[Show abstract][Hide abstract] ABSTRACT: Background
Cystic echinococcosis (CE), caused by Echinococcus granulosus metacestode, invokes a serious public health concern. Early diagnosis has great impacts on reduction of disability-adjusted life years. Several antigen B-related molecules (EgAgB; EgAgB1-5) are known to be immunopotent, but detection of EgAgB is variable in many patients and may not allow reliable interpretation of its immunological relevance. More importantly, the immunoproteome profile of hydatid fluid (HF) has not been addressed.Methods
We conducted a proteome analysis of the HF of a single fertile cyst of CE1 and CE2 stages through two-dimensional electrophoresis (2-DE). Each protein spot was analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). We subsequently determined the immunoproteome profile employing patient sera of entire disease spectrum from CE1 to CE5 stages.ResultsWe identified 40 parasite proteins, of which EgAgB (28 spots) and antigen 5 (EgAg5; 5 molecules) were abundant. EgAgB proteoforms constituted the majority, mostly EgAgB1 (24 spots), followed by EgAgB2 and EgAgB4 (2 spots each). EgAgB3 was detected only by liquid chromatography-MS/MS. EgAgB5 was not recognized. We also detected 38 host proteins, which were largely composed of serum components, antioxidant/xenobiotic enzymes, and enzymes involved in carbohydrate metabolism. CE1 and CE2 HF exhibited comparable spotting patterns, but CE2 HF harbored greater amounts of EgAgB and EgAg5 complexes. CE sera demonstrated complicated immune recognition patterns according to the disease progression; CE2 and CE3 stages exhibited strong antibody responses against diverse EgAgB and EgAg5 proteoforms, while CE1, CE4, and CE5 stages mainly reacted to EgAg5 and cathepsin B. Patient sera of alveolar echinococcosis (AE) cross-reacted with diverse EgAgB isoforms (36%). EgAg5 and cathepsin B also demonstrated cross-reactions with sera from neurocysticercosis and sparganosis.Conclusions
Our results demonstrated that detection of a single defined molecule may not properly diagnose CE, since specific immunodominant epitopes changed as the disease progresses. Immunoproteome analysis combined with imaging studies may be practical in the differential diagnosis of CE from AE and other cystic lesions, as well as for staging CE, which are pertinent to establish appropriate patient management.
[Show abstract][Hide abstract] ABSTRACT: Objectives
Neurocysticercosis (NC), an infection of the central nervous system with Taenia solium metacestodes (TsM), invokes a formidable neurological disease. A bundle of antigens is applicable for serodiagnosis of active cases, while they demonstrate fairly low reactivity against sera of chronic NC. Identification of sensitive biomarkers for chronic NC is critical for appropriate management of patients.Methods
Proteome analysis revealed several isoforms of 65- and 83-kDa TsM fasciclin-like proteins (TsMFas) to be highly reactive with sera of chronic NC. A cDNA encoding one of the 83-kDa TsMFas (TsMFas1) was isolated from a cDNA library. We expressed a recombinant protein (rTsMFas1) and evaluated its diagnostic potential employing sera from chronic NC (n = 80), tissue-invasive cestodiases (n = 169) and trematodiases (n = 80) and those of normal controls (n = 50).ResultsSecretory TsMFas1 was composed of 766 amino acid polypeptide and harboured fasciclin and fasciclin-superfamily domains. The protein was constitutively expressed in metacestode and adult stages, with preferential locality in the scolex. Bacterially expressed rTsMFas1 exhibited 78.8% sensitivity (63/80 cases) and 93% specificity (278/299 samples) in diagnosing chronic NC. Some cross-reactivity was observed with sera of cystic echinococcosis (10/56, 17.8%) and sparganosis (4/50, 8%). Positive and negative predictive values were 75% and 95.5%, respectively.Conclusion
TsM fasciclin-like protein may be useful for differential diagnosis of chronic NC in clinical settings, especially where both NC and other infectious cerebral granulomatoses are prevalent.
Tropical Medicine & International Health 03/2014; · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Human alveolar echinococcocosis (AE) is a highly pathogenic zoonotic disease caused by the larval stage of the cestode E. multilocularis. Its life-cycle includes more than 40 species of small mammal intermediate hosts. Therefore, host biodiversity losses could be expected to alter transmission. Climate may also have possible impacts on E. multilocularis egg survival. We examined the distribution of human AE across two spatial scales, (i) for continental China and (ii) over the eastern edge of the Tibetan plateau. We tested the hypotheses that human disease distribution can be explained by either the biodiversity of small mammal intermediate host species, or by environmental factors such as climate or landscape characteristics. METHODOLOGYFINDINGS: The distributions of 274 small mammal species were mapped to 967 point locations on a grid covering continental China. Land cover, elevation, monthly rainfall and temperature were mapped using remotely sensed imagery and compared to the distribution of human AE disease at continental scale and over the eastern Tibetan plateau. Infection status of 17,589 people screened by abdominal ultrasound in 2002-2008 in 94 villages of Tibetan areas of western Sichuan and Qinghai provinces was analyzed using generalized additive mixed models and related to epidemiological and environmental covariates. We found that human AE was not directly correlated with small mammal reservoir host species richness, but rather was spatially correlated with landscape features and climate which could confirm and predict human disease hotspots over a 200,000 km(2) region. CONCLUSIONSSIGNIFICANCE: E. multilocularis transmission and resultant human disease risk was better predicted from landscape features that could support increases of small mammal host species prone to population outbreaks, rather than host species richness. We anticipate that our study may be a starting point for further research wherein landscape management could be used to predict human disease risk and for controlling this zoonotic helminthic.
[Show abstract][Hide abstract] ABSTRACT: To clone and express EgCyP gene of Echinococcus granulosas and analyze EgCyP using bioinformatics.
Total RNAS of adult E. granulosus was extracted and reversedly transcripted to cDNA. EgCyP gene was amplified from cDNA and inserted into vector pET28a. Recombinant plasmid pET28a-EgCyP was transformed into E. coli BL21 (DE3) for expression under the induction of IPTG. The expressed product was identified by SDS-PAGE and Western blotting. EgCyP was analyzed by the bioinformatics software.
The EgCyP gene was successfully amplified from cDNA of adult E. granulosus and a fusion protein was expressed in E .coli BL21 (DE3). The molecular weight of the expressed protein was about 22 kDa. The Western blotting indicated that the antigenicity of the protein was specific. The bioinformatics analysis revealed that there were 7 antigen epitopes in EgCyP.
EgCyP of E. granulosus is cloned and expressed in E. coli BL21 (DE3) successfully, which might be the foundation for the further study of its immunogenicity.
Zhongguo xue xi chong bing fang zhi za zhi = Chinese journal of schistosomiasis control. 06/2012; 24(3):294-7, 302.
[Show abstract][Hide abstract] ABSTRACT: To analyze the epidemiological status of echinococcosis among children in three different zones of Qinghai Province.
B-ultrasound and ELISA were used in the survey to evaluate echinococcus infection among 6 to 15 year old children in the three zones, namely, Qinghai southern plateau, Qilian mountain-Hehuang valley and Chaidamu basin.
The prevalence of echinococcus infection scanned by B-ultrasound and sera positive rate detected by ELISA in children were 1.5% (320/20 730) and 6.5% (1024/15 762) respectively, and the prevalence of cystic echinococcosis and alveolar echinococcosis by ultrasound were 1.0% (206/20730) and 0.6% (114/20730), respectively. The prevalence and sera positive rate were 9.5% (707/7 453) and 2.3% (269/11 618) in Qinghai southern plateau, 3.8%(289/7 544) and 0.6% (50/8 275) in Qilian mountain-Hehuang valley, and 3.7% (28/765) and 0.1% (1/837) in Chaidamu basin, respectively. The southern plateau showed the highest prevalence, with a significant statistical difference among the three areas (P < 0.01). The prevalence of alveolar echinococcosis was 1.0% (114/11 618) in the southern plateau, but no alveolar echinococcosis patients were found in other two areas. Correlation analysis showed that the prevalence increased with the altitude (r(s-e) = 0.96, P < 0.05, R2 = 0.93; r(p-e) = 0.82, P < 0.05, R2 = 0.67).
The prevalence of echinococcosis among children shows an apparent geographical difference in Qinghai Province.
Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 04/2012; 30(2):127-30.
[Show abstract][Hide abstract] ABSTRACT: The Qinghai-Tibetan Plateau (QTP, in western China), which is the largest and highest plateau on Earth, is a highly epidemic region for Echinococcus spp. We collected 70 Echinococcus samples from humans, dogs, sheep, yaks, plateau pikas, and voles in eastern and southern Qinghai and genotyped them using the mitochondrial DNA marker cytochrome oxidase subunit I gene and maximum parsimony and Bayesian reconstruction methods. Based on the 792-bp sequence matrix, we recorded 124 variable sites, of which, 115 were parsimony-informative. Thirty-four haplotypes (H1-H34) were detected, of which H1-H15, H16-H17, and H18-H34 belonged to Echinococcus shiquicus, Echinococcus multilocularis, and Echinococcus granulosus, respectively. Within 26 human isolates, three were identified as E. multilocularis and 23 were E. granulosus. We also detected a dual infection case in a dog with E. multilocularis and E. granulosus. The intraspecific haplotype (Hd ± SD) and nucleotide (Nd ± SD) diversity of E. shiquicus (0.947 ± 0.021; 0.00441 ± 0.00062) was higher than that for E. granulosus (0.896 ± 0.038; 0.00221 ± 0.00031) and E. multilocularis (0.286 ± 0.196; 0.00036 ± 0.00025). Moreover, the haplotype network of E. shiquicus showed a radial feature rather than a divergent feature in a previous study, indicating this species in the QTP has also evolved with bottleneck effects.
Parasitology Research 01/2012; 111(1):179-84. · 2.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To clone and express EPCl gene of Echinococcus granulosus, and investigate its immunogenicity and diagnostic value.
Total RNA was extracted from hydatid cyst protoscoleces and EPC1 gene of Echinococcus granulosus was amplified by RT-PCR. The PCR product was cloned into pGEM-T vector, and then subcloned into the prokaryotic expression vector PET28a(+). The positive recombinants were transformed into Escherichia coli BL21 (DE3), and followed by expression of the protein induced by IPTG. The recombinant protein was identified by SDS-PAGE and Western blotting, and used to establish ELISA. Serum samples from patients with cystic echinococcosis (60 cases), alve-olar echinococcosis (37 cases), cysticercosis (16 cases), clonorchiasis sinensis (7 cases), schistosomiasis japonica (4 cases) and healthy persons (33 cases) were examined.
The recombinant plasmid PET28a-EgEPC1 was identified by restriction enzyme digestion and sequencing. SDS-PAGE result showed that the recombinant containing recombinant plasmid PET28a-EgEPC1 expressed a soluble fission protein of EgEPC1 (about M, 11 000). The protein was recognized by pool sera of cystic echinococcosis patients. The overall sensitivity and specificity of diagnosis by ELISA for cystic echinococcosis were 78.3% (47/60), and 98.3% (59/60), respectively. The cross reaction with sera of alveolar echinococcosis was 40.5% (15/37).
The recombinant EgEPC1 antigen has diagnostic value in cystic echinococcosis.
Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 06/2011; 29(3):167-71.
[Show abstract][Hide abstract] ABSTRACT: To clone and express Echinococcus granulosus Eg18 gene and evaluate the immunoreactivity of the recombinant protein.
Eg18 coding sequence was amplified by RT-PCR using primers designed according to the sequence of Eg18 in GenBank from total RNA extracted from Echinococcus granulosus protoscoleces isolated from infected sheep in Qinhai Province and cloned into the prokaryotic expression vector pET-28a(+). The recombinant expression vector was transformed to Escherichia coli BL21 (DE) and induced to express by IPTG. The expressed products were analyzed by SDS-PAGE and purified with Ni-IDA agarose affinity chromatography. rEg18 was evaluated for its reactivity with the sera from the patients infected with hydatid and other helminthes by Western blotting and ELISA.
The sequence of cloned Eg18 was completely identical wit the original sequences of Eg18 and Em18 deposited in GenBank. The recombinant protein strongly reacted to the sera from the patients with alveolar echinococcosis, cystic echinococcosis, and cysticercosis, and weakly reacted to the sera from the patients with nematodiasis, schistosomiasis and clonorchiasis. The detection of specific IgG4 was much more specific than that of IgG.
Eg18/Em18 is the common antigen of cestode and its specific IgG4 is a serum marker of alveolar echinococcosis.
Zhongguo xue xi chong bing fang zhi za zhi = Chinese journal of schistosomiasis control. 04/2011; 23(2):192-6.
[Show abstract][Hide abstract] ABSTRACT: Objective
To identify full length cDNA sequence of lactate dehydrogenase (LDH) from adult Echinococcus granulosus (E. granulosus) and to predict the structure and function of its encoding protein using bioinformatics methods.
Asian Pacific Journal of Tropical Medicine 10/2010; 3(10):757-761. · 0.93 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cystic echinococcosis (CE) or hydatid disease is known to be cosmopolitan in its global distribution, while alveolar echinococcosis (AE) is a much rarer though more pathogenic hepatic parasitic disease restricted to the northern hemisphere. Both forms of human echinococcosis are known to occur on the Tibetan Plateau, but the epidemiological characteristics remain poorly understood. In our current study, abdominal ultrasound screening programs for echinococcosis were conducted in 31 Tibetan townships in Ganze and Aba Tibetan Autonomous Prefectures of northwest Sichuan Province during 2001-2008. Hospital records (1992-2006) in a major regional treatment centre for echinococcosis in Sichuan Province were also reviewed. Of 10,186 local residents examined by portable ultrasound scan, 645 (6.3%) were diagnosed with echinococcosis: a prevalence of 3.2% for CE, 3.1% for AE and 0.04% for dual infection (both CE and AE). Human cystic and alveolar echinococcosis in pastoral areas was highly co-endemic, in comparison to much lower prevalences in semi-pastoral or farming regions. The high ultrasound prevalence in these co-endemic areas in northwest Sichuan Province was also reflected in the hospital study, and hospital records furthermore indicated another possible highly co-endemic focus in Guoluo Prefecture of Qinghai Province, located at the border of northwest Sichuan. These chronic cestode zoonoses constitute an unparalleled major public health problem for pastoral Tibetan communities, and pose great difficulties for adequate treatment access and effective transmission control in such remote regions.
[Show abstract][Hide abstract] ABSTRACT: The genetic polymorphisms of Echinococcus spp. in the eastern Tibetan Plateau and the Xinjiang Uyghur Autonomous Region were evaluated by DNA sequencing analyses of genes for mitochondrial cytochrome c oxidase subunit 1 (cox1) and nuclear elongation factor-1 alpha (ef1a). We collected 68 isolates of Echinococcus granulosus sensu stricto (s.s.) from Xinjiang and 113 isolates of E. granulosus s. s., 49 isolates of Echinococcus multilocularis and 34 isolates of Echinococcus shiquicus from the Tibetan Plateau. The results of molecular identification by mitochondrial and nuclear markers were identical, suggesting the infrequency of introgressive hybridization. A considerable intraspecific variation was detected in mitochondrial cox1 sequences. The parsimonious network of cox1 haplotypes showed star-like features in E. granulosus s. s. and E. multilocularis, but a divergent feature in E. shiquicus. The cox1 neutrality indexes computed by Tajima's D and Fu's Fs tests showed high negative values in E. granulosus s. s. and E. multilocularis, indicating significant deviations from neutrality. In contrast, the low positive values of both tests were obtained in E. shiquicus. These results suggest the following hypotheses: (i) recent founder effects arose in E. granulosus and E. multilocularis after introducing particular individuals into the endemic areas by anthropogenic movement or natural migration of host mammals, and (ii) the ancestor of E. shiquicus was segregated into the Tibetan Plateau by colonising alpine mammals and its mitochondrial locus has evolved without bottleneck effects.
International journal for parasitology 09/2009; 40(3):379-85. · 3.39 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To make an epidemiological investigation on echinococcosis in Darlag County, Guoluo Tibetan Autonomous Prefecture of Qinghai Province.
Local participants were screened for echinococcosis using B ultrasound from August to September in 2007. IHA was carried out to detect antibody in human sera. Indirect ELISA using recombinant-AgB and Em18 antigen was conducted to further diagnose the human alveolar echinococcosis (AE) and cystic echinococcosis (CE) . The prevalence of echinococcosis in wild rodents, yaks, sheep and dogs was examined either by dissection or through slaughterhouses. Adult worms and hydatid cysts were detected by PCR-RFLP to discriminate 3 Echinococcus species and genotype of Eg. Dog feces were tested by double antibody sandwich ELISA method to confirm the coproantigen positive rate.
1723 people were examined by ultrasound and 236 were clinically diagnosed as hydatid diseases with an overall morbidity of 13.7%, of which 5.5% were CE and 8.2% were AE. Prevalence in males and females was 11.6% and 15.6% respectively with a statistically significant difference (chi2=7.0, P< 0.05) . Prevalence in herdsmen and monks/nuns was as high as 23.0% and 13.2%, respectively. Adult worms were found in 5 out of 9 stray dogs by necropsy. 27 of 239 pikas (Ochotona curzoniae) were infected by E. shiquicus (11.3%). CE prevalence in sheep and yaks was 26.4% and 5/16 respectively. The Eg genotype was G1 (sheep strain).
Three species of Echinococcus, e.g. E. granulosus, E. multilocularis and E. shiquicus are all prevalent in Darlag County. The prevalence of both AE and CE in human population is high in the area. Domestic dogs are the primary definitive host for both E. granulosus and E. multilocularis.
Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 03/2009; 27(1):22-6.
[Show abstract][Hide abstract] ABSTRACT: Local inhabitants in 8 towns/townships of the counties Yushu, Zhiduo and Chengduo were examined with serology and ultrasound in 2006. Among 2 251 people tested by indirect hemagglutination (IHA), 207 showed anti-hydatid IgG positive (9.2%). Ultrasound examination found 106 cases out of 2581 people, with a morbidity of 4.1%. Females showed higher sero-positive rate (11.3%) and morbidity rate (5.0%) than males (6.6% and 3.0% respectively). The highest sero-positive rate was in the group of 40-49 years old (16.4%). The morbidity rate increased with age, with the highest rate (15.5%) in the group of 60 years old and above. Occupationally, those involved in semi-agriculture and semi-animal husbandry showed highest sero-positive rate and morbidity rate, 21.5% and 11.4% respectively. The survey demonstrated that the prevalence of hydatid disease in human population is at a high level in Yushu Tibetan Autonomous Prefecture.
Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 01/2009; 26(6):480-1, 484.
[Show abstract][Hide abstract] ABSTRACT: An epidemiological investigation on echinococcosis was made in Jiuzhi County of Qinghai Province, western China. Ultrasonography and an indirect hemagglutination test revealed a morbidity of 8.0% (124/1,549) and a seroprevalence of 25.8% (287/1,113), respectively, in the Tibetan population. The morbidity in herdsmen (16.6%) and Buddhist priests (15%) was significantly higher than that in other occupation groups (3.2%), and it was higher in females (9.8%) than in males (6.2%). The ultrasound images showed a coexistence of cystic echinococcosis (CE) and alveolar echinococcosis (AE), occupying 69 and 31% of the cases, respectively. An Echinococcus Western blot assay was performed as a serological backup test for differentiating CE and AE. The assay revealed that serum samples from most cases with a positive AE image showed a specific antibody against antigen bands at 16/18 kDa. Autopsy proved that 9 out of 12 stray dogs were infected with Echinococcus granulosus (n = 8) and E. multilocularis (n = 1). Inspection at the abattoirs demonstrated a hydatid rate of 78.5% in yaks and 82.6% in sheep. The data indicate that Jiuzhi County is an important endemic area for both CE and AE, in both human and animal populations.
Japanese journal of infectious diseases 06/2008; 61(3):242-6. · 1.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Parasitological and dissection methods were used to confirm Echinococcus infection and echinococcosis in animals, and to understand the epidemiological situation in final and intermediate animal hosts in Qinghai Province. Three Echinococcus species were found in the Province including E. granulosus, E. multilocularis and E. shiquicus. The prevalence of hydatid disease showed a tendency of increase with an average rate of over 50% in yaks and sheep. Higher prevalence in wild animals as intermediate host was found in the southern plateau, while that in final hosts such as dogs and wolves, the prevalence was higher in the area of Qilian mountain. There is a complex chain of life cycle of the parasites among animals, which poses a threat to the health of human population. Effective measures to control the disease in animals are urgently needed.
Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 09/2007; 25(4):350-2.
[Show abstract][Hide abstract] ABSTRACT: The survey was carried out in July, 2006 in Zhiduo County. The IHA and ELISA positive rate in human population was 4.5% (42/933) and 8.2% (76/931) respectively. Ultrasonography revealed a morbidity of 3.4% (33/979) with 3.2% Echinococcus granulosus and 0.2% of E. multilocularis respectively. Animal dissection showed an infection rate of 15.1% (14/93) in pikas with one infected by E. shiquicus proved by molecular biology. Coproantigen rate by ELISA was 62% (12/193) in dogs and 35.7% (5/14) in wolves. The results indicated that Zhiduo County is a mixed endemic area for echinococcosis.
Zhongguo ji sheng chong xue yu ji sheng chong bing za zhi = Chinese journal of parasitology & parasitic diseases 07/2007; 25(3):229-31.