Hoplotylus femina s’Jacob, 1960 (Nematoda: Pratylenchidae) from Spain with molecular phylogenetic relationships inferred by D2-D3 expansion fragments of 28S and the partial 18S rRNA gene sequences

This is the first report of Hoplotylus femina in Spain. It was collected from the rhizosphere and roots of common alder (Alnus glutinosa) in La Alberca, Salamanca Province, in Spain. The morphology and morphometrics of the Spanish population of H. femina agree closely with the original description of the species and other descriptions from The Netherlands, USA, New Zealand, Poland, Japan and Mexico. Only small differences in some morphological characters were found probably due to geographical intraspecific variability and/or different methods of fixing nematodes. This study provides new molecular markers for species identification (D2-D3 expansion regions of 28S and ITS-rRNA genes) and a new phylogenetic position for the D2-D3 region marker. Maximum likelihood analysis using the Shimodaira-Hasegawa test did not support the inclusion of Radopholus and Hoplotylus in the Pratylenchidae and both genera were more related phylogenetically to Hoplolaimidae.

Download Full-text

Phylogenetic relationships of Scelimeninae genera (Orthoptera: Tetrigoidea) based on COI, 16S rRNA and 18S rRNA gene sequences

Zootaxa ◽

10.11646/zootaxa.4482.2.11 ◽

2018 ◽

Vol 4482 (2) ◽

pp. 392

Author(s):

YA-ZHEN CHEN ◽

WEI-AN DENG ◽

JIA-MIN WANG ◽

LI-LIANG LIN ◽

SHAN-YI ZHOU

Keyword(s):

16S Rrna ◽

Genetic Distance ◽

Phylogenetic Relationships ◽

18S Rrna ◽

18S Rrna Gene ◽

Rrna Genes ◽

Rrna Gene ◽

Average Genetic Distance ◽

Molecular Phylogenetic ◽

18S Rrna Genes

Scelimeninae is an important subfamily of Tetrigoidea; however, the phylogenetic relationships within Scelimeninae are poorly understood, and its generic classification has remained unstable. In this study, the COI, 16S rRNA and 18S rRNA genes from 24 species in 9 genera within Scelimeninae were amplified and sequenced, the base composition and inter-species genetic distance of the combined sequence of COI, 16S rRNA and 18S rRNA genes were analyzed, and the molecular phylogenetic relationships were reconstructed using Maximum Likelihood (ML) and Bayesian inference (BI) methods. The results of sequence analysis showed that the total length of the combined COI, 16S rRNA and 18S rRNA gene sequence was 3507 bp, including 2345 conservative sites, 1144 variable sites and 901 parsimony-informative sites. The average A+T content was 63.5% and 78.1% in the COI, 16S rRNA sequences, respectively, indicating A+T bias. The average genetic distance between all species was 0.134, and the average genetic distance in the inner group (Scelimeninae) was 0.126. A phylogenetic tree based on the combined sequences of the COI, 16S rRNA and 18S rRNA genes showed that the phylogenetic relationships among 9 Scelimeninae genera were as follows: Criotettix + (((Zhengitettix + Hebarditettix) + (Falconius + (Scelimena + Paragavialidium))) + ((Eucriotettix + Thoradonta) + Loxilobus)). The molecular phylogenetic results generally support the morphological taxonomy; at the genus level, Criotettix, Scelimena, Paragavialidium, Thoradonta and Eucriotettix are monophyletic groups, Scelimena and Paragavialidium form sister groups, and Thoradonta and Eucriotettix also form sister groups, but the relationship between Hebarditettix and Zhengitettix needs further study. At the species level, synonyms may exist between Thoradonta spiculoba and Thoradonta transpicula and Thoradonta nodulosa and Thoradonta obtusilobata, but more studies are required to confirm this inference.

Download Full-text

Hemicriconemoides phoenicis Van den Berg et al., 2015 (Nematoda: Criconematidae) from Iran: a morphological and molecular phylogenetic study

Nematology ◽

10.1163/15685411-00003342 ◽

2020 ◽

Vol 22 (7) ◽

pp. 815-824

Author(s):

Sedighe Azimi ◽

Majid Pedram

Keyword(s):

Date Palm ◽

Phylogenetic Analyses ◽

Original Description ◽

Original Data ◽

18S Rrna Gene ◽

Phylogenetic Study ◽

Rrna Gene ◽

28S Rrna ◽

Khuzestan Province ◽

Molecular Phylogenetic

Summary A population of Hemicriconemoides phoenicis was recovered from Khuzestan province, south-western Iran, in association with date palm. The recovered population was characterised by 518-645 μm long females having a 76-82 μm long stylet, rounded to oblong spermatheca filled with sperm, a 28.0-39.8 μm long tail, juveniles common, with 14 longitudinal rows of rounded scales, and males absent. Compared to the original data, no morphological and morphometric differences were observed. In molecular phylogenetic analyses using the D2-D3 expansion segments of the 28S rRNA gene and a near-full-length fragment of the 18S rRNA gene sequences using Bayesian inference (BI) and maximal number of species of the genus, the two newly generated 28S sequences of the Iranian population formed a maximally supported clade with two original sequences of the species; and the 18S sequence formed a maximally supported clade with an unidentified isolate of the genus in the corresponding phylogeny. This is the second report of the species since its original description, Iran representing a new geographical record and supporting the suggestion that date palm could be its preferred host.

Download Full-text

Redescription and molecular characterisation of Xiphinema barense Lamberti et al., 1986 (Nematoda: Longidoridae) from wild olive trees in southern Italy

Nematology ◽

10.1163/15685411-00002836 ◽

2014 ◽

Vol 16 (9) ◽

pp. 1079-1089 ◽

Cited By ~ 8

Author(s):

Francesca De Luca ◽

Antonio Archidona-Yuste ◽

Alberto Troccoli ◽

Elena Fanelli ◽

Nicola Vovlas ◽

...

Keyword(s):

Southern Italy ◽

Phylogenetic Analyses ◽

Original Description ◽

Rrna Genes ◽

Phylogenetic Position ◽

Species Differentiation ◽

Rrna Gene ◽

28S Rrna ◽

Wild Olive ◽

Olive Trees

A population of Xiphinema barense from wild olive trees in Torre Pozzella, Brindisi province, southern Italy, is described using both morphological and molecular studies and compared with the description of the type specimens. The wild olive nematode population agrees very well with all morphometrics provided in the original description. However, detailed observations of the lumen of the tubular portion of the uterus in paratypes and specimens of the new population revealed a clear pseudo-Z-organ with small granules mixed with crystalloid bodies which were previously undetected. Photomicrographs of adult paratypes, which were lacking in the original description, and of specimens of the new population from wild olive trees are provided. The results of the phylogenetic analyses based on the sequences of the D2-D3 expansion regions of the 28S rRNA gene and ITS rRNA genes confirm the species differentiation and indicate the phylogenetic position of X. barense and its relationship with closely related species.

Download Full-text

Molecular Characterization of Ciliate Diversity in Stream Biofilms

Applied and Environmental Microbiology ◽

10.1128/aem.01438-07 ◽

2008 ◽

Vol 74 (6) ◽

pp. 1740-1747 ◽

Cited By ~ 52

Author(s):

Andrew Dopheide ◽

Gavin Lear ◽

Rebecca Stott ◽

Gillian Lewis

Keyword(s):

18S Rrna ◽

Pcr Primers ◽

18S Rrna Gene ◽

Sequence Diversity ◽

Rrna Genes ◽

Rrna Gene ◽

Sequencing Analysis ◽

Specific Pcr ◽

Stream Biofilms

ABSTRACT Free-living protozoa are thought to be of fundamental importance in aquatic ecosystems, but there is limited understanding of their diversity and ecological role, particularly in surface-associated communities such as biofilms. Existing eukaryote-specific PCR primers were used to survey 18S rRNA gene sequence diversity in stream biofilms but poorly revealed protozoan diversity, demonstrating a need for protozoan-targeted primers. Group-specific PCR primers targeting 18S rRNA genes of the protozoan phylum Ciliophora were therefore designed and tested using DNA extracted from cultured protozoan isolates. The two most reliable primer combinations were applied to stream biofilm DNA, followed by cloning and sequencing analysis. Of 44 clones derived from primer set 384F/1147R, 86% were of probable ciliate origin, as were 25% of 44 clones detected by primer set 121F/1147R. A further 29% of 121F/1147R-detected clones matched sequences from the closely related phylum Apicomplexa. The highly ciliate-specific primer set 384F/1147R was subsequently used in PCRs on biofilm DNA from four streams exhibiting different levels of human impact, revealing differences in ciliate sequence diversity in samples from each site. Of a total of 240 clones, 73% were of probable ciliate origin; 54 different putative ciliate sequences were detected from throughout seven taxonomic ciliate classes. Sequences from Oligohymenophorea were most commonly detected in all samples, followed by either Spirotrichea or Phyllopharyngea. Restriction fragment length polymorphism profile-based analysis of clones suggested a potentially higher level of diversity than did sequencing. Nevertheless, newly designed PCR primers 384F/1147R were considered to provide an effective molecular basis for characterization of ciliate diversity in stream biofilms.

Download Full-text

Steinernema borjomiense n. sp. (Rhabditida: Steinernematidae), a new entomopathogenic nematode from Georgia

Nematology ◽

10.1163/15685411-00003167 ◽

2018 ◽

Vol 20 (7) ◽

pp. 653-669 ◽

Cited By ~ 4

Author(s):

Oleg Gorgadze ◽

Elena Fanelli ◽

Manana Lortkhipanidze ◽

Alberto Troccoli ◽

Medea Burjanadze ◽

...

Keyword(s):

New Species ◽

18S Rrna ◽

Entomopathogenic Nematode ◽

Phylogenetic Analyses ◽

Tail Length ◽

Morphological Characters ◽

18S Rrna Gene ◽

The Body ◽

Rrna Gene ◽

A New Species

Summary A new species of entomopathogenic nematode, Steinernema borjomiense n. sp., was isolated from the body of the host insect, Oryctes nasicornis (Coleoptera: Scarabaeidae), in Georgia, in the territory of Borjomi-Kharagauli. Morphological characters indicate that the new species is closely related to species of the feltiae-group. The infective juveniles are characterised by the following morphological characters: body length of 879 (777-989) μm, distance between the head and excretory pore = 72 (62-80) μm, pharynx length = 132 (122-142) μm, tail length = 70 (60-80) μm, ratio a = 26.3 (23.0-29.3), H% = 45 (40-51), D% = 54 (47-59), E% = 102 (95-115), and lateral fields consisting of seven ridges (eight incisures) at mid-body. Steinernema borjomiense n. sp. was molecularly characterised by sequencing three ribosomal regions (the ITS, the D2-D3 expansion domains and the 18S rRNA gene) and the mitochondrial COI gene. Phylogenetic analyses revealed that S. borjomiense n. sp. differs from all other known species of Steinernema and is a member of the monticolum-group.

Download Full-text

Stenamoeba dejonckheerei sp. nov., a Free-Living Amoeba Isolated from a Thermal Spring

Pathogens ◽

10.3390/pathogens9070586 ◽

2020 ◽

Vol 9 (7) ◽

pp. 586

Author(s):

Manuel Alejandro Borquez-Román ◽

Luis Fernando Lares-Jiménez ◽

Libia Zulema Rodriguez-Anaya ◽

Jose Reyes Gonzalez-Galaviz ◽

Paul A. Fuerst ◽

...

Keyword(s):

Thermal Spring ◽

Small Subunit ◽

18S Rrna Gene ◽

Rrna Gene ◽

Likelihood Analysis ◽

Ribosomal Ribonucleic Acid ◽

Pcr Products ◽

Northwestern Mexico ◽

Rrna Gene Sequencing ◽

A New Species

Two amoeboid organisms were obtained from water samples taken from a thermal spring, "Agua Caliente", in Northwestern Mexico. The isolates were obtained when samples were cultivated at 37 °C on non-nutrient agar coated with Escherichia coli. The initial identification of the isolates was performed morphologically using light microscopy. The samples were found to have trophozoite morphology consistent with members of the genus Stenamoeba, a genus derived in 2007 from within the abolished polyphyletic genus Platyamoeba. Further analysis was performed by sequencing PCR products obtained using universal eukaryotic primers for the small subunit ribosomal ribonucleic acid (SSU rRNA) gene. Sequencing primers were designed to allow the comparison of the 18S rRNA gene sequences of the new isolates with previous sequences reported for Stenamoeba. Phylogenetic relationships among sequences from Stenamoeba were determined using Maximum Likelihood analysis. The results showed the two "Agua Caliente" sequences to be closely related, while clearly separating them from those of other Stenamoeba taxa. The degrees of sequence differentiation from other taxa were considered sufficient to allow us to propose that the Mexican isolates represent a new species.

Download Full-text

Development of a Denaturing High-Performance Liquid Chromatography Method for Detection of Protist Parasites of Metazoans

Applied and Environmental Microbiology ◽

10.1128/aem.02131-07 ◽

2008 ◽

Vol 74 (14) ◽

pp. 4336-4345 ◽

Cited By ~ 17

Author(s):

Christofer Troedsson ◽

Richard F. Lee ◽

Vivica Stokes ◽

Tina L. Walters ◽

Paolo Simonelli ◽

...

Keyword(s):

High Performance Liquid Chromatography ◽

Liquid Chromatography ◽

High Performance ◽

18S Rrna ◽

18S Rrna Gene ◽

Rrna Genes ◽

Rrna Gene ◽

Chromatography Method ◽

Factors Affecting ◽

Triethylammonium Acetate

ABSTRACT Increasingly, diseases of marine organisms are recognized as significant biotic factors affecting ecosystem health. However, the responsible disease agents are often unknown and the discovery and description of novel parasites most often rely on morphological descriptions made by highly trained specialists. Here, we describe a new approach for parasite discovery, utilizing denaturing high-performance liquid chromatography (DHPLC) reverse-phase ion-paring technology. Systematic investigations of major DHPLC variables, including temperature, gradient conditions, and target amplicon characteristics were conducted to develop a mechanistic understanding of DNA fragment separation by DHPLC. As a model system, 18S rRNA genes from the blue crab (Callinectes sapidus) and a parasitic dinoflagellate Hematodinium sp. were used. Binding of 18S rRNA gene PCR amplicons to the DNA separation column in the presence of triethylammonium acetate (TEAA) was inversely correlated with temperature and could be predicted based on the estimated DNA helicity of the PCR amplicon. Amplicons of up to 498 bp were resolved as single chromatographic peaks if they had high (>95%) DNA helicity. Amplicons that differed by as few as 2 bp could be resolved. Separation of 18S rRNA gene PCR amplicons was optimized by simultaneous manipulation of both temperature and solvent gradients. The optimal conditions included targeting regions of high DNA helicity (>95%), temperatures in the range of 57 to 63°C, and a linear acetonitrile gradient from 13.75 to 17.5% acetonitrile in 0.1 M TEAA (55 to 70% buffer B) over a 9-min period. Under these conditions, amplicons from a variety of parasites and their hosts can be separated and detected by DHPLC.

Download Full-text

Design and Validation of Four New Primers for Next-Generation Sequencing To Target the 18S rRNA Genes of Gastrointestinal Ciliate Protozoa

Applied and Environmental Microbiology ◽

10.1128/aem.01644-14 ◽

2014 ◽

Vol 80 (17) ◽

pp. 5515-5521 ◽

Cited By ~ 22

Author(s):

Suzanne L. Ishaq ◽

André-Denis G. Wright

Keyword(s):

18S Rrna ◽

18S Rrna Gene ◽

Rrna Genes ◽

Rrna Gene ◽

Content Type ◽

Hypervariable Regions ◽

Blast Database ◽

Primer Sets ◽

18S Rrna Genes ◽

Ciliate Protozoa

ABSTRACTFour new primers and one published primer were used to PCR amplify hypervariable regions within the protozoal 18S rRNA gene to determine which primer pair provided the best identification and statistical analysis. PCR amplicons of 394 to 498 bases were generated from three primer sets, sequenced using Roche 454 pyrosequencing with Titanium, and analyzed using the BLAST database (NCBI) and MOTHUR version 1.29. The protozoal diversity of rumen contents from moose in Alaska was assessed. In the present study, primer set 1, P-SSU-316F and GIC758R (amplicon of 482 bases), gave the best representation of diversity using BLAST classification, and the set amplifiedEntodinium simplexandOstracodiniumspp., which were not amplified by the other two primer sets. Primer set 2, GIC1080F and GIC1578R (amplicon of 498 bases), had similar BLAST results and a slightly higher percentage of sequences that were identified with a higher sequence identity. Primer sets 1 and 2 are recommended for use in ruminants. However, primer set 1 may be inadequate to determine protozoal diversity in nonruminants. The amplicons created by primer set 1 were indistinguishable for certain species within the generaBandia,Blepharocorys,Polycosta, andTetratoxumand betweenHemiprorodon gymnoprosthiumandProrodonopsiscoli, none of which are normally found in the rumen.

Download Full-text

Reclassification of Trichlorobacter thiogenes as Geobacter thiogenes comb. nov.

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY ◽

10.1099/ijs.0.63408-0 ◽

2007 ◽

Vol 57 (3) ◽

pp. 463-466 ◽

Cited By ~ 28

Author(s):

Kelly P. Nevin ◽

Dawn E. Holmes ◽

Trevor L. Woodard ◽

Sean F. Covalla ◽

Derek R. Lovley

Keyword(s):

Type Strain ◽

Original Description ◽

16S Rrna Genes ◽

Rrna Genes ◽

Phylogenetic Position ◽

Temperature Optimum ◽

Ph Optimum ◽

Donor And Acceptor ◽

The Family ◽

Electron Donor And Acceptor

Reclassification of the species Trichlorobacter thiogenes as Geobacter thiogenes comb. nov. is proposed on the basis of physiological traits and phylogenetic position. Characteristics additional to those provided in the original description revealed that the type strain (strain K1T=ATCC BAA-34T=JCM 14045T) has the ability to use Fe(III) as an electron acceptor for acetate oxidation and has an electron donor and acceptor profile typical of a Geobacter species, contains abundant c-type cytochromes, and has a temperature optimum of 30 °C and a pH optimum near pH 7.0; traits typical of members of the genus Geobacter. Phylogenetic analysis of nifD, recA, gyrB, rpoB, fusA and 16S rRNA genes further indicated that T. thiogenes falls within the Geobacter cluster of the family Geobacteraceae. Based on extensive phylogenetic evidence and the fact that T. thiogenes has the hallmark physiological characteristics of a Geobacter species, Trichlorobacter thiogenes should be reclassified as a member of the genus Geobacter.

Download Full-text

Molecular identification ofSarcocystisspp. helped to define the origin of green pythons (Morelia viridis) confiscated in Germany

Parasitology ◽

10.1017/s0031182013001960 ◽

2013 ◽

Vol 141 (5) ◽

pp. 646-651 ◽

Cited By ~ 9

Author(s):

GASTÓN MORÉ ◽

NIKOLA PANTCHEV ◽

DALAND C. HERRMANN ◽

MAJDA GLOBOKAR VRHOVEC ◽

SABINE ÖFNER ◽

...

Keyword(s):

18S Rrna ◽

Sequence Data ◽

18S Rrna Gene ◽

Rrna Genes ◽

Rrna Gene ◽

Intermediate Hosts ◽

Apicomplexan Parasites ◽

Large Numbers ◽

Wide Range ◽

18S Rrna Genes

SUMMARYSarcocystisspp. represent apicomplexan parasites. They usually have a heteroxenous life cycle. Around 200 species have been described, affecting a wide range of animals worldwide, including reptiles. In recent years, large numbers of reptiles have been imported into Europe as pets and, as a consequence, animal welfare and species protection issues emerged. A sample of pooled feces from four confiscated green pythons (Morelia viridis) containingSarcocystisspp. sporocysts was investigated. These snakes were imported for the pet trade and declared as being captive-bred. Full length 18S rRNA genes were amplified, cloned into plasmids and sequenced. Two differentSarcocystisspp. sequences were identified and registered asSarcocystissp. fromM. viridisin GenBank. Both showed a 95–97% sequence identity with the 18S rRNA gene ofSarcocystis singaporensis.Phylogenetic analysis positioned these sequences together with otherSarcocystisspp. from snakes and rodents as definitive and intermediate hosts (IH), respectively. Sequence data and also the results of clinical and parasitological examinations suggest that the snakes were definitive hosts forSarcocystisspp. that circulate in wild IH. Thus, it seems unlikely that the infected snakes had been legally bred. Our research shows that information on the infection of snakes withSarcocystisspp. may be used to assess compliance with regulations on the trade with wildlife species.

Download Full-text