Characterization of the Accessory Layer of the Cuticle of Muscle Larvae of Trichinella spiralis

Abstract Background: The main targets of the host’s immune system in Trichinella spiralis infection are the adult worms (AW), at the gut level, and the migrant or newborn larvae (NBL), at systemic and pulmonary levels. Most of the studies carried out in the gut mucosa have been performed on the Payer’s patches and/or the mesenteric lymph nodes but not on the lamina propria, therefore, knowledge on the gut immune response against T. spiralis remains incomplete. Methods This study aimed at characterizing the early mucosal immune response against T. spiralis, particularly, the events taking place between 1 and 13 dpi. For this purpose, Wistar rats were orally infected with muscle larvae of T. spiralis and the humoral and cellular parameters of the gut immunity were analysed, including the evaluation of the ADCC mechanism exerted by lamina propria cells. Results A marked inflammation and structural alteration of the mucosa was found. The changes involved an increase in goblet cells, eosinophils and mast cells, and B and T lymphocytes, initially displaying a Th1 profile, characterised by the secretion of IFN-γ and IL-12, followed by a polarization towards a Th2 profile, with a marked increase in IgE, IgG1, IL-4, IL-5 and IL-13 levels, which occurred once the infection was established. In addition, the helminthotoxic activity of lamina propria cells demonstrated the role of the intestine as a place of migrant larvae destruction, indicating that not all the NBLs released in the gut will be able to reach the muscles. Conclusions The characterization of the immune response triggered in the gut mucosa during T. spiralis infection showed that not only an effector mechanism is directed toward the AW but also towards the NBL as a cytotoxic activity was observed against NBL exerted by lamina propria cells.

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Characterization of a Novel Glutamine Synthetase From Trichinella spiralis and Its Participation in Larval Acid Resistance, Molting, and Development

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.729402 ◽

2021 ◽

Vol 9 ◽

Author(s):

Tong Xu Zhuo ◽

Zhen Wang ◽

Yan Yan Song ◽

Shu Wei Yan ◽

Ruo Dan Liu ◽

...

Keyword(s):

Escherichia Coli ◽

Life Cycle ◽

Glutamine Synthetase ◽

Enzymatic Activity ◽

Post Infection ◽

Catalytic Domain ◽

Trichinella Spiralis ◽

Acid Resistance ◽

Muscle Larvae

Trichinella spiralis is a major foodborne parasite worldwide. After the encapsulated muscle larvae (ML) in meat are ingested, the ML are liberated in the stomach of the host and activated into intestinal infectious larvae (IIL), which develop into adult worm after molting four times. A novel glutamine synthetase (TsGS) was identified from T. spiralis IIL at 10 h post-infection, but its biological role in T. spiralis life cycle is not clear. The aim of this study was to investigate the biological characteristics of TsGS and its functions in larval acid resistance, molting, and development. TsGS has a glutamine synthetase (GS) catalytic domain. Complete TsGS sequence was cloned and expressed in Escherichia coli BL21. rTsGS has good immunogenicity. qPCR and Western blotting showed that TsGS was highly expressed at IIL stage, and immunofluorescence revealed that TsGS was principally localized at the cuticle and intrauterine embryos of this nematode. rTsGS has enzymatic activity of natural GS to hydrolyze the substrate (Glu, ATP, and NH4+). Silencing of TsGS gene significantly reduced the IIL survival at pH 2.5, decreased the IIL burden, and impeded larval molting and development. The results demonstrated that TsGS participates in T. spiralis larval acid resistance, molting and development, and it might be a candidate vaccine target against Trichinella molting and development.

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Characterization of Spanish Trichinella isolates by random amplified polymorphic DNA (RAPD)

Journal of Helminthology ◽

10.1017/s0022149x00015637 ◽

1996 ◽

Vol 70 (4) ◽

pp. 335-343 ◽

Cited By ~ 10

Author(s):

E. Rodríguez ◽

J. Nieto ◽

J.A. Castillo ◽

T. Gárate

Keyword(s):

Copy Number ◽

Rapd Markers ◽

Trichinella Spiralis ◽

Random Amplified Polymorphic Dna ◽

Arbitrary Primers ◽

Cross Hybridization ◽

Muscle Larvae ◽

Genetic Clusters ◽

Species Specific

AbstractThe random amplified polymorphic DNA (RAPD) assay was used to find molecular markers able to distinguish Trichinella spiralis from T. britovi, the two recognized Spanish Trichinella species. Fourteen Spanish Trichinella isolates, as well as reference Trichinella isolates representing the five species T. spiralis (T1), T. nativa (T2), T. britovi (T3), T. pseudospiralis (T4) and T. nelsoni (T7) and the three other taxa Trichinella T5, Trichinella T6 and Trichinella T8 of the genus, were characterized by RAPD using both purified and crude DNAs from infective muscle larvae (ML) and seven arbitrary primers. Three primers yielded diagnostic RAPD markers for the Spanish T. spiralis and T. britovi isolates as well as for the Trichinella reference isolates analysed, and in the case of crude DNAs the results were obtained in few hours. In addition, the species-specificity of the diagnostic RAPD markers from Spanish Trichinella isolates was studied by cross-hybridization assays. These assays confirmed that the selected diagnostic DNA fragments were not species-specific, but showed potential differences in the copy number among the examined Trichinella genetic clusters.

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Influence of chemically and biosynthesized silver nanoparticles on in vitro viability and infectivity of Trichinella spiralis muscle larvae

10.1101/2020.12.17.423206 ◽

2021 ◽

Author(s):

ahmed kamal dyab ◽

Salwa Mahmoud Abd-ELrahman ◽

Abeer El-sayed Mahmoud ◽

Nahed Ahmed Elossily ◽

Fahd mohammed Alsharif ◽

...

Keyword(s):

Trichinella Spiralis ◽

In Vitro Study ◽

Transmission Electron ◽

Muscle Larvae ◽

Principal Finding ◽

Vitro Model ◽

Biosynthesized Agnps ◽

Vitro Effect

Background Trichinellosis is a serious worldwide parasitic zoonosis. The available therapy for the treatment of Trichinella spiralis is not satisfactory. This work aimed at evaluating of the in vitro effect of silver Therefore, the recovery of effective treatment is required.nanoparticles (AgNPs) on muscle larvae of Trichinella. Methodology / principal finding The present study investigated the larvicidal properties of chemical and myrrh AgNPs on muscle larvae (ML) of T. spiralis. The used AgNPs were chemically prepared using NaBH4 as reducing agent and biosynthesized using methanolic myrrh extract. Characterization of synthesized AgNPs was monitored via UV-Vis spectrophotometry, Fourier transform infrared spectroscopy and transmission electron microscopy (TEM) studies. The ML incubated with AgNPs at concentrations ranged from 1μg/ml to 20μg/ml. Conclusions /Significance Chemical and biosynthesized AgNPs revealed marked larvicidal effect against ML of Trichinella. Additionally, this in vitro study showed degenerative changes affecting the cuticle of AgNPs treated ML. The effectiveness of AgNPs on the infectivity of Trichinella ML was also assessed. The results showed complete inhibition of the infectivity of ML exposed to sublethal doses of chemical and myrrh prepared AgNPs when used to infect animal models. This is the first report where myrrh synthesized AgNPs have been tested for their anthelminthic activity against Trichinella in an in vitro model.

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Trichinella spiralis: Partial Characterization of Antigens Isolated by Immuno-Affinity Chromatography from the Large-Particle Fraction of the Muscle Larvae

Journal of Parasitology ◽

10.2307/3280552 ◽

1981 ◽

Vol 67 (3) ◽

pp. 332 ◽

Cited By ~ 19

Author(s):

Dickson D. Despommier ◽

Anthony Laccetti

Keyword(s):

Affinity Chromatography ◽

Large Particle ◽

Trichinella Spiralis ◽

Partial Characterization ◽

Particle Fraction ◽

Muscle Larvae

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Preparation and characterization of monoclonal antibodies against ES antigens of Trichinella spiralis

Helminthologia ◽

10.2478/s11687-014-0238-3 ◽

2014 ◽

Vol 51 (4) ◽

pp. 253-261 ◽

Cited By ~ 2

Author(s):

J. Cui ◽

F. Jing ◽

G. Fu ◽

H. Ren ◽

L. Liu ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Western Blot ◽

Post Infection ◽

Trichinella Spiralis ◽

Internal Organs ◽

Cross Reactions ◽

Muscle Larvae ◽

Somatic Antigens

AbstractHybridomas secreting monoclonal antibodies (MAbs) against excretory-secretory (ES) antigen of Trichinella spiralis muscle larvae were produced. The 12 monoclones (designated 3A11, 7D9, 7F6, 5A7, 6H7, 39F3, 38H2, 37E7, 35B9, 29A11, 39C9 and 39A6) secreted IgM, IgG3 and IgG2b, respectively. Western blot using T. spiralis ES antigens showed that ten of 12 MAbs recognized the bands between 119.8 − 19.3kDa (mainly 68.4 − 28.7kDa) and two MAbs (3A11, 37E7) did not recognize any protein constituents of ES antigens. Nine of 12 MAbs also recognized the larval antigens of T. nativa, T. pseudospiralis and T. nelsoni at 40 days post-infection (dpi). No cross-reactions were found with the somatic antigens of adult worms of P. westermani, C. sinensis, and T. solium cysticercus. However, three (3A11, 6H7 and 39A6) were cross-reacted with the somatic antigens of adult worms of S. japonicum. Immunofluorescent location showed that the nine MAbs reacted with the cuticle and internal organs of T. spiralis larvae. Additionally, five and eight MAbs generated in this study can recognize the early antigens of pre-encapsulated T. spiralis larvae at 11 dpi and 13 dpi, respectively. The generation and characterization of the MAbs against T. spiralis ES antigens provide foundation for the development of specific early serodiagnostic techniques for trichinellosis.

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Characterization of a novel cysteine protease in Trichinella spiralis and its role in larval intrusion, development and fecundity

Veterinary Research ◽

10.1186/s13567-021-00983-1 ◽

2021 ◽

Vol 52 (1) ◽

Author(s):

Yuan Yuan Hu ◽

Ru Zhang ◽

Shu Wei Yan ◽

Wen Wen Yue ◽

Jia Hang Zhang ◽

...

Keyword(s):

Cysteine Protease ◽

Vaccine Development ◽

Trichinella Spiralis ◽

Biological Properties ◽

Dependent Manner ◽

Intestinal Epithelial ◽

Muscle Larvae ◽

Candidate Target ◽

Dose Dependent

AbstractThe aim of this study was to investigate the biological properties of a novel gut-specific cysteine protease in Trichinella spiralis (TsGSCP) and its role in larval intrusion, development and fecundity. TsGSCP has a functional C1 peptidase domain; C1 peptidase belongs to cathepsin B family. The TsGSCP gene cloned and expressed in Escherichia coli BL21 showed intensive immunogenicity. qPCR and Western blotting revealed that TsGSCP mRNA and protein were expressed at various T. spiralis stages, but their expression levels in intestinal infectious larvae (IIL) were clearly higher than those in muscle larvae (ML), adult worms (AWs) and new-born larvae (NBL). Indirect immunofluorescence (IIF) analysis showed that TsGSCP was primarily located at the outer cuticle and the intrauterine embryos of this parasite. rTsGSCP showed the ability to specifically bind with IECs, and the binding site is within the IEC cytoplasm. rTsGSCP accelerated larval intrusion into host intestinal epithelial cells (IECs), whereas anti-rTsGSCP antibodies suppressed larval intrusion; the acceleration and suppression was induced by rTsGSCP and anti-rTsGSCP antibodies, respectively, in a dose-dependent manner. When ML were transfected with TsGSCP-specific dsRNA, TsGSCP expression and enzymatic activity were reduced by 46.82 and 37.39%, respectively, and the capacity of the larvae to intrude into IECs was also obviously impeded. Intestinal AW burden and adult female length and fecundity were significantly decreased in the group of mice infected with dsRNA-transfected ML compared to the control dsRNA and PBS groups. The results showed that TsGSCP plays a principal role in gut intrusion, worm development and fecundity in the T. spiralis lifecycle and might be a candidate target for vaccine development against Trichinella intrusion and infection.

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Molecular cloning and characterization of a serine proteinase inhibitor from Trichinella spiralis

Parasitology ◽

10.1017/s0031182001008010 ◽

2001 ◽

Vol 123 (1) ◽

pp. 77-83 ◽

Cited By ~ 27

Author(s):

I. NAGANO ◽

Z. WU ◽

T. NAKADA ◽

A. MATSUO ◽

Y. TAKAHASHI

Keyword(s):

Recombinant Protein ◽

Proteinase Inhibitors ◽

Trichinella Spiralis ◽

Serine Proteinase ◽

Predicted Amino Acid Sequence ◽

Serpin Gene ◽

Specific Expression ◽

Crude Extracts ◽

Muscle Larvae

We produced a recombinant protein from a cDNA library from muscle larvae of Trichinella spiralis which had proteinase inhibitory activity. The predicted amino acid sequence of the clone had an identity of only 30% to the serine proteinase inhibitors (serpins) from Caenorhabditis elegans or Brugia malayi. At the putative reactive region, however, the identity was about 50%. The recombinant protein expressed in Escherichia coli inhibited 82% of the activity of the serine proteinase (trypsin). Stage-specific expression of this protein was suggested from the following experiments. Antibody against the recombinant protein could stain proteins migrating at about 42 kDa (which is the expected size from the sequence) in crude extracts from newborn larvae and 18-day post-infection (p.i.) muscle larvae, but it failed to stain any proteins in crude extracts from 30-day p.i. muscle larvae. Production of mRNA transcript for the serpin gene was restricted largely to the newborn larvae and to 18-day p.i. muscle larvae. The antibody reacted with the stichocytes of the larvae at 18 days p.i., but did not react with the muscle larvae at 24 days and 30 days p.i.

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Development of a rapid and sensitive immunochromatographic strip based on EuNPs-ES fluorescent probe for the detection of early Trichinella spiralis-specific IgG antibody in pigs

Veterinary Research ◽

10.1186/s13567-021-00951-9 ◽

2021 ◽

Vol 52 (1) ◽

Author(s):

Xinyu Wang ◽

Bin Tang ◽

Ying Zhao ◽

Jing Ding ◽

Nan Wang ◽

...

Keyword(s):

Test Line ◽

Trichinella Spiralis ◽

Parasite Infection ◽

Igg Antibody ◽

Immunochromatographic Strip ◽

Muscle Larvae ◽

Zoonotic Parasite ◽

Circulating Antibodies ◽

Novel Method ◽

Specific Igg

AbstractTrichinellosis, which is caused by nematodes of the genus Trichinella, is one of the most important zoonotic parasite diseases in the world. A rapid and sensitive immunochromatographic strip (ICS) based on Eu (III) nanoparticles (EuNPs) was developed for the detection of Trichinella spiralis (T. spiralis) infection in pigs. T. spiralis muscle larvae excretory secretory or preadult worm excretory secretory (ML-ES or PAW-ES) antigens were conjugated with EuNPs probes to capture T. spiralis-specific antibodies in pig sera, after which the complex bound to mouse anti-pig IgG deposited on the test line (T-line), producing a fluorescent signal. In the pigs infected with 100, 1000 and 10 000 ML, seroconversion was first detectable for the EuNPs-ML-ES ICS at 30, 25 and 21 days post-infection (dpi) and for the EuNPs-PAW-ES ICS at 25, 21 and 17 dpi. These results show that EuNPs-PAW-ES ICS detects anti-Trichinella IgG in pigs 4–5 days earlier that test using ML-ES antigens. Our ICS have no cross reaction with other parasite infection sera. Furthermore, the detection process could be completed in 10 min. This study indicated that our ICS can be used for the detection of the circulating antibodies in early T. spiralis infection and provide a novel method for on-site detection of T. spiralis infection in pigs.

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