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Research Article
Mazus motuoensis (Mazaceae), a new species from Xizang, China
expand article infoWen-Bin Ju§|, Xiong Li|, Heng-Ning Deng|, Meng Li, Xing-Jin He§, Xin-Fen Gao|, Bo Xu|
‡ Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu, China
§ Sichuan University, Chengdu, China
| University of Chinese Academy of Sciences, Beijing, China
¶ Nanjing Forestry University, Nanjing, China
Open Access

Abstract

Mazus motuoensis W.B.Ju, Bo Xu bis & X.F.Gao is a newly described species found in Xizang Autonomous Region, China. Morphologically, this species differs from all the other known Mazus species by having erect perennial herb form with a rhizome, presence of multicellular hairs, without basal leaves, opposite arrangement of stem leaves, and corolla lobes with erose-toothed margins. Molecular phylogenetic analysis using nuclear and cpDNA genes suggests that this new species occupies a basal position within Mazus. In conclusion, both morphological evidence and molecular phylogenetic analyses support that this species belongs to Mazus and represents an as-yet-unreported new species with distinct differences from other species within the genus.

Key words

Mazus, molecular phylogenetics, morphology, taxonomy

Introduction

Mazus Loureiro is the largest genus within the family Mazaceae Reveal (2011), comprising 38 accepted species (POWO 2022). Most of these species are found in eastern and southeastern Asia, Australia, and New Zealand (Li 1954; Hsieh 2000). The genus is characterized by a distinct two-lipped corolla (3/2-bilabiate), a palate with two longitudinal plaits, and a capsule enclosed in a persistent calyx (Fischer 2004; Deng et al. 2019). In China, there are approximately 31 species and three varieties have been recognized (Hong et al. 1998; Hsieh 2000; Deng et al. 2016; Ying 2019; Xiang et al. 2021; Li et al. 2022), which is the distribution and diversity center of the genus (Hsieh 2000). Originally categorized under Scrophulariaceae through morphological studies (Von Wettstein 1891; Thieret 1954, 1967; Hong et al. 1998), molecular phylogenetic analyses unveiled a robustly supported clade uniting Mazus and Lancea Hook.f. & Thomson, recognized as the subfamily Mazoideae within Phrymaceae (Beardsley and Olmstead 2002). Nevertheless, subsequent phylogenetic studies confirmed that Mazus should be separated from Phrymaceae (Oxelman et al. 2005; Albach et al. 2009; Xia et al. 2009; Schäferhoff et al. 2010), leading to the establishment of a new family called Mazaceae within the Lamiales. The latest phylogenetic studies and morphological evidence indicate that M. lanceifolius Hemsly is a distinct species, positioned at the most basal branch within the Mazaceae family, and is the sister genus to the three recognized genera Dodartia, Lancea, and Mazus (Xia et al. 2012; Deng et al. 2019; Xiang et al. 2021).

In 2022, a field survey was conducted in Motuo County, located within the Xizang Autonomous Region of southwest China, the authors discovered an unknown species of Mazaceae in an evergreen broad-leaved forest. Through careful comparison with specimens, related literature, and phylogenetic analysis of Mazaceae, it was concluded that this species represents a new addition to the Mazus.

Materials and methods

Morphological analysis

One population of this new species was rediscovered in Mar 2022 in Xizang Autonomous Region, China. Morphological observations of the new species were conducted using living plants collected from the type locality, as well as type specimens deposited at CDBI. Detailed photographs of morphological features, such as rhizomes, multicellular hairs, stems, leaves, inflorescences, and flowers, were taken using a digital camera and stereoscope. Measurements were carried out on both wild plants and pressed specimens using a ruler and a metric vernier caliper. Digital herbarium images of Mazus specimens were sourced from diverse outlets, including JSTOR Global Plants (http://plants.jstor.org/), the Global Biodiversity Information Facility (https://www.gbif.org/zh/), the Chinese Virtual Herbarium (https://www.cvh.ac.cn/), and Europeana (https://www.europeana.eu/en/search). A thorough examination and comparison of these images with the new species ensued. Subsequently, the morphological attributes of the species were meticulously described in accordance with the guidelines provided by the Flora of China (Hong et al. 1998).

Assessment of conservation status

In the field, we conducted an estimation of the population size of the new species and evaluated the factors posing threats to its existence. In order to determine the conservation status of the new species, we applied the established criteria as outlined by the International Union for Conservation of Nature (IUCN 2019) Red List.

DNA sequencing and outgroup selection

We extracted total DNA from silica gel-dried leaves of the new species using a modified CTAB protocol (Doyle and Doyle 1987). To determine the phylogenetic position of the new species within the Mazus genus, we employed two datasets for our analysis. The first dataset consisted of a combined matrix of two cpDNA regions (rbcL, trnL-trnF), while the second dataset was nrITS. The DNA sequences were amplified and sequenced following the methods described by Deng et al. (2019), using the primers specified in their study. Based on previous phylogenies (Deng et al. 2019; Xiang et al. 2021), 20 species with 28 accession of the relatives of M. motuoensis were selected as ingroups. Additionally, we chose five species from three different genera as outgroups. The related sequences were obtained from NCBI (https://www.ncbi.nlm.nih.gov/). The GenBank accession numbers for the new species are OQ383430 (trnL-F), OQ383431 (rbcL) and OP720888 (ITS). A comprehensive list of all species included in the phylogenetic analysis, along with their respective accession numbers, can be found in Table 1.

Table 1.

Information of samples used for phylogenetic inference in this study.

Taxa rbcL trnL-F ITS
Mazus alpinus Masamune 1 KX783481 KX783520 MK192641
Mazus alpinus Masamune 2 KX783480 KX783519 MK192642
Mazus caducifer Hance 1 KX783477 KX783516 MK192664
Mazus caducifer Hance 2 KX783487 KX783526 MK192659
Mazus celsioides Handel-Mazzetti KX783486 KX783525
Mazus fruticosus Bo Li, D.G.Zhang & C.L.Xiang 1 KX783470 KX783509 MK192660
Mazus fruticosus Bo Li, D.G.Zhang & C.L.Xiang 2 KX783471 KX783510 MK192649
Mazus gracilis Hemsley FJ172729 FJ172687 FJ172738
Mazus humilis Handel-Mazzetti MK266421 MK192667
Mazus japonicus (Thunburg) O. Kuntze FJ172728 FJ172686
Mazus japonicus var. delavayi (Bonati) Tsoong KX783482 KX783521
Mazus longipes Bonati KX783474 KX783513 MK192652
Mazus miquelii Makino 1 KX783475 KX783514 MK192637
Mazus miquelii Makino 2 KX783476 KX783515 MK192655
Mazus miquelii Makino 3 KX783483 KX783522 MK192656
Mazus motuoensis W.B.Ju, Bo Xu bis & X.F.Gao OQ383431 OQ383430 OP720888
Mazus novaezeelandiae W.R.Barker KX783469 KX783508 MK192676
Mazus omeiensis H. L. Li 1 KX807209 KX807208 MK192636
Mazus omeiensis H. L. Li 2 FJ172731 FJ172688 MK192663
Mazus procumbens Hemsley KX783478 KX783517 MK192647
Mazus pulchellus Hemsley KX783472 KX783511 MK192638
Mazus pumilio R.Brown KX783468 KX783507 MK192671
Mazus pumilus (N. L. Burman) Steenis 1 MK266346 KX807206 MH711724
Mazus pumilus (N. L. Burman) Steenis 2 HM850162 KX807207 FJ172737
Mazus reptans N.E. Brown HQ384872 AF479004 AF478940
Mazus spicatus Vaniot FJ172730 FJ172689 FJ172740
Mazus sunhangii D. G. Zhang & T. Deng 1 KX783485 KX783524
Mazus sunhangii D. G. Zhang & T. Deng 2 KX783484 KX783523
Mazus xiuningensis X. H. Guo & X. L. Liu MK266349 MK266430
OUTgroup
Puchiumazus lanceifolius (Hemsly) Bo Li, D. G. Zhang & C. L. Xiang 1 MW373737 MW373741 MW364623
Puchiumazus lanceifolius (Hemsly) Bo Li, D. G. Zhang & C. L. Xiang 2 MW373738 MW373742 MW364624
Dodartia orientalis Linnaeus JQ342984 JQ342981 JQ342980
Lancea tibetica J. D. Hooker & Thomson 1 KX783467 KX807205 MK192678
Lancea tibetica J. D. Hooker & Thomson 2 MF786661 FJ172685 FJ172736

Phylogenetic analysis

The sequence chromatograms were visually inspected on Sequencher 5.2.4 (Gene Codes Corporation) and integrated into a single sequence. All sequences were then aligned with MUSCLE in MEGA 7.0.14 (Kumar et al. 2016) and manually adjusted. Phylogenetic analyses were performed based on the combined cpDNA dataset (rbcL and trnL-trnF) and the nrITS dataset using both the maximum likelihood (ML) and Bayesian inference (BI), respectively. We did not combine the cpDNA and nrITS datasets for analysis because of the different sampling of taxa in the datasets. Settings of parameters during analysis follow those presented in Deng et al. (2019) and Xiang et al. (2021).

Results and discussion

Morphological analysis

Morphologically, the new species has intermediate characteristics of Mazus and Puchiumazus. The new species has characteristics such as rhizomes, erect stems, and stem leaves opposite similar to Puchiumazus. However, the plant of this new species is covered with multicellular hair and has stems that are not quadrangular and leaf blade elliptic-ovate, which distinguishes it from the only known species, Puchumazus lancefolius (Hemsley) Bo Li, D.G.Zhang, and C.L.Xiang (Xiang et al. 2021). There are also perennial herb species with erect stems and opposite cauline leaves in the genus of Mazus, such as M. caducifer Hance (1882). But the new species has a series of ray characteristics not commonly seen in Mazus, including single erect unbranched stems without basal leaves, stem leaves many and opposite, petioles nearly absent, lobes margin erose-toothed. In conclusion, based on the morphological key provided by Hong et al. (1998), the new species is classified morphologically within the Mazus and represents an anomalous existence.

Phylogenetic analysis

The phylogenetic tree was generated using a combined cpDNA matrix, consisting of 34 aligned sequences and comprising 2197 characters (rbcL: 1318 bp; trnL-trnF: 879 bp). Additionally, the nrITS matrix included 28 aligned sequences and comprised 703 characters. Due to differences in taxon sampling between the cpDNA and nrITS datasets, they were not combined for analysis. Both maximum likelihood (ML) and Bayesian inference (BI) methods yielded congruent topologies. Therefore, only the results of the ML trees are presented (Figs 1, 2: MLBS: 100%, BIPP: 1.00; all support values follow this order hereafter).

Figure 1. 

Phylogenetic relationships of M. motuoensis and related species inferred from ML and BI analyses based on the nrITS dataset. Numbers on the branches indicate the bootstrap support of the ML and the posterior probability of BI analyses.

The results of nrITS and cpDNA phylogenies (Figs 1, 2) show that Dodartia-Lancea formed the basal clade with Puchiuomazus clade as sister to Mazus (100%, 0.99 in nrITS tree; 100%, 1 in cpDNA tree). The new species was located at the base of the genus Mazus with other sampled species of Mazus together forming monophyletic clade with strong support (Fig. 1: 85%, 0.99; Fig. 2: 100%, 1). This phylogenetic result is broadly consistent with previous studies (Xiang et al. 2021), indicating that the new species belongs to the Mazus representing a unique presence of the genus.

Figure 2. 

Phylogenetic relationships of M. motuoensis and related species inferred from ML and BI analyses based on the combined dataset of rbcL and trnL-trnF. Numbers on the branches indicate the bootstrap support of the ML and the posterior probability of BI analyses.

Taxonomic treatment

Mazus motuoensis W.B.Ju, Bo Xu bis & X.F.Gao, sp. nov.

Figs 3, 4, 5

Diagnosis

The new species is distinguished from congeneric species by its rhizomes, perennial herb covered with multicellular white villus, erect and unbranched stems, having no basal leaves, stem leaves opposite, subsessile, lower lobes margins erose-toothed.

Type

China, Xizang, Motuo County, DeXing town, Nibi Valley, ditch edge in the forest, 29°22'27.98"N, 95°10'0.88"E, alt. 2253 m. 31 Mar 2022, WenBin JU & XIONG LI, YLZB08519 (holotype: CDBI0279767; isotypes: CDBI0279765, CDBI0279766)

Description

Perennial herbs, 15–25 cm tall, the whole plant is covered with long white soft multicellular hairs. Rhizome white. Stems erect, unbranched. Leaves opposite, numerous, petiole inconspicuous to nearly absent; lower leaf blade scalelike and small, obovate-oblong, apex obtuse, middle and upper leaves with leaf blade elliptic to ovate, papery, 0.8–4.0 × 0.4–1.8 cm, adaxially clothed with multicellular hairs, abaxially subglabrous, multicellular hairs on veins, base cuneate, margin serrate, lateral veins 3–5 pairs. Racemes terminal, ascending to 5 cm long, lax, fewer than 5; pedicels 4–6 mm, glabrous or with a few multicellular hairs; bracts tiny, narrowly lanceolate to linear, glabrous. Calyces broadly campanulate, ca. 6 mm long, 5–veined, glabrous outside and inside, lobes 5, triangular-lanceolate, as long as tube, apex acute, midrib conspicuous, lateral veins inconspicuous. Corolla 1.2–1.5 cm long, white, but often purple on upper lobes, glabrous outside and inside apart from clavate hairs on palate; tube 0.3–0.5 cm long, shorter than calyx; limb 2–lipped, upper lip bilobed, slightly upwarp, lobes triangular ovate, apex subacute, sometimes weakly obtuse or retuse; lower lip trilobed, lobes margins erose-toothed, middle lobe usually rounded, smaller than lateral lobes, yellow palate comprising 2 longitudinal elevations extending from point of filament fusion to the base of lower lobes, with erect clavate hairs. Stamens 4, didynamous, glabrous, inserted at the same level in distal part of tube, inserted at the distal end of the tube at the same level, included; anterior pair longer, curved, appressed to corolla tube, posterior pair spreading; anthers bithecal, locules divergent, apically connivant, positioned adjacent to corolla tube on upper lip; filaments filiform, glabrous. Ovary ca. 2 mm long, glabrous, ovoid; styles ca. 7 mm long, included, glabrous, exserted beyond anthers, stigma bilobed. Fresh capsule and calyx light green, included by persistent calyx.

Figure 3. 

Living images of M. motuoensis A habit B inflorescence in frontal view C inflorescence in rear view D rhizome.

Figure 4. 

Morphology of M. motuoensis A plant B leaves C old stems D multicellular hairs attached to stem E flower F unfolded corolla, showing limb upper lip and lower lip G bract H, I calyx J anthers K ovary and style.

Figure 5. 

Line drawings of M. motuoensis A whole plant B multicellular hairs attached to stem C leaves D inflorescence E flower F unfolded corolla G calyx H ovary and style. Drawn by Mr. Zhen-long Liang.

Distribution and habitat

Mazus motuoensis is currently known from Nibi Valley, Motuo County, Xizang, China. It can be found under evergreen broad-leaved forest at altitudes of 2253 m.

Phenology

Flowering was observed from May to June.

Etymology

The specific epithet “motuoensis” refers to the locality, Motuo County, Xizang, China.

Vernacular name

Simplified Chinese: 墨脱通泉草; Chinese pinyin: Mòtuō Tōngquáncǎo.

Conservation status

Currently, the authors have discovered only one population of Mazus motuoensis from one single locality in Nibi Valley of Motuo County in Xizang Province, China, and ca. 30 individuals from the type locality. Evergreen broad-leaved forests are widely distributed in this area, so we speculate that this new species has a relatively wide distribution range. Due to insufficient field survey, the natural distribution of this species in the wild is not clear. Following the IUCN Red List criteria (2019), we suggest this species placement in the Data Deficient.

Acknowledgements

We would like to express our sincere thanks to Mr. Zhenlong Liang (Chengdu Institute of Biology, Chinese Academy of Sciences) for his help with the DNA extraction, PCR amplification and the line drawing. We are grateful to Dr. Fei Zhao for providing valuable suggestions during the phylogenetic analysis process of the new species.

Additional information

Conflict of interest

The authors have declared that no competing interests exist.

Ethical statement

No ethical statement was reported.

Funding

This research was supported by the Biological Resources Programme, Chinese Academy of Sciences, BRP CAS (Grant No. KFJ-BRP-017-102), and the Wild Plants Sharing and Service Platform of Sichuan Province.

Author contributions

Conceptualization: XFG. Investigation: ML, HND. Methodology: XJH. Project administration: BX. Software: XL. Writing – original draft: WBJ.

Author ORCIDs

Wen-Bin Ju https://orcid.org/0000-0002-5665-6290

Xiong Li https://orcid.org/0000-0002-1450-8679

Heng-Ning Deng hhttps://orcid.org/0000-0002-0581-0649

Meng Li https://orcid.org/0000-0003-2085-3388

Xing-Jin He https://orcid.org/0000-0003-2064-0112

Xin-Fen Gao https://orcid.org/0000-0002-5703-1639

Bo Xu https://orcid.org/0000-0003-3507-9321

Data availability

All of the data that support the findings of this study are available in the main text.

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