Research Article |
Corresponding author: Peter Decker ( decker@myriapoden-info.de ) Academic editor: Pavel Stoev
© 2018 Peter Decker.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Decker P (2018) Phylogenetic and morphological discord indicates introgressive hybridisation in two genera of Australian millipedes (Diplopoda, Polydesmida, Paradoxosomatidae). ZooKeys 809: 1-14. https://doi.org/10.3897/zookeys.809.30087
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Discord between molecular and morphological datasets was observed in two pairs of species of Australian millipedes in the family Paradoxosomatidae using morphological and molecular phylogenetic analysis (mitochondrial COI rDNA and 16 rRNA, and nuclear 28S rRNA). Close to the presumed distributional boundary between Pogonosternum nigrovirgatum (Carl, 1912) and Pogonosternum jeekeli Decker, 2017, near Dargo in Central Gippsland, Victoria, Pogonosternum specimens were collected which are phylogenetically closer to P. jeekeli in COI and 16S sequences, but are morphologically closer to P. nigrovirgatum. At Mount Osmond, Adelaide, South Australia, eight morphologically typical Somethus castaneus (Attems, 1944) specimens were collected are phylogenetically closer to S. castaneus in 28S genealogy, but three of the eight are closer to S. lancearius Jeekel, 2002 in COI genealogy. These two cases are discussed in terms of hybridisation, past introgressive hybridisation events and aberrant morphology.
Aberrant morphology, Arthropoda , COI, hybridisation, introgression, Myriapoda , 16S, 28S
While many cases of hybridisation in plants, fungi and animals are well known (
In Illinois, United States, the monotypic polydesmidan Illiniurus beattyi Shear, 1968 was later described as “transitional” in gonopod structure between Euryurus leachii Gray, 1832 and Auturus evides Bollman, 1887, both of which occur near the I. beattyi type locality (
The introduction of genes from the gene pool of one species into that of another during hybridization, especially near species boundaries, is called introgressive hybridisation and can affect nuclear or mitochondrial DNA (
Two cases of disagreement between relationships inferred from morphological similarity and molecular phylogenetics were observed in recent taxonomic studies of the Australian paradoxosomatid genera Pogonosternum Jeekel, 1965 (
Pogonosternum cf. nigrovirgatum (Carl, 1912) “Dargo”: 7 males, 1 female, 1 juvenile (NMV K-12202, K-13866, K-13867, K-13474, K-13868, SMNG VNR018276, VNR018277) were collected by hand in forest on Dargo Road, SSW of Dargo, central Gippsland, Victoria, 37.595S, 147.193E by P. Decker, K. Voigtländer and R. Mesibov on 14 August 2014.
Somethus castaneus (Attems, 1944): specimens were collected by hand at two localities in Mount Osmond Reserve, Adelaide, South Australia by P. Decker and K. Voigtländer : 1 male (SMNG VNR016973) and 3 females (SAM OM2149, SMNG VNR016975 and VNR016976) on a southwestern slope (34.969S, 138.654E, 27 August 2014, site number S110), and 5 males (SAM OM2138, SAM registration in progress, SMNG VNR018275) and 1 female (SMNG VNR018274) on a northern slope (34.962S, 138.659E, 23 August 2014, site number S90).
Specimens were killed and stored in 95% ethanol, with a change of ethanol after 1–2 months. One male of Somethus castaneus from Mt. Osmond (SAM OM2138) was found dead in the field. DNA was not obtained from this specimen. The material is deposited in the Museums Victoria, Melbourne, Victoria, Australia (NMV), the South Australian Museum, Adelaide, Australia (SAM) and the Senckenberg Museum of Natural History Görlitz, Görlitz, Germany (SMNG).
Preserved specimens were imaged with a Leica Z6 Apo stereo microscope and Leica DFC420 camera. Focus-stacked images were assembled from 25–40 source images using the software package Leica Application Suite 4.5. All images were later edited using Adobe Photoshop CS4 and assembled into plates. The distribution maps were created with ArcMap 10.
DNA was extracted from 2–4 legs from each of four male Pogonosternum cf. nigrovirgatum “Dargo” and nine Somethus castaneus specimens from Mt Osmond (Table
Site numbers, localities, GenBank accession numbers and repository accession numbers for all specimens analysed. NMV = Museum Victoria, Melbourne, Victoria, Australia; SAM = South Australian Museum, Adelaide, Australia; SMNG = Senckenberg Museum of Natural History Görlitz, Görlitz, Germany; SA = South Australia; VIC = Victoria.
Species | Site No. | Locality | Sex | GenBank Acc. No. COI | GenBank Acc. No. 16S | GenBank Acc. No. 28S | Voucher |
---|---|---|---|---|---|---|---|
Somethus castaneus | S110-1 | SA, Mt. Osmond, SW slope | male | KT948668 | KT964470 | SMNG VNR016973 | |
S110-2 | SA, Mt. Osmond, SW slope | female | MK170142 | MK142784 | SAM OM2149 | ||
S110-3 | SA, Mt. Osmond, SW slope | female | MK170143 | MK142785 | SMNG VNR016975 | ||
S110-4 | SA, Mt. Osmond, SW slope | female | MK170144 | MK142786 | SMNG VNR016976 | ||
S90-1 | SA, Mt. Osmond, N slope | male | MK170145 | MK142787 | SAM | ||
S90-2 | SA, Mt. Osmond, N slope | male | MK170146 | MK142788 | SAM | ||
S90-3 | SA, Mt. Osmond, N slope | male | MK170147 | MK142789 | SAM | ||
S90-4 | SA, Mt. Osmond, N slope | female | MK170148 | MK142790 | SMNG VNR018274 | ||
S90-5 | SA, Mt. Osmond, N slope | male | MK170149 | MK142791 | SMNG VNR018275 | ||
Pogonosternum nigrovirgatum | Dargo-1 | VIC, SSW of Dargo | male | MK170150 | MK170154 | NMV K-12202 | |
Dargo-2 | VIC, SSW of Dargo | male | MK170151 | MK170155 | NMV K-13866 | ||
Dargo-3 | VIC, SSW of Dargo | male | MK170152 | MK170156 | SMNG VNR018276 | ||
Dargo-4 | VIC, SSW of Dargo | male | MK170153 | MK170157 | SMNG VNR018277 |
Phylogenetic hypothesis was inferred for COI+16S, COI and 28S by using the maximum likelihood method conducted in MEGA6 (
The final alignments consisted of 618 bp of COI mtDNA and 1206 bp of 28S rRNA in Somethus, and 1158 bp for COI+16S in Pogonosternum. Individual alignments are available upon request from the author. The best-fit model of nucleotide substitution selected using MrModelTest 2 was the General Time Reversible model with gamma distribution and proportion of invariant sites (
Pogonosternum nigrovirgatum and P. jeekeli are very similar in somatic morphology, and the “Dargo” form agrees with both species in size, colouration, spiracle morphology and form of the leg pair 2 coxa in females. The “Dargo” form (Fig.
Pogonosternum cf. nigrovirgatum “Dargo”, male, right gonopod (NMV K-12202). A Posterior view B Lateral view C Anterior view D Mesal view. Abbreviations: fp1 = femoral process 1; fp2 = femoral process 2; lp = lateral process; prof = prolongation of femorite; S = solenomere; F = femorite. Scale bar: 0.5 mm.
Pogonosternum nigrovirgatum (Carl, 1902), ♂, right gonopod. A, B AMS KS96017 Ferntree Gully C, DNMV K-10248 from Sandringham and Brighton. Pogonosternum jeekeli Decker, sp. n., ♂, right gonopod E, FNMV K-10252 from Dyer Creek G, HNMV K-10250 from Bemm River. A, C Posterior view B, D Mesal view. Scale bar: 0.5 mm.
Bootstrap consensus tree for the combined mt COI+16S dataset in Pogonosternum; maximum likelihood, 1000 bootstrap replicates. Coloured dotted lines indicate specimens with similar gonopod morphology. Coloured bars indicate last male leg pairs with tibial and tarsal brushes (leg pair 7 = blue, leg pair 9 = green).
All six male specimens from Mt Osmond fully agree in gonopod morphology with S. castaneus and lack a medial prefemoral process (
However, two specimens from site S90 and one from site S110 form a separate, well-supported clade (100% bootstrap support) within Somethus, with genetic p-distances of 4.8–6.9% to S. lancearius, 5.3–6.9% to S. castaneus and 7.2–8.8 % to S. inflatus. The two Mt Osmond localities are in the centre of the S. castaneus distribution (Fig.
Despite the clear phylogenetic placement of this form within P. jeekeli as indicated by COI+16S gene trees (Fig.
Pogonosternum nigrovirgatum and P. jeekeli form a sister clade (
Somethus castaneus was sampled for sequencing at 13 localities covering most of the known species range, and genetic variability was shown to be low (up to 3.8% in uncorrected p-distances in COI) with three phylogenetic lineages (
Several of the paratypes of S. inflatus were collected in the Adelaide suburb of Glen Osmond, near Mt Osmond, in 1969 (
The three discordant S. castaneus found on Mt Osmond might be evidence for past introgression of mitochondrial DNA following hybridisation with another South Australian Somethus species. Alternatively, the anomalous individuals might represent a distinct and distantly related S. castaneus lineage which is either naturally occurring on Mt Osmond or introduced from another locality within the Adelaide Hills. There is no support for both hypotheses, but it seems that the likelihood of the presence (or former existence) on Mt Osmond of the in COI and 28S variable S. lancearius or S. inflatus is higher than that of a fourth distinct COI and 28S lineage in S. castaneus.
Future investigations with additional molecular markers and more individuals of from S. lancearius may not assist in clarifying the situation, as much of the former genetic variation of S. lancearius has probably been lost due to habitat loss and local extinctions. If the missing paratypes of S. inflatus are found in future, it might be possible to extract DNA and obtain sequences from them which could reveal whether S. inflatus in the Mt Osmond area has contributed mitochondrial COI to the local S. castaneus population.
The results presented here suggest that introgressive hybridisation may have occurred in the paradoxosomatid millipede genera Pogonosternum and Somethus in southeastern Australia. With the increasing use of molecular data in taxonomy and in barcoding projects, similar cases are likely to be found elsewhere. Interestingly, no evidence of introgressive hybridisation was found in more than 2000 COI sequences from Central European millipedes during the German Barcoding of Life Project (GBOL) (Wesener, Spelda, Reip, Decker pers. comm.). The phenomenon may be rare, or limited to narrow parapatric zones, as appears to be the case in Pogonosternum.
For loans of material I am very grateful to Kate Sparks (formerly at SAM), Peter Lillywhite and Catriona McPhee (NMV). Also sincere thanks to Karin Voigtländer (SMNG), Ulrich Burkhardt (SMNG), and Thomas Wesener (Bonn) for technical advice as well as interesting discussions. I am very grateful to Robert Mesibov (West Ulverstone), Meike Schuppenhauer (SMNG), Paul Marek (Blacksburg), and Henrik Enghoff (Copenhagen) for their helpful and constructive comments on earlier versions of the manuscript. The field trip of the author to Australia 2014 was financially supported by the ‘Förderkreis Naturkundemuseum Görlitz’.