The Chloroplast Phylogenomics and Systematics of Zoysia (Poaceae)

Abstract

The genus Zoysia Willd. (Chloridoideae) is widely distributed from the temperate regions of Northeast Asia-including China, Japan, and Korea-to the tropical regions of Southeast Asia. Among these, four species-Zoysia japonica Steud., Zoysia sinica Hance, Zoysia tenuifolia Thiele, and Zoysia macrostachya Franch. & Sav.-are naturally distributed in the Korean Peninsula. In this study, we report the complete plastome sequences of these Korean Zoysia species (NCBI acc. nos. MF953592, MF967579~MF967581). The length of Zoysia plastomes ranges from 135,854 to 135,904 bp, and the plastomes have a typical quadripartite structure, which consists of a pair of inverted repeat regions (20,962~20,966 bp) separated by a large (81,348~81,392 bp) and a small (12,582~12,586 bp) single-copy region. In terms of gene order and structure, Zoysia plastomes are similar to the typical plastomes of Poaceae. The plastomes encode 110 genes, of which 76 are protein-coding genes, 30 are tRNA genes, and four are rRNA genes. Fourteen genes contain single introns and one gene has two introns. Three evolutionary hotspot spacer regions-atpB~rbcL, rps16~rps3, and rpl32~trnL-UAG-were recognized among six analyzed Zoysia species. The high divergences in the atpB~rbcL spacer and rpl16~rpl3 region are primarily due to the differences in base substitutions and indels. In contrast, the high divergence between rpl32~trnL-UAG spacers is due to a small inversion with a pair of 22 bp stem and an 11 bp loop. Simple sequence repeats (SSRs) were identified in 59 different locations in Z. japonica, 63 in Z. sinica, 62 in Z. macrostachya, and 63 in Z. tenuifolia plastomes. Phylogenetic analysis showed that the Zoysia (Zoysiinae) forms a monophyletic group, which is sister to Sporobolus (Sporobolinae), with 100% bootstrap support. Within the Zoysia clade, the relationship of (Z. sinica, Z japonica), (Z. tenuifolia, Z. matrella), (Z. macrostachya, Z. macrantha) was suggested.

Keywords: Zoysia; plastome evolution; rpl3-trnL-UAG spacer; simple sequence repeats; small inversion.

Figure 1

Genome map of the complete chloroplast sequences of Zoysia japonica and three other Zoysia species sequenced in this study. The gene order and structure among Zoysia species are the same. The gray region within the inner circle shows the GC content ratio.

Figure 2

Comparison of the borders of the large single-copy (LSC), small single-copy (SSC), and inverted repeat (IR) regions among the plastomes of five Poaceae species. The IR regions of Zoysia species extend into the ndhF gene, which is characteristic of the PACMAD clade of Poaceae. Four comparative species were selected from closely related genera to Zoysia.

Figure 3

Sliding window analysis of the whole plastomes of six Zoysia species. The x-axis represents the midpoint position of a window, and the y-axis represents the nucleotide diversity of each window. Three noncoding regions (atpB~rbcL, rpl16~rps3, and rpl32~trnL-UAG) show more than 2% sequence divergences among the six Zoysia species.

Figure 4

The sequence difference between the rpl32 and trnL-UAG genes among six Zoysia species. All six species of Zoysia have an 11 bp long loop region, but this region has a different orientation in Z. japonica, Z. sinica, and Z. macrostachya compare to that of Z. tenuifolia, Z. matrella, and Z. macrantha. This difference is due to a small inversion at the 22 bp long stem region (orange color).

Figure 5

A maximum likelihood (ML) tree constructed from the plastomes of 26 species within the subfamily Chloridoideae (Poaceae) with Danthonia californica as an outgroup. The numbers at each node indicate the ML bootstrap values/Bayesian posterior probability. The ML tree was constructed from 76 protein-coding and 4 rRNA genes by RaxML. The newly sequenced species in this study are indicated in red. The short internal nodes among Zoysia species are expanded in the box at the upper left corner.