The genus Primula is extremely diverse in the east Himalaya-Hengduan Mountains (HHM) in China as a result of rapid radiation. In order to overcome the difficulty of morphological classification of this genus, we surveyed three plastid regions (rbcL, matK, and trnH-psbA) and two nuclear markers (ITS and ITS2) from 227 accessions representing 66 Primula species across 18 sections, to assess their discriminatory power as barcodes. We found that ITS alone or combined with plastid regions showed the best discrimination across different infrageneric ranks and at species level. We suggest rbcL + matK + ITS as the first choice at present to barcode Primula plants. Although the present barcoding combination performed poorly in many closely related species of Primula, it still provided many new insights into current Primula taxonomy, such as the underlying presence of cryptic species, and several potential improper taxonomic treatments. DNA barcoding is one useful technique in the integrative taxonomy of the genus Primula, but it still requires further efforts to improve its effectiveness in some taxonomically challenging groups.

To investigate the mechanisms responsible for the high diversity in central and adjacent southwestern China, we inferred the phylogeographic history of Primula ovalifolia from chloroplast DNA data. One hundred and thirty five individuals from 13 natural populations (including one Primula tardiflora population) were analyzed. A total of 23 haplotypes were identified. Most of them were exclusive to a single population. Strong phylogeography structure was detected, with N ST (0.936) significantly higher than G ST (0.784). Phylogenetic analysis showed that all haplotypes were clustered into three lineages (clades I, II, and III). High genetic diversity was revealed, possibly due to the interglacial contraction and glacial expansion. The three identified lineages may have undergone different historical processes after the mid-Pleistocene, due to their early divergent time. Multiple refugia were inferred for the three lineages. Pleistocene climatic oscillations, combined with the complex local topography, were responsible for the strong phylogeographic pattern of P. ovalifolia. Our analysis supports the hypothesis that the central and adjacent southwestern region of China was an important refugium for the survival, persistence, and further speciation of most East Asia flora, which has led to high species diversity in this region.

Even though ladybirds are well known as economically important biological control agents, an integrative framework of DNA barcoding research was not available for the family so far. We designed and present a set of efficient mini-barcoding primers to recover full DNA barcoding sequences for Coccinellidae, even for specimens collected 40 years ago. Based on these mini-barcoding primers, we obtained 104 full DNA barcode sequences for 104 species of Coccinellidae, in which 101 barcodes were newly reported for the first time. We also downloaded 870 COI barcode sequences (658 bp) from GenBank and BOLD database, belonging to 108 species within 46 genera, to assess the optimum genetic distance threshold and compare four methods of species delimitation (GMYC, bPTP, BIN and ABGD) to determine the most accurate approach for the family. The results suggested the existence of a 'barcode gap' and that 3% is likely an appropriate genetic distance threshold to delimit species of Coccinellidae using DNA barcodes. Species delimitation analyses confirm ABGD as an accurate and efficient approach, more suitable than the other three methods. Our research provides an integrative framework for DNA barcoding and descriptions of new taxa in Coccinellidae. Our results enrich DNA barcoding public reference libraries, including data for Chinese coccinellids. This will facilitate taxonomic identification and biodiversity monitoring of ladybirds using metabarcoding. As species are a fundamental biological category, accurately identifying them is an essential premise of biological studies. Tautz et al. 1 proposed DNA sequences as a species identification system for the first time. Subsequently, the 5' end of the mitochondrial cytochrome c oxidase subunit I gene (COI) has been suggested as a standardized DNA "barcode" for identifying species of all groups of animals, launching the DNA barcoding technology 2. The success of barcode identification based upon genetic distances ultimately depended on differences between intraand interspecific divergences 2-4. DNA barcoding using the 658 bp 5' region of mtCOI DNA sequence as a tool, has turned out as very efficient and reliable for identifying specimens of unknown origin and taxonomic status, and also for the identification of different developmental stages. The approximately 1600 base-pairs comprise a range of different functional domains showing heterogenous substitution patterns 5,6. In addition to its utility for distinguishing known species, the COI region has also been found suitable for revealing cryptic species 7-11 , and biogeographic and phylogeographic patterns, and also species level phylogenetic relationships 12-14. The family Coccinellidae is placed in the superfamily Coccinelloidea within the Coleopteran suborder Polyphaga 15,16. Coccinellids are well known as economically important biological control agents, but this family is ecologically and morphologically very diverse. It comprises about 490 genera and nearly 6000 described species worldwide 17. Due to relatively small body size ...

Five species of the genus Horniolus Weise, 1901 from China are revised, including the description of a new species, Horniolus hainanensis Chen & Ren, sp. n. Horniolus sonduongensis Hoàng, 1979 is reported from China for the first time. A key to the species from China is provided. Nomenclatural history, diagnoses, detailed descriptions, illustrations, and distribution for each species have been provided. A checklist of all known species of this genus is also presented.

Obtaining genetic information from museum specimens is a fundamental component of many fields of research, including DNA barcoding, population genetics, conservation genetics, and phylogenetic analysis. However, acquiring genetic information from museum specimens is challenging because of the difficulty in amplifying the target sequences due to DNA damage and degradation. Different pretreatments can significantly impact the purity and concentration of genomic DNA from museum specimens. Here, we assessed four pretreatment methods—use of 0.9% NaCl buffer, phosphate-buffered saline (PBS), Saline Tris-EDTA (STE) buffer, and sterile water—to determine which pretreatment is most suitable for DNA extraction from dried specimens of ladybird beetles. We completed a comprehensive phylogenetic analysis to test whether the sequences obtained from dried specimens enable proper phylogenetic inference. Our results showed that pretreatment can improve the quality of DNA from dried specimens. The pretreatment effects of 0.9% NaCl buffer and STE buffer were better than those of PBS buffer and sterile water. The phylogenetic analyses results showed that museum specimens can be used to generate cogent phylogenetic inferences. We report the optimum pretreatment methods for DNA extraction from dried ladybird beetles specimens as well as provide evidence for accurately determining phylogenetic relationships for museum specimens.