ABSTRACT
Objetivo Describir la presencia de fluido subretiniano (FSR), fluido intrarretiniano (FIR) y fluido sub-epitelio pigmentario de la retina (FSEPR) en pacientes näive con degeneración macular asociada a la edad (DMAE) neovascular exudativa en el momento basal y al año de seguimiento y tratamiento, en nuestra práctica clínica, y realizar un análisis de concordancia entre médicos residentes. Método Se realizó un análisis retrospectivo de los pacientes näive que acudieron a nuestro servicio durante 6meses, entre 2016 y 2017, por DMAE neovascular. Las tomografías de coherencia óptica (OCT), del momento basal y al año de seguimiento, fueron analizadas de forma independiente por dos médicos residentes, determinando la presencia o no de FSR, FIR y FSEPR. Un oftalmólogo especialista en retina intervino en los casos en que no hubo consenso entre los médicos residentes. Se realizó un análisis descriptivo y de concordancia interobservador. Resultados Se evaluaron 28 ojos de 24 pacientes, siendo el 20,8% hombres y el 79,16% mujeres, con una edad media de 78,57±8años. El 32,14% de los ojos presentaron los tres tipos de fluido antes del inicio del tratamiento. La frecuencia de los diferentes fluidos en el inicio y al final del seguimiento fueron, respectivamente: FSR, 82,1 y 50%; FIR, 57,1 y 41,7%, y FSEPR, 67,9 y 79,2%. El análisis kappa de concordancias interobservador en la evaluación de los diferentes fluidos en el inicio y al final del seguimiento fueron, respectivamente: FSR, 0,88 y 0,67; FIR, 0,86 y 0,91, y FSEPR, 0,65 y 0,78. Conclusiones La presencia de FSR, FIR y FSEPR en práctica clínica en el debut de la DMAE neovascular tiene una distribución similar a la presentada en ensayos clínicos internacionales. La concordancia entre médicos residentes es muy buena para el FSR y FIR y buena para el FSEPR en el debut de la enfermedad y buena para el FSR y FIR y muy buena para el FSEPR al año de tratamiento (AU)
Purpose To analyze the presence of subretinal fluid (SRF), intraretinal fluid (IRF) and subretinal pigment epithelial fluid (SRPEF) in näive patients with exudative neovascular AMD at baseline and at one year follow-up and treatment, in clinical practice, and perform a concordance analysis between resident physicians. Methods A retrospective analysis of the näive patients who attended our service for 6months between 2016-2017 by neovascular AMD was performed. Optical coherence tomography (OCT), at baseline and at one year follow-up, were analyzed from independently by two resident doctors, determined the presence or not of SRF, IRF, SRPEF. A retina specialist ophthalmologist intervened in cases where there was no consensus among resident physicians. A descriptive and interobserver concordance analysis was performed. Results 27 eyes of 24 patients were evaluated, 20.8% being men and 79.16% women, with a mean age of 78.57±8years. 32.14% of the eyes presented the three types of fluid before the start of treatment and the frequency of the different fluids at the beginning and at the end of the follow-up were respectively: SRF, 82.1% and 50%; IRF, 57.1% and 41.7%, and SRPEF, 67.9% and 79.2%). The Kappa analysis of interobserver concordance in the evaluation of the different fluids at the beginning and at the end of the follow-up were respectively: SRF, 0.88 and 0.67; IRF, 0.86 and 0.91, and SRPEF, 0.65 and 0.78. Conclusions The presence of SRF, IRF, RPEF in clinical practice, in the debut of neovascular AMD has a similar distribution to that presented in international clinical trials. The agreement between resident physicians is very good for SRF and IRF and good for SRPEF in the debut of the disease and good for SRF and IRF and very good for SRPEF at one year of treatment (AU)
Subject(s)
Humans , Male , Female , Aged , Aged, 80 and over , Internship and Residency , Clinical Competence , Macular Degeneration/diagnostic imaging , Retrospective Studies , Follow-Up Studies , Tomography, Optical Coherence , Macular Degeneration/drug therapy , Prognosis , Observer VariationABSTRACT
PURPOSE: To analyze the presence of subretinal fluid (SRF), intraretinal fluid (IRF) and subretinal pigment epithelial fluid (SRPEF) in näive patients with exudative neovascular AMD at baseline and at one year follow-up and treatment, in clinical practice, and perform a concordance analysis between resident physicians. METHODS: A retrospective analysis of the näive patients who attended our service for 6months between 2016-2017 by neovascular AMD was performed. Optical coherence tomography (OCT), at baseline and at one year follow-up, were analyzed from independently by two resident doctors, determined the presence or not of SRF, IRF, SRPEF. A retina specialist ophthalmologist intervened in cases where there was no consensus among resident physicians. A descriptive and interobserver concordance analysis was performed. RESULTS: 27 eyes of 24 patients were evaluated, 20.8% being men and 79.16% women, with a mean age of 78.57±8years. 32.14% of the eyes presented the three types of fluid before the start of treatment and the frequency of the different fluids at the beginning and at the end of the follow-up were respectively: SRF, 82.1% and 50%; IRF, 57.1% and 41.7%, and SRPEF, 67.9% and 79.2%). The Kappa analysis of interobserver concordance in the evaluation of the different fluids at the beginning and at the end of the follow-up were respectively: SRF, 0.88 and 0.67; IRF, 0.86 and 0.91, and SRPEF, 0.65 and 0.78. CONCLUSIONS: The presence of SRF, IRF, RPEF in clinical practice, in the debut of neovascular AMD has a similar distribution to that presented in international clinical trials. The agreement between resident physicians is very good for SRF and IRF and good for SRPEF in the debut of the disease and good for SRF and IRF and very good for SRPEF at one year of treatment.
ABSTRACT
Trichuris trichiura is a nematode considered as the whipworm present in humans and primates. The systematics of the genus Trichuris is complex. Morphological studies of Trichuris isolated from primates and humans conclude that the species infecting these hosts is the same. Furthermore, numerous molecular studies have been carried out so far to discriminate parasite species from humans and Non-Human Primates using molecular techniques, but these studies were not performed in combination with a parallel morphological study. The hypothesised existence of more species of Trichuris in primates opens the possibility to revise the zoonotic potential and host specificity of T. trichiura and other putative new species of whipworms. In the present work, a study of Trichuris Roederer, 1761 (Nematoda:Trichuridae) parasitizing C. g. kikuyensis, P. ursinus, Macaca sylvanus, Pan troglodytes, and Sus scrofa domestica has been carried out using modern morphometric techniques in order to differentiate populations of Trichuris isolated from four species of captive NHP from different geographical regions, and swine, respectively. The results obtained revealed strong support for geometrical morphometrics as a useful tool to differentiate male Trichuris populations. Therefore, morphometrics in combination with other techniques, such as molecular biology analyses, ought to be applied to further the differentiation of male populations. On the other hand, morphometrics applied to female Trichuris species does not seem to contribute new information as all the measurements combinations of obtained from females always showed similar results.
ABSTRACT
The human flea Pulex irritans Linnaeus, 1758 (Siphonaptera: Pulicidae) is one of the most studied species together with the cat flea Ctenocephalides felis Bouché, 1835, because they have a cosmopolitan distribution and are closely related to humans. The present study aimed to carry out a comparative morphometric and molecular study of two different populations of P. irritans (Spain and Argentina). Accordingly, internal transcribed spacer (ITS)1 and ITS2 of rDNA and the partial cytochrome c oxidase subunit 1 (cox1) and cytochrome b (cytb) mtDNA genes of these taxa were sequenced. Furthermore, the taxonomy, origin, evolution and phylogeny of P. irritans was assessed. The morphometric data obtained did not show significant differences between P. irritans specimens from Spain and Argentina, even when these two populations were collected from different hosts; however, there was a considerable degree of molecular divergence between both populations based on nuclear and mitochondrial markers. Thus, it is proposed that P. irritans, in contrast with other generalist fleas, maintains a certain degree of morphological similarity, at least between Western Palearctic and Neotropical areas. Furthermore, two well defined geographical genetic lineages within the P. irritans species are indicated, suggesting the existence of two cryptic species that could be discriminated by a polymerase chain reaction-linked restriction fragment length polymorphism.
Subject(s)
Biological Evolution , Siphonaptera/classification , Animals , Argentina , DNA, Ribosomal Spacer/analysis , Electron Transport Complex IV/analysis , Female , Insect Proteins/analysis , Male , Phylogeny , Siphonaptera/anatomy & histology , Siphonaptera/genetics , SpainABSTRACT
Flea identification is a significant issue because some species are considered as important vectors of several human pathogens that have emerged or re-emerged recently, such as Bartonella henselae (Rhizobiales: Bartonellaceae) and Rickettsia felis (Rickettsiales: Rickettsiaceae). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been evaluated in recent years for the identification of multicellular organisms, including arthropods. A preliminary study corroborated the usefulness of this technique for the rapid identification of fleas, creating a preliminary database containing the spectra of five species of flea. However, longterm flea preservation in ethanol did not appear to be an adequate method of storage in the context of specimen identification by MALDI-TOF MS profiling. The goal of the present work was to assess the performance of MALDI-TOF MS in the identification of seven flea species [Ctenocephalides felis (Siphonaptera: Pulicidae), Ctenocephalides canis, Pulex irritans (Siphonaptera: Pulicidae), Archaeopsylla erinacei (Siphonaptera: Pulicidae), Leptopsylla taschenbergi (Siphonaptera: Ceratophyllidae), Stenoponia tripectinata (Siphonaptera: Stenoponiidae) and Nosopsyllus fasciatus (Siphonaptera: Ceratophyllidae)] collected in the field and stored in ethanol for different periods of time. The results confirmed that MALDI-TOF MS can be used for the identification of wild fleas stored in ethanol. Furthermore, this technique was able to discriminate not only different flea genera, but also the two congeneric species C. felis and C. canis.
Subject(s)
Dog Diseases/classification , Flea Infestations/veterinary , Hedgehogs , Insect Vectors/classification , Siphonaptera/classification , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/veterinary , Algeria , Animals , Dogs , Ethanol , Flea Infestations/classification , Spain , Specimen Handling/veterinary , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methodsABSTRACT
In the present work, we carried out a morphological, biometrical and molecular study of the species Archaeopsylla erinacei (Bouché, 1835) and their subspecies: Archaeopsylla erinacei erinacei (Bouché, 1835) and Archaeopsylla erinacei maura (Jordan & Rothschild, 1912) isolated from hedgehogs (Erinaceus europaeus) from different geographical regions (Seville and Corse). We have found morphological differences in females of A. erinacei from the same geographical origin that did not correspond with molecular differences. We suggest that some morphological characters traditionally used to discriminate females of both subspecies should be revised as well as we set the total length of the spermatheca as a valid criterion in order to discriminate between both subspecies. The Internal Transcribed Spacers 1 and 2 (ITS1, ITS2) and partial 18S rRNA gene, and partial cytochrome c-oxidase 1 (cox1) and cytochrome b (cytb) mtDNA gene sequences were determined to clarify the taxonomic status of these taxa and to assess intra-specific and intra-population similarity. In addition, a phylogenetic analysis with other species of fleas using Bayesian and Maximum Likelihood analysis was performed. All molecular markers used, except 18S, showed molecular differences between populations corresponding with geographical origins. Thus, based on the phylogenetic and molecular study of two nuclear markers (ITS1, ITS2) and two mitochondrial markers (cox1 and cytb), as well as concatenated sequences of both subspecies, we reported the existence of two geographical genetic lineages in A. erinacei corresponding with two different subspecies: A. e. erinacei (Corse, France) and A. e. maura (Seville, Spain), that could be discriminated by polymerase chain reaction-linked random-fragment-length polymorphism.
Subject(s)
Siphonaptera/anatomy & histology , Siphonaptera/genetics , Animals , DNA, Mitochondrial/analysis , DNA, Ribosomal Spacer/analysis , Female , France , Insect Proteins/analysis , Male , Phylogeny , RNA, Ribosomal, 18S/analysis , Sequence Analysis, DNA , Sequence Analysis, RNA , Siphonaptera/classification , Siphonaptera/enzymology , SpainABSTRACT
In the present work, we carried out a comparative molecular study of Stenoponia tripectinata tripectinata isolated from Mus musculus from the Canary Islands, Spain. The Internal Transcribed Spacers 1 and 2 (ITS1, ITS2) and 18S ribosomal RNA partial gene and cytochrome c-oxidase 1 (cox1) mitochondrial DNA partial gene sequences of this subspecies were determined to clarify the taxonomic status of this subspecies and to assess inter-population variation and inter-specific sequence differences. In addition, we have carried out a comparative phylogenetic study with other species of fleas using Bayesian, Maximum Parsimony, Maximum Likelihood and Neighbor-Joining analysis. A geographical signal was detected between the cox1 partial gene sequences of S. t. tripectinata isolated from M. musculus from different islands and those isolated from Apodemus sylvaticus from the Iberian Peninsula. Our results assess the monophyletic origin of Stenoponiinae and a different genetic lineage from Ctenophthalmidae. Thus, the elevation of subfamily Stenoponiinae to family level (Stenoponiidae) is suggested.
Subject(s)
Genetic Variation , Phylogeny , Siphonaptera/genetics , Animals , DNA, Intergenic/chemistry , DNA, Mitochondrial/chemistry , Electron Transport Complex IV/chemistry , Electron Transport Complex IV/genetics , Likelihood Functions , Mice , Phylogeography , RNA, Ribosomal, 18S/chemistry , Siphonaptera/anatomy & histology , Spain , Species SpecificityABSTRACT
At the present work, we carried out a morph-biometrical and molecular study of Trichuris species isolated from Camelus dromedarius from Iran and from Ovis aries from South Africa comparatively with other species of Trichuris from different herbivorous hosts and geographical regions. The population from camels from Iran was identified as Trichuris globulosa. Two different morphometrically populations of Trichuris sp. from sheep from South Africa were identified: Trichuris ovis and Trichuris skrjabini. Ribosomal data did not reveal significate differences in the ITS2 sequences between T. ovis and T. globulosa to assess a specific determination. The mitochondrial data suggest that T. globulosa constitute a different genetic lineage to T. ovis. Cytochrome c-oxidase and cytochrome b partial gene sequences corroborated the existence of a different genetic lineage of T. ovis from sheep of South Africa that would be closely related to the populations of T. globulosa from camels from Iran. The cytochrome c-oxidase and cytochrome b partial gene sequences of T. globulosa have been reported for the first time.
Subject(s)
Sheep Diseases/parasitology , Trichuriasis/veterinary , Trichuris/classification , Animals , Camelus/parasitology , Cytochromes b/genetics , DNA, Helminth/genetics , DNA, Mitochondrial/genetics , DNA, Ribosomal/genetics , Electron Transport Complex IV/genetics , Female , Genes, Helminth , Herbivory , Male , Phylogeny , Sequence Analysis, DNA , Sheep , Species Specificity , Trichuriasis/parasitology , Trichuris/anatomy & histology , Trichuris/geneticsABSTRACT
In the present work, a comparative morphological, biometrical and molecular study of Ctenocephalides spp. isolated from dogs (Canis lupus familiaris) from different geographical regions (Spain, Iran, and South Africa) has been carried out. The internal transcribed spacer 1 (ITS1) sequences of Ctenocephalides felis and Ctenocephalides canis collected from dogs from different geographical regions have been determined to clarify the taxonomic status of these species and to assess intraspecific variation and interspecific sequence differences. In addition, a phylogenetic analysis based on ITS1 sequences has been performed. Four different morphological populations were observed in the individuals of C. felis collected from dogs from different geographical locations. Nevertheless, the comparative study of the ITS1 sequences of the different morphological populations observed in C. felis did not show molecular differences. The results showed clear molecular differences between C. felis and C. canis and some specific recognition sites for endonucleases were detected between both species. Thus, BfrBI and DraI sites have diagnostic value for specific determination in C. felis. The phylogenetic tree based on the ITS1 sequences of C. felis and C. canis revealed that all the populations of C. felis from different geographical regions clustered together and separated, with high bootstrap values, from C. canis. We conclude that ITS1 region is a useful tool to approach different taxonomic and phylogenetic questions in Ctenocephalides species.
Subject(s)
Biometry , Ctenocephalides/anatomy & histology , Ctenocephalides/genetics , Dog Diseases/parasitology , Genetic Variation , Phylogeography , Animals , Cluster Analysis , DNA, Ribosomal Spacer/chemistry , DNA, Ribosomal Spacer/genetics , Dogs , Ectoparasitic Infestations/parasitology , Ectoparasitic Infestations/veterinary , Iran , Molecular Sequence Data , Sequence Analysis, DNA , Sequence Homology, Nucleic Acid , South Africa , SpainABSTRACT
Comparative morphological, biometrical and molecular studies of Trichuris discolor isolated from Bos taurus from Spain and Iran was carried out. Furthermore, Trichuris ovis isolated from B. taurus and Capra hircus from Spain has been, molecularly, analyzed. Morphological studies revealed clear differences between T. ovis and T. discolor isolated from B. taurus but differences were not observed between populations of T. discolor isolated from different geographical regions. Nevertheless, the molecular studies based on the amplification and sequencing of the internal transcribed spacers 1 and 2 ribosomal DNA and 16S partial gene mitochondrial DNA showed clear differences between both populations of T. discolor from Spain and Iran suggesting two cryptic species. Phylogenetic studies corroborated these data. Thus, phylogenetic trees based on ITS1, ITS2 and 16S partial gene sequences showed that individuals of T. discolor from B. taurus from Iran clustered together and separated, with high bootstrap values, of T. discolor isolated from B. taurus from Spain, while populations of T. ovis from B. taurus and C. hircus from Spain clustered together but separated with high bootstrap values of both populations of T. discolor. Furthermore, a comparative phylogenetic study has been carried out with the ITS1and ITS2 sequences of Trichuris species from different hosts. Three clades were observed: the first clustered all the species of Trichuris parasitizing herbivores (T. discolor, T. ovis, Trichuris leporis and Trichuris skrjabini), the second clustered all the species of Trichuris parasitizing omnivores (Trichuris trichiura and Trichuris suis) and finally, the third clustered species of Trichuris parasitizing carnivores (Trichuris muris, Trichuris arvicolae and Trichuris vulpis).
Subject(s)
Cattle Diseases/parasitology , DNA, Mitochondrial/genetics , DNA, Ribosomal Spacer/genetics , RNA, Ribosomal, 16S/genetics , Trichuriasis/veterinary , Trichuris/genetics , Animals , Base Sequence , Cattle , Cattle Diseases/epidemiology , Demography , Female , Genetic Markers , Iran/epidemiology , Male , Molecular Sequence Data , Spain/epidemiology , Trichuriasis/epidemiology , Trichuriasis/parasitology , Trichuris/physiologyABSTRACT
In this paper, a morphological and biometrical study by optical microscopy and scanning electronic microscopy (SEM) of Trichuris suis isolated from different hosts (Sus scrofa domestica and Sus scrofa scrofa) and Trichuris trichiura isolated from chimpanzee, has been carried out. Our results demonstrate the existence of typical pericloacal papillae in both species. Biometrical parameters of T. suis and T. trichiura overlapped but males and females of T. trichiura tended to be shorter and thinner than those of T. suis. Our results suggest that T. suis and T. trichiura cannot be differentiated using standard procedures as morphological and biometrical determinations. Thus, the ITS1-5.8S-ITS2 region of the ribosomal DNA was sequenced to allow a differentiation between T. suis and T. trichiura on genetic level. The ITS1 and ITS2 sequences derived from T. trichiura eggs isolated from feces of primates (Colobus guereza kikuyensis and Nomascus gabriellae) showed clear differences to the respective sequences of T. suis derived from eggs of different porcine hosts. The 5.8S gene was similar between the two species. Sequences obtained from different populations of the same species showed no significant differences indicating that the ITS1-5.8S-ITS2 sequences reported in this study are representative for T. trichiura and T. suis, respectively. Phylogenetic relationships have been determined attending to the ITS1 and ITS2 sequences from different species of the genus Trichuris. In conclusion, T. trichiura and T. suis are considered to be closely related but genetically different species. Both species can be easily and reliably distinguished by a PCR-RFLP analysis of the ITS1 and ITS2 sequences with different restriction enzymes.
Subject(s)
Trichuriasis/veterinary , Trichuris/anatomy & histology , Trichuris/classification , Animals , Base Sequence , Cluster Analysis , DNA, Helminth/chemistry , DNA, Helminth/genetics , DNA, Intergenic/chemistry , DNA, Intergenic/genetics , Female , Male , Microscopy/methods , Microscopy, Electron, Scanning/methods , Molecular Sequence Data , Pan troglodytes/parasitology , Phylogeny , RNA, Ribosomal, 5.8S/chemistry , RNA, Ribosomal, 5.8S/genetics , Sequence Analysis, DNA , Sequence Homology , Sus scrofa/parasitology , Trichuriasis/parasitology , Trichuris/genetics , Trichuris/isolation & purificationABSTRACT
The partial mitochondrial cytochrome c-oxidase subunit 1 gene (cox 1) and partial mitochondrial 16S ribosomal DNA of Trichuris skrjabini (Baskakov 1924) isolated from Capra hircus have been amplified and sequenced. The analyses of multiple sequence alignments of mitochondrial 16S rDNA and cox 1 of T. skrjabini revealed high homology with those of Trichinella species. For the first time, the mitochondrial DNA gene sequences of one species of trichurid nematode have been cited.
Subject(s)
DNA, Helminth/genetics , DNA, Mitochondrial/genetics , DNA, Ribosomal/genetics , Electron Transport Complex IV/genetics , Trichuris/genetics , Animals , DNA, Helminth/chemistry , DNA, Mitochondrial/chemistry , DNA, Ribosomal/chemistry , Goats/parasitology , Molecular Sequence Data , RNA, Ribosomal, 16S/genetics , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology , Trichinella/genetics , Trichuris/enzymology , Trichuris/isolation & purificationABSTRACT
Adults of Trichuris skrjahini have been isolated from the cecum of caprine hosts (Capra hircus), Trichuris ovis and Trichuris globulosa from Ovis aries (sheep) and C. hircus (goats), and Trichuris leporis from Lepus europaeus (rabbits) in Spain. Genomic DNA was isolated and the ITS1-5.8S-ITS2 segment from the ribosomal DNA (rDNA) was amplified and sequenced by polymerase chain reaction (PCR) techniques. The ITS1 of T. skrjabini, T. ovis, T. globulosa, and T. leporis was 495, 757, 757, and 536 nucleotides in length, respectively, and had G + C contents of 59.6, 58.7, 58.7, and 60.8%, respectively. Intraindividual variation was detected in the ITSI sequences of the 4 species. Furthermore, the 5.8S sequences of T. skrjabini, T. ovis, T. globulosa, and T. leporis were compared. A total of 157, 152, 153, and 157 nucleotides in length was observed in the 5.8S sequences of these 4 species, respectively. There were no sequence differences of ITS1 and 5.8S products between T. ovis and T. globulosa. Nevertheless, clear differences were detected between the ITS1 sequences of T. skrjabini, T. ovis, T. leporis, Trichuris muris, and T. arvicolae. The ITS2 fragment from the rDNA of T. skrjabini was sequenced. A comparative study of the ITS2 sequence of T. skrjabini with the previously published ITS2 sequence data of T. ovis, T. leporis, T. muris, and T. arvicolae suggested that the combined use of sequence data from both spacers would be useful in the molecular characterization of trichurid parasites.
Subject(s)
DNA, Ribosomal Spacer/chemistry , Goat Diseases/parasitology , Rabbits/parasitology , Sheep Diseases/parasitology , Trichuriasis/veterinary , Trichuris/genetics , Animals , Base Sequence , Cecum/parasitology , Consensus Sequence , DNA, Helminth/chemistry , Goats , Male , Molecular Sequence Data , Polymerase Chain Reaction/veterinary , RNA, Ribosomal, 5.8S/genetics , Sheep , Spain , Trichuriasis/parasitology , Trichuris/classificationABSTRACT
The complete internal transcribed spacer 1 (ITS1), 5.8S rDNA and ITS2 region of the ribosomal DNA from 11 species of rhinonyssid mites ( Tinaminyssus columbae, T. minisetosum, T. sartbaevi, T. bubulci, T. melloi, T. streptopelioides, Sternostoma fulicae, S. boydi, S. strandtmanni, S. turdi, Rhinonyssus tringae) were sequenced to assess the utility of this genomic region in resolving taxonomic questions in this group and to estimate phylogenetic relationships between species. Two different geographic locations of T. melloi and T. streptopelioides were analyzed to detect intraspecies variation. Our study shows that ribosomal sequences can help to discriminate between T. melloi and T. sartbaevi, which are morphologically very close and difficult to separate by classic methods. The resulting phylogenetic tree shows some differences from the current taxonomy of the family Rhinonyssidae. This study appeals for the revision of the taxonomic status of S. boydi and closely related species which parasitize aquatic birds and suggests the synonymy of S. boydi and S. strandtmanni, despite the different hosts of the two mites.
Subject(s)
Acaridae/genetics , DNA, Ribosomal/genetics , Acaridae/classification , Animals , Base Sequence , Molecular Sequence Data , Phylogeny , Polymerase Chain Reaction , Sequence Alignment , Sequence Homology, Nucleic Acid , Species SpecificityABSTRACT
Trichuris muris has been isolated from murid hosts ( Apodemus sylvaticus and Mus musculus) and Trichuris arvicolae from arvicolid rodents in Barcelona, Spain. Genomic DNA was isolated and the ITS1-5.8S-ITS2 segment from the ribosomal DNA (rDNA) was amplified and sequenced using polymerase chain reaction techniques. The ITS2 of both populations isolated from Apodemus and Mus was 382 nucleotides in length and had a GC content of about 60.73%, while the ITS2 of T. arvicolae was 442 nucleotides in length and had a GC content of about 59.8%. Furthermore, the ITS1 of Trichuris from murids was 448 nucleotides in length and had a GC content of about 56.47%, while T. arvicolae was 446 nucleotides in length and had 57.62% of GC content. A total of 161 and 173 nucleotides were observed along the 5.8S gene of T. murisand T. arvicolae, respectively; This difference in nucleotides was due to the insertion of a DNA segment (transposon) in the 5.8S sequence of the latter species. Slight intraindividual and intraspecific variations were detected in the rDNA of both species. The presence of microsatellites was observed in all of the individuals assayed. Sequence analysis of the internal transcribed spacers and the 5.8S gene demonstrated no sequence differences between T. muris isolated from both of its murid hosts. Nevertheless, clear differences were detected between the ITS2, ITS1 and 5.8S gene of T. muris and T. arvicolae. This corroborates the existence of two separate Trichuris species in murid and arvicolid hosts. Furthermore, a phylogenetic analysis was carried out and endonucleases restriction maps were elaborated for both species.
Subject(s)
Arvicolinae/parasitology , DNA, Ribosomal Spacer/analysis , Muridae/parasitology , RNA, Ribosomal, 5.8S/genetics , Trichuris/isolation & purification , Animals , Base Sequence , DNA, Helminth/analysis , Mice , Models, Genetic , Molecular Sequence Data , Nucleic Acid Amplification Techniques , Phylogeny , Polymerase Chain Reaction , Sequence Alignment , Sequence Homology, Nucleic Acid , Species Specificity , Trichuris/geneticsABSTRACT
A 390 bp region of the 16S rDNA gene was sequenced from six species of rhinonyssid mites (Tinaminyssus columbae, T. minisetosum, Sternostoma turdi, S. sternahirundo, S. fulicae and Ptilonyssus euroturdi) and two subspecies (Tinaminyssus melloi melloi and Tinaminyssus melloi streptopeliae) to examine the level of sequence variation and the taxonomic levels to show utility in phylogeny estimation. Furthermore, two different geographic locations of T. m. melloi and T. m. streptopeliae were analyzed to detect variation between populations. Molecular data revealed the existence of two distinct groups in the genus Tinaminyssus parasitic on columbiform birds. These results are in agreement with those reported by some authors using morphological characters. Sternostoma turdi parasitizing aerial birds appeared to be phylogenetically separated from other species of this genus isolated from aquatic birds. Moreover, our study addresses the validity of the subspecies status of T. melloi streptopeliae. This region of the mitochondrial 16S rDNA gene is a useful marker for inferring phylogenetic relationships among closely related rhinonyssid species, but not for more distantly related taxa.
Subject(s)
Birds/parasitology , DNA, Mitochondrial/genetics , DNA, Ribosomal/genetics , Mites/genetics , Animals , Base Sequence , DNA, Mitochondrial/chemistry , DNA, Ribosomal/chemistry , Mites/classification , Molecular Sequence Data , Phylogeny , RNA, Ribosomal, 16S/chemistry , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA , Sequence Homology, Nucleic Acid , SpainABSTRACT
Adult worms of Trichuris ovis and T. globulosa were collected from Ovis aries (sheep) and Capra hircus (goats). T. suis was isolated from Sus scrofa domestica (swine) and T. leporis was isolated from Lepus europaeus (rabbits) in Spain. Genomic DNA was isolated and a ribosomal internal transcribed spacer (ITS2) was amplified and sequenced using polymerase-chain-reaction (PCR) techniques. The ITS2 of T. ovis and T. globulosa was 407 nucleotides in length and had a GC content of about 62%. Furthermore, the ITS2 of T. suis and T. leporis was 534 and 418 nucleotides in length and had a GC content of about 64.8% and 62.4%, respectively. There was evidence of slight variation in the sequence within individuals of all species analyzed, indicating intraindividual variation in the sequence of different copies of the ribosomal DNA. Furthermore, low-level intraspecific variation was detected. Sequence analyses of ITS2 products of T. ovis and T. globulosa demonstrated no sequence difference between them. Nevertheless, differences were detected between the ITS2 sequences of T. suis, T. leporis, and T. ovis, indicating that Trichuris species can reliably be differentiated by their ITS2 sequences and PCR-linked restriction-fragment-length polymorphism (RFLP).
Subject(s)
DNA, Helminth/genetics , DNA, Ribosomal Spacer/genetics , Trichuris/classification , Trichuris/genetics , Animals , Base Composition , Base Sequence , DNA Restriction Enzymes , DNA, Helminth/chemistry , DNA, Ribosomal Spacer/chemistry , Genes, rRNA , Genetic Variation , Goats/parasitology , Molecular Sequence Data , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , Rabbits/parasitology , Restriction Mapping , Sheep/parasitology , Species Specificity , Swine/parasitologyABSTRACT
The glucose-6-phosphate dehydrogenase (G6PD, EC.1.1.1.49), glucose phosphate isomerase (GPI, EC.5.3.1.9), and malate dehydrogenase (MDH, EC.1.1.1.37) isoenzymatic patterns of Chabertia ovina were determined by starch-gel electrophoresis. The G6PD and GPI isoenzymatic patterns were characterized by the existence of three phenotypes: (1) a single and slow anodic band, (2) a single and fast anodic band, and (3) a large spot matching its migration with bands 1 and 2. These three phenotypes may be explained as the existence of only one gene locus for the G6PD and GPI in C. ovina. Allelic frequencies and the Hardy-Weinberg test were determined. This test indicated that the population was not in Hardy-Weinberg equilibrium. The MDH isoenzymatic pattern of C. ovina was characterized by the presence of two bands with anodic and cathodic migration. Furthermore, comparative isoenzyme studies were carried out between Oesophagostomum venulosum and C. ovina. The different G6PD, GPI, and MDH isoenzymatic patterns observed for the two species allowed us to distinguish them and, therefore, to use isoenzymatic patterns as a diagnostic tool to discriminate these species.
Subject(s)
Isoenzymes/analysis , Oesophagostomum/classification , Oesophagostomum/enzymology , Strongyloidea/classification , Strongyloidea/enzymology , Animals , Electrophoresis, Starch Gel/methods , Glucose-6-Phosphate Isomerase/analysis , Glucose-6-Phosphate Isomerase/genetics , Glucosephosphate Dehydrogenase/analysis , Glucosephosphate Dehydrogenase/genetics , Goats/parasitology , Isoenzymes/genetics , Malate Dehydrogenase/analysis , Malate Dehydrogenase/genetics , Oesophagostomum/genetics , Strongyloidea/geneticsABSTRACT
Four different morphological and biometrical populations of Oesophagostomum have been identified, using classical taxonomy methods, from Sus scrofa domestica (pigs): O. dentatum (Od), O. quadrispinulatum (Oq), O. granatensis (Og), and a fourth population, including individuals with morphological and biometric parameters overlapping these three species that were clasified as Oesophagostomum sp. The G6PD and MDH isoenzymatic patterns did not discriminate between the three species, while GPI showed a diagnostic isoenzymatic pattern for Oq. Og showed identical G6PD, GPI and MDH isoenzymatic pattern as Od. Furthermore, after rDNA amplification by polymerase chain reaction (PCR), the uncut PCR product showed that the ITS2 of these three species had a similar size of 320 base pairs (bp). Restriction-fragment-length polymorphisms (RFLP) were analyzed after digestion of the ITS2 with 13 different restriction enzymes. After electrophoretic separation of the digested PCR products, only one unique differentiating pattern of bands was observed for Od and Oq. This was when Sau3AI was used, while Og showed an identical band pattern to Od. Thus, our studies provided no evidence for the existence of Og and Od as differentiated populations. O. venulosum was isolated from sheep and goat; G6PD and MDH isoenzymatic patterns discriminated this species from porcine species of Oesophagostomum. The ITS2 region appeared as a different band of 380 bp from those observed for porcine Oesophagostomum species.