Chinese Journal of Schistosomiasis Control

Expression and identification of immune activity of cytosolic superoxide dismutase of Schistosoma japonicum

Yu Chuan-xin,Yin Xu-ren,Li Jian,Xu Yong-liang,Xu Ming, Gao Qi,Liang You-sheng

2007, 19(3): 161-164.

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Objective To express the cytosolic superoxide dismutase (SOD) of Schistosoma
japonicum and analyze its antigenicity. Methods The DNA sequence of Schistosoma japonicum
gene of cytosolic SOD was amplified by reverse transcription polymerase chain reaction (RT-PCR)
and subcloned into the expression vector pGEX-4T-3 to construct a recombinant plasmid Sj SOD-
pGEX-4T-3. This recombinant plasmid was transformed into component cell of E. coli BL21(DE3).
The fusion protein of GST-SOD was expressed by inducing with IPTG and purification by affinity
chromatography. The specific antiserum was prepared by immunizing the BALB/c mouse with
GST-SOD fusion protein, and the immnuogenicity of GST-SOD was investigated by Western blot
analysis. Results The gene of cytosolic SOD was amplified successfully and subcloned into expres-
sion vector pGEX-4T-3 forming the recombinant expression plasmid Sj SOD-pGEX-4T-3. The fu-
sion protein GST-SOD was expressed after the recombinant containing Sj SOD-pGEX-4T-3 being
induced by IPTG. Immunizing the BALB/c mouse with the fusion protein GST-SOD produced high
titer specific antiserum of l : 12 800 and the fusion protein GST-SOD was recognized by sera of se-
vere infection rabbits. Conclusions The cytosolic SOD has been expressed successfully and has a
preferable immunogenicity.

Cloning and expression of the gene encoding hypoxanthine-guanine phosphoribosyltransferase of Schistosoma japonicum

Shen Yu-juan, Xia Chao-ming, Cao Jian-ping, Xu Yu-xin,Li Xiao-hong, Liu Hai-peng

2007, 19(3): 165-169.

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Objective To perform the cloning of the gene encoding Schistosoma japonicum Chi-
nese-strain hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and its expression in Es-
cherichia coli. Methods A couple of primers were designed with the BamHI restriction endonucle-
ase site introduced in forward primer and SalIm reverse primer. Total RNA was isolated from adult
worms of S. japonicum Chinese-strain(Anhui-strain, Sjc-A)and the SjcHGPRT gene was amplified
by reverse transcriptase-polymerase chain reaction (RT-PCR). The PCR product and the prokary-
otic expression vector pET28a were digested by both restriction endonucleases BamHI and Sall. The
target DNA fragments were purified and cloned properly into pET28a. After identification by endonucleases digestion, PCR and sequencing, the recombinant plasmid pET28a-SjcHG PRT was transformed into competent E.coli BL21 and expressed in the presence of IPTG. Results pET28a-
SjHGPRT was sequenced and shown to be 99% and 83% identical in deduced amino acid sequence
to that of S. japonicum Chinese-strain (Hunan-strain, Sjc-H) and S. mansoni HGPRT, respectively.
The results of SDS-PAGE and Western blot revealed that the molecular weight of expressed protein
was around 30 kDa and could be recognized by anti-His-G-HPR antibody and sera from mice and hu-
man with schistosomasis japonica. Conclusion The recombinant plasmid containing SjcHGPRT cD-
NA is successfully constructed and its expression protein (reSjcHGPRT) is also successfully puri-
fied.

Investigation on Schistosoma hematobium infection among pupils in Boghe and Kaedi of Mauritania

Shu Jing| Dong Yun-xia, Zhang Xiao-li, Li Yi-bao

2007, 19(3): 169-174.

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Cloning and expression of the gene encoding Schistosoma japonicum Glyceraldehyde-3-phosphate dehydrogenase |Wang

Wang Wen-qin. Li Xiao-hong', Liu Shu-xian', Song Guang-cheng', Xu Yu-xin', Cao

2007, 19(3): 170-174.

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Objective To clone and express the gene encoding Schistosoma japonicum (Chinese
,train) Glyceraldehyde-3-phosphate dehydrogenase (SjcGAPDH) and predict the protein structure
and antigen determinants. Methods After digestion of pBluescript-SjcGAPDH with Xho I and Sac
I, the target DNA fragment was purified and subcloned into the proper prokaryotic expression vec-
tor pET28b. After identification of sequencing, the recombinant plasmid pET28a-SjcGAPDH was
transformed into competent E. coli BL21 and expressed in the presence of isopropyl-;3-D-thiogalac-
topyranoside (IPTG). Results The results of SDS-PAGE showed that the molecular weight of ex-
pressed protein was around 42.7 kDa. The recombinant protein was recognized by Western blot
with the sera from rabbits immunized with attenuated cercariae of S. japonicum. According to the
prediction, the N-terminal No. 50- 70, 102-122,135-148,165 -175,187 - 205,254- 272,278-
285 may be antigen determinants. Conclusion The gene encoding SjcGAPDH has been coloned and
expressed successfully, which will be helpful for identification of its function and antigenicity.

Studies on the optimal bivalent DNA vaccine immunization of Schistosoma japonicum

Ai Xi.Xia Li-xia, Xie Jiao,Hu Yuan, Shi You-en

2007, 19(3): 175-178.

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Objective To study the optimal immunization of bivalent DNA vaccine encoding
Schistosoma japonicum 14 kDa (Sj14) and 23 kDa (Sj23) with the different immunized dosages, in-
oculated times, inoculated routes and challenge time after the last immunization. Methods Con-
struction and preparation of the bivalent DNA vaccine of Schistosoma japonicum 14 kDa (Sj14) and
23 kDa(Sj23) were porformed. A total of 110 male BALB/c mice, about 5-6 weeks old, were ran-
domly divided int0 11 groups,2 were the control groups and 9 were the experimental groups, de-
signed with four factors in orthogonal trial. Each mouse of the groups was infected with (40± 2)
cercariaes of Schistosoma japonicum in the 30 th day after last immunization, and 45 days after the
infection, the mice were sacrificed, and the burden of the adult worms was invetigated. Results
There was no significant difference in the impact on immunity protection excited by the different fac-
tors and different levels of each factor (P>O. 05). Conclusion According to this experiment, the
optimal immunization for this vaccine is 50 ptg per mouse, twice immunization and intradermal injec-
tion, and the period of efficacy is at least 12 weeks after the challenge.

Surveillance on snail status after schistosomiasis has been interrupted in Huzhou City

Xia Di-ming

2007, 19(3): 178-178.

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Comparison of the molluscicidal effect of three formulations of niclosamide against Oncomelania snails

Dai Jian-rong, Liang You-sheng, Li Hong-jun, Tao Yong-hui, Tang Jian-xia

2007, 19(3): 179-182.

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Objective To compare the molluscicidal effect of suspension concentrate of 25%
niclosamide (SCN) and emulsifiable concentrate of 25% niclosamide (ECN) and wettable powder of
50% niclosanide ethanolamide salt (WPN) against Oncomelania snails, in order to give the reference
for schistosomiasis control in the field. Methods SCN, WCN and WPN were weighed and the so-
lutions of 0. 250, 0. 125, 0. 063, 0. 032 and 0. 016 mg/L of the active ingredient (niclosamide) pre-
pared in dechlorinated water. The 100 ml flasks were filled with the solutions and 10 adult or imma-
ture snails added and the flasks were covered with the plastic gauze to prevent snails from crawling
out the solutions. The controls only had dechlorinated water. After being immersed for 24, 48 h
and 72 h at 25 'C , the snails were washed with dechlorinated water and fed for a further 48 h. The
death of snails was determined by the knocked method and the LCso was calculated. Results In the
0.125 mg/L or more active content of the three formulations of niclosamide, there were high moI-
luscicidal effects in laboratory, and there were no significant differences among the three formula-
tions (P>O. 05). In 0. 063 mg/L active content of the three formulations, the death rates of SCN
and ECN were higher than that of WPN, there was a significant difference between SCN and WPN
or between ECN and WPN (P<O. 05). LCso of SCN and ECN at 24 h were the half of WPN, other
LCso of SCN and ECN were lower than that of WPN. The price of SCN was the half of ECN. Con-
clusion Compared with ECN and WPN, SCN has the best molluscicidal effect against Oncomelania
snails, and is the easiest and the cheapest for use in the field.

Effect of drug mud method for killing Oncomelania snails in honeycomb stone ditches

He Jian-gong, He Qiong-guang

2007, 19(3): 182-182.

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Study on the risk factors of schistosomiasis transmission in the
Three Gorges Reservoir Areas |]i Influence of the socioeconomic develop-
ment on schistosomiasis transmission

2007, 19(3): 183-187.

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Objective To understand the impacts of socioecononuc development on schistosomi-
asis transmission in the Three Gorges Reservoir Areas (TGRA). Methods Approaches including
epidemiology, immunology and socio-medical survey were applied'to investigate the potential risk
factors which would involve the importation of infectious resources of the mobile and migrant popu-
lation, and livestock, and the import of Oncomelania snails, and to explore the potential impact of
the socioeconomic development on schistosomiasis transmission in the reservoir areas. Results It is
possible that snail and arumal infectious resources will be introduced into TGRA by importing flow-
ers, trees and livestock. Among the migrant population in the survey, 31. 43% had infected with
schistosomiasis, 37 27% had contacted with the infested water, and the positive rate of immunolo-
gy tests was l.18%. The migrant population will be the mainly infection source. Orange industry,
travel industry, animal husbandry and fishery will develope preferentially in the future of TGRA.
However, the development of animal husbandry, fishery and travel industry will increase the risk of
the import of infection source and snails to TGRA. Conclusions TGRA would become a potential
schistosomiasis endemic areas since the changes of socioeconomic condition. We should insist on the
surveillance and supervising to prevent the transmission of schistosomiasis in TGRA.

Changes of schistosomiasis epidemic after the construction of boat factory in marshland

Zhang Zhen-qiu, Gao Yang, Tang Ming-liang, Li Ding, Tian Bin, Zhang Tian-shun

2007, 19(3): 187-191.

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Acute toxicity of LDS, a molluscicide

Yuan Yi', Xu Xing-jian, Wei Feng-hua, Li Gui-ling, Zhao Yun-bin, Liu Min

2007, 19(3): 188-191.

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Objective To investigate the acute toxicity of LDS, a molluscicide. Methods Ac-
cording to "the method of toxicity test for agriculture register" GB15670- 1995, an acute toxicity
test was carried out via mouth, skin and inbreathe with Wistar rat. Meanwhile, the eye and skin
stimulating test with rabbit as well as skin stimulating test with guinea pig were also conducted.
Results Based on the classification of appraising criterion on acute toxicity test via mouth, skin and
inbreathe with Wistar rat, LDS belonged to a feebleness toxicity degree. Based on intension classifi-
cation criterion of acute skin stimulation, LDS was of no acute skin stimulation to the rabbit. Based
on intension classification criterion of acute eye stimulation, LDS was of no acute eye stimulation to
the rabbit. Based on intension classification criterion of impressible rate, LDS was feebly impress-
ible. Conclusion LDS has a feeble toxicity degree to mammal.

An exploration of the conditions for isolating peripheral blood mononuclear cells from pig's blood using Ficoll density gradient centrifugation

Zhang Su-hua,Lin Dan-dan,Zhang Mei-juan,Chen Xue-song,Xu Jin-mei,Shan Hao|etc

2007, 19(3): 192-195.

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Objective To explore the best condition isolating the peripheral blood mononuclear
cells(PBMC)from pig's blood in order to provide an effective method for parasite immunology.
Methods PBMCs from pig's blood were isolated with pig's lymphocyte separation medium (LSM)
at different rotation speeds for different periods, the thickness of cloudlike PBMC layer was ob-
served and the yield of obtained PBMC and their activity were caculated. Simultaneously, PBMCs
from pig's blood were isolated with human LSM under the conditions reported by previous papers
(1 500 r/ min,30 min) and the isolating efficiency was compared with that of pig's LSM. Results
The best result was got with pig's LSM(density:l.llo ng/L ) by first centrifuging the samples at
1 500 r/ min (radius:15 cm) for 30 minutes at room temperature(20～25 'C), followed by washing
the cell pellets twice with D-Hank's medium at 1 500 r/min for 10 minutes at 4 'C. PBMC that
were isolated with human LSM from pig's blood under the same condition were impure and mixed
with other cells such as red cells and other cell debris, and both the yield of obtained PBMC and
their activity were low. Conclusion This study offered a method to get a large quantity of active
PBMC.

Advanced schistosomiasis combined with liver cancer:a report of 51 cases

Du Hua

2007, 19(3): 195-199.

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Effects of culture supernatant of Toxoplasma gondii on CD4+CD25+ T cells in mice in vitro

Zhang Gai, Ge Yi-yue, Wu Jiang-ping, Wang Yong, Hu Wei, Tan Ming-juan

2007, 19(3): 196-199.

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Objective To explore the effect of culture supernatant of Toxoplasma gondii on
CD4+CD25+ T cells in nuce in vitro. Methods CD4+CD25+ T cells were separated from spleens of
C57BL/6 mice and incubated in the culture superntant of Toxoplasma gondii. The apoptosis per-
centage of the CD4+CD25+ T cells were detected by FACScan, and the suppression of the CD4+
CD25+ T regulatory cells were examined by 3H_TdR incorporation. Results Compared with the
CD4+CD25+ T cells incubated with RPMI-1640, (36. 90±0. 36)% CD4+CD25+ T cells took apop-
tosis after incubated with the culture supernatant of Toxoplasma gondii for 10 hours,and the An-
nexin-v positive rate increased by (13. 60士2. 15) %. Compared with RPMI-1640, the culture super-
natant of Toxoplasma gondii incubating the CD4+CD25+ T regulatory cells for 5,10 hours reduced
their suppressive potential on the proliferation of the CD4+CD25- T cells significantly. Conclusions
Some composition of the culture supernatant of Toxoplasma gondii might cause apoptosis of
CD4+CD25+ T cells and as a result, could reduce their suppression on CD4+CD25- T cells.

CD34 expression and microvessel density in liver tissue from fibrotic patients with Schistosoma japonicum infection

Chen Jia-lin, Wen Xiao-wei, Lu Ning, Zhang Yan-ping,Wang Wei, Yang Hong-jie

2007, 19(3): 200-202.

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Objective To explore the relationship between CD34 expression or microvessel den-
sity and hepatic fibrosis degree in advanced schistosomiasis patients. Methods Thirty-five ad-
vanced schistosomiasis patients and 5 control patients were included in this study. CD34 expression
in liver tissue was measured with immunohistochemistry while microvessel density (MVD) of liver
tissue was evaluated under a light microscope. Results CD34 expression and MVD value were sig-
nificantly correlated with hepatic fibrosis degree in liver tissue from the advanced schistosomiasis pa-
tients. Conclusions CD34 expression and MVD value are two histological parameters representing
the liver sinusoid changes during fibrogenesis in advanced schistosonuasis patients.

Application of two dot immunogold filtration assay reagent boxes in hilly and mountainous areas of Yunnan Province

Fang Wen, Qiu Zong-ze|Chen feng

2007, 19(3): 202-207.

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Impact of vegetation changes on distribution of Oncomelania hupensis in marshland and lake regions

Cui Dao-yong, Ni Ying, Zhang ZhI-JIe, Peng Wen-xiang

2007, 19(3): 203-207.

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Objective   To study the impact of vegetation changes on the distribution of Oncomela-
nia hupensis in the marshland and lake regions. Methods    A bottomland from the 14 bottomlands a-
long the Qiupu River in the Guichi District, Chizhou City, Anhui Province was selected, and a 50 m
X 50 m area of uniform vegetation was choosed as the research field. Low-grass group, boundary-
grass group, hay group and control group were designed to represent four types of vegetation's sta-
tus and the snail density, soil temperature and soil moisture were surveyed half a month after the
design. The change of Oncmnelania hupensis's distribution due to the vegetation change was ana-
lyzed statistically. Results   The snail density of the hay group was nearly O with high soil moisture
and low soil temperature. The snail density of the low-grass group was obviously decreased (32.1/
O.l mz), and the snail density of the boundary group slightly increased, which had no statistical
significance compared with the snail density of the control group. Conclusions   Oncmnelania hupen-
sis moves from the low-grass group to the boundary group after the vegetation is cut down, which
changes the distribution of snail. Hay group is the worst environment for Oncamelania hupensis to
survive and the method of cut-overlay grass may be used to improve the effect of molluscicide.

Analysis of acute infection of Schistosoma japonicum in Hubei Province

Tu Zu-wu,Dai Yu-hai,Mao Guan-xiang,Zhu Hui-guo, Zhang Ying-hao

2007, 19(3): 208-211.

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Objective To understand the epidemic situation of acute infection of Schistosama
japonicum in Hubei Province in order to provide the basis of control strategy. Methods The data of
acute schistosome infection from 2004 t0 2005 in Hubei Province were collected. The indexes of
sex, age, occupation, infection date, morbidity, ways of infection were analysed with Excel. Re-
suIts In 20049 160 acute schistosomiasis cases were reported, and 83 cases (51. 87%) were prima-
ry and middle school students, and 68 cases (42. so%) were farmers. The main ways of contact in-
fested water were swimming (80 cases, 50. 00%) and agricultural production (39 cases, 24. 38%).
The peak of the infection was in June and July. The peak of the incidence was in August. Four out-
break epidemics including 55 cases occurred in the same time. In 2005, 157 acute schistosomiasis
cases were reported, and 61 cases (38. 85%) were primary and middle school students, and 72 cases
(45.86%) were farmers. The main ways of contact infested water were swimming (61 cases,
38. 85%) and agricultural production (48 cases, 30. 70%). The peak of the infection was in June.
The peak of the incidence was in July. Five outbreak epidemics including 44 cases occurred in the
same time. Conclusions Most acute cases were primary and middle school students and farmers,
so we need to take schistosomiasis control health education to high risk populations for the aim to
avoid contacting infested water, strengthen strategies to control and manage infection sources of
schistosome, eventually control and reduce the acute infection.

Molluscicidal effect of Rongbao Powder in schistosomiasismountainous |areas

Zhu Hong-qing| Zhong Bo, Cao Chun-li| Bao Zi-Ping| Wan Xue-xiang, etc

2007, 19(3): 212-216.

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Objective To evaluate the molluscicidal effect of Rongbao Powder in schistosomiasis
mountainous areas. Methods A total of 64 134 mz in Yuanping Village, Duoyue Township and 50
934 m2 in Wuxing Village, Funiu Township with exempting ploughing farmland of Meishan City,
Sichuan Province were sampled as the study group. Rongbao Powder mixtured with fine soil was
sprinkled in the field of the study group. With the sameness of landform and vegetation, 36 600 m2
of Yuanping Village and 4 333 m2 0f Wuxing Village were sampled as the control group. There was
no snail control measure in the canals and ditches of study and control groups in Yuanping Village,
and niclosamide mud was used in those places of study and control groups in Wuxing Village. The
appearance rate of snail frames, the change of average density of living snails and the result of envi-
ronment intervention were observed, and the fertilization and harvest of the field were investigated.
Results As the short- or long-term results, the molluscicidal effectiveness of Rongbao Powder,
the study group in Wuxing Village, Funiu Township was better than that in Yuanping Village,
Duoyue Township. After one planting duration, the appearance rate of snail frames of Yuanping
Village's study group declined by 68. 64% (X² =136. 58,P<0.01), the average density of living
snails of Yuanping Village decreased by 35. 78% (Z = - 2. 22, P<0. 05), the appearance rate of
snail frames of Wuxing Village reduced by 97. 03% (X² = 349. 68,P<0.01), the average density of
living snails of Wuxing Village reduced by 96. 64% (Z = - 7. 65,P<0.01). On the cost-benefit,

Analysis of factors on abnormal liver function in chronic schistosomiasis japonica patients

Chen Si-xi, Hou Xun-ya,Yan Fei-xiong

2007, 19(3): 217-219.

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Objective To analyze the associated factors on abnormal liver function (LF) in
chronic schistosomiaisis japonica patients. Methods The clinical data of 120 inpationts with chronic
schistosomiasis were collected and analyzed by logistic regression. The relevant treatment methods
were summarized. Results The associated factors on abnormal LF of the patients were hepatitis B,
alcohol drinking, hyperlipidemia, nonalcoholic fatty liver disease (NAFLD), and obesity, respec-
tively. The ordinary medication for improving liver function was effective to these causes. Prazi-
quantal can be adnunistrated successfully to these patients after LF normalized. Conclusion
Chronic schistosomiasis patients with abnormal LF but without severe vital organ diseases could be
treated with praziquantal after their LF improving.

Analysis of malaria prevalence in national malaria surveillance sites of Jiangsu Province

Wang Wei-ming, Jin Xiao-lin, Zhou Hua-yun, Li Ju-Iin, She Gui-zhi, Tang Yue-e

2007, 19(3): 220-223.

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Objective To understand the current status of malaria prevalence,its impacts and
malaria transmission trends in Jiangsu Province. Methods The malaria incidence, and its symp-
toms, mosquito preventive measures, Anopheles density, mosquito biting rates and related factors
were investigated in malaria epidemic villages as surveillance sites such as Guanshan Town of Suin-
ing County, Tangzhang Town of Tongshan County, Tianganghu town of Sihong County, Qiuji
Town of Xuyi County and Zhangzhu Twon of Yixing City. Results In 2006,the bednet coverage
was 20. 29% and bednet-using rate was 34. 32% in 27 903 residents of 7 921 households of 15 ad-
ministrative villages. In 2005, 750 Anopheles sinensi were captured from 50 bednets in the early
morning, the human biting ratio was 0.17/person per night, and in 2006, 1 927 Anopheles sinensi
were captured from 50 bednets in the early morrung, human biting ratio was 0. 38/person per night.
The average blood smear checking rate was 2. 72% (6 089 people) and the average positive rate was
0. 48% (29 plasmodium positive individuals). Conclusion The lower blood smear microscopy test-
ing,few positive rates of febrile patients and increased Anopheles sinensis density were the main fac-
tors of malaria relapses in the area of the North Huaihe River in Jiangsu Prownce.