QD. Immunogenicity of plasmid DNA encoding the 62 kDa fragment of Schistosoma japonicum myosin. Vaccine 2000;18:2102–9.