*Article* **Macro-Morphological and Ecological Variation in** *Rosa sericea* **Complex**

**Fazal Ullah 1,2 , Yundong Gao 1,\* , ˙Idris Sari 3 , Rui-Fang Jiao 1,4, Saddam Saqib 2,5 and Xin-Fen Gao 1,\***

**Abstract:** Taxon delimitation and identification are important in many areas of biology, especially in taxonomy and conservation. Taxonomic treatment is required to establish and justify recommendations in conservation biology for the group being considered. Imperfect and controversial taxonomy can undermine conservation assessment. We studied 71 populations; 665 individuals corresponding to the morphology of the *Rosa sericea* complex (including six taxa, one of which has extremely narrow distributions) were collected from sympatric, parapatric, and allopatric populations distributed in China. This study aims to investigate whether the complex species are macromorphologically different species and evaluate the rare taxa of the complex for conservation priority. The morphological characters and principal component analysis (PCA) of the *R. sericea* complex showed that the complex species have overlapping characters but can distinguish morphologically. The species of *R. sericea* complex systematics status based on previous DNA sequencing is controversial. The ecological habitat's current morphological characters only delimit the *R. morrisonensis* (in Taiwan). To evade mistakes in species conservation, we recommend that taxonomical knowledge be needed to ensure success in protecting target species. Thus, the complementarity of systemic and conservation assurance makes conservation actions more necessary for the complex's rare taxa. The ecological niche modelling (ENM) results showed that habitats of these conspecific taxa would be shrunken. With the presence of snapshots in time, the geography of taxa might decrease rapidly in representative entirely of the Geographic space (G-space) and Environmental space (E-space) that such taxa are bright to inhabit. So far, the significant inferences meant for the niche occupy the most incredible comparative research, taking the impermanent nature of taxa distributions and undertaking that such species are at a state of stability. If the artificially identified species (rarely distributed) are based on morphological identification, they must be conserved.

**Keywords:** species boundaries; species concept; taxonomic relationship; conservation; future climate change; ecology

### **1. Introduction**

The concept of species is crucial in evolutionary research and all biological thoughts. Species are the fundamental unit of life and biological diversity with a specific karyotype, morphology, DNA sequence, ecological niche, or behaviour [1]. Taxonomists made several efforts to describe species, beginning from simple morphology to genetics [2]. Earlier

**Citation:** Ullah, F.; Gao, Y.; Sari, ˙I.; Jiao, R.-F.; Saqib, S.; Gao, X.-F. Macro-Morphological and Ecological Variation in *Rosa sericea* Complex. *Agronomy* **2022**, *12*, 1078. https:// doi.org/10.3390/agronomy12051078

Academic Editor: Marco Scortichini

Received: 25 January 2022 Accepted: 27 April 2022 Published: 29 April 2022

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**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

taxonomists such as Linnaeus defined species based on what they saw. Later on, this was formalized as a morphological or typological species concept. Reproductive isolation is considered the key standard for delimiting the species [3–5]. Yet, the limitation between closely related taxa becomes unclear with hybridization or continued gene flow [6]. Delimitation of species is important to distinguish between the species of the same nature, and diagnostic characters are helpful to separate all sorts of species boundaries in biological sciences. The idea of delimitation is known as the species concept [7,8]. Though, the species concept itself is under disputation. To understand the taxonomy of biodiversity, especially conservation biology, the delimitation of species is vital for the controversial taxa. Furthermore, species are important, especially in conserving the existing biodiversity in the face of the extinction disaster of the Anthropocene [9–11].

The complex group of plants exhibits diverse forms. Taxonomic boundaries between putative taxa are often concealed by a lack of identified fixed morphological differences, potential hybridization, and a lack of informative collections. The taxonomically complex taxa are sometimes difficult to differentiate and delimit due to their complex characters and distribution. The artificial taxonomy or the species delimit through confused characters sometimes leads the taxa in danger for biological conservation [12,13]. Misidentification could lead to underestimating or overestimating species richness, and these difficulties could entirely compromise the investigation. The poor taxonomy could risk understanding ecological patterns since they are based on richness and measurement of species turnover between sites [14,15]. Conservation actions are taken without accurate taxonomic identifications may impair the effective conservation of the target taxa. The goal of taxonomic studies should not be restricted to distinguishing and describing living things, and it seems impossible to talk about conservation without taxonomy [15]. For species delimitation, taxonomic variations are important [15].

Though systematic is usually regarded as the science of diversity, it played a limited role in developing current approaches to conservation biology. In contrast, ecological values and population genetic concepts are more widely incorporated into conservation theory and management strategies [16,17]. Different studies examined species delimitation based on morphological and geographical distribution [18–20]. However, these studies were limited to one or two factors, whereas species limitation is based on various drivers such as interaction with climate, evolutionary traits, morphological characteristics, and biogeographical characteristics. In other words, species delimitation and conservation can be addressed by focusing on different levels, from genes to populations, ecosystems, and species concepts [21,22]. Scientists favour biodiversity as more integrative based on genetic and genomic approaches [23]. Systematics offers a foundation of information for conservation biology equally valuable as that of population genetics [19,20]. Systematic is vital due to the linkage of a taxon to other relative species based on taxonomic and molecular characterizations [24]. Species delimitation without an accurate taxonomic identification is incomplete. However, funding agencies have neglected taxonomy nowadays because of being descriptive discipline [15]. Difficulties and hurdles in systematics, ecology, and taxonomic determination among species hamper research in areas of species definition [25]. For species delimitation, identification is important for the complex taxa [15]. Funding agencies need to provide financial assistance for taxonomy and ecology to conduct extensive field works to explore rare and endangered species, especially in megadiverse countries like China. The technique red lists would be viewed and used for poorly known species, depleted species, population decline, restricted ranges, and rarity based on all these criteria. Procedures associated with priority situations and the progress of national red lists are essential to justify some expectations in formulating the criteria [26]. Taxonomy, ecological distribution threats, and population knowledge are important for the red-list assessments and adequate capacity to process and analyze data. Both capacity and data are absent for numerous species-rich taxa, despite their great ecological importance [27,28].

The taxa of the *R. sericea* complex occur at a high latitudinal gradient from 1000 to more than 4000 m above sea level [6,29]. Based on the previous study, we hypothesized that the species of *R. sericea* complex showed few distinct macro-morphological characteristics. Our observation demonstrates that the studied species have clear morphological characters to differentiate the taxa in the complex. So, in this research, we applied population morphological and ecological data (precipitation, temperature, and nineteen bio factors) to observe the variation in the focal taxa and their distribution. This paper aims to distinguish between the species of the *R. sericea* complex using morphological and ecological characteristics and conservation status to attain the following objectives: (1) To delineate species boundaries in *R. sericea* complex by adopting more comprehensive approaches based on populations sampling; (2) To exclude vulnerably, threatened taxa utilizing the above process; and (3) To determine how accurate and efficient morphological variation is possible based on proper species delimitation.

### **2. Materials and Methods**

### *2.1. Study Area, Populations Morphology, and Record of Samples*

A total of 665 individuals of 71 populations of six taxa of *R. sericea* complex (*R. sericea* Lindley (RS) 36 populations, 311 individuals; *R. omeiensis* Rolfe (RO) 24 populations, 285 individuals; *R. sikangensis* T. T. Yu and T. C. Ku (RSK) 8 populations, 29 individuals; *R. mairei* H. Léveillé (RM), *R. morrisonensis* Hayata (RMO) with 10, 21 individual, respectively; *R. zhongdianensis* T. C. Ku (RZ) 1 population, 9 individuals) were collected from China (Table S1) and studied at the CDBI herbarium. RMO is the only taxon with one population collected from Taiwan (Figure 1). This work examined fifty morphological characteristics for each individual: twenty-six leaf characters, twenty-one flower and fruit characters, and three other traits. The mean values of the quantitative morphological characters were used for principal component analysis (PCA). Our studied populations showed that the species of this complex individuals sometimes share and grow in the same ecological habitat, except the species RMO (from Taiwan) and RM, and RZ from southwest China. During the collection, we press the samples in the newspapers in the field and bring them to CDBI for a detailed study. The population records and codes information are given in Table 1. The detailed morphological features were studied in the herbarium of the Chengdu Institute of Biology (CDBI). We used a binocular dissecting microscope to distinguish the study taxa based on detailed characters studied for a concise and clear character. The macro-morphological characters of each specimen studied herein are described to determine the species macromorphologically. We also selected some taxonomically important characters to distinguish the complex taxa species. We constructed dichotomous keys and taxonomic descriptions to identify these taxa easily (Table 2). Some characters were noted differently within the same population, while some features were observed overlap between the different species populations. Various qualitative and quantitative characters have been examined in detail.

### *2.2. PCA Analysis of Morphological and Environmental Factor*

The PCA analysis was carried out for the morphological and environmental factors to understand the relationship between the studied taxa. The morphological characters were analyzed using qualitative, quantitative, and ecological characters to see the complex taxa's structure, relationship, and species boundaries. We did PCA analysis for macro morphological characteristics, qualitative and quantitatively, and the environmental factors (Figure S1).

### *2.3. Correlation between Quantitative Characters and Environmental Factors*

The quantitative data and the 19 bio parameters (https://www.worldclim.org/, (accessed on 15 June 2021) were subjected to analysis of variance in R studio to evaluate the difference between quantitative characters and environmental factors relationship. We used the Pearson linear correlation bivariate between the quantitative and ecological characters of different morphological characters.

**Figure 1.** The distribution of different taxa of the *Rosa sericea* complex based on different herbaria samples. The map is based on different Herbaria records i.e., CDBI, PE, QTPMB, NAS, KUN, LZH, and KATH. **Figure 1.** The distribution of different taxa of the *Rosa sericea* complex based on different herbaria samples. The map is based on different Herbaria records i.e., CDBI, PE, QTPMB, NAS, KUN, LZH, and KATH.

*RS* Tibet, Yadong 27.55 89.00 3455.09 GXF-17513 2020 *RS* Tibet, Yadong 27.51 88.95 3232.9 GXF-17514 2020 *RS* Tibet, Jilong 28.78 85.30 3955.38 GXF-17538 2020

**Table 1.** *Cont.*

### *2.4. Distribution and Ecological Characters*

We pursued to relate and contrast the geographic ranges and environmental tolerance of putative taxa within the studied taxa. Our studied samples were only belonging to the geographical ranges of China. We obtained the environmental data while using our field collection data of given populations. The distribution data of 71 populations and the spatially unique data at 2.5 arc minutes (for future distribution) resolution (5 ∗ 5 km2 at the equator) of 63 recorded points. We used the ENMSDM package and China elevation data (30 arcs second for current), taken from the SRTM elevation data of 63 records. We took and used nineteen temperature, seasonality, and perception variables from WorldClim [30]. **Table 2.** Dichotomous taxonomic key of the Taxa of *Rosa sericea* complex based on macromorphological features.

### *2.5. Modelling the Species Complex Current and Future Distribution* 2.5.1. Records of the Species Complex and Bioclimatic Variables

We collected 63 presence points (records) of the species complex during fieldworks from 2010 to 2020 (Table 1). After thinning records of the species using the enmSdm package [31] and China's elevation data (30 arc seconds) derived from Worldclim (https://www.worldclim.org, accessed on 15 June 2021), we obtained 62 records of the species complex. Nineteen bioclimatic variables are crucial in defining species' climatic niches downloaded from Worldclim dataset v2.1 (30 arc-seconds) [32] for the current period. Because the high correlation between variables is expected to affect model performance and increase uncertainty in model results. Spearman rank correlation (*r*s) was calculated using R packages (scales and legendary) in R v4.04 to evaluate multicollinearity among the bioclimatic variables. In the analysis output, positive correlations (*rs* ≥ 0.7) between variable pairs were drawn in black and negative correlations (*r*s ≤ −0.7) were drawn in red.

### 2.5.2. MaxEnt Modelling of the Species Complex Distribution

We modelled the current and future distribution of the species complex using the MaxEnt method [33] implemented through Maxent v3.4.4 in the SDMtune package. SDMtune uses a particular object to compile the data for the analysis. This object, called SWD, bundles all the information related to each record, thereby reducing the risk of mistakes in further investigations [34]. After creating the SWD object, we trained a model using all 19 climatic variables. Then we considered the variable importance of the Jackknife test of the first model and correlations among bioclimatic variables to model species complex distribution with fewer variables. Because a study [35] revealed that seed dormancy breaking requirements and timing of seedling emergence of the species have a chilling need, Mean Temperature of Coldest Quarter (Bio11) was chosen as the predictor variable among highly correlated variables. In the end, paying regard to all of these predictions obtained, we decided to model the distribution of the species complex with Temperature Seasonality (Bio04), Bio11, and Precipitation of Coldest Quarter (Bio19) (Figure S2).

The results of SDMs under future climatic conditions are affected by a range of factors, including the choice of the statistical model, climate model range, and emission scenarios [36,37]. Although the Intergovernmental Panel on Climate Change (IPCC) considers all Global Climate Models (GCMs) equal, certain GCMs better represent some climate

types and regions. For example, among the GCMs, BCCCSM (Beijing Climate Centre, Climate System Model) has higher reliability and has been better studied in China. The GCM used in the study was the BCC-CSM2-MR GCM [34] (2.5 arc minutes) obtained from (https://www.worldclim.org/data/cmip6/cmip6\_clim2.5m.html, accessed on 15 June 2021). While the scenarios used were the Shared Socioeconomic Pathways (SSPs; ssp2 4.5 and ssp5 8.5) (CMIP6), which are now being used as important inputs for the latest climate models, feeding into the Intergovernmental Panel on Climate Change (IPCC) sixth assessment report is due to be published in 2021 (https://tntcat.iiasa.ac.at/SspDb/dsd?Action=htmlpage& page=welcome, accessed on 22 June 2021; https://www.ipcc.ch/assessment-report/ar6, accessed on 22 June 2021). The species complex's future distribution was predicted for 20-year periods (2021–2040, 2041–2060, 2061–2080, and 2081–2100).

### 2.5.3. Tuning Model Hyperparameters and Evaluation Model Performance

Tuning model hyperparameters is a long process, as it requires testing many combinations to identify the best-performing model [34]. To get the model with high predictive power, we tuned model parameters using 'gridSearch' function implemented in SDMtune. Since model evaluation measures the capacity of a given model to reflect "truth" and whether it can be applied under other conditions [38] to assess the performance of the models. We used receiver operating characteristic (ROC) curve analysis and the true skill statistic (TSS). Thresholds for interpreting the TSS values were defined as follows: Value ≥ 0.9 = best, 0.9 > Value ≥ 0.8 = very good, 0.8 > Value ≥ 0.7 = good and Value < 0.7 = weak [39].

In creating distribution maps of the species, five threshold values were used. Accordingly, '0–0.25' denotes unsuitable areas, '0.25–0.5' of very low suitability, '0.75–1' highly suitable. We also calculated the size of moderate and highly suitable (we call them suitable here) predicted by models to quickly assess future distribution areas of the species complex and to see if the species complex would have gain or loss in the size of suitable habitat areas in the future. To do that, first, we reclassified highly suitable sites as '1' and other areas as '0' using ArcMap v10.8. Second, we converted raster files into shape format. Lastly, we calculated the size of the regions from shape data. Distribution maps of the species complex and cartography works were created in QGIS v3.18.

To grasp more information about the Rsc climatic niche and distribution in China, we used the "rmaxent" and "ENMTools" that have some excellent features implemented in functions about the species niches (URL: https://github.com/johnbaums/rmaxent, accessed on 9 June 2021) [40]. In the R maxent, we modelled the species' climatic niche using all bioclimatic variables to reveal which variable/variables affect the species' complex distribution in China. In the ENMTools, we measured the spatial heterogeneity of the distribution of suitability scores from the model results we obtained. This feature returns Levins' two metrics of niche breadth (B1 and B2) [40].

### **3. Results**

Different morphological characters have been studied in this article for the six taxa of the *R. sericea* complex (RO, RS, RMO, RM, RSK, and RZ) (Figure 2). In addition, we focused on the conservation of rare species of the complex.

ins' two metrics of niche breadth (B1 and B2) [40].

focused on the conservation of rare species of the complex.

**3. Results**

**Figure 2.** PCA analysis of quantitative morphological characteristics of *R. sericea* complex (**A**), PCA analysis of qualitative morphological features of *R. sericea* complex (**B**), PCA analysis of environmental characteristics population-based of *R. sericea* complex (**C**), PCA analysis of morphological characters and environmental factors of *R. sericea* complex (**D**). **Figure 2.** PCA analysis of quantitative morphological characteristics of *R. sericea* complex (**A**), PCA analysis of qualitative morphological features of *R. sericea* complex (**B**), PCA analysis of environmental characteristics population-based of *R. sericea* complex (**C**), PCA analysis of morphological characters and environmental factors of *R. sericea* complex (**D**).

To grasp more information about the Rsc climatic niche and distribution in China, we used the "rmaxent" and "ENMTools" that have some excellent features implemented in functions about the species niches (URL: https://github.com/johnbaums/rmaxent, accessed on 9 June 2021) [40]. In the R maxent, we modelled the species' climatic niche using all bioclimatic variables to reveal which variable/variables affect the species' complex distribution in China. In the ENMTools, we measured the spatial heterogeneity of the distribution of suitability scores from the model results we obtained. This feature returns Lev-

Different morphological characters have been studied in this article for the six taxa of the *R. sericea* complex (RO, RS, RMO, RM, RSK, and RZ) (Figure 2). In addition, we

#### *3.1. Morphological Variation 3.1. Morphological Variation*

Variation has been observed in different morphological characteristics of the studied taxa. The characters investigated in the focal taxa samples were somehow morphologically significant for delimiting the taxa into distinct taxon. We identified different morphological characters to differentiate the study taxa of the complex. A detailed morphological description of the study taxa based on qualitative and quantitative features was revised. In the present work, we studied 50 qualitative and quantitative characters of each individual, which showed some variation in the studied species. Moreover, these characters were analyzed with the environmental characters, and the correlation is given in (Figure 3). The vegetative morphology was mainly focused on the leaf morphological Variation has been observed in different morphological characteristics of the studied taxa. The characters investigated in the focal taxa samples were somehow morphologically significant for delimiting the taxa into distinct taxon. We identified different morphological characters to differentiate the study taxa of the complex. A detailed morphological description of the study taxa based on qualitative and quantitative features was revised. In the present work, we studied 50 qualitative and quantitative characters of each individual, which showed some variation in the studied species. Moreover, these characters were analyzed with the environmental characters, and the correlation is given in (Figure 3). The vegetative morphology was mainly focused on the leaf morphological characters. One of the most important characteristics was the leaflet's morphology and leaflets in different individuals (species). The species boundaries in the complex taxa were evaluated based on morphological characters (Appendix A).

Each individual's qualitative and quantitative characteristics were examined in the CDBI herbarium and statistically analyzed. The description of these species was mainly based on the morphological characteristics of collected populations specimens from different geographical distribution ranges of the *R. sericea* complex. Population information is given in (Table 1), and morphological characters descriptions are given in (Appendix A). The studied species of this complex have tetramerous perianth. The height of these shrubby plants' species ranged from 1 to 4 m.

evaluated based on morphological characters (Appendix A).

**Figure 3.** Correlation between the 19 bio and quantitative characteristics (1 = Length of leaf minimum, 2 = Length of leaf maximum, 3 = length of the first leaflet on apex minimum, 4 = length of the first leaflet on apex maximum, 5 = width of the first leaflet on apex minimum, 6 = width of the first leaflet on apex maximum, 7 = length of the first leaflet on-base minimum, 8 = length of the first leaflet on-base maximum 9 = width of the first leaflet on-base minimum, 10 = width of the first leaflet onbase maximum, 11 = length of pedicel minimum, 12 = length of pedicel maximum, 13 = length of sepal minimum, 14 = length of sepal maximum, 15 = width of sepal minimum, 16 = width of sepal maximum, 17 = bio1, 18 = bio2, 19 = bio3, 20 = bio4, 21 = bio5, 22 = bio6, 23 = bio7, 24 = bio8, 25 = bio9, 26 = bio10, 27 = bio11, 28 = bio12, 29 = bio13, 30 = bio14, 31 = bio15, 32 = bio16, 33 = bio17, 34 = bio18, 35 = bio19) the bio 19 at 2.5 arc minute resolution and projected based on present-day climatic conditions the codes of bio 19 is according to https://www.worldclim.org/, accessed on 15 June 2021. The graphical representation showed a correlation between −1 and 1 from brown and red to bluish. Each individual's qualitative and quantitative characteristics were examined in the CDBI herbarium and statistically analyzed. The description of these species was mainly **Figure 3.** Correlation between the 19 bio and quantitative characteristics (1 = Length of leaf minimum, 2 = Length of leaf maximum, 3 = length of the first leaflet on apex minimum, 4 = length of the first leaflet on apex maximum, 5 = width of the first leaflet on apex minimum, 6 = width of the first leaflet on apex maximum, 7 = length of the first leaflet on-base minimum, 8 = length of the first leaflet on-base maximum 9 = width of the first leaflet on-base minimum, 10 = width of the first leaflet on-base maximum, 11 = length of pedicel minimum, 12 = length of pedicel maximum, 13 = length of sepal minimum, 14 = length of sepal maximum, 15 = width of sepal minimum, 16 = width of sepal maximum, 17 = bio1, 18 = bio2, 19 = bio3, 20 = bio4, 21 = bio5, 22 = bio6, 23 = bio7, 24 = bio8, 25 = bio9, 26 = bio10, 27 = bio11, 28 = bio12, 29 = bio13, 30 = bio14, 31 = bio15, 32 = bio16, 33 = bio17, 34 = bio18, 35 = bio19) the bio 19 at 2.5 arc minute resolution and projected based on present-day climatic conditions the codes of bio 19 is according to https://www.worldclim.org/, accessed on 15 June 2021. The graphical representation showed a correlation between −1 and 1 from brown and red to bluish.

characters. One of the most important characteristics was the leaflet's morphology and leaflets in different individuals (species). The species boundaries in the complex taxa were

### *3.2. Ecological Niche Modelling*

The complex taxa except for RS, and RO, while the taxa RSK, RM, RMO, and RZ have narrow ecological amplitude, and hence their distribution is rare. Based on our field observations and herbarium records, the population of the one taxon RZ (with nine individuals with one population) compiled as taxon of very narrow ecological distribution, followed by RM, RMO, and RSK. Some types specimens of this taxon were available in the herbarium. Phylogenetically these species are complex, and the evolutionary tree of these taxa quite complicated. This species are closely interrelated, and morphologically synapomorphies have been examined (Appendix A).

These taxa have phylogenetically complicated systematics. The species boundaries were difficult to differentiate [6,29], while the morphological characters showed that the species have distinct characteristics to define species boundaries. Due to the narrow ecological amplitude and rarity, these taxa should be conserved. The phylogenetic classification is essential if we are thinking of organizing the biodiversity in such a way as to establish conservation significance and improve informed conservation policies [41,42]. Additionally, revised classification is dreaded by conservation practitioners and other taxonomy users. The new classification of plants is based on molecular or phylogenetic analyses considered the basic basis of practical taxonomy [43,44].

From our fieldwork and analysis of morphological characters, and work previously done on the species of the complex [45], these taxa were clustered in the same clade [46]. These taxa's field observations and geographical distribution showed that RS and RO have a broad ecological amplitude. In contrast, RSK and RM have moderate distribution, while the taxonRZ have a narrow ecological amplitude and few populations in nature. The RMO has occurred in the isolated habitat in Taiwan. We have recorded one population with 21 individuals (Figure 1). Morphologically these species could be differentiated because they have shown morphological species concepts to distinguish them. We assessed the quantitative characteristics of these taxa showed variation in the studied taxa. Taxonomic distinctness based on morphological characters showed that and fulfilled the conservation criteria due to their narrow biogeographical distribution and rarity in nature. Populations extinction is one of the significant threats to plant diversity, foremost to range reductions, disintegration, and isolation, which reduces species abundance.

The current SDMs of the *R. sericea* complex predicted through the model provide very high success rates with training and test AUC values of 0.90 and 0.97, respectively. These findings indicate that the predictor variables used for SDMs were suitably selected, thus leading to very high prediction success. The AUC curves in developing *R. sericea* complex SDM under current environments are given (Figure 4). The TSS, AUC, and variable importance values for the complex conspecific under future climatic are given (Figure 4). The present finding obtained from the model, the sum of the three first potential distribution variables was 59.407%. The higher the percent contribution, the more important that a variable is for predicting the occurrence of the taxa. Bio 11 had the highest predictive contribution in the present study, 28.44%.

The current potential distribution of the taxa showed that the area with red indicates a highly suitable area for the complex species. In contrast, the blue indicates the site is not suitable for taxa (Figure 5). The BCC-CSM2-MR GCM and the SSPs (ssp2 4.5 and ssp5 8.5) (CMIP6) based models predicting future habitat suitability of the *R. sericea* complex taxa are given in (Figures 6 and 7). Concluding from these futures modelling, the results indicate that the species may have difficulties shortly. Some of the rare taxa of the complex will be extinct due to their narrow ecological amplitude.

**Figure 4.** The response of *R. sericea* complex to three climatic variables and ROC curve of the model. **Figure 4.** The response of *R. sericea* complex to three climatic variables and ROC curve of the model. cate that the species may have difficulties shortly. Some of the rare taxa of the complex will be extinct due to their narrow ecological amplitude.

*Agronomy* **2022**, *12*, x FOR PEER REVIEW 11 of 26

**Figure 5.** The current potential distribution model of *R. sericea* complex. **Figure 5.** The current potential distribution model of *R. sericea* complex.

**Figure 5.** The current potential distribution model of *R. sericea* complex.

**Figure 6.** Future suitability areas and the size of the highly suitable areas predicted by models BCC-CSM2-MR ssp245 (**left**) and BCC-CSM2-MR ssp585 (**right**) for 2100. **Figure 6.** Future suitability areas and the size of the highly suitable areas predicted by models BCC-CSM2-MR ssp245 (**left**) and BCC-CSM2-MR ssp585 (**right**) for 2100.

**Figure 7.** The size and placement of highly suitable areas predicted by the future distribution models BCC-CSM2-MR ssp245 (**left**) and BCC-CSM2-MR ssp585 (**right**) for 2100. **Figure 7.** The size and placement of highly suitable areas predicted by the future distribution models BCC-CSM2-MR ssp245 (**left**) and BCC-CSM2-MR ssp585 (**right**) for 2100.

### *3.3. Environmental Factors, Morphological Variation, and Correlation*

The ecological species concept of the complex taxa somehow showed variation, and some of the populations occurred parapatrically. Our present work's geographical distribution of RS populations showed a wide distribution range from 1380 to 4168 m in altitude. In comparison, RO has altitudinal variation from 1800 to 4250 m, while RSK has altitude distribution from 2797 to 3851 m. Some RS, RO, and RSK shared the same ecological habitats and grow parapatrically in the study area. We collected each population specimen of RM (2185 m Yunnan, Qujing) and RZ (3007 m Yunnan Diqing), and one population of RMO from an altitude of 3180 m, from Taiwan, Hualian (Table 1, Figure 1). RMO has a unique ecological boundary among the studied taxa and occurs in Taiwan. RZ could also be isolated easily from the rest of the taxa of the complex allopatrically.

The correlation between the quantitative data and 19 bio data is given in (Figure 7). The correlation between the environmental factors and quantitative morphological features shown with single star \* correlation is significant at the 0.05 level, while two-star \*\* correlation is significant at the 0.01 level (Figure 3). The positive and negative correlations between different parameters are shown in (Figure 3). In detail, some of the quantitative characters showed a negative correlation represented with the negative sign (-), while some of the variables have a positive correlation at 0.05 and 0.01 levels. Some variables don't show any significant correlation represented in the Figure without the star \* symbol.

### **4. Discussion**

### *4.1. Ecotypes Plasticity and Species Boundaries*

Different populations of the *R. sericea* complex were studied here. The individuals' morphological characters showed variation in characters and morphologically distinct taxa. Some populations shared the same environmental and ecological habitat of the complex species. Due to similar characteristics in different species populations and individuals with different characteristics, the complex species had a wide range of character's plasticity (Figure 2A–C). The phenotypic plasticity could activate together through ecotype formation, further increasing suitability. The modest level of phenotypic plasticity may facilitate a population's expansion into novel environments. The trait may place the populations on the slope of an adaptive peak from which natural selection can advance [47]. The RMO adapted to a novel environment and isolated to a new habitat in Taiwan. The limitations to the entirely isolated territory and fundamental asymmetry in migration due to differences in density-dependent viability both redirect selection on a phenotypically plastic trait. Together, these environmental factors might cause the appearance of feature values and genetic constitution that seem specialized to one of the extreme environments of the territory [48]. In the study by Gao et al. [6]), RS and RO were observed parapatrically. However, we observed that the RS, RO, and RSK occur parapatrically, sympatrically, and allopatrically (Figure 1).

Documentation of morphological, environmental, and distribution data provides a basis for the species status of divergent populations of some complex taxa. Conversely, the morphological and ecological characteristics showed that the taxa are distinct taxon based on distinctive features (Table 2). The morphological characters showed that the taxa of the complex are different species. In contrast, predictable morphological features, and environmental and distribution data accept and support different taxon statuses. In our study, the divergent parapatric would stand and be treated as species by default like RMO, RM, and RZ occur parapatrically. Taxon RS has extensive qualitative morphological variation identified in our populations, while the RO has a wide range of characters quantitatively. The PCA analysis showed combinedly that the characters of RO populations are dominated and almost share similar morphological characters with other taxa of the complex (Figure 2A,B,D). Our environmental data showed that the species RMO and RZ have unique environmental characteristics and are isolated from the other taxa of the complex. The factors include low temperature and high precipitation. The morphological characters retain the status of these taxa to distinguish taxon and prove that the taxa in

the complex are morphologically different [49–51]. Species of the complex may be a single hyper-diverse taxon or contrasted that the speciation or evolution has happened or is ongoing. If the speciation occurred or is ongoing, we would assume to detect apparent morphological and genetic variations, which may show more robust evidence if speciation is completed, or weaker if the speciation is ongoing or confounded by introgression [52,53]. Overall, however, the morphological characteristics support the status of the species as different taxon.

### *4.2. PCA Analysis of Environmental and Morphological Characteristics*

The PCA analysis of environmental and morphological features demonstrated some significant information about the complex species. We used PCA analysis of ecological features and quantitative and qualitative morphological characters to visualize the relationships and differences among the six species of the complex. For the ecological parameters, we used 2.5 arc minutes resolution (∼5 km2 at the equator), for which we obtained 19 temperature, perception, and seasonality variables from WorldClim [30,32].

The PCA analysis of only quantitative morphological characteristics showed more variation in the studied specimens of the same species from different populations (Figure 2A and Table 3). The qualitative morphological characters PCA analysis of the studied specimens of diverse populations shows overlap and difficulty in distinguishing the taxa from each other in the studied taxa (Figure 2B and Table 4). We studied the PCA analysis of nineteen bio factors and 12-months temperature and precipitation. Environmental aspects of the studied specimens and populations showed significant importance in differentiating some of the studied taxa (Figure 2C). We also determined the PCA analysis of 19 bio factors, 12-month temperature minimum, maximum, and precipitation combined with quantitative morphological characters (Figure 2D). The PCA analysis of environmental factors and quantitative features of various populations showed the association between the studied taxa (Figure 2D, Table 5).

**Table 3.** PCA analysis of quantitative morphological characteristics.

**Table 4.** PCA analysis of qualitative morphological characteristics.

**Table 5.** PCA analysis of environmental characteristics of the studied populations.

### *4.3. Morphological Characters Evaluation*

Morphological characters for species delimitation are usually directly dependent on the field observation and herbarium specimens. These closely interrelated taxa showed dissimilarity in their morphological observation. For the morphological study, we focus on two parts of a plant: (i) the vegetative part and (ii) the reproductive part of the specimens. The vegetative morphology is generally referred to as leaf morphology and is related to different size measures of a species [54]. Morphological features used for taxa delimitation and identification is dependent directly on the studied species specimens.

The present research finding showed that the morphological characters of the *R. sericea* complex sometimes have complicated taxonomic characters, which is challenging to differentiate the complex species. This estimation on the morphological characters and species collection localities, where some taxa co-occur over the geographical regions investigated (Table 1). The study species here are capable of hybridizing with each other. Some of the individuals are commonly possible to assign to species in mixed populations. The

species RS and RO in a previous study [6] differentiated through leaflet numbers. The RS showed fewer leaflets, while RO has more leaflets morphologically. Several ongoing or historical evolutionary processes clarify the morphological dissimilarity among entities within the studied taxa. One possibility is that the species of *R. sericea* complex represents a single hyperdiverse species phylogenetically. Due to this, these species display genetic panmixia or high levels of gene flow due to less or more constant contact between the populations [6,55,56]. The hyper-diverse species' morphological characters may be intergrading or demonstrating an approximate relationship with the regional environment [52,53].

### *4.4. Conservation Strategy*

According to Darwin, rarity is a significant precursor of extinction [57,58]. Precise assessments of rare taxon populations and the significant threats are vital to conservation planning and resource allocation for recovery action [59]. As phylogenetically, these species are quite difficult to differentiate, and maybe the species are under ongoing speciation. Our group team has been working on these species for more than a decade [45,46,51,60], and we have information about very few populations, such as the demographic status of RSK, RM, and RZ. The RZ taxon have few numbers of individuals as compared to RSK and RM. As the populations and individual numbers of the one taxon RZ are limited, there is a great chance of drift and inbreeding, altering the genetic structure [61]. Many studies of genetic variation in rare plant taxa have demonstrated that the proportion of polymorphic loci and the number of alleles per locus significantly reduced in small-sized populations [62].

The results of our ENMs indicated that the complex species would have difficulty shortly if climate change drives the taxa into extinction. According to [63], climate change is quantitatively considered in Red list assessments for only a few species. The authors of [64] highlighted that determining the application of SDMs to Red Lists is challenging due to model uncertainties and many biotic and abiotic factors that cannot be studied in such models. They suggested that SDMs and Red List assessments might play a complementary role in conservation actions, such as Red List provides evidence on both current and future risk of extinction for the species, while SDMs warn the magnitude of future extinction risk. Due to the shrinking habitat of the complex in the future climate scenarios, some of the taxa of the complex will lead to extinction due to their rarity and narrow ecological amplitude (Tables 6–8).

**Table 6.** PCA analysis of environmental and quantitative characteristics.

**Table 7.** TSS and AUC values obtained from the models implemented in this study.

**Table 8.** Variable importance of the first model carried out with all climatic variables.

The rare species of this complex should be conserved in natural habitats where the species grow. Direct population management will help conserve these rare species of the *R. sericea* complex in the appropriate habitat. Given current land uses (and other pressures of the Anthropocene), however, human interference may be required to maintain the habitat suitable for conserving these rare taxa. As a result, it is not only species that are conservation reliant, but entire ecosystems and the associated disturbance regimes and ecological succession pathways that define them need to be conserved. However, this requires sufficient data on species' distribution and abundance. The limitation between closely related taxa becomes unclear with hybridization between the same species within the genus and cross genus taxa. Understanding the taxonomy, biodiversity, and conservation biology, the delimitation of species are controversial for biologists, so several species concepts have been evaluated. Reproductive isolation is considered the key standard for delimiting the species [3,5,65].

### *4.5. Ecology of R. sericea Complex*

The *R. sericea* complex showed apparent macro-morphological variation like some other species of different families [66,67]. Our field investigation observed intergrading morphological characteristics; even the populations were geographically intermixed or proximal, except few entities. However, morphological characters only have been inadequately considerable to determine taxonomic problems within the *R. sericea* complex. These characters might characterize acclimatization instead of evolutionary variation. Given lim-

ited genetic adaptability among the recognized species and the infrequent individual with intermediate characters, these are particularly factual. So, only through integrating ecology will we imply current or ongoing speciation within the *R. sericea* complex and display assistance for the present taxonomic opinion that the *R. sericea* complex signifies unique, divergent, or diverging entities. The characteristics that differentiate the *R. sericea* complex can have ecological significance as variations alongside the environmental gradient over which these two taxa are diverging.

Morphologically *R. sericea* complex taxa have a smaller number of leaflets or have high leaflet numbers may characterize a variation in temperature, which is higher in the lower elevations of some taxa where the species arises and, therefore, improves the possibility for water loss. Such plants can mitigate water loss in high-temperature environments (niche) with smaller leaf surface areas [68], such as from the smaller number and leaflet size.

### **5. Conclusions**

Through our integrative approach using the field data, ecological factors, and population morphology, we elucidated that qualitative and quantitative characters provide taxonomic descriptions and keys to delimit the taxa. The macromorphological characters showed clear species boundaries, and the taxonomic keys and descriptions provide sufficient evidence as six separate entities. The previous work on the genetics method failed to define species boundaries of the complex. However, species concepts need cohesive evidence such as ecology, morphology, and so on to delineate evolutionarily unique entities since evolution is continuous and leaves different, detectable footprints within different groups of organisms. Additionally, the present knowledge of the conservation status of rare species and the IUCN and its robust network of voluntary experts around the globe is undoubtedly in the best position to guide such work. The present work is based on the collection of samples from more than a decade, suggesting that the species RZ have quite rare populations of individuals and should be conserved. Recognition of unique entities, such as *R. sericea* complex rare taxa, has vital implications for conservation decisions and robust biodiversity estimates. As the one taxon RZ has narrow ecological amplitude, our niche modelling results suggest that the habitat of these taxa is shrinking with the future climate changes. There is a chance of the extinction of the complex's rare taxa, which are morphologically different entities.

**Supplementary Materials:** The following supporting information can be downloaded at: https:// www.mdpi.com/article/10.3390/agronomy12051078/s1. Table S1. The morphological characteristics of different populations of *Rosa sericea* complex from different geographical ranges of Southwest China. Figure S1. The correlation lines among the variables used in the modelling study (Black and red lines indicate a high correlation between variables). Figure S2. The most limiting variables of the species complex distribution in China.

**Author Contributions:** F.U. and X.G.: Conceptualization, Data curation, Formal analysis, Methodology, Writing—original draft preparation, Writing—review and editing. Y.G. and R.J., Data curation, Formal analysis, Methodology. ˙I.S. and S.S., Methodology, Writing—review and editing. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was supported by the Second Tibetan Plateau Scientific Expedition and Research (STEP) program (2019QZKK0502) and the National Natural Sciences Foundation of China (Project No. 31670192), and National Natural Science Foundation of China 32171605. We are thankful to the Chinese Government Scholarship Program (CSC No. 2018GXZ017484).

**Data Availability Statement:** All data generated and analyzed during this study are included in this published article.

**Acknowledgments:** The authors are thankful to the curator and team of CDBI for their support in the examination of samples.

**Conflicts of Interest:** The authors declare no conflict of interest.

### **Appendix A**

Qualitative and quantitative morphological description of the *Rosa sericea* complex based on Table S1.