Daniel Ebert

Chloroplast simple sequence repeats (cpSSRs): technical resources and recommendations for expanding cpSSR discovery and applications to a wide array of plant species.

Chloroplast microsatellites, or simple sequence repeats (cpSSRs), are typically mononucleotide tandem repeats. When located in the noncoding regions of the chloroplast genome (cpDNA), they commonly show intraspecific variation in repeat number. Despite the growing number of studies applying cpSSRs, studies of economically important plants and their relatives remain over‐represented. Thus, the potential of cpSSRs to offer unique insights into ecological and evolutionary processes in wild plant species has yet to be fully realized. This review provides an overview of the technical resources available to aid cpSSR discovery including a list of cpSSR primer sets available and cpDNA sequencing resources. Our updated analysis of 99 whole chloroplast genomes downloaded from GenBank confirms that potentially variable cpSSRs are abundant in the noncoding cpDNA of plants. Overall variation in the frequency of cpSSRs was extreme, ranging from one to 700 per genome (median = 93), while in 81 vascular plants, between 35 and 160 cpSSRs were detected per genome (median = 86). We offer five recommendations to aid wider development and application of cpSSRs: (i) When genus‐specific cpSSR primers are available, cross‐species amplification can often be fruitful. (ii) While potentially useful, universal cpSSR primers at best provide access to only a small number of variable markers. (iii) De novo sequencing of noncoding cpDNA is the most effective and efficient way to develop cpSSR markers in wild species. (iv) DNA sequencing of cpSSR alleles is essential, given the complex nature of the genetic variation associated with hypervariable cpDNA regions. (v) The reliability of cpSSR length based genetic assays need to be validated in all studies.

Comparative genetic study confirms exceptionally low genetic variation in the ancient and endangered relictual conifer, Wollemia nobilis (Araucariaceae)

The Wollemi pine, Wollemia nobilis (Araucariaceae), was discovered in 1994 as the only extant member of the genus, previously known only from the fossil record. With fewer than 100 trees known from an inaccessible canyon in southeastern Australia, it is one of the most endangered tree species in the world. We conducted a comparative population genetic survey at allozyme, amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) loci in W. nobilis, Araucaria cunninghamii and Agathis robusta — representatives of the two sister genera. No polymorphism was detected at 13 allozyme loci, more than 800 AFLP loci or the 20 SSR loci screened in W. nobilis. In Ag. robusta only one of 12 allozyme loci, five of 800 AFLP loci and none of the 15 SSR loci were variable. For A. cunninghamii, 10 of > 800 AFLP loci and five of 20 SSR loci were variable. Thus low genetic diversity characterizes all three species. While not ruling out the existence of genetic variation, we conclude that genetic diversity is exceptionally low in the Wollemi pine. To our knowledge this is the most extreme case known in plants. We conclude that the combination of small population effects, clonality and below‐average genetic variation in the family are probable contributing factors to the low diversity. The exceptionally low genetic diversity of the Wollemi pine, combined with its known susceptibility to exotic fungal pathogens, reinforces current management policies of strict control of access to the pines and secrecy of the pine locations.

A new set of universal de novo sequencing primers for extensive coverage of noncoding chloroplast DNA: new opportunities for phylogenetic studies and cpSSR discovery

We present a new set of universal de novo sequencing primers targeting noncoding chloroplast DNA. The set of 107 polymerase chain reaction (PCR) primers span approximately 86% of the noncoding nucleotides in the large single copy region of Nicotiana tabacum, Oryza sativa and the orchid Phalaenopsis aphrodite. PCR tests confirmed the primers are effective in a wide range of monocots and dicots. More than 19.5 kb of cpDNA sequence was obtained across representative orchid genera with up to 82 chloroplast simple sequence repeats (cpSSRs) detected per genus. This primer set will facilitate both phylogenetic studies and rapid discovery of cpSSRs for plants, such as orchids, where there are limited genomic resources.

Phylogeny and evolution of the Meliphagoidea, the largest radiation of Australasian songbirds

The Meliphagoidea comprises the largest radiation of Australasian passerines. Here we present the first detailed molecular phylogenetic analysis of its families and genera, particularly the Acanthizidae, using sequences from nine gene regions including both mitochondrial and nuclear DNA. Our results support some suggested relationships but challenge other groupings, particularly in Meliphagidae and Acanthizidae. Maluridae is sister to all other members of the superfamily. With appropriate taxon sampling and multilocus data, we provide the first strong molecular evidence supporting earlier recognition of bristlebirds, Dasyornis, as a separate family, Dasyornithidae. We further clarify its position as sister to Acanthizidae+Pardalotidae+Meliphagidae. Pardalotidae is sister to Acanthizidae, and thus its retention as a separate family is arbitrary. The meliphagid genus Lichenostomus is polyphyletic. We find no support for the current subfamily structure within Acanthizidae but recognise a clade that includes members of the subfamily Sericornithinae excluding Oreoscopus and Acanthornis. Subfamily Acanthizinae is paraphyletic. Surprisingly, the Tasmanian island endemic Acanthornis magna of mesic habitats is sister to the Aphelocephala whitefaces of mainland Australian xeric zones. This is one of several unexpected alignments of taxa as sisters that probably reflects the age of the Meliphagoidea. We find no evidence for separate radiations of New Guinean and Australian members of the Meliphagoidea.

Genetic evidence for cooperative polyandry in reverse dichromatic Eclectus parrots

Group size in cooperatively polyandrous birds appears to be limited by the diminishing probability that any one male will achieve paternity as group size increases. Here we use molecular techniques to analyse the mating system of Eclectus parrots, a species with a surprisingly large number of males (up to seven) in each group. We show that their social groups are not composed of kin, and that all males seek direct fitness. Most broods of two nestlings had just one father. However, further group males gained paternity in other nesting attempts within and between breeding seasons. Although the likelihood of multiple paternity increased with the number of males in the group, only a subset of group males gained paternity over the eight-year study period. Some males gained paternity at irregular intervals with the same female over multiple years, and many also mated polygynandrously at widely dispersed nests. Our data also confirm that females monopolise scarce breeding hollows over multiple years. We suggest that Eclectus parrots occur in larger group sizes than other cooperatively polyandrous species as a result of a severely biased operational sex ratio brought about by a scarcity of breeding females and a male-biased adult sex ratio.

Chloroplast simple sequence repeat markers for evolutionary studies in the sexually deceptive orchid genus Chiloglottis

The orchids in the genus Chiloglottis are pollinated exclusively by sexual deception. Extensive sequencing (> 19.5 kb) of noncoding chloroplast DNA revealed that simple sequence repeats (cpSSRs) were abundant, enabling a set of 41 cpSSR markers to be developed. All markers were polymorphic across the genus. Polymorphism reflected variation at both mononucleotide repeats and indels. For a subset of four taxa with 40 samples each, locus polymorphism varied from 46 to 81%, while the number of haplotypes ranged from seven to 21 per taxon. Extensive differentiation among the taxa was detected. These cpSSRs markers will enable novel insights into the evolution of this unique genus.

Rhodopsin population genetics and local adaptation: variable dim‐light vision in sand gobies is illuminated

The visual pigments of fish are thought to be adapted to the variable spectral qualities of aquatic light environments. Most research on the role of natural selection on the evolution of rhodopsins and dim‐light vision in fish has focused on variation among species and higher taxa. In this issue, Larmuseau et al. reveal substantial intraspecific sequence variation in RH1 (the rhodopsin gene) in sand gobies (Pomatoschistus minutus). Using population genetics and molecular evolution approaches, they detect positive selection on RH1 and find evidence for adaptation to local light conditions.