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The protein C-type lectin is secreted by the secondary-stage larve of Toxocara canis (T. canis). Its antigenic characteristics have been the subject of research. The recombinant pET-32a (+) plasmid containing the 660 bp sequence of T. canis C-type lectin gene was successfully synthesized, cloned, and expressed in Escherichia coli (DE3+). Serum samples were collected from 56 pet owners. Forty-four samples were negative and twelve samples were positive which were performed with a commercial ELISA kit. The same samples were also tested using dot blot and Western blot analysis prepared with recombinant CTL antigen (rCTL). The expression was confirmed by Sodium Dodecyl Sulfate (SDS-PAGE) and Western blot. The study results revealed that two samples, which tested negative in the ELISA analysis, were found to be positive when using the recombinant C-type lectin (rCTL) antigen in both dot blot and Western blot analyses. The overall test results showed a high level of agreement between the three methods, with Kappa coefficients of 1 and 0.9 for the comparisons. This indicates the potential of using recombinant CTL antigen, to detect positive samples that may be missed by E-S Antigen. The recombinant rCTL demonstrated higher sensitivity than the excretory-secretory Antigens, indicating its potential for improved serodiagnosis of human toxocariasis. The study proposes that the rCTL could be further evaluated in supplementary studies to improve and develop the diagnostic kits for toxocariasis. [ABSTRACT FROM AUTHOR]

BACKGROUND: Toxocara canis C-type lectin (T. canis-CTL) is the main protein part of the secretory-excretory product secreted by Toxocara canis infective larvae. T. canis-CTL can stimulate immune response-mediated regulatory T lymphocytes, increase the FOXP3+ cells population, and reduce severe inflammatory responses. T. canis-CTL is a promising candidate for immune modulation in some autoimmune diseases, deserving further investigation. OBJECTIVES: The current research aimed to purify the recombinant T. canis-CTL under denaturation and native condition to increase exploration and maintain biological activity. METHODS: The expression vector, pET32a was constructed with the partial sequence 660bp of T. canis-CTL and expressed in E. coli BL21 (DE3). T. canis-CTL protein expression was induced by IPTG (1 mM) at 37°C after 6 h. In this study, different buffers were used for cell explosion and recombinant protein solubilization, including lysis buffer with urea (8 M, pH=8) and lysozyme enzyme as well as lysis buffer with Imidazole (0.01 M) and lysozyme enzyme, and previous buffers in addition to sonication. The effect of these buffers was evaluated in bacterial cells explosion, using Gram-staining and microscopic examination. Recombinant T. canis-CTL protein was extracted and purified under denaturation and native condition using Ni-NTA affinity chromatography by agarose and sepharose resin. A New Zealand male rabbit was immunized with the recombinant protein to evaluate the bioactivity of the protein. RESULTS: Lysis buffer with urea (8 M, pH=8) and lysozyme enzyme, in addition to sonication, provided acceptable results, and an additional amount of recombinant T. canis-CTL protein was secreted in the buffer. Protein purification under denaturation conditions with Ni-NTA agarose affinity chromatography also provides further recombinant protein. Most of the induction of recombinant T. canis-CTL with 41 KDa molecular weight was collected 6 h after induction at 37°C. Dot-blot results illustrate the brown dot, which showed a 1:500 titer of specific IgG polyclonal antibody has developed in the sera of rabbits immunized with T. canis-CTL recombinant protein. CONCLUSIONS: The denaturation condition did not affect the biological activity of the T. canis-CTL recombinant protein and can recover a further amount of recombinant proteins. [ABSTRACT FROM AUTHOR]

Echinococcoses require the involvement of specialists from nearly all disciplines; standardization of the terminology used in the field is thus crucial. To harmonize echinococcosis terminology on sound scientific and linguistic grounds, the World Association of Echinococcosis launched a Formal Consensus process. Under the coordination of a Steering and Writing Group (SWG), a Consultation and Rating Group (CRG) had the main missions of (1) providing input on the list of terms drafted by the SWG, taking into account the available literature and the participants’ experience; and (2) providing independent rating on all debated terms submitted to vote. The mission of the Reading and Review Group (RRG) was to give an opinion about the recommendation paper in terms of readability, acceptability and applicability. The main achievements of this process were: (1) an update of the current nomenclature of Echinococcus spp.; (2) an agreement on three names of diseases due to Echinococcus spp.: Cystic Echinococcosis (CE), Alveolar Echinococcosis (AE) and Neotropical Echinococcosis (NE), and the exclusion of all other names; (3) an agreement on the restricted use of the adjective “hydatid” to refer to the cyst and fluid due to E. granulosus sensu lato; and (4) an agreement on a standardized description of the surgical operations for CE, according to the “Approach, cyst Opening, Resection, and Completeness” (AORC) framework. In addition, 95 “approved” and 60 “rejected” terms were listed. The recommendations provided in this paper will be applicable to scientific publications in English and communication with professionals. They will be used for translation into other languages spoken in endemic countries.

Background: Echinococcosis/hydatidosis is one of the most important zoonotic diseases. The definitive hosts for Echinococcus granulosus (E. granulosus) include a wide variety of the Canidae and dogs. Early detection of Echinococcosis in dogs is the most influential factor in improving the prevention and control of hydatidosis. The primary purpose of the present study was to optimize the real-time quantitative PCR to diagnose E. granulosus infection in dogs before the disposal of eggs. Materials and Methods: Three puppies were selected to be inoculated by 70000 protoscoleces. Normal saline was inoculated to the other two puppies chosen as an experimental negative control group. Ten privately owned healthy puppies were selected for the natural negative control group. Stool samples were collected on days 7, 14, 21, 28, and 35 post-infection, DNA was extracted, then a 287bp fragment of tandem repeat region gene was amplified and cloned into linearized TA vectors. Serial dilutions of recombinant plasmid DNA and a standard curve were established. The copy amount of DNA in each sample was determined based on the standard curve. Results: The minimum time copro -DNA could be detected in the stool sample was found on the 7th day post-infection, which was equal to 9750×10-8 copy number or 9.75 pg of DNA. The assay was linear in 105 to 108 copies of the recombinant plasmid per microliter. Conclusion: The real-time PCR assay diagnosed the infection eight days earlier than the copro antigen ELISA method (7 days versus 15 days, respectively). [ABSTRACT FROM AUTHOR]

Iran is one of the endemic areas of Dirofilariasis, and also one of the most important zoonotic infections. Dirofilaria immitis causes a severe and fatal disease called heartworm disease in dog. It also produces pulmonary nodules in humans. The worm is to be investigated as a potential infection of humans and animals in various provinces in Iran. In this research, the samples were studied with modified Knott's test and molecular method. The results of the modified Knott's test method indicated that 14.00% of sheepdogs were infected with filarial microfiler. The microfilers were characterized with basic morphological features, the length of the infective larva and tail ending. There was an estimated prevalence of 4.45% for Dirofilaria immitis and 9.55% for Acanthocheilonema reconditum microfiler. To verify the differential diagnosis, molecular method was performed using PCR with Dirofilaria specific primers for amplification of ITS2 locus. Gene locus sequencing results of D. immitis and sequence alignment recorded in GeneBank showed 97.00% similarity, and relatively 98.00% similarity was observed in A. reconditum. The results of the molecular method confirmed the result of modified Knott's test method. Low infection with D. immitis was observed the region, probably due to the fact that the annual temperature and precipitation in this area were not suitable for the proliferation of the vector mosquitoes. In general, there was less infection in the region compared to regions with relatively similar climatic conditions. Hence, the results suggested that alternative diagnostic tests are required to determine the occult infections. [ABSTRACT FROM AUTHOR]