To our knowledge,few reports on Demodex studied at the molecular level are available at present.In this study our group,for the first time,cloned,sequenced and analyzed the chitin synthase(CHS) gene fragments of Demodex folliculorum,Demodex brevis,and Demodex canis(three isolates from each species) from Xi'an China,by designing specific primers based on the only partial sequence of the CHS gene of D.canis from Japan,retrieved from GenBank.Results show that amplification was successful only in three D.canis isolates and one D.brevis isolate out of the nine Demodex isolates.The obtained fragments were sequenced to be 339 bp for D.canis and 338 bp for D.brevis.The CHS gene sequence similarities between the three Xi'an D.canis isolates and one Japanese D.canis isolate ranged from 99.7% to 100.0%,and those between four D.canis isolates and one D.brevis isolate were 99.1%-99.4%.Phylogenetic trees based on maximum parsimony(MP) and maximum likelihood(ML) methods shared the same clusters,according with the traditional classification.Two open reading frames(ORFs) were identified in each CHS gene sequenced,and their corresponding amino acid sequences were located at the catalytic domain.The relatively conserved sequences could be deduced to be a CHS class A gene,which is associated with chitin synthesis in the integument of Demodex mites.
Ya-e ZHAOZheng-hang WANGYang XUJi-ru XUWen-yan LIUMeng WEIChu-ying WANG
For a long time,classification of Demodex mites has been based mainly on their hosts and phenotypic characteristics.A new subspecies of Demodex folliculorum has been proposed,but not confirmed.Here,cox1 partial sequences of nine isolates of three Demodex species from two geographical sources(China and Spain) were studied to conduct molecular identification of D.folliculorum.Sequencing showed that the mitochondrial cox1 fragments of five D.folliculorum isolates from the facial skin of Chinese individuals were 429 bp long and that their sequence identity was 97.4%.The average sequence divergence was 1.24% among the five Chinese isolates,0.94% between the two geographical isolate groups(China(5) and Spain(1)),and 2.15% between the two facial tissue sources(facial skin(6) and eyelids(1)).The genetic distance and rate of third-position nucleotide transition/transversion were 0.0125,2.7(3/1) among the five Chinese isolates,0.0094,3.1(3/1) between the two geographical isolate groups,and 0.0217,4.4(3/1) between the two facial tissue sources.Phylogenetic trees showed that D.folliculorum from the two geographical isolate groups did not form sister clades,while those from different facial tissue sources did.According to the molecular characteristics,it appears that subspecies differentiation might not have occurred and that D.folliculorum isolates from the two geographical sources are of the same population.However,population differentiation might be occurring between isolates from facial skin and eyelids.
Ya-e ZHAOJun-xian MALi HULi-ping WUManuel DE ROJAS
