Molecular and morphological characterisation of Xiphinema americanum-group species (Nematoda: Dorylaimida) from California, USA, and other regions, and co-evolution of bacteria from the genus Candidatus Xiphinematobacter with nematodes

in Nematology
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The Xiphinema americanum-group is a large species complex containing more than 50 nematode species. They are economically important because they are vectors of nepoviruses. The species differentiation of X. americanum-group is problematic because the species share similar morphological characters. In the present study we collected nematode samples from different locations in the USA, Italy and Russia. Six valid species, X. americanum s. str., X. brevicolle, X. californicum, X. pachtaicum, X. rivesi and X. simile, and four unidentified putative Xiphinema species were characterised by morphology and sequencing of D2-D3 of 28S rRNA, ITS1 rRNA and mitochondrial COI genes. New nematode sequences generated totalled 147. Phylogenetic relationships of the X. americanum-group species reconstructed by Bayesian inference for D2-D3 of 28S rRNA gene sequences did not provide clear species delimitation of the samples studied, although the mtDNA presented interspecific variations useful for demarcation among species. Xiphinema americanum s. str., X. californicum, X. pachtaicum, X. rivesi, and two unidentified Xiphinema species were found in 72 soil samples from California. We also reconstructed the phylogenetic relationships using partial 16S rRNA gene sequences within endosymbiotic bacteria of the genus Candidatus Xiphinematobacter and provided solid evidence for distinguishing 17 species of this genus based on the analysis of new and previously published sequences. Fifty-five new bacterial sequences were obtained in the present study and comparison of the bacterial 16S rRNA and nematode COI phylogenies revealed a high level of co-speciation events between host and symbiont.

Molecular and morphological characterisation of Xiphinema americanum-group species (Nematoda: Dorylaimida) from California, USA, and other regions, and co-evolution of bacteria from the genus Candidatus Xiphinematobacter with nematodes

in Nematology



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    Photomicrographs of anterior region of females. A-D: Xiphinema pachtaicum (CD49, V1, V2a, V2c); E, F: X. americanum s. str. (CD48, CD1074); G, H: Xiphinema sp. 1 (CD96, CD50). (Scale bar = 10 μm.)

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    Photomicrographs of posterior region of females. A-D: Xiphinema pachtaicum (V1, V2a, CD49, CD49); E-H: X. americanum s. str. (CD48, XA39, CD1244, XA34); I-L: Xiphinema sp. 1 (CD47, CD50, CD91, XA20). (Scale bar = 10 μm.)

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    Photomicrographs of anterior region of females. A: Xiphinema californicum (XA44); B: X. rivesi (CD1715); C: X. americanum s. str. (CD82b); D: X. rivesi (CD1101). (Scale bar = 10 μm.)

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    Photomicrographs of posterior region of females. A-D: Xiphinema californicum (CD1247, XA41, XA44, CD3); E, F: X. rivesi (CD1715); G: Xiphinema sp. 2 (CD152); H: X. americanum s. str. (CD82b); I-L: X. rivesi (CD1165, CD1101, CD1101, CD1096). (Scale bar = 10 μm.)

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    Phylogenetic relationships within the Xiphinema americanum-group. Bayesian 50% majority rule consensus tree as inferred from the analysis of the D2 and D3 expansion segments of 28S rRNA sequence alignment under a GTR + I + G model. Newly obtained sequences are indicated in bold.

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    Fragment of the D2 segment of 28S rRNA gene alignment with signature sequences for some Xiphinema americanum-group species. Numbers in brackets after species name indicates other putatively identified species under similar sequence fragment: (1) X. diffusum; (2) X. rivesi and X. thornei; (3) X. californicum and X. pacificum; (4) X. citricolum; (5) X. tarjanense. Numbers in brackets after accession number is the number of sequences.

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    Phylogenetic relationships within the Xiphinema americanum-group. Bayesian 50% majority rule consensus tree as inferred from the analysis of the partial COI sequence alignment under a HKY + I + G model. Posterior probabilities more than 70% are given for appropriate clades. New sequences obtained in this study are in bold. Identification of some species made in this study are given in the right from vertical bars. Original species identification are provided in appropriate clades.

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    Phylogenetic relationships within species of the clade II of the Xiphinema americanum-group as defined by Archidona-Yuste et al. (2016). Bayesian 50% majority rule consensus tree as inferred from the analysis of the ITS1 rRNA sequence alignment under a GTR + I + G model. Newly obtained sequences are indicated in bold.

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    Phylogenetic relationships within Candidatus Xiphinematobacter. Bayesian 50% majority rule consensus trees as inferred from 16S rRNA gene sequences alignment under a GTR + I + G model. Newly obtained sequences are indicated in bold.

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    Co-phylogenetic relationships between the endosymbionts Candidatus Xiphinematobacter (right, 16S rRNA tree) and their hosts Xiphinema americanum-group species (left, COI tree). Bayesian 50% majority consensus trees with posterior probability values more than 70% for appropriate clades. Dotted lines show the association between a X. americanum-group species and its symbiont.

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    Reconciliation between Candidatus Xiphinematobacter (16S rRNA tree) and Xiphinema (COI tree) phylogenies. One of 291 isomorphic solutions with 17 co-speciations, four duplications and host switches, and two losses (total cost = 10). The reconciliation of bacteria and host trees was generated with Jane 4. Thin and thick lines represent bacteria and their nematode hosts, respectively. Empty circles represent co-speciations, arrows represent host switches, and dashed lines represent sorting events.

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    Contribution of each XiphinemaCandidatus Xiphinematobacter association to a general co-evolution. Each bar represents a jack-knifed squared residual and error bars represent upper 95% confidence intervals. The dashed line indicates the median squared residual value.

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