RESUMEN
P-glycoproteins (Pgps) are adenosine triphosphate-binding transporter proteins thought to be associated with multi-drug resistance in mammals and protozoans and have been suggested to be involved in the mechanism of ivermectin (IVM) resistance in Haemonchus contortus. Until now, resistance to IVM has not been reported in cyathostomins in horses in spite of its widespread and frequent use. Reasons for this might be differences in the molecular mechanism of the development of resistance. Based on this hypothesis, the present study was carried out to find homologues of Pgp in cyathostomins. A 416-bp polymerase chain reaction (PCR) product was generated using complementary DNA (cDNA) of Cylicocyclus elongatus and Cylicocyclus insigne and degenerate primers, located in the conserved Pgp nucleotide-binding domains. Resulting PCR products showed interspecific nucleotide and amino acid sequence identities of 73.3 and 76.8%, respectively. Specific primers were designed based on the Cc. elongatus sequence, and a PCR product of 268-bp was amplified from cDNA of single adults of Cylicocyclus radiatus, Cc. insigne, Cylicocyclus nassatus, Cc. elongatus, Cylicostephanus hybridus (2 individuals), Cylicostephanus goldi, Cyathostomum pateratum, Cyathostomum coronatum, and Cyathostomum catinatum. Two clusters of sequences were found representing 2 different internucleotide-binding domains (IBDs). A further distinct IBD is represented by the 416-bp PCR product of Cc. insigne. Therefore, a total of 3 clearly different sequences of the IBD were cloned and sequenced, suggesting that at least 2 Pgp genes exist in cyathostomins.
Asunto(s)
Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Variación Genética , Strongyloidea/genética , Subfamilia B de Transportador de Casetes de Unión a ATP/química , Secuencia de Aminoácidos , Animales , Antihelmínticos/farmacología , Secuencia de Bases , Bencimidazoles/farmacología , ADN Complementario/química , ADN de Helmintos/química , Resistencia a Medicamentos/genética , Caballos , Ivermectina/farmacología , Reacción en Cadena de la Polimerasa/veterinaria , ARN de Helminto/genética , ARN de Helminto/aislamiento & purificación , ARN Mensajero/genética , ARN Mensajero/aislamiento & purificación , Alineación de Secuencia/veterinaria , Infecciones Equinas por Strongyloidea/parasitología , Strongyloidea/química , Strongyloidea/efectos de los fármacosRESUMEN
The diversity of the beta-tubulin cDNAs of the cyathostominae and the occurrence of further isotypes were examined in adult worms isolated from an anthelmintic-naïve horse. cDNAs encoding beta-tubulin from Cyathostomum catinatum, Cylicocyclus nassatus, Cylicocyclus insigne, Cylicocyclus radiatus, Cylicocyclus elongatus, Cyathostomum coronatum, and Cyathostomum pateratum were characterized using specific primers developed from the cDNA sequence of Cc. nassatus. The cDNA sequences span 1,429 bp and show identities ranging from 95.6 to 100%. The deduced protein sequences span 448 amino acids and were 98-100% identical. The amino acid sequences of the 7 species varied within and between species at 10 positions. A 3' Rapid Amplification of cDNA ends using a degenerate forward primer was carried out with cDNA from Cy. pateratum, Cy. coronatum, Cy. catinatum, and Cc. nassatus to investigate the occurrence of further beta-tubulin isotypes. The expected polymerase chain reaction (PCR) product of 400 bp, including 306 bp of coding sequence, was amplified, as was an additional fragment of 600 nucleotides in the case of Cy. pateratum, Cy. coronatum, and Cy. catinatum. Sequencing of the PCR products revealed no evidence for the existence of a second beta-tubulin isotype in cyathostomes. The variation in size was caused by a length polymorphism within the 3' untranslated region, and 2 functional mRNAs seem to be transcribed from the same gene.
Asunto(s)
ADN Complementario/química , ADN de Helmintos/química , Strongyloidea/genética , Tubulina (Proteína)/genética , Regiones no Traducidas 3'/química , Secuencia de Aminoácidos , Animales , Enfermedades de los Caballos/parasitología , Caballos , Datos de Secuencia Molecular , Polimorfismo Genético , Isoformas de Proteínas/química , Alineación de Secuencia , Homología de Secuencia de Ácido Nucleico , Infecciones Equinas por Strongyloidea/parasitología , Strongyloidea/química , Strongyloidea/aislamiento & purificaciónRESUMEN
The Strongylocentrotus purpuratus genome contains a single ten-gene Hox complex >0.5 megabase in length. This complex was isolated on overlapping bacterial artificial chromosome and P1 artificial chromosome genomic recombinants by using probes for individual genes and by genomic walking. Echinoderm Hox genes of Paralog Groups (PG) 1 and 2 are reported. The cluster includes genes representing all paralog groups of vertebrate Hox clusters, except that there is a single gene of the PG4-5 types and only three genes of the PG9-12 types. The echinoderm Hox gene cluster is essentially similar to those of the bilaterally organized chordates, despite the radically altered pentameral body plans of these animals.
Asunto(s)
Evolución Molecular , Genes de Helminto , Genes Homeobox , Familia de Multigenes , Filogenia , Strongyloidea/genética , Secuencia de Aminoácidos , Animales , Cromosomas Bacterianos , Secuencia de Consenso , Drosophila , Genoma , Datos de Secuencia Molecular , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Strongyloidea/química , VertebradosRESUMEN
The gape nematode, Syngamus trachea, has been used as a model to study nematode secretions. Individual and intact pairs of amphidial glands, pharyngeal glands and pairs of excretory-secretory gland cells have been dissected and their secretory products analysed. The protein profiles of each gland and the total nematode secretions were analysed on 12.5% homogeneous SDS-PAGE minigels. The protein analyses revealed that the structural protein profile of each gland is different. The amphidial gland secretes two major proteins of 36.0 and 41.5 kDa, the excretory-secretory gland cell secretes a protein of 28.2 kDa and a protein of 14.3 kDa, and the pharyngeal gland secretes proteins of 41.5 and 14.6 kDa. Analysis of the total nematode secretions revealed all of the above major secretory proteins and an additional protein of 49.3 kDa. Syngamus trachea secretes acetylcholinesterases and its secretions contain multiple proteases.