Use of specific immunoglobulin G antibody avidity in the differential diagnosis of active and chronic Toxocara canis infections

To differentiate between acute and past Toxocara canis infections, a modified enzyme-linked immunosorbent assay, based on the dissociation of antigen-antibody complex with 6 M urea solution was used to measure levels of avidity of specific immunoglobulin G antibody against Toxocara canis excretory-secretory antigen. IgG avidity index was determined in the sera of 212 patients, divided into two groups with various stages of Toxocara infection. Patients were classified according to time period, from the onset of clinical symptoms or first positive serology result. Low IgG avidity index was found in the sera of 6.6% of patients in the acute phase, and all samples from the chronic stage of infection were characterized by high avidity values above 0.5 (mean 0.74). It was documented, that the avidity test for specific IgG antibodies is a valuable diagnostic method that helps to distinguish the early from the later phase of Toxocara canis infection. Indications for anti-parasitic treatment according to immunological activity of Toxocara infection were discussed.     

Antibody response of Echinococcus granulosus infected mice: Protoscolex specific response during infection is associated with decreasing specific IgG1/IgG3 ratio as well as decreasing avidity

The antibody response was followed during 68 weeks in 17 Balb/c mice intraperitoneally (i.p.) infected with Echinococcus granulosus protoscoleces (PSC) and in three mice i.p. immunized with dead PSC. Titres of antibodies recognizing peptidic and glucidic PSC epitopes, as well as their isotypic and avidity profiles were followed by ELISA. In addition, antigen recognition patterns were analysed by immunoblot. The response against carbohydrate epitopes was dominant in infected and immunized mice but stronger in the first group. Infected mice showed similar profiles of specific IgG and IgM with maximum titres from week 38 to 53. Although IgG1 and IgG3 were the predominant antibody subclasses, the ratio of IgG1/IgG3 antibody titres as well as antibody avidity decreased during the experiment, encompassing a decrease in recognition of peptidic epitopes. Immunized mice did not show significant levels of specific IgM and, after week 15, showed IgG titres lower than the infected mice. IgG1 was the predominant IgG subclass during all the experiment with background levels of IgG3. The mean Ab avidity was high and showed no significant changes during immunization. Different patterns of response were thus produced by dead and developing live parasites. Although high avidity IgG1 antibodies were early found in both cases, lower avidity IgG3 antibodies were increasingly produced afterwards only in infected animals. The isotype switch and avidity decrease observed only during infection are consistent with a possible parasitic mechanism to evade host immunity .     

新疆北部双峰驼细粒棘球蚴感染调查

目的：确定我国双峰驼中细粒球蚴的感染状况和寄生特征。方法：在屠宰场检查来自新疆北部各地骆驼体内的感染情况。测量棘球蚴囊的大小，检查内容物的性质并镜检有无原头节。结果：在３７５只骆驼中检出细粒棘球蚴感染者１８５只，感染率４９．３％。主要寄生在肝和肺，肝内较多，肝∶肺＝１∶０．６４。育囊携带率３４．８％，育囊率３９．２％， 囊指数７．５３。棘球蚴囊位于肝表面，单个存在，囊壁较薄，内无子囊。育囊平均直径在肝为５．６±２．５６ｃｍ，在肺为４．８±２．０３ｃｍ，钙化率高，单纯携带钙化病灶的骆驼占感染骆驼的６４．３％。结论：新疆北部双峰驼细粒棘球蚴的感染率很高。其寄生特征与本地区牛、羊中的细粒棘球蚴有明显区别，与非洲单峰驼亦有不同，应进一步深入研究。     

Serological reactivity to heat shock protein 70 in patients with hydatid disease

Heat shock protein 70 (hsp70) was chosen as a model antigen with which to investigate autoantibody production in humans with cystic hydatid disease. Levels of specific serum antibody were assessed in the sera of patients with surgically confirmed infection with E. granulosus and sera from non-infected controls. Antigens used were human hsp70, obtained from K562 cells grown in culture, and E. granulosus hsp70 obtained by expression of the full length protein in Escherichia coli following cloning of the associated mRNA. Antibody reactivity to human hsp70 was detected in the sera of only a small proportion of hydatid patients (10%) as well as a similar proportion of sera from age matched controls. Specific antibodies reactive with E. granulosus hsp70 were detected in 60% of hydatid patients, although some samples (21%) from healthy controls also reacted with E. granulosus hsp70, the level of reactivity was significantly higher in hydatid patients. This report identifies E. granulosus hsp70 as an immunogen during human hydatid infection but, despite its having a predicted 81% protein sequence homology with human hsp70, it does not appear to induce autoimmune reactivity against the homologous human protein.     

青海高原不同宿主源细粒棘球蚴的病原生物学考察

本文对我国青海高原牦牛和绵羊体内原发性细粒棘球蚴的病原生物学进行了考察。结果表明牦牛细粒棘球蚴的感染率和钙化率显著高于绵羊源(P<0.01);牦牛肝脏感染和肺脏感染无明显差异(各占34.14％和31.20％),绵羊则绝大多数寄生于肝脏(50.76％),肺脏较少(13.40％),肝、肺均感染者与牦牛相近(34.26％和32.58)。牦牛肝脏棘球蚴的钙化率高(73.00％),育囊率低(69.51％),但肺脏棘球蚴与绵羊肝、肺脏棘球蚴的钙化率相近(分别为20.15％、28.00％和18.04％),育囊率均高(分别为94.58％、91.76％和90.00％)。人体棘球蚴的育囊率达100.00％。对各源棘球蚴内的原头节测量结果表明不论原头节寄生在何种宿主,其肺脏原头节总是大于肝脏原头节(P<0.01);同源肝、肺脏原头节的吻钩大小无显著差异(P>0.05)。牦牛棘球蚴体积较绵羊的大,但原头节却小的多(P<0.01),且二者原头节吻钩的多项测量指标之间存在显著的差异(P<0.01或P<0.05)。上述结果提示对这种差异不应仅以棘球蚴寄生在不同宿主来解释,还应考虑到该地区是否存在不同宿主的细粒棘球绦虫虫株。     

Native and recombinant antigens in the immunodiagnosis of human cystic echinococcosis

To evaluate the diagnostic sensitivity and specificity of immunoelectrophoresis (IEP), indirect haemagglutination (IHA), enzyme-linked immunosorbent assay (ELISA) and immunoblotting (IB), we compared their ability in detecting IgG antibodies to a hydatid fluid fraction (HFF) and to native and recombinant antigen B of Echinococcus granulosus. We tested sera from patients who had cystic echinococcosis (CE) grouped according to their type of cysts (n = 204), from patients with other parasitic diseases (n = 21), lung or liver carcinomas (n = 6) or serous cysts (n = 26) and from healthy controls (n = 90). HFF-IB gave the highest sensitivity (80%) followed by ELISA (72%), IHA (54%) and IEP (31%), respectively. The diagnostic sensitivity significantly (P < 0.01) decreased as cysts matured from type I-II to type VII. Recombinant and native antigen B-IB yielded similar sensitivity (74%). A large number of clinically or surgically confirmed CE patients (20%) resulted negative. In these patients' sera, IB to assess the usefulness of the recombinant E. granulosus elongation factor-1 beta/delta in detecting IgE antibodies yielded 33% of positivity. Our findings underline the need to standardize techniques and antigenic preparations and to improve the performance of immunodiagnosis by characterizing new antigens and detecting distinct immunoglobulin classes.     

Antibody response in CD4-depleted mice after immunization or during early infection with Echinococcus granulosus

The aims of this work were to investigate the existence of T-independent antigens in Echinococcus granulosus protoscoleces and to evaluate the relative contribution of T-independent stimulation to the overall antibody response in early infection. Mice depleted of CD4(+)-cells were immunized with protoscolex somatic antigens (PSA) or infected with E. granulosus protoscoleces (PSC). Results showed that the response of CD4-depleted immunized mice had the expected characteristics of a T-independent stimulation and that such T-independent stimulation was important mainly during primary response. During infection absence of CD4(+)-cells affected mainly the secretion of all IgG subclasses with the exception of IgG3 and IgM. To carry out a preliminary isolation of PSC T-independent antigens we prepared a carbohydrate enriched fraction from protoscolex antigens, using a monoclonal antibody specific for the carbohydrate moiety Gal alpha(1,4)Gal highly expressed in PSC. This fraction was mitogenic for naive mouse splenocytes and was recognized by a high percentage of the specific antibodies secreted by CD4-depleted immunized or infected mice. In summary, these results suggest that E. granulosus protoscoleces contain immunogenic T-independent antigens. Primary antibody responses to protoscolex somatic antigens and the production of IgM and IgG3 in early infection would be mainly stimulated by a T-independent mechanism.     

''Arc 5'' antibodies in sera of sheep infected with Echinococcus granulosus, Taenia hydatigena and Taenia ovis

The 'arc 5' precipitin band, formed when test human serum is reacted against immunoelectophoresed hydatid cyst fluid antigen, has provided a positive diagnosis of Echinococcus granulosus infection. These antibodies to 'arc 5' antigen have now been found in the sera of sheep. They appear 2 weeks after infection with Taenia ovis, after 3 weeks with T. hydatigena and after 16 weeks with E. granulosus. An antigen similar to the 'arc 5' antigen of E. granulosus cyst fluid was also demonstrated in cyst fluid from T. hydatigena, but it would not be positively identified in T. ovis cyst fluid. The presence of 'arc 5' in immunoelectrophoresis tests is not suitable for specific immunodiagnosis of E. granulosus infections in sheep in New Zealand. 'Arc 5' antibodies were only associated with living E. granulosus cysts and were not present if cysts were dead. The location of the cysts in either liver or lungs and the onset of brood capsule production did not influence the presence of 'arc 5' antibodies.     

An enzyme-linked immunosorbent assay for detection of IgG1 antibodies specific to human cystic echinococcosis in Egypt

Human cystic hydatidosis (cystic echinococcosis) is a chronic zoonotic disease that results from infection with the dog tapeworm Echinococcus granulosus. In Egypt, cystic echinococcosis (CE) is recognized in slaughtered livestock by veterinarians, however, there is little information about human CE infection rates. We describe an immunological assay useful for the diagnosis of human cystic hydatidosis. Sera were collected from surgically confirmed hydatid cases (34), nonendemic subjects free from parasitic infection (20) and from subjects (109) infected with other helminths (Hymenolepis nana, Schistosoma mansoni, Fasciola hepatica and Ancylostoma duodenale). Hydatid cyst fluid (HCF) of camel origin was used as antigen in an ELISA format to measure total E. granulosus specific IgG antibodies and IgG subclasses. Sensitivity measurements of total IgG, and IgG1-4 were 100, 100, 79.4, 61.8 and 55.9%, respectively, whereas respective specificity reached 65.1, 97.7, 98.4, 96.1 and 83. 7%. The diagnostic value of measuring IgG1 (97.7%), as assessed by a rating index (J) for combined sensitivity and specificity, was superior to total IgG (65.1%) and IgG2-4 (77.8, 57.9 and 39.6%, respectively). These findings set the stage for field evaluation of the IgG1 assay in areas endemic with human cystic hydatidosis.     

细粒棘球蚴重组抗原B 8-kDa亚单位1对囊型包虫病的血清学诊断价值

目的通过基因工程技术获得细粒棘球蚴抗原B8-kDa亚单位1重组蛋白(rEgAgB8/1),探讨其对囊型包虫病(CE)的血清学诊断价值。方法将构建的rEgAgB8/1原核表达质粒(pET32b-rEgAgB8/1)转化至E.coli BL-21(DE3)中,用IPTG诱导表达,经亲和层析纯化获得高纯度rEgAgB8/1,以rEgAgB8/1为抗原,应用ELISA和Immuno blotting方法对31例手术确诊的囊型包虫病病人血清进行了回顾性检测与分析。结果ELISA和Immuno blotting方法检测CE病人血清阳性率均为90.3%(28/31),3例血清学检测阴性的CE病人均为初次诊断为CE及单纯性肝脏单发感染的病人;血清抗体水平随着病人棘球蚴囊数目增加而有所增加,棘球蚴囊的数目与血清抗体水平的比较用单因素方差分析有显著性差异(F=5.06,P=0.0142),1个囊与2个囊/3个囊组间血清抗体水平有显著差异,2个囊与3个囊组间差异无统计学意义。结论rEgAgB8/1重组蛋白抗原对囊型包虫病有较高的血清学诊断价值,多囊型包虫病人血清抗体水平高于单囊型包虫病人。     

棘球蚴3种抗原在包虫病斑点酶联免疫吸附试验中特异性和敏感性比较

用羊源细粒棘球蚴囊液、原头蚴及生发层３种抗原,用酶标记羊抗人ＩｇＧ,对９４例包虫病患者、４４例非包虫患者及５０例健康人血清进行了斑点酶联免疫吸附试验。结果用囊液、原头蚴、生发层３种抗原的敏感性分别为９２．６％、９３．６％和９３．６％,５０例健康人血清均未出现假阳性反应,与各种肿瘤、各种炎症、各种肝病及结核患者血清有一定的交叉反应     

包虫病人血清特异性抗包虫IgG和IgM的检测及其临床意义探讨

本文报告应用IFAT法检测包虫病人血清中特异性包虫IgG和IgM的初步结果。44例正常人血清特异性IgG和IgM的假阳性率均为0％,83例包虫病患者血清特异性IgG和IgM的阳性检出率分别为96.36％和19.28％,二者有极显著性差异(P<0.01)。提示现症包虫病患者血清中的特异性抗包虫抗体主要为IgG。在16例特异性IgM阳性病人中,有9例存在包虫囊破裂,7例为肺包虫或肝肺混合包虫病患者,表明特异性IgM在包虫囊破裂时升高,肺包虫病人比肝包虫病人检出率高。提示包虫病人血清中的特异性IgM抗体水平与包虫囊的生活状态有关。     

Cloning and expression of a cDNA encoding an elongation factor 1β/δ protein from Echinococcus granulosus with immunogenic activity

A cDNA clone (Eg EF-1beta/delta) of Echinococcus granulosus has been isolated by an expression library screened with immunoglobulin (Ig)E of sera from patients with cystic echinococcosis (CE). The Eg EF-1beta/delta was identified on the basis of sequence homology to the subunits beta or delta of the elongation factor-1. The amino acid sequence deduced from this open reading frame is 244 residues long with a predicted molecular mass of 31 kDa. In Southern blot under high stringent condition, Eg EF-1beta/delta hybridized to genomic DNA of E.granulosus at two bands of 4 and 2.5 Kb. In immunoblotting analysis, the Eg EF-1beta/delta protein shows immunological reactivity with sera from CE patients: 51.7% of sera contained IgE, 41.7% IgG and 18.3% IgG4 specific to the recombinant protein. We identify the Eg EF-1beta/delta by immunoblotting with specific monoclonal antibody both in protoscoleces and in sheep hydatid fluid. The higher percentage of humoral immune response against Eg EF-1beta/delta observed in CE patients with calcified cysts than in patients with active cysts indicates the possible release of the protein in the hydatid fluid after protoscoleces degeneration suggesting the possible use of this antigen in the immunosurveillance of CE. Overall, these findings seem to assign to Eg EF-1beta/delta a key role in the allergic disorders and in the complex host-parasite relationship in CE.     

Rapid dot-ELISA for the detection of specific antigens in the cyst fluid from human cases of cystic echinococcosis

A rapid dot-ELISA was developed for the detection of specific antigens in samples of cyst fluid from human cases of Echinococcus granulosus infection, permitting the confirmation of cystic echinococcosis (CE). Peroxidase-conjugated antibodies against antigen B (derived from the fluid in the cysts of sheep infected with E. granulosus) were used to test cyst-fluid samples from 22, surgically confirmed cases of human CE and 21 domestic animals (horses, sheep, buffalo, a cow and a camel) with CE. All of these samples were found to be strongly positive in the dot-ELISA, by direct reading with the naked eye. In contrast, fluid samples from seven, non-parasitic liver cysts of human origin were all negative, and a sample taken from the residual cavity left in a patient after previous CE surgery showed a weakly positive reaction. The antigen-detection assay, which can be completed within 10 min, may be a useful 'bedside' test for the differential diagnosis of cysts during surgery or percutaneous treatment for suspected CE.     

Serological monitoring of protection of sheep against Echinococcus granulosus induced by the EG95 vaccine

Although immunity to Echinococcus granulosus in sheep has been shown to be antibody-mediated and complement-dependent and can be passively transferred in colostrum, in animals vaccinated with EG95, the relationship between protection against an oral challenge infection with E. granulosus eggs and anti-EG95 IgG ELISA absorbance values at the time of challenge has not been satisfactorily proven. Using a combination of results from three EG95 vaccination trials, we have found that the IgG ELISA absorbance at the time of challenge infection explains approximately 50% (P ≤ 0·001) of the variability in the percentage protection against an oral challenge with E. granulosus eggs (transformed with arcsin).     

Human cystic echinococcosis: epidemiologic, zoonotic, clinical, diagnostic and therapeutic aspects

This review represents an updated scenario on the transmission cycle, epidemiology, clinical features and pathogenicity, diagnosis and treatment, and prevention and control measures of a cestode parasite Echincoccus granulosus (E. granulosus) infection causing cystic echinococcosis (CE) in humans. Human CE is a serious life-threatening neglected zoonotic disease that occurs in both developing and developed countries, and is recognized as a major public health problem. The life cycle of E. granulosus involves a definitive host (dogs and other canids) for the adult E. granulosus that resides in the intestine, and an intermediate host (sheep and other herbivores) for the tissue-invading metacestode (larval) stage. Humans are only incidentally infected; since the completion of the life cycle of E. granulosus depends on carnivores feeding on herbivores bearing hydatid cysts with viable protoscoleces, humans represent usually the dead end for the parasite. On ingestion of E. granulosus eggs, hydatid cysts are formed mostly in liver and lungs, and occasionally in other organs of human body, which are considered as uncommon sites of localization of hydatid cysts. The diagnosis of extrahepatic echinococcal disease is more accurate today because of the availability of new imaging techniques, and the current treatments include surgery and percutaneous drainage, and chemotherapy (albendazole and mebendazole). But, the wild animals that involve in sylvatic cycle may overlap and interact with the domestic sheep-dog cycle, and thus complicating the control efforts. The updated facts and phenomena regarding human and animal CE presented herein are due to the web search of SCI and non-SCI journals.     

Kinetics of the SARS-CoV-2 Antibody Avidity Response Following Infection and Vaccination

Serological assays capable of measuring antibody responses induced by previous infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been critical tools in the response to the COVID-19 pandemic. In this study, we use bead-based multiplex assays to measure IgG and IgA antibodies and IgG avidity to five SARS-CoV-2 antigens (Spike (S), receptor-binding domain (RBD), Nucleocapsid (N), S subunit 2, and Membrane-Envelope fusion (ME)). These assays were performed in several cohorts of healthcare workers and nursing home residents, who were followed for up to eleven months after SARS-CoV-2 infection or up to six months after vaccination. Our results show distinct kinetic patterns of antibody quantity (IgG and IgA) and avidity. While IgG and IgA antibody levels waned over time, with IgA antibody levels waning more rapidly, avidity increased with time after infection or vaccination. These contrasting kinetic patterns allow for the estimation of time since previous SARS-CoV-2 infection. Including avidity measurements in addition to antibody levels in a classification algorithm for estimating time since infection led to a substantial improvement in accuracy, from 62% to 78%. The inclusion of antibody avidity in panels of serological assays can yield valuable information for improving serosurveillance during SARS-CoV-2 epidemics.     

Epidemiology of hydatidosis/echinococcosis in Ouarzazate, the pre-Saharian region of Morocco

In the Ouarzazate province of Southern Morocco, 1085 cattle and 358 sheep were examined for hydatid cysts. The prevalence was 44.6% (range 8.3-83.4%) in cattle and 5.3% (range 1.3-28.6%) in sheep. The prevalence increased with the age of the animals. The lung was the predominant site of infection, followed by the liver. In cattle, 14.2% of hydatid cysts were fertile and 16.4% had degenerative changes. The mean loss per head of cattle slaughtered was about 1 kg of liver and 900 g of lungs in the urban abattoirs. In monetary terms at the current price of 1980, this meant a loss of U.S. +4.5 at urban and +1.6 at rural abattoirs. The mean prevalence of infection in 61 stray dogs was 50.8%, with a mean worm burden of 413. The urban dogs had a lower infection rate (42.9%) than the rural dogs (61.5%). There were 130 dogs per 1000 inhabitants. The human population was ignorant of the life-cycle of Echinococcus granulosus and was not aware of risk of infection through dogs. Dogs become infected at abattoirs, at home slaughter and in the field by consuming dead carcasses. The maintenance and transmission of E. granulosus in animals and man is related to social, religious and cultural factors.     

细粒棘球蚴重组抗原rEgferritin诱导宿主抗感染免疫应答的初步研究

目的 初步探讨细粒棘球蚴重组铁蛋白rEgferritin免疫小鼠产生的抗细粒棘球蚴感染的免疫机制.方法 rEgferritin免疫ICR小鼠3次、同时设立佐剂组和对照组,第12w用细粒棘球蚴原头蚴对3组小鼠进行攻击感染,5个月后剖杀小鼠,检查各组小鼠腹腔内包囊个数,根据公式计算保护力.分别于0w,12w,32w取血,制...     

T-cell activity associated with secondary infections and implanted cysts of Echinococcus granulosus in BALB/c mice

Selected cytokine profiles of lymphocytes were assessed in BALB/c mice infected with protoscoleces of Echinococcus granulosus . Late stages of the infection (three months +) were characterized by more dominant Th2 activity with elevated IL-4 and IL-10 and reduced IFN-γ output by Con-A and antigen stimulated splenocytes. Circumparasitic leucocytes produced mainly IL-10 by five months post infection. A peak in IFN-γ production in the first month of infection may suggest Th1 or Th0 activity at this time and this may be correlated with initial protoscolex death. In addition, cytokine profiles from mice implanted with intact hydatid cysts were also assessed. At two weeks post implantation all cysts were still viable and cytokine production was characterized mainly by elevated IL-10 production. However, at four months post implantation, some of the cysts from two mice had been killed whilst all cysts in the remaining mouse remained viable. In the mice where dead cysts were present, elevated levels of IFN-γ were detected from splenocytes and circumparasitic cells. Elevated IL-4 was also evident with the splenocytes. In the mouse with viable cysts IFN-γ production was reduced. Results indicate that IFN-γ (Th1) activity may be correlated with killing of both protoscoleces and established cysts of E. granulosus.