Immunohistochemical characterization of polyclonal antibodies against <i><scp>E</scp>nteromyxum leei</i> and <i><scp>E</scp>nteromyxum scophthalmi</i> (<scp>M</scp>yxozoa: <scp>M</scp>yxosporea), intestinal parasites of fish

Estensoro, Itziar; Redondo, María J.; Álvarez-Pellitero, Pilar; Sitjà‐Bobadilla, Ariadna

doi:10.1111/jfd.12168

Cited by 5 publications

(6 citation statements)

References 30 publications

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“…was found in Portugal (Rangel et al 2011). E. leei spores were demonstrated to be immunogenic for rabbits, as the aPabEleei specifically detected surface epitopes on spore valves by immunohistochemistry (Estensoro et al 2013). The immunogenicity of myxospores in organisms different than the corresponding hosts (mice and rabbits) has already been observed in other studies (Clouthier et al 1997, Chase et al 2001, 2003.…”

Section: Discussionmentioning

confidence: 89%

“…have been detected with the aPabEleei, but Myxobolus spp. have not been tested for cross-reactivity (Estensoro et al 2013). The glycoproteic antigens found in the present study (15, 34 and 165 kDa) (Appendix 1) may represent further common antigens shared between E. leei and other myxo sporeans.…”

Section: Discussionmentioning

confidence: 99%

“…The production and characterization of the Pab raised against Enteromyxum leei (Pab Eleei) is described in a previous work (Estensoro et al 2013). This antiserum was adsorbed with normal GSB intestinal scrapings to avoid background noise due to host cell detection.…”

Section: Polyclonal Antibodiesmentioning

confidence: 99%

“…This antibody was obtained and titred by the Fish Pathology group, IATS-CSIC, Spain(Estensoro et al 2013); b This antibody was produced by the Department of Cell and Systems Biology, University of Toronto, Canada (Ringuette et al 2011) Table 1. Detector molecules used in Western blots.…”

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confidence: 99%

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Antigenic characterization of Enteromyxum leei (Myxozoa: Myxosporea)

Estensoro¹,

Álvarez-Pellitero²,

Sitjà‐Bobadilla³

2013

Dis. Aquat. Org.

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Enteromyxum leei, an intestinal myxozoan parasite affecting a wide range of fish, was partially purified, and the immunogenic composition of its glycoproteins as well as the proteolytic activity were studied. Parasite extracts, mainly containing spores, were separated by SDS-PAGE, and thereafter, immunoblots were carried out with a polyclonal antiserum (Pab) raised against E. leei. Periodic acid/Schiff staining of gels, periodate- and Proteinase K-treated Western blots and Lectin blots were performed to analyse the terminal carbohydrate composition of the parasite's antigens. Additionally, the cross-reaction of the parasite extracts with a Pab raised against the polar filament of the myxozoan Myxobolus pendula was studied. Both Pabs detected proteic epitopes on antigenic proteins and glycoproteins of E. leei, ranging between 15 and 280 kDa. In particular, 2 glycoproteic bands (15 and 165 kDa), immunoreactive to both Pabs and with glucose and mannose moieties, could correspond to common antigens shared among myxozoans. The 165 kDa band also presented galactose, N-acetyl-galactosamine and N-acetyl-glucosamine, pointing to its possible origin on chitin-built spore valves and to its possible involvement in host-parasite interactions. The molecular weight of the 15 kDa glycoproteic antigen matches that of minicollagen, a cnidarian-specific protein of nematocysts with a myxozoan homologue. Several proteases with apparent molecular weights ranging between 43 and 245 kDa were found in zymographies of E. leei extracts, and these may have a potential role in the parasite's pathogenesis. This is a useful approach for further studies to detect targets for antiparasitic therapy.

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Section: Discussionmentioning

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Section: Polyclonal Antibodiesmentioning

confidence: 99%

mentioning

confidence: 99%

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Antigenic characterization of Enteromyxum leei (Myxozoa: Myxosporea)

Estensoro¹,

Álvarez-Pellitero²,

Sitjà‐Bobadilla³

2013

Dis. Aquat. Org.

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“…Immunological methods based on antibodies or lectins (Knaus & El-Matbouli 2005, Estensoro et al 2013) have also been developed. Nevertheless, these methods are generally restricted by cross-reactivity between different species (Lu et al 2002, Estensoro et al 2014, Jia et al 2016. In recent years, PCR-based molecular detection and identification with higher sensitivity, specificity and rapidity has been utilised widely in myxo sporean examination (Qureshi et al 2002, Cavender et al 2004, Clark 2006, Grabner et al 2012, Jeon et al 2014.…”

Section: Introductionmentioning

confidence: 99%

Development of a multiplex PCR method for the simultaneous detection of four myxosporeans infecting gibel carp Carassius auratus gibelio

Li¹,

Zhai²,

Gu³

et al. 2017

Dis. Aquat. Org.

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Gibel carp Carassius auratus gibelio (Bloch), a commercially important freshwatercultured fish in China, is threatened by myxosporeans, particularly Thelohanellus wuhanensis, Myxobolus honghuensis, M. wulii and M. turpisrotundus. Here, we developed a multiplex PCR assay for simultaneous detection of these 4 myxosporeans. The specific primers for each species were designed based on the 28S rDNA gene of T. wuhanensis, the ITS-5.8S rDNA of M. honghuensis and M. wulii, and the 18S rDNA gene of M. turpisrotundus. Specificity testing confirmed that the 4 primer sets have no cross-reactivity with other related myxosporean species tested. Detection limits of the multiplex PCR assay were 0.2, 0.3, 3.1 and 3.8 spores for T. wuhanensis, M. honghuensis, M. wulii and M. turpisrotundus, respectively. Following screening of 104 field samples, the analytical sensitivity of the present multiplex PCR assay was found to be similar to the sensitivity obtained by the singleplex PCR assays and was higher than that of microscopic examination. Moreover, Kappa analysis showed a strong agreement between the results of the singleplex and multiplex PCR assays, indicating that the developed multiplex PCR assay was an efficient approach for the diagnosis of the 4 myxosporeans infecting gibel carp. KEY WORDS: Multiplex PCR · Molecular detection · Myxosporeans · Gibel carp · rDNA Resale or republication not permitted without written consent of the publisherDis Aquat Org 124: [31][32][33][34][35][36][37][38][39] 2017 forms plasmodia in the skin of the host and is responsible for chronic mortality and poor growth (Liu et al. 2014b). Minimizing the severe negative impact of myxosporidiosis on commercially important gibel carp is considered a priority in China. Early and accurate detection of myxosporeans could help fish farmers to carry out prompt drug therapy and minimize the likelihood of a myxo sporidiosis outbreak (Jia et al. 2015). Therefore, a simple and efficient assay for detection of myxo sporean pathogens is needed.Microscopic examination of squash preparations or histological sections is the most direct and commonly used method to detect and distinguish myxosporean parasites (St-Hilaire et al. 1997, Kelley et al. 2004). However, this approach can be time-and laborconsuming, and detection and identification is mainly based on the mature spore stages, with the early parasite stages likely to be neglected (Grabner et al. 2012, Mahony et al. 2015. Immunological methods based on antibodies or lectins (Knaus & El-Matbouli 2005, Estensoro et al. 2013) have also been developed. Nevertheless, these methods are generally restricted by cross-reactivity between different species (Lu et al. 2002, Estensoro et al. 2014, Jia et al. 2016. In recent years, PCR-based molecular detection and identification with higher sensitivity, specificity and rapidity has been utilised widely in myxo sporean examination (Qureshi et al. 2002, Cavender et al. 2004, Clark 2006, Grabner et al. 2012, Jeon et al. 2014. Given the multiple copies and moderate i...

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