The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species

Monzalvo, René; Escorcia-Guerrero, Diana Lizbeth; García-Montes, Mario Adolfo; Rewicz, Agnieszka; Rewicz, Tomasz; Manríquez-Morán, Norma L.

doi:10.3390/d16020087

Open AccessEditor’s ChoiceArticle

The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species

by

René Monzalvo

^1,2,3,*,

Diana Lizbeth Escorcia-Guerrero

⁴,

Mario Adolfo García-Montes

²

,

Agnieszka Rewicz

⁵

,

Tomasz Rewicz

⁶

and

Norma L. Manríquez-Morán

^1,*

¹

Laboratorio de Sistemática Molecular, Universidad Autónoma del Estado de Hidalgo, Mineral de la Reforma 42184, Hidalgo, Mexico

²

Laboratorio de Genética, Universidad Autónoma del Estado de Hidalgo, Mineral de la Reforma 42184, Hidalgo, Mexico

³

School of Biological Sciences, Southern Illinois University, 1125 Lincoln Drive, Carbondale, IL 62901-6501, USA

⁴

Laboratorio de Ecología de Poblaciones, Universidad Autónoma del Estado de Hidalgo, Mineral de la Reforma 42184, Hidalgo, Mexico

⁵

Department of Geobotany and Plant Ecology, University of Lodz, Banacha 12/16, 90-237 Łódź, Poland

⁶

Department of Invertebrate Zoology and Hydrobiology, University of Lodz, Banacha 12/16, 90-237 Łódź, Poland

^*

Authors to whom correspondence should be addressed.

Diversity 2024, 16(2), 87; https://doi.org/10.3390/d16020087

Submission received: 30 December 2023 / Revised: 20 January 2024 / Accepted: 24 January 2024 / Published: 31 January 2024

(This article belongs to the Special Issue Recent Advances and Future Perspectives for Balsaminaceae Research)

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Abstract

:

Impatiens mexicana is a native balsam restricted to the cloud forests of central and southwest Mexico, which is currently known to exist in four states (Zacatecas, Veracruz, Puebla, and Oaxaca), and is probably threatened by geographic restriction. Morphological and distributional information about this species is still scarce, with only a couple of investigations since its first description in 1910, which means the phylogenetic position of this species has yet to be placed. In order to contribute to knowledge of I. mexicana, morphological and molecular studies were carried out with material collected in the localities of Hidalgo, Veracruz, and Oaxaca, during expeditions from July to October 2021 and in 2022. The specimens selected were measured, photographed, and compared with morphological information from the six American balsams. At the same time, phylogenetic studies were carried out by using two regions of the cpDNA and one of the nrDNA. We report I. mexicana for the first time in Hidalgo state, and identify new registers in the Veracruz and Oaxaca localities, thus increasing the knowledge in its geographic distribution. We also redescribe the Mexican balsam by using new and complementary traits: we note that while the species is morphologically similar to I. capensis, it is distinguished by differences in flower color, the distribution of spot patterns on the upper petal, lower sepal, and lateral petals, and geographic information such as endemic geographic distribution in Mexico’s Tropical Montane Cloud Forest (TMCF). We also carried out phylogenetic analyses by using ITS and ITS + atpb-rbcL, which showed the Mexican balsam was independent from its Asian-American congeners. Interestingly, our genetic distance analyses reveal differences of 0.01–0.16% between I. mexicana and the other North American congeners that use ITS, atpb-rbcL and trnL-F, both independently and as concatenated genes: this low divergence most likely occurred because of the recent diversification of the group. However, both future redescriptions and phylogenetic studies in American species are imperative, as this will enable better discrimination.

Keywords:

Impatiens mexicana; redescription; morphology; phylogenetics; geographical distribution; land-use change; Tropical Montane Cloud Forest; México

Graphical Abstract

1. Introduction

Impatiens is one of two genera that constitute the Balsaminaceae. After first discovering I. balsamina in India in 1753, Linnaeus proposed the genus [1]. From this moment, the number of species expanded to more than 1000 (e.g., [2,3,4,5,6,7]), as new species (e.g., [8,9,10,11,12,13,14]) were frequently discovered. Impatiens are distinguished by the variation of their morphological attributes [2,6,15], and this is mainly evidenced in the richness of their floral and seed sculpture morphology [16].

The origin of Impatiens was postulated to be 50 Mya [2], with a vast and abrupt diversification of the genus during the Pliocene-Pleistocene [17]. It is assumed that it originated in western China before gradually dispersing to India and Africa. Today, the rich Impatiens species are grouped in five hotspots (Africa, Madagascar, South India, Sri-Lanka, Sino-Himalayas, and Southeast Asia), see [18,19,20] that extend across Old World tropics and subtropics). However, while the genus includes native species of temperate zones in Europe [21] and temperate, tropical, and subtropical regions of North America and Central America [22,23,24,25], there are no Australia or South America representatives [17,19].

The expansion of Impatiens to the Americas has been hypothesized to have occurred in the Pleistocene, with the species traveling from Southeast China (across the Bering Strait or the Aleutian Islands) some 1.26 million years ago [26]. Six species have been recognized: five North American species (I. aurella [25], I. capensis [27], I. ecornuta [28,29], I. pallida [30], and I. mexicana [24]), and a Central American counterpart (I. turrialbana [22,23]). However, I. noli-tangere [21], a predominantly Eurasian species, has also been reported in North America, in the Pacific northwest, southern Canada, British Columbia, South Alaska, and northwest Washington state [31,32]. American Impatiens include the hybrid (I. × pacifica [31]), a product of recent hybridization between I. capensis and I. ecornuta, and several naturalized and exotic species have been documented (see [33]).

Phylogenetic studies of Impatiens have been carried out in different world regions; for example, [15,18,19,20] principally used the cpDNA and nrDNA regions, and others have combined DNA sequences and morphological attributes [15]. However, recent investigations have included phylogenetic proposals with complete plastomes (e.g., [26,34,35,36,37,38,39]). Different phylogenetic studies have been conducted, including by [19], who carried out the most comprehensive phylogenetic study of American species, which encompassed six (I. aurella, I. capensis, I. ecornuta, I. noli-tangere, I. turrialbana, and I. pallida) of seven species, showing their aggregation with seven species from central China. However, most of the sequences used in this study are not available.

Finally, recent studies [26] that use plastomes have supported the claim that two North American species (I. capensis and I. pallida) are the sister group of the boreal I. noli-tangere; the grouping of these species in an independent clade showed they are the sister group of several Impatiens spp. from East China. To date, the Mexican balsam and the hybrid I. × pacifica are the American taxa that have not been sampled. The distinctiveness of I. mexicana has also been questioned, both because of its remarkable similarity to other species, (such as I. capensis), and its geographical proximity to I. turrialbana, its southeastern congener.

Impatiens mexicana is an endemic species from central-south Mexico with a known restricted distribution that spans four states (Puebla, Veracruz, Oaxaca and Zacatecas—see Table A1). Rydberg described it for the first time in 1910, drawing on a specimen deposited in the New York Botanical Garden Herbarium that was previously collected by Friederich Muller in Aserradero Santa Cruz, Orizaba, Veracruz, Mexico, during his explorations of flora in Mexico (1853–1854) [40,41,42]. Subsequent morphological and distributional work on the species was carried out by Barringer in 1991 [43], which documented new localities from Cofre de Perote in Veracruz, including a series of new characteristics for the species. Other posterior and documental works have been addressed by [44,45,46], who consider I. mexicana as an endemic species restricted to the Tropical Mountain Cloud Forest (TMCF); some current records are also available in international and national online databases (e.g., iNaturalist [33], Enciclovida [47], GBIF. Global Biodiversity Information Facility [48], and eFloraMEX [49] (Mexico)—see Table A1).

In seeking to contribute to knowledge of the Mexican balsam, this study aims to: (1) elucidate the phylogenetic position of I. mexicana within the North American–Asiatic clade; and (2) contribute new morphological and geographical information about the species through a redescription that uses material collected from different sites of its distribution. In acknowledging that I. mexicana has an isolated distribution in the TMCF of Mexico, and a remarkable lack of connectivity with its other congeners, we hypothesize that it is a valid species.

2. Materials and Methods

2.1. Biological Material Sampling

Expeditions from July to October 2021 and from July to September 2022 were carried out in TMCF in 18 localities in three states (Hidalgo, Veracruz, and Oaxaca, Mexico—see Figure 1). Some complete specimens were collected and prepared for herbarium collection, and leaf material was dried in silica gel and stored at −80 °C for later DNA-based studies. The TMCF in Mexico, the northernmost extension of the New World’s tropical montane cloud forests, is home to around 2800 vascular plants (10% of the estimated total in Mexico) [50]. It is located between 800 and 2500 m above sea level (hereafter referred to as ‘masl’) [51], has a discontinuous distribution pattern, and is fragmented by alterations in land use across moist mountain slopes, ravines, and gorges. These ecosystems only cover 0.5–1% of the country [52], and exhibit a high species richness in a small region. It is considered threatened because of its fragility and small size, and communities with a few or scarce individuals, who are usually seen in a few isolated patches [53].

2.2. Morphological Study

Mexican Impatiens specimens from 18 populations were observed and measured in the field by using measuring tape and a digital caliper, and several morphological structures measures and photographs were carried out by using a 3.5×–180× Trinocular Stereo Zoom Microscope AMSCOPE^®, accompanied by a stage micrometer. The mean, range, and standard deviation were calculated for each quantitative trait. The distinct color variations of chasmogamous and cleistogamous flowers were assessed by using the Munsell^® plant tissue color charts, which are a valuable tool in plant taxonomy and ecology, as they provide a standardized and precise system for describing and communicating the colors of plant tissues across the three main dimensions of hue, intensity, and color brightness [54]. Complementary information about geographical conditions, surrounding vegetation, and the presence of visitors in I. mexicana populations was collected at the same time.

2.3. Phylogenetic and Genetic Distance Analysis

DNA from nine individuals (three individuals per state) was obtained from fresh leaves by using a modified CTAB method, with the aim of avoiding the possible presence of secondary metabolites and resuspension in the TE buffer [55]. A second purification was made by using 500 μL of sodium chloride (1% w/v) and 100 μL of SDS (20% w/v), which were stirred at room temperature for one hour to obtain good quality DNA, before precipitation was later carried out with cooled isopropanol. In order to confirm the amount and quality of DNA, the material was measured to 260 and 280 nm absorbance on a NanoGenious spectrophotometer (Shangai MAPADA^® Instruments Co., Ltd., Shangai, China, 2017). In addition, DNA extractions were visualized by using a 1% agarose gel.

The phylogenetic analyses were accomplished using three regions of the cpDNA and nrDNA. In order to amplify the genes, different primers used in previous studies were selected (Table 1), including those used to infer phylogenies in the Balsaminaceae family across the world, and establish representative (although scarce) sequences in the American clade. Polymerase chain reactions (PCRs) were carried out in 25 μL reaction mixture; 1.8 mL of buffer; 1.5 to 1.4 μL MgCl2, 1 μL of DNA; 1 μL of dNTPs; 0.2 μL of each primer; 0.15 μL of Taq polymerase; and 19.2 μL of distilled water. These elements were then subject to the following conditions: Initial denaturation at 94 °C for 5 min; 38 cycles of denaturation at 94 °C for 30 s; primers annealing at 45–60 °C for 30 s; and extension at 72 °C for 60 s; before a final extension at 72 °C for 7 min. Subsequently, PCR products were purified by using the polyethylene glycol (PEG) method. Bidirectional sequence reads were generated in the Laboratorio Nacional de Genómica para la Biodiversidad (LANGEBIO, IPN), where sequences were assembled and edited, in pregap 4 and gap 4, respectively, from Staden Package [56]. To determine the phylogenetic position of I. mexicana, 127 sequences of Impatiens were selected according to representatives of the subgenus Clavicarpa and Impatiens, in the last one including: the Impatiens (America and Asia) sect., the Fasciculatae and Scorpioidae (South-West China) sect., the Uniflorae (South-West China, Africa, and South-East Asia) sect., the Racemosae (Himalayas) sect., and the Semeiocardium (South-West China) sect. In drawing on the phylogenetic proposals of some authors [15,16,18], two available sequences that belonged to the external group (Hydrocera triflora) were selected. We made sure to use the majority of sequences per species that came from a similar individual, and only did not do this when lack of availability prevented it (Table A2). All information (specimen details, photographs, and sequences) is available in BOLD systems [57], under project DS-IMMEX Impatiens mexicana (Process IDs: IMMEX001-23, IMMEX002-23, IMMEX003-23, IMMEX004-23, IMMEX005-23, IMMEX006-23, IMMEX007-23, IMMEX008-23, IMMEX009-23); and in GenBank [58], under the following accession numbers: atpB-rbcL: OR255132, OR525133, OR525134, OR525135, OR525136, OR525137, OR525138, OR525139, ITS: OR525067, OR525068, OR525069, OR525070, OR525071, OR525072, OR525073, OR525074, OR525075, trnL-F; OR525076, OR525077, OR525078, OR525079, OR525080, OR525081, OR525082, OR525083.

Sequences of each gene were aligned using the MAFFT/Q-INS-i alignment online version 7 (https://mafft.cbrc.jp/alignment/software/ accessed on 23 September 2023) [59] and were subsequently trimmed in MEGA X [60]. The alignment was constituted by 858, 978, 1001, and 2836 bp when using the internal transcriptional spacer (ITS), atpb-rbcL, trnL-F, and concatenated regions, respectively. GTR+G (ITS), TVM+G (atpb-rbcL), and T92+G (trnL-F) were found to be the best evolutionary models for each gene after the jModelTest 2.1.10 [61] software and the Akaike information criteria (AIC) were applied. The models were then used to correct genetic distances and generate phylogenetic trees, the model GTR+G was employed to analyze the ITS + atpB-rbcL and ITS + atpB-rbcL + trnL-F matrices. Phylogenetic analyses were carried out using Bayesian inference and the Maximum likelihood approach, Bayesian analysis was performed in MrBayes 3.1.2 [62], using two independent analyses of 30 million generations and four Markov Monte Carlo Chains. Sampling was conducted every 1000 generations; of the topologies generated at the start of the analysis, 25% were eliminated. Nodes with a posterior probability (PP) > 0.95 were considered significantly supported. Maximum likelihood and genetic analyses were conducted in MEGA X [60], to evaluate the branch support of the ML phylogenetic proposals; the bootstrap method was used with 1000 pseudoreplicates.

Table 1. Primers used to amplify the genes of Impatiens mexicana.

Primer Name	Region-Primer Sequence	Source
ITS
ITS5	5′-GGAA GTA GAA GTC GTA ACA AGG-3′	[63]
ITS4	5′-TCC TCC GCT TAT TGA TAT GC-3′	[63]
trnL-F
B49317	5′-CGAAATCGGTAGACGCTACG-3′	[64]
A50272	5′-ATTTGAACTGGTGACACGAG-3′	[64]
atpB-rbcL
RBCL1	5′-GAATCCAACACTTGCTTTAGTCTCT-3′	[65]
S2R	5′ AGAAGTAGTAGGATTGATTCTCATA-3′	[65]

3. Results

3.1. Redescription

Measurements are given in millimeters, unless otherwise indicated. The range of measurements is presented at the beginning, followed by the mean and the standard deviation. The description is based on nine populations from the nine localities of Hidalgo (3), Oaxaca (3), and Veracruz (3). Annual plants, terrestrial and epiphyte (this was rare—one was found growing on a tree trunk—see (Figure 4J) of 24–235 cm (111.75 ± 41.22, n = 191); glabrous, light or medium green stem that was tinged and aqueous, and that had thickened nodes and, occasionally, purple pigmentation. Petiole glabrous measuring 1–69 (18.19 ± 11.72, n = 182). Alternate leaves, with reticulate green venation; ovate blades with margin serrate; dentate with obliquely triangular mucronate teeth that were dark green on the adaxial face, pale green on the abaxial face, 6.07–69 (32.69 ± 15.26, n = 269) long, 4–42 (17.98 ± 8.10, n = 260) wide, whose apex was most often acuminate (and rarely truncate), and which was base acute (Figure 2E–H). Inflorescence mainly occurs in flowers that grow in pairs (Figure 4E). Flowers may present variations of yellow color that extend from strong to pale yellow: Munsell’s color codes include 5Y 9/12 (Figure 2I), 7.5Y 9/12 (Figure 2J); and vivid to vivid-pale: Munsell’s color codes are 7.5Y 9/12 (Figure 2K), 7.5Y 9/10 (Figure 2L); and with intense to pale red spots: Munsell’s color codes are 5R 4/14 (Figure 2I), 5R 5/14 (Figure 2J) and 7.5R 5/16 (Figure 2K,L), covering the dorsal surface (approximately ¾ of the region) (Figure 3A–C) and lateral regions of the lower sepal (Figure 3D–K); meanwhile, in the spur the dots fade as they reach the distal zone, and, in part of the lobes, spots do not cover everything (internal face, ¾ of basal lobe and ½ or less of distal lobe). And both red spot-mark lines and individuals vary, presenting diffuse or condensed and intense patterns (Figure 2I–L). Stipe 2–47 (25.84 ± 12.24, n = 35); two lateral sepals ovate with dichotomous to longitudinal venation, and the main vein in the middle is greener than the secondary venation, while the sepals are pale green-white (greener in the middle zone, sometimes with gray spots), 2.5–7 (4.81 ± 1.08, n = 44) long, and 2.5–7 (4.06 ± 1.02, n = 44) wide. The upper petal is orbicular, yellow with occasionally some red spots in the mid-posterior region, and is 4.5–10 (6.28 ± 1.72, n = 16) long, and 3.5–11.5 (7.23 ± 2.18, n = 16) wide. The upper petal, which has a white or green hump, continues into a white and large horn that terminates with a concave tip, whose horn may be of different lengths and orientate in different directions. The lower sepal is elongated with a trumpet-funnel form that is spotted, and is 9–21.5 (16.93 ± 3.70, n = 15) long, and 6–15 (9.5 ± 2.23, n = 31) wide. The spur is 4–16 (11.43 ± 2.95, n = 23) long, with a ranging angle that is slightly recurved (110°) (Figure 3E,F), medium (180°) (Figure 3J), highly (225°) (Figure 3D) and extremely (270°) (Figure 3H), reflexed (45°) (Figure 3I), recurved with a hook end (Figure 3G) and perpendicular (90°) (Figure 3K); the spur may present, at its distal tip, a slightly or accentuated bifurcation (Figure 4B) and, occasionally, two bulges on the extremes (Figure 3H). The lower sepal + spur are 19–40 (29.54 ± 5.44, n = 31).

The lateral united petals orbicular, reniform or obcorded lobes mostly do not overlap each other (Figure 5B), and occasionally overlap themselves (Figure 5C). They are 10–24 (16.21 ± 3.64, n = 30) long, 4–13 (7.49 ± 2.01, n = 30) wide, and have two lobes: a distal lobe, doblariform 6–17 (11.62 ± 3.09, n = 29) long, and 5–15.5 (8.41 ± 2.75, n = 30) wide; and a basal lobe orbicular, oblong, and subequal 1.5–6 (3.39 ± 0.98, n = 35) long, and 1–4.5 (2.47 ± 0.80, n = 34) wide. Cleistogamous flowers that are bright to pale yellow are (5Y 9/12, 5Y 9/8, 5Y 9/10) 13–16 (14.33 ±, 1.52 n = 3) long, 7–8 (7.66 ± 0.57, n = 3) wide, and have a reflexed spur that is 9–10 (9.33 ± 0.57, n = 3) long, and are occasionally green in the distal region (5GY 9/12, 5GY 9/10), mottling patterns can be diffused or concentrated. The lower sepal + spur are (23.66 ± 1.15, n = 3) long.

The capsule fusiform (usually elongated fusiform and rarely short fusiform), which is green and glabrous, is 5.14–26.5 (14.35 ± 4.91, n = 60) long, and 1.13–6 (3.22 ± 1.18, n = 46) wide. Ellipsoid green seeds (approximately four per capsule or less) have a brown external tegument and are characterized by the possession of a series of warts. Their seeds possess ornamentation, and they are generally found in a polar region in a rectangular-square form; however, they may rarely present this ornamentation in both poles of the seed 2–5.5 (3.82 ± 0.76, n = 110) long, 0.75–4 (1.87 ± 0.62, n = 110) wide (Figure 6E).

The primary root is 110–150 (129.33 ± 20.03, n = 3) long, 6–10 (7.66 ± 2.08, n = 3) wide, and also has several secondary roots (see Figure 4F,G).

3.1.1. Distribution, Habitat, and Ecology

I. mexicana has been reported to inhabit primary and secondary forests (oak tree forests), and to be a product of cultivation in TMCF ecosystems at 1906–2657 masl (Table 2). The Hidalgo state records are reported for the first time, along with complementary new registers for Veracruz and Oaxaca. In Hidalgo state, we detected four populations inhabiting conserved forests close to water bodies (e.g., waterfalls, rivers, ponds), and several companion plant species were detected nearby: I. mexicana populations, highlighting Pinus greggii, Cuphea sp., Viburnum sp., Salvia sp., Clethra mexicana; some Miconia sp., Cosmos bipinnatus, Peperomia sp., Cyathea sp., Dicksonia sp., Solanaceae, Phaseolus sp., Lopezia racemosa, Rubus sp., Cuscuta sp., Arthrostemma sp. Phytolacca sp., Araceae, Salvia sp.; and a few Persea americana var. drymifolia-introduced trees. In the Veracruz sites, we reported thirteen populations growing in secondary forests: Pinus patula and P. hartwegii were found in forest clearings, and Baccharis conferta and Salvia tiliifolia, Rubus sp., Achyranthes aspera, Cunila sp., Salvia spp., Phytolacca sp., Lonicera japonica were common adjacent species. Some ferns, Blechnaceae, some Asteraceae, Poaceae, and Cucurbitaceae were also found. Finally, the only population reported in Oaxaca was found at an elevation of 2657 m, in conserved TMCF where Pinus, Quercus, Clethra, Saurauia, Magnolia, Liquidambar, Alnus, Ternstroemia, Arbutus, Chusquea, Oreopanax, Styrax, Cyathea, Miconia, Calyptranthes, Bejaria, Mollinedia, Baccharis, Crataegus, Cestrum, abundant Salvia sp, and some creepers grew. Endemism is also high in this ecosystem, and many rare, endangered plant species are exposed to habitat fragmentation, land use change, and overlogging.

Conservation status: The recorded I. mexicana specimens were found to have a maximum of under 70 individuals per population, growing near roadsides or inside primary and secondary forests. Here we particularly emphasized the Hidalgo and Veracruz populations, which were affected by decreasing numbers and the risk of gradually disappearing [66]; in particular, we emphasized the role of livestock (Figure 4D), the persistent presence of common cattle, and ‘chapeo’, the traditional practice of removing vegetation for different purposes (e.g., constructing open roads and agriculture). While fungal plagues were also observed in Veracruz populations (Figure 4I), further studies are necessary to determine the impact of the aftermath on the populations.

3.1.2. Phenology

The flowering and fruiting of Hidalgo specimens occurred from the beginning of June to the end of September, lasting approximately three months; while in the Veracruz localities, flowering and fruiting continued up until the middle of October. In the Oaxaca specimens, phenology occurs earlier, approximately extending from the end of April/the beginning of May to the end of July [67].

3.1.3. Floral Visitors

Different organisms were observed in the populations of I. mexicana, such as Apis mellifera, Bombus spp., Phasmatodea, Coccinellidae, Eugenes fulgens, and Cynanthus spp.

3.1.4. Studied Localities

A total of 18 localities in three Mexican states are reported as follows; Hidalgo, Agua Blanca de Iturbide. Population 1. Las Golondrinas Waterfalls, San Pedrito (20°22.501′ N, 98°20.201′ W). Population 2. Ranchería de San Pedrito (20°22.864′ N, 98°20.699′ W). Population 3. La Laguna (20°23.130′ N, 98°20.546′ W). Population 4. 300 m Velo de Novia Waterfalls (20°22.880′ N, 98°21.242′ W). Veracruz, Orizaba, Municipality of Nogales, Santa Cruz Muyuapan. Population 5. Las Lajas (18°52.237′ N, 97°13.222′ W). Population 11. Manzano (18°51.846′ N, 97°12.627′ W). Population 15. Casco de Hacienda (18°52.468′ N, 97°12.367′ W). Population 16. Carboncillo (18°51.877′ N, 97°12.674′ W). Veracruz, Orizaba, Municipality of Nogales, Chicahuaxtla. Population 6. Sótano de Tula (18°52.072′ N, 97°12.916′ W). Population 7. La Roca (18°52.100′ N, 97°12.774′ W). Population 8. La Cueva (18°52.050′ N, 97°12.672′ W). Population 9. Ladera y Carbón (18°51.987′ N, 97°12.595′ W). Population 10. Soldado fantasma (18°51.868′ N, 97°12.427′ W). Population 12. Las Tuzas (18°51.736′ N, 97°12.408′ W). Population 13. Las Tuzas II (18°51.816′ N, 97°12.370′ W). Population 14. Leoneros II (18°51.632′ N, 97°12.630′ W). Population 17. Paso del Toro (18°51.974′ N, 97°12.361′ W). Oaxaca, Santa María Yalina. Population 18. Santa Maria Yalina (17°17′23.04″ N, 96°21′4.74″ W) (Figure 1, Table A1).

3.1.5. Specimens Examined and Deposited

Due to restricted distribution, and possible endangered status, nine individual samples (three per state; for molecular-morphological studies) were selected and deposited in the HGOM, Herbario del Centro de Investigaciones Biológicas, Universidad Autónoma del Estado de Hidalgo (UAEH), with the following collection numbers: (OR0007UAEH-HGOM, OR0001UAEH-HGOM, OR0002UAEH-HGOM, OA0026UAEH-HGOM, AB0025UAEH-HGOM, OA0027UAEH-HGOM, AB0026UAEH-HGOM, AB0002UAEH-HGOM, OA0029UAEH-HGOM). In addition, specimens with the following collection numbers (OR0004UAEH-HGOM, OR0003UAEH-HGOM, OR0005UAEH-HGOM, OR0012UAEH-HGOM, OR0011UAEH-HGOM, OR0017UAEH-HGOM, OR0018UAEH-HGOM, OR0009UAEH-HGOM, OR0021UAEH-HGOM, OR0019UAEH-HGOM, OR0028UAEH-HGOM, AB0013UAEH-HGOM, AB0014UAEH-HGOM) were studied and also deposited. Other organisms mentioned in the results were also analyzed during fieldwork.

3.1.6. Remarks and Differential Diagnosis

Impatiens mexicana is one of seven Impatiens species that occur in North America (I. aurella, I. capensis, I. ecornuta, I. pallida, I. turrialbana, along with I. noli-tangere, which is predominantly Eurasiatic). The Mexican species first described by Rydberg in 1910 [24] was based on a specimen deposited in the New York Botanical Garden Herbarium that is currently located in Aserradero, Nogales Veracruz, very close to the Orizaba populations sampled in this study (Santa Cruz Muyuapan: 97°15′44.432″ 18°50′16.528″, see (Table A1). In the original description, several morphological features were considered (e.g., morphology of the flower, leaves, stem, and distributional data). Barringer subsequently (in 1991) [43] reported new records from Cofre de Perote, San Juan Xico, Veracruz, and included a short description with some additional characteristics and data, including descriptions of distal and basal lobes, the reported presence of “espolon” (without a horn in the upper petal), stem length, flowering seasonality and altitude data (Table 2).

Specimens reported in the present study complement several characteristics addressed by Rydberg’s and Barringer’s descriptions by adding new measures, morphological details in flower structures, and geographical and phenological information (see Table 2). Some data contrasts with the previous research, including the variation in total height (24–235), which diverges from the (40+) and (60) cm reported by Rydberg [24] and Barringer [43], respectively; in addition, the lateral united petals (at 10–22 × 4–10 mm) are longer and wider than those reported by Barringer, which measured 8–10 mm. Another essential feature that the first description highlights is the yellow color of the flower, which, according to Munsell color codes, varies in both the Veracruz and Hidalgo localities, ranges between vivid to vivid-pale and strong to pale yellow (7.5Y 9/12, 7.5Y 9/10, 5Y 9/12, 7.5Y 9/12). New characteristics included in the present redescription correspond to the description of seed morphology, measures, and coloration in cleistogamous flowers, spur variations and angles, as well as the disposition of spots-mark lines in the different structures that compound the chasmogamous flowers (Table 2).

I. mexicana was morphologically compared to the six other Impatiens taxa that occur in North America by drawing on information from original descriptions and available literature, including morphological traits and ecological information, as well as verified photographs of the referred species published online, which can provide helpful information that is not considered in the literature: I. mexicana [24,43]; I. aurella [3,24,25]; I. capensis [3,24,68,69,70,71,72,73,74,75,76,77,78,79]; I. ecornuta [3,24,28,29]; I. pallida [24,30]; I. turrialbana [22,23,24]; I. noli-tangere [1,21] and websites [33,80]. I. mexicana is separated from these six species by four general variations of yellow color, and the presence, shape, and disposition of spot patterns in the upper petal, lateral sepals, and lower sepal. In contrast, the North American species I. aurella exhibits a pale-yellow color, while I. ecornuta presents a yellow or orange; both species lack spots or mottling, and I. pallida’s light sulfur-yellow flowers are usually unspotted or contain minute reddish or brownish dots [24]. Some photographic evidence demonstrated individual variations in this species, including a white or cream color in lateral petals and part of the lower sepal. In the other Impatiens, spots occur inside the lower sepal and cover parts of the basal and distal lobes (see I. pallida in [80]); for example, I. turrialbana presents a flower with scarlet color petals [23], which may have yellow tones inside and outside the upper petal and (rarely) dark spots in the middle of the lower sepal and on the spur [81]. I. noli-tangere exhibits yellow flowers with small brownish or red spots [21]. And I. capensis, a species with more remarkable morphological similarities, is different because its flowers are orange-brown, red, or purple-spotted, in addition to being mottled [3,24] and sometimes wholly unspotted [24]. I. capensis forma citrina (I. biflora citrina) exhibits a perianth lemon-yellow with crimson spots, while the flowers of forma albiflora (I. capensis albiflora) are white or cream in color, with spots often paler than in the typical form, which is pink or brownish red in color, with white petals and a cream color spur [68]. However, wild and cultivated populations of the orange jewelweed show evidence of several color variations; the same applies to the shape and disposition of spot patterns reported in available web pages but not in formal descriptions (e.g., [82]).

Identifying phenotypes consist of diverse polymorphisms such as: 1.—Yellow perianth with an intense orange mottling that covers a significant part of the lateral petals; 2.—Yellow perianth with a uniform light orange mottling that completely covers lateral petals and the inner upper petal; 3.—Yellow perianth, with orange spots in the inner upper petal and spots that mostly do not intrude on half of the lateral petals surface; and 4.—Yellow perianth with dark orange (uniform or dispersed) spots that cover the lateral petal and inner upper petal, as well as new phenotypes established in small populations with new mottling patch patterns [83]. However, despite the reported variations, formal investigations that seek to expand and describe the color of perianth and the disposition of mark-spots in the morphotypes are essential, as this will help to achieve better discrimination.

Geographical distribution is another attribute that allow the six American species to be separated from I. mexicana, which is restricted to TMCF from central (Puebla, Hidalgo, and Veracruz states), and southwest (Oaxaca) Mexico; in contrast, the four North American species are distributed in the USA, Canada, and Europe, across a great variety of ecosystems, and in wet or humid places (Table 3); I. aurella and I. ecornuta are found in Canada, the Pacific region, British Columbia, and the North-West USA; I. pallida is found in central Canada, the Atlantic Region, and Mid-West, South-West and South-central USA. I. noli-tangere is widely spread across Canada (northern territories, North-West Territories, the Pacific region, and prairie provinces, western USA, and extensive parts of Europe and Asia. I. capensis is the North American species with the widest native distribution in America, including most of the Canadian provinces (northern territories, prairie provinces, central Canada, and the Atlantic region) and the USA (West, North-East, South-central). It was introduced in the USA in Washington and Oregon, before spreading out to British Columbia in Canada and Coahuila in Mexico [84,85].

Finally, I. turrialbana, the unique Central American native balsam, is restricted to the tropical forests of Costa Rica (in the provinces of Alajuela, Cartago, San José and Panamá (in the province of Chiriqui) [23,33].

The Mexican balsam is different from I. capensis (setting aside the possible exception of I. aurella, I. ecornuta and I. pallida whose masl is still unclear because of a lack of information) because it is distributed above 1900 masl (1925–2657); in contrast, the altitude distribution of orange balsam (I. capensis) oscillates between 39–1344 masl (see [3,24,25,68,69,70,71,74,76,77,78,79,86]).

3.2. Phylogenetic and Distance Genetic Analyses

The phylogenetic analyses that use the ITS spacer (Figure 7A) and implement the maximum likelihood (ML) and Bayesian inference (BI) methods reveal that the I. mexicana complex from central (Hidalgo), eastern (Veracruz), and southeastern (Oaxaca) Mexico forms a monophyletic group, which is supported by posterior probability (PP = 95) and bootstrap values (BS = 92). I. mexicana is from an independent lineage and is grouped within the Impatiens sect. Impatiens, along with the species occurring in North America (I. capensis, I. pallida, and I. noli-tangere, which is predominantly Eurasiatic) are, along with the Asian species I. yilligniana, sister groups (PP = 1, BS = 100), which together form the “Asiatic-American clade”. Phylogenetic reconstructions using the ITS spacers and atpb-rbcL show a congruent topology when placing and showing the independence of the I. mexicana complex (PP = 0.92) and (BS = 92) and between congeners (I. capensis, I. pallida, I. noli-tangere, and I. yilligniana) (PP = 1, BS = 100) (Figure 7B). The concatenated analyses employing ITS, atpb-rbcL, and trnL-F showed high support (PP = 1) for I. mexicana and its sister group (I. capensis, I. pallida, I. noli-tangere, and I. yilligniana) (Figure 7C).

Genetic distances between the species recovered in the phylogenetic reconstruction with I. mexicana ranged from 0.01–21.65% with ITS, 0.01–6.51% with atpb-rbcL, 0.03–10.03% with trnL-F, and 0.06–14.53% with the concatenated matrix. Divergence values for the I. mexicana complex and its sister congeneric clade varied between 0.01–0.07%, 0.01–0.16%, 0.03–0.07%, and 0–0.07% with ITS, atpb-rbcL, trnL-F, and concatenated regions respectively; genetic distances between I. mexicana individuals from Hidalgo, Veracruz, and Oaxaca were 0–0.03% (with ITS), 0–0.06% (with atpb-rbcL), 0–0.03% (with trnL-F), and 0–0.02% (with concatenated regions).

4. Discussion

4.1. A Morphological Discrimination of I. mexicana with American Balsam Congeners

The redescription of I. mexicana was carried out after considering detailed characteristics proposed in previous studies [24,43], and the incorporation of new characteristics (see Table 2), taking into account the distribution of different populations from the northern (Veracruz, Hidalgo) to the southern distribution of the species (Oaxaca). It was possible to distinguish the Mexican species from the other American species by using morphological and geographic characteristics (see Table 3). One of the species with the greatest morphological overlap that has given rise to confusion is the orange balsam, I. capensis, which has features that resemble those of I. mexicana. However, in referring to the tonalities of the flowers, the speckled pattern, the geographical distribution and the altitude, we were able to identify four characteristics that made it possible to separate them.

Within the studied populations, different forms of mottling were found, as well as variations in the shape of the distal tip and direction of the spur (e.g., individuals from Veracruz and Hidalgo), which helped to increase the knowledge of phenotypic scarcity in known species and improve discrimination. However, some recommendations for future research that seek to improve the separation of species have been put forward in morphological studies of color and mottling variations, which present in the flowers of the species’ southern populations, and which have been hitherto encountered in the state of Oaxaca, and potential distribution areas.

The compilation of new features in North American balsams, including pictures and drawings in redescription study papers are necessary, especially for the older descriptions (e.g., I. aurella, I. capensis, I. pallida, and I. turrialbana) or highly varied morphotypes; particular attention should be paid to coloration and mottling patterns (e.g., I. capensis), and to both native territories and to distribution in populations established in America and Europe (e.g., [84,85]). These detailed studies would make it possible to anticipate a better separation through the application of different multivariate discrimination methods, such as those successfully to the North American species I. capensis from I. pallida [88]. Likewise, it is notable that, in various species of Impatiens, micromorphological attributes have been of particular use in the species´ separation, especially those that lie in pollen and seeds (e.g., [4,15,89,90,91,92,93]). Future exploratory studies of the Mexican balsam and American species should acknowledge this, and take into account the fact that knowing the variations will improve the separation process.

4.2. Impatiens mexicana: Current Known Status

Impatiens mexicana has been reported in conserved TMCF, but mostly in secondary vegetation, which has been impacted by human activities such as deforestation and livestock and land use changes rooted in the use of different practices (Figure 4D). In the reported populations there are differences in the number of individuals, and also conditions that may favor their abundance or eventual disappearance. In the largest populations that have hitherto been found in the state of Veracruz, the population numbered much more than 70 individuals per population, which was favored by territorial extension and population connectivity; however, some of the biggest problems found on the sites were the substitution of native flora, which was achieved by planting important timber species such as Pinus patula, traditional charcoal production practices (Figure A1); and the presence of a fungus plague that promotes the senescence of individuals (Figure 4I).

Veracruz is considered to be one of the Mexican states with the most significant environmental deterioration, which has resulted from different practices introduced since “The Spanish colonization”, including extensive agriculture (cane cultivation practice and the cultivation of agricultural products from different regions of the world), livestock (local and extensive) (see [94]), and current practices that affect ecosystems, contributing to agricultural and pasture expansion, forest exploitation and species extraction [94,95,96].

In Hidalgo, the populations fluctuated between 40 or less individuals. Populations are mostly associated with water bodies, such as waterfalls, ponds, and streams, but can also be commonly found along roads that connect human communities. In these localities, the populations are subject to a potential risk of disappearance, mainly due to deforestation for commercial purposes and “chapeo“, the practice of cutting flora to open roads and building settlements for agricultural purposes, which eliminates potential individuals that have not yet been pollinated or self-pollinated. Similarly, in the case of discovered populations, the aforementioned factors have been reported to cause an obvious lack of connectivity. The TMCF of Agua Blanca de Iturbide, Hidalgo, has been identified by some authors as “patches of forest” located on slopes or near rivers, which are affected by maize and bean crops [97]; however, the TMCF in this region is now affected by agricultural practices such as the introduction of species of economic value (coffee, and the cultivation of fruits from tropical climates) but mainly by forest management of vulnerable ecosystems, with practices authorized by Mexican government institutions [81].

Finally, populations from Oaxaca, who live at the highest altitudes reported in Mexico, are located in conserved forests in locations that are challenging to reach, both because of their topography and established community practices in the region, which include regulating the entry of people not native to the communities and maintaining constant surveillance of the surrounding forests [98]. According to government reports, 80% of the population in the municipality of Santa Maria Yalina [99] is engaged in agriculture, primarily for self-consumption or “traspatio“. The remaining 20% is involved in various other activities, including farming, non-extensive livestock farming, and timber harvesting. In seeking to address this last activity, the Zapotec community has implemented alternatives that seek to contribute to the conservation of forests by working to achieve social organization with sustainable use; implement environmental education; and initiate forest restoration programs [98,99].

Considering the few known records and the distribution of the species in TMCF patches (threatened ecosystem), the population size, and the adjacent dangers for most of the populations (especially evident in Hidalgo and Veracruz), it is presumably a species with some degree of vulnerability, which deserves to be engaged by other fields of study, such as population genetics and demographic studies. These studies will provide insights that can be used to advise on the creation of conservation measures, which is particularly important because, up until now, the species has not been considered at risk in the national categories established in the NOM-059-SEMARNAT-2010, or included in TMCF flora in states where the species has been registered: Veracruz (e.g., [95]), Hidalgo (e.g., [97,100]), Puebla (e.g., [101]) and Oaxaca (e.g., [102]) have also been overlooked, perhaps because previous studies provided little information about them (e.g., [24,43]).

4.3. Molecular Studies: Phylogenetic Position and Genetic Distances of Impatiens mexicana

The genus Impatiens is known for its taxonomic difficulty, and molecular methods have been used to study the divergence between species, resulting in a more recent classification, in which this genus has been divided into two subgenera (Impatiens and Clavicarpa) [15]. ITS sequences from [18] provided insight into the phylogeny of the Impatiens species, showing that the existing species originated in Southeast Asia and spread to boreal Eurasia and North America, Central Asia, and Eastern Europe via the Himalayas, as well as to India and Africa. According to [17], who uses a phylogeny based on the atpB-rbcL spacer sequence, Impatiens may have originated in South China, and then, after colonizing neighboring areas, spread to North America, Africa, India, the South-East Asian peninsula, and the Himalayan region.

In this study, phylogenetic proposals that used the ITS ribosomal region showed the best support values in both probabilistic analyses, highlighting the independence of the Mexican complex at three different points of its distribution. This was also reiterated by its inclusion within the “Asian-American clade”. This clade belongs to the section Impatiens, which is characterized by two-flowered inflorescences, a linear or cylindrical capsule, ellipsoid seeds, and a seed coat with reticulate ornamentation [15], The “Asian-American clade” share a geographic continuity that extends from the coasts of the East China Sea (Zhejiang; I. yilligniana) (see [103]) to the New World (North American species in Canada, USA, and Mexico, namely I. capensis, I. pallida, I. noli-tangere and I. mexicana). It is characterized by the possession of orange (I. capensis), yellow (I. mexicana, I. noli-tangere, I. pallida, I. yilligniana, and I. aurella, I. ecornuta [the last two species are not included in the analysis]), red (I. turrialbana [not included in the analyses]), pink and white flowers (I. capensis, I. noli-tangere); the presence (I. mexicana, I. noli-tangere, I. pallida, and I. capensis), and absence (I. aurella, I. ecornuta, I. capensis and I. yilligniana) of spots; and elliptic, ovate-ovoid and lanceolate leaves, that may present (crenate or coarsely dentate) leaf margin morphology (Table 3). I. mexicana is placed as a sister group of the North American and Asiatic species but is not closely related to I. capensis (a sister group), as would perhaps be expected, given its similar morphology.

The phylogenetic position of I. mexicana suggests an independent speciation event distinguished it from its sister group, occurring after the migration of populations from East Asia through the Bering Strait or Aleutian Islands 1.26 MY ago, during the Pleistocene; and after the Eurasian migration (I. noli-tangere); and the formation and diversification of the Northamerican clade, which was established between 1.26 to 0.44 MY (see I. capensis and I. pallida [26]).

The restricted distribution of I. mexicana, which has hitherto been known to be in the TMCF patches of central and south Mexico, suggests the presence of probable extinct or existing relictual populations, which were distributed in areas of the north of Mexico during the Pleistocene-Holocene (e.g., TMCF distributed in Tamaulipas, Mexico), in similar conditions to the TMCF, as part of the process of diversification of North American Impatiens. The TMCF patches are considered to be relictual ecosystems, with expansion and contraction dynamics that are mediated by climatic oscillations [104], who hosted widely distributed temperate flora in the past, after the eventual increase in temperature that occurred in the Holocene [105]. TMCF’s geography, orography, and changes of area were probably crucial for speciation processes in the clade (I. mexicana), which was apparently a strictly distributed species with an affinity for TMCF microconditions (humidity, rain, and fog).

The ITS gene has been widely used as an individual region in phylogenetic proposals in balsams because of its relatively high divergence, compared to the more conserved regions (such as the atpB-rbcL and trnL-F cpDNA) seen, both in other balsams applied around the world and other plant groups [19,106,107,108,109,110]. While ITS has helped resolve relationships between closely related lineages, it lacks the power to resolve relationships between major lineages (see [18,19]). This study’s phylogenetic proposals, which use ITS + atpB-rbcL and ITS + atpB-rbcL+ trnL-F genes, support this assertion, showing a better resolution in polytomies present in large groups (compared to the proposal using ITS) and also decreasing the support of the Mexican complex, with both probabilistic approaches (PP:0.92, BS: 92; PP:0.67, BS:- respectively). On the other hand, separate analyses in atpb-rbcL and trnL-F cpDNA regions did not provide information about separating I. mexicana from its North American and Asiatic congeners [19] showed similar patterns when inferring the relationships between six American species I. aurella, I. capensis, I. ecalcarata, I turrialbana, I. pallida, and boreal distributed (I. noli-tangere). The low support values and lack of resolution of American species when atpb-rbcL and trnL-F are used separately, also suggests regions are not convenient when inferring phylogenetic positions in species with probable recent evolutionary histories. In confronting these situations, there is a need to, in future, include other types of elements, so as to gain a better overview of both types of relationships; one example is the use of complete chloroplast genomes, which several proposals in balsams have already addressed [26,36,37,38,39]. Likewise, the species delimitation of Impatiens could be improved by using current techniques, including the multispecies coalescent method (MSC), which offers a framework for estimating evolutionary parameters, such as migration rates or intensity of introgression, and provides a useful way to apply empirical criteria to delimit species or generate hypotheses about the current state of species, and integrate the findings with evidence obtained from other fields, such as morphology and ecology. Many contemporary species delimitation studies have been too quick to accept results based on particular genetic models and assumptions, even after other analyses have provided strongly conflicting information [110,111,112,113].

The genetic distances between the Mexican complex and its congeners of the Asian-American clade were, after the three regions were used, found to vary between 0.01–0.16%; the distances observed between species are low compared to those obtained in this study which, after comparing the distance between the Mexican balsam and the different geographically distant tribes of Impatiens, found values that ranged between 0.01–21.65% (e.g., ITS matrix). These essentially low patterns in nucleotide differences between American congeners probably occurred due to their recent diversification (see [17,26]), and America being the last point for large-scale settlement of the genus.

4.4. Limitations of the Study and Recommended Additional Research

In the present study: 1.-morphological discrimination according to other congeners was limited, mainly because of the scarcity of attributes and age of the descriptions, and the constant reporting of variations in known species that have not been formally studied (e.g., I. capensis, I. noli-tangere). This also occurs in the Mexican species, where the existence of potential phenotypes has been suggested (particularly phenotypes from the south (Oaxaca), along with TMCF areas in Mexico such as Tamaulipas and Chiapas, the northernmost and southernmost patches of these forests in Mexico, respectively).

To improve morphological discrimination, the following studies are recommended: 2.-Investigations of the morphological variations of potential TMCF populations in Mexico, which identify their presence by using tools such as potential distribution models. 3.-Morphometric studies that engage between Mexican species and American congeners. This will involve redescriptions (mainly of I. aurella, I. capensis, and I. turrialbana), and will also consider possible variations in their distribution areas. 4. Comparative micromorphological studies (pollen and seeds) should be undertaken with information from unknown species (I. aurella, I. ecalcarata, I. mexicana, I. × pacifica, and I. turrialbana) that support the morphological separation.

The information included in the phylogenetic analyses needed to be improved, mainly due to the insufficient number of publicly available or sequenced species (particular emphasis needs to be placed on the Asian-American clade) and insufficient markers in the species. Although, new phylogenetic proposals should prevail if they include all representatives of Impatiens species from America with the aim of supporting the status of Impatiens mexicana as a valid species, they should principally be favored if they are concerned with Asian-American species relationships. One way to better delimit species is by applying a model based on Bayesian methods that has been developed in recent years by using the multispecies coalescent (MSC) algorithm.

In order to know more about the current status of species conservation, it is necessary to carry out studies that focus on population dynamics and structure, as this will help to provide insight into essential demographic parameters and contribute to the emergence of studies that include indicators of population genetic diversity and structure. By drawing on this information and incorporating the IUCN assessments, we will be in a better position to help the species be placed in a risk category. Likewise, the Extinction Risk Assessment Method for Wild Species in Mexico (MER in Spanish), which is guided by Mexican conservation laws, could be an excellent tool that can be applied to evaluate the risk of extinction the taxon faces.

4.5. Final Comments

After examining phylogenetic studies, the use of morphological attributes and geographic distribution, and considering North American and Asiatic congeners, this study has concluded that I. mexicana is a valid species. However, further investigations, which will work to complete the phylogenetic proposals and evolutionary history of all American congeners, are necessary and should be undertaken in the future.

Author Contributions

Conceptualization, R.M., A.R., T.R., D.L.E.-G., M.A.G.-M. and N.L.M.-M. methodology, R.M., A.R., T.R., M.A.G.-M., D.L.E.-G. and N.L.M.-M.; labwork, R.M. and M.A.G.-M., formal analyses, R.M. and N.L.M.-M.; investigation, R.M., A.R., T.R., D.L.E.-G., M.A.G.-M. and N.L.M.-M.; resources, R.M. and D.L.E.-G.; data curation, R.M. and D.L.E.-G.; writing—original draft preparation, R.M, A.R., T.R., D.L.E.-G., M.A.G.-M. and N.L.M.-M.; writing—review and editing, R.M., A.R., T.R., D.L.E.-G., M.A.G.-M. and N.L.M.-M.; visualization, R.M., A.R., T.R., D.L.E.-G., M.A.G.-M. and N.L.M.-M.; supervision, R.M.; project administration, R.M. All authors have read and agreed to the published version of the manuscript.

Funding

This research received no external funding.

Institutional Review Board Statement

Not applicable.

Data Availability Statement

Data related to alignments used in the phylogenetic proposals and genetic distance analyses are available and can be downloaded at https://zenodo.org/records/10397628 (accessed on 20 December 2023). DOI 10.5281/zenodo.10397627.

Acknowledgments

We would like to thank the following people who made this research possible: Monserrat Monzalvo, for her support with the drawings; the biologist Jocelyn Briseño (from UAEH), for her help with the expeditions in the state of Hidalgo. The biologists Aura Ávila (UAEH), Loranca (from UNAM), and especially Fidel Rodolfo Ramírez Rosas for his kind aid during the samplings in the communities of the state of Veracruz; the biologist Kenia Velasco, Israel Santiago and Israel Santiago Jr., for providing their assets to us in the Zapotec communities of Yalina, Oaxaca; Tien Nguyen (from UMass) for her valuable support in providing information about I. capensis; Kurt Neubig and Kayla Grey from SIU for their helpful suggestions about the manuscript; and Wojciech Adamowski for his constant interest and support, and valuable comments about this manuscript and project. Finally, we would like to dedicate this article to the memory of Diana Elizabeth Hernández-Cerón, a young scientist and our closest friend.

Conflicts of Interest

The authors declare no conflicts of interest.

Appendix A

Table A1. Occurrences of Impatiens mexicana, based on the literature review and fieldwork.

State	Distribution. Locality-Town/Municipality/State	Coordinates/Longitude-latitude/% Altitude (masl) (If Available)	Collection-Institution Deposited/	Source
Hidalgo	* Las Golondrinas Waterfalls, San Pedrito, Agua Blanca de Iturbide Population 1.	98°20.201′, 20°22.501′ (−98.33, 20.37) %1925	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). AB0002UAEH-HGOM.	Present study
	* Rancheria, San Pedrito, Agua Blanca de Iturbide. Population 2.	98°20.699′, 20°22.864′ (−98.34, 20.38) %1906	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). AB0013UAEH-HGOM.	Present study
	* La Laguna, Agua Blanca de Iturbide. Population 3.	98°20.546′, 20°23.130′ (98.35, 20.38) %2070	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). AB0014UAEH-HGOM.	Present study
	* Velo de Novia, Agua Blanca de Iturbide. Population 4.	98°21.242′, 20°22.880′ (−98.34, 20.38) %1932	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). AB0025UAEH-HGOM. AB0026UAEH-HGOM.	Present study
Oaxaca	10 km South of Talea, Distrito de Villa Alta, Villa Talea de Castro.	96°16. 7999′, 17°21′30″ (−96.26, 17.35) %2370	Xalapa (INECOL) Herbarium. 14229.	[48]
Oaxaca	10 km South of Talea, San Andres Solaga.	# 98°21.242′, 20°22.880′ (% −96.26, 17.35) %2370	Mexico City (IBUNAM) 422421.	[48]
Oaxaca	* Santa María Yalina Population 18.	96°21′4.74″, 17°17′23.04″ (−96.35, 17.28) %2657	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OA0026UAEH-HGOM. OA0027UAEH-HGOM. OA0029UAEH-HGOM.	Present study
Oaxaca	Santa Maria Yalina.	96°21′11.9″, 17°17′23.5″ (−96.35, 17.28) %2649	Mexico City (IBUNAM).	[67]
Puebla	Chilchotla.	97°10′28.829″, 19°14′20″ (−97.17, 19.23) %2160	No deposited material in collection or institutions (Observation).	[33]
Puebla	Chilchotla waterfalls, Quimixtlan.	97°10′16.99″, 19°15′2.65″ (−97.17, 19.23) %2100	Xalapa (INECOL) Herbarium. XAL0145838.	[48]
Puebla	Chilchotla.	97°10′23.887″, 19°15′6.635″ (−97.17, 19.25) %2110	No deposited material in collection or institutions (Observation).	[33]
Veracruz	Near San Juan, Cofre del Perote, Xico.	97°8′27.695″, 19°29′7.121″ (−97.14,19.48) %1150	Xalapa (INECOL) Herbarium.	[43,47,48]
Veracruz	Southeast of Rinconada, near Cerro La Tolva, Las Minas.	97°7′59.99″, 19°39′30″ (−97.13, 19.65) %1900	IEB (INECOL) Herbarium. 14228.	[47,48]
Veracruz	Near Cofre de Perote Region, Las Minas.	97°8′48.009″, 19°41′26.98″ (−97.14, 19.69) %1971	No deposited material in collection or institutions (Observation).	[44,47,48]
Veracruz	500 m Southwest of Maquistla, Road to Jacal, Calcahualco.	97°10′59.999″ 19°7′ (−97.18, 19.11) %1960	Xalapa (INECOL) Herbarium. V094128. 973417.	[47,48]
Veracruz	Cerro la Tolva, Las Minas.	97°8′1.999″, 19°38′55″ (−97.13, 19.64) %2408	Xalapa (INECOL) Herbarium.	[47,48]
Veracruz	Near San Juan, Cofre de Perote, Xico.	97°8′27.695″, 19°29′7.121″ (% −97.14, 19.48) %2300	Mexico City (IBUNAM).	[43,48]
Veracruz	Xico.	# 97°0′38.836″, 19°25′27.307″ (−97.01, 19.42) %2130	Xalapa (INECOL) Herbarium.	[48]
Veracruz	Aserradero de Santa Cruz Muyuapan, Orizaba.	97°15′44.432″ 18°50′16.528″ (−97.26, 18.83) %1830	The New York Botanical Garden.	[24,48].
Veracruz	Nogales.	97°12′19.378″ 18°51′54.414″ (−97.20, 18.86) %2205	No deposited material in collection or institutions (Observation).	[33]
Veracruz	Northeast of Pico de Orizaba, Alpatlahuac and Calcahualco.	97°5′15.681″, 19°7′13.549″ (−97.08, 19.12) %1750	No deposited material in collection or institutions (Observation).	[47,48]
Veracruz	500 m Southwest of Maxistla Road to Jacal, Calcahualco.	97°8′, 31.999″, 19°7′10″ (−97.14, 19.11) %2650	Xalapa (INECOL) Herbarium.	[47,48]
Veracruz	500 m Southwest of Maxistla Road to Jacal, Calcahualco.	97°10′58.8″, 19°7′1.2″ (−97.18, 19.11) %2510	Mexico City (IBUNAM).	[47,48]
Veracruz	500 m Southwest of Maxistla road to Jacal, Calcahualco.	97°4′30″, 19°8′30″ (−97.07, 19.14) %1890	IEB (INECOL) Herbarium.	[47,48]
Veracruz	Acajete, Xalapa.	97°2′29″,19°34′36″ (−97.04, 19.57) %2389	No deposited material in collection or institutions (Observation).	[33]
Veracruz	* Las Lajas Population 5.	97°13.222′, 18°52.237′ (−97.22, 18.87) %2334	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0009UAEH-HGOM.	Present study
Veracruz	* Sótano de Tula Population 6.	97°12.916′, 18°52.072′ (−97.2152667, 18.8678667) %2308	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0007UAEH-HGOM.	Present study
Veracruz	* La roca Population 7.	97°12.774′18°52.100′ (−97.2129000, 18.8683333) %2325	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0021UAEH-HGOM.	Present study
Veracruz	* La cueva Population 8.	97°12.672′18°52.050′ (−97.2112, 18.8675) %2275	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0028UAEH-HGOM.	Present study
Veracruz	* Ladera y Carbón Population 9.	97°12.595′, 18°51.987′ (−97.2099167, 18.8664500) %2348	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0011UAEH-HGOM.	Present study
Veracruz	* Soldado fantasma Population 10.	97°12.427′, 18°51.868′ (−97.2071167, 18.8644667) %2233	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0019UAEH-HGOM.	Present study
Veracruz	* El Manzano Population 11.	97°12.627′18°51.846′ (−97.2104500, 18.8641000) %2269	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0003UAEH-HGOM.	Present study
Veracruz	* Las Tuzas Population 12.	97°12.408′, 18°51.736′ (−97.2068, 18.8622667) %2290	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0017UAEH-HGOM.	Present study
Veracruz	* Las Tuzas II Population 13.	97°12.370′, 18°51.816′ (−97.2061667, 18.8636000) %2245	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0018UAEH-HGOM.	Present study
Veracruz	* Leoneros II Population 14.	97°12.630′, 18°51.632′ (−97.2105000, 18.8605333) %2398	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0001UAEH-HGOM. OR0002UAEH-HGOM.	Present study
Veracruz	* Casco de hacienda Population 15.	97°12.367′, 18°52.468′ (−97.206531,18.874339) %2199	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0005UAEH-HGOM.	Present study
Veracruz	* Carboncillo Population 16.	97°12.674′, 18°51.877′ (−97.21123333, 18.8646167) %2208	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0012UAEH-HGOM.	Present study
Veracruz	* Paso del Toro Population 17.	97°12.361′, 18°51.974′ (−97.20601667, 18.8662333) %2204	Herbario de la Universidad Autónoma del Estado de Hidalgo (UAEH-HGOM). OR0004UAEH-HGOM.	Present study
Zacatecas	? Villa de Cos.	102°31′58.389″, 23°57′1.67″ (−102.5328861, 23.9504638) %2204	BM Herbarium NHM Natural History Museum, London.	[48]

* New registers. # Georeferenced. ? Suspicious record.

Table A2. Accession numbers and complementary information about the species used in the phylogenetic proposals.

Species	Specimen Voucher Deposited	Origin	atpB-rbcL	ITS	trnL-F
Subgen. Clavicarpa
Impatiens omeiana Hook.f	* Ruchisansakun 236 (QBG) ** S.X. Yu 4093	* China origin, cult. Chiang Mai, Thailand ** China: Sichuan	* KC905619	* KC905505	** KP776152
I. tubulosa Hemsl. ex F. B. Forbes & Hemsl	* S.X. Yu 3762 (PE)	* China: Guangxi	* KP776056	* KP776108	* KP776172
Subgen. Impatiens, sect. Fasciculatae
I. arguta Hook. f. & Thomson	* Yuan CN2k-74 (NEU), ** S.X. Yu 5406 (PE)	* China: Yunnan, ** Xizang	* DQ147812	* AY348746	** KP776116
Subgen. Impatiens, sect. Impatiens
I. aurea Hook. f. Thomson (I. pallida)	* Janssens SJ008 (LV) ** L0388636 (L)	North American origin, cult. Holden arboretum	* DQ147813	** MH377175	/
I. barbata H.F. Comber	* CN2k2-178 (NEU) **/	* China: Yunnan **/	* DQ147818	* AY348750	** AB043658
I. capensis Meerb.	* Janssens SJ009 (LV) ** W. Fritsch 1772(CAS)	* North American origin: Holden Arboretum (Cult.) ** Strybing Arboretum, San Francisco, U.S.A., bed 40B;P.	* DQ147823	* AY348759	**AF396206
I. chekiangensis Y.L. Chen	* Qin et al. 20141	* China: Zhejiang	* KP776014	* KP776064	* KP776121
I. chiulungensis Y.L. Chen	* S.X. Yu et al. 3989	* China: Sichuan	* KP776016	* KP776066	* KP776124
I. corchorifolia Franch.	* Chassot & Yuan, 99-173 (NEU) ** S.X. Yu et al. 4596 (PE)	* China: Yunnan, ** Sichuan	* DQ147831	* AY348767	** KP776127
I. davidii Franch.	* H.N. Qin et al. 19960 (PE)	* China: Fujian, Anhui	* KP776020	* KP776070	* KP776129
I. delavayi Franch.	* Chassot and Yuan 99–154 (NEU) ** S.X. Yu et al. 4783 (PE)	* China: Yunnan, ** Sichuan	* DQ147836	* AY348773	** KP776130
I. faberi Hook. f.	* Song S 007 (NEU) ** S.X. Yu 4091	China: Sichuan	* DQ147841	* AY348778	** KP776132
I. fissicornis Maxim.	* Wang SH-003 (NEU)	* China: Shaanxi	* DQ147844	* AY348782	/
I. forrestii Hook. f. ex W.W. Smith	* Yuan, CN2k-79 (NEU)	* China: Yunnan	* DQ147847	* AY348784	/
I. imbecilla Hook. f.	* Hao 426 (NEU)	* China: Sichuan	* DQ147851	* AY348796	/
I. macrovexilla Y.L. Chen	* S.X. Yu s. n. (PE)	*China: Guangxi	* KP776034	* KP776082	* KP776142
I. mexicana Rydb.	* AB0025UAEH-HGOM AB0026UAEH-HGOM AB0002UAEH-HGOM OR0007UAEH-HGOM,OR0001UAEH-HGOMOR0002UAEH-HGOM * OA0026UAEH-HGOM,OA0027UAEH-HGOM,OA0029UAEH-HGOM	Mexico: * Hidalgo, Veracruz, * Oaxaca	* OR525134,OR525135,OR525136, OR525137, OR525138, OR525139 * OR525133, OR525132, OR525131	* OR52572, OR525071, OR525070 OR525075, OR525074, OR525073 * OR525069, OR525068, OR525067	* OR525080, OR525079 OR525083, OR525082, OR52581 * OR525078, OR525077, OR525076
I. microstachys Hook. f.	* S.X. Yu et al. 5043 (PE)	* China: Sichuan	/	* KP776085	* KP776144
I. neglecta Y.L. Xu & Y.L. Chen	* H.N. Qin et al. 19942 (PE)	* China: Anhui	* KP776038	* KP776087	* KP776147
I. noli-tangere L.	* KUMA:K08	* Japan: Kumamoto, Gokanosho	* LC465215	* LC465177	* LC465233
I. nubigena W.W. Smith	* S.X. Yu et al. 4800	* S.X. Yu et al. 4800	* KP776040	* KP776089	* KP776149
I. lateristachys Y.L. Chen & Y.Q. Lu	* S.X. Yu et al. 4097	* China: Sichuan	* KP776030	* KP776078	/
I. lecomtei Hook. f.	* Yuan CN2k2-202 (NEU)	* China: Yunnan	* DQ147855	*AY348802	/
I. leptocaulon Hook. f.	* S.X. Yu et al. 4019 (PE)	* China: Guangxi	* KP776032	* KP776080	* KP776140
I. oxyanthera Hook. f.	* Song S008 (NEU)	* China: Sichuan	* DQ147865	* AY348814	/
I. platychlaena Hook. f.	* Song, S009 (NEU) ** S.X. Yu 4081 (PE)	China: Sichuan	* DQ147867	* AY348818	** KP776154
I. platysepala Y.L. Chen	* H.N. Qin et al. 19983A	* China: Anhui	* KP776044	* KP776095	* KP776155
I. pterosepala Hook. f	* S.X. Yu s.n (PE)	* China: Hubei	* KP776046	* KP776097	* KP776158
I. poculifer Hook. f	** Yuan CN2K2-209 (NEU)	** China: Yunnan	** DQ147870	** AY348820	/
I. soulieana Hook. f	* Yuan CN2k2-163 (NEU) ** S.X. Yu et al. 5053	* China: Sichuan **/	* DQ147880	* AY348833	** KP776164
I. tayemonii Hayata	* Zhengyu Jiang T2 (NEU)	* China: Taiwan	/	* AY348839	/
I. textorii Miquel	* KUMA:K29	* Japan: Kumamoto, Takamori-machi, Nojiri	* LC465213	* LC465175	* LC465231
I. tienmushanica Y.L. Chen	* S.X. Yu et al. 20089 (PE)	* China: Zhejiang	* KP776053	* KP776105	* KP776169
I. tortisepala Hook. f	*S.X. Yu et al. 4925	*China: Sichuan	*KP776054	*KP776106	*KP776170
I. yillingniana X.F.Jin, S.Z.Yang & L.Qian	* X. F. Jin 1901 (HTC).	* China: Zhejiang, Lin’an, Mt. Tianmu, Dashuwang.	* MN974549	* MN974566	* MN974583
Subgen. Impatiens, sect. Racemosae
I. scullyi Hook. F.	* Y.S. Chen et al. 652 (PE)	* China: Xizang	* KP776048	* KP776100	* KP776163
I. radiata Hook. f.	* S.X. Yu et al. 4760 (PE)2	* China: Sichuan	* KC905573	* KC905523	* KP776160
I. harae H. Ohba & S. Akiyama.	* Y.S. Chen et al. 392 (PE)	* China: Xizang	* KP776025	* KP776075	* KP776136
Subgen. Impatiens, sect. Scorpioidae
I. hunanensis Y.L. Chen	* S.X. Yu 3759 (PE)	* China: Guangxi	* KP776028	* KP776077	* KP776137
I. rubrostriata Hook. f.	* Yuan CN2k1-4 (NEU) ** S.X. Yu 3753 (PE)	China: Yunnan	* DQ147876	* AY348828	** KP776161
Subgen. Impatiens, sect. Semeiocardium
I. angulata S.X. Yu, Y.L. Chen & H.N. Qin	* S.X. Yu 3777 (PE)	* China: Guangxi	* KP776010	* KP776060	* KP776113
I. obesa Hook. f.	* S.X. Yu 3775 (PE)	* China: Guangxi	* KP776042	* KP776091	* KP776151
I. wenshanensis S.H. Huang	* S.X. Yu 4044 (PE)	* China: Guangxi	* KP776057	* KP776110	* KP776175
Subgen. Impatiens, sect. Uniflorae
I. walleriana Hook.f.	* O. Phaehler & M. Schnell I08 (NEU) S3926 (BR) * H. Fujihashi 3 (TI)	* Kenya, African origin: Nat. Bot. Gard. Meise (Cult.), * Tokyo (Cult.).	*AY348849	**DQ147892	***AB043641
I_napoensis Y.L. Chen	* Yuan CN2k1-61 (NEU), ** S.X. Yu 3049 (PE)	China: * Yunnan, ** Guangxi	* DQ147861	** AY348811	** KP776146
I. gongshanensis Y.L. Chen	* PT-ET 975 (PE)	* Myanmar: Putao	* KP776024	* KP776074	* KP776135
I. chinensis L.	* Yuan CN2k1-49 (NEU), ** S.X. Yu 3656 (PE).	China: * Yunnan, ** Guangxi	* DQ147825	* AY348761	** KP776122
Outgroup
Hydrocera triflora (L.) Wight & Arn.	** Robyns 7260, 0249369 (L)	** Sri Lanka	** DQ147895	** AY348853	/

/ No information available. “*”, “**”, “***” Same voucher utilized.

Figure A1. Traditional charcoal production practices in Veracruz, México. Photographs: Mario Adolfo García-Montes.

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Figure 1. Sampling of I. mexicana populations was carried out in three different states (Hidalgo, Veracruz, and Oaxaca). The numbers and red dots indicate collected sites and new records; the yellow triangles show known records see (Table A1); and the question mark indicates that the reporting in Zacatecas needs further identification.

Figure 2. Impatiens mexicana growing in three different sites. (A) Las Golondrinas Waterfalls, Agua Blanca, Hidalgo, (B) La roca, (C) Ladera y Carbón, Santa Cruz Muyuapan, Orizaba, Veracruz, (D) Santa Maria Yalina, Oaxaca; (E) Adaxial and (F) Abaxial surface of leaf with apex truncate; (G) Adaxial and (H) Abaxial leaf with apex acuminate; (I,J) Front view of Agua Blanca specimens with strong to pale yellow color, (K,L) Front view of Orizaba specimens with a yellow color that varies between vivid and pale-vivid. Munsell plant tissue color chart codes: (5Y 9/12 (I), 7.5Y 9/12 (J), 7.5Y 9/12 (K), 7.5Y 9/10 (L)). The color of the mottling is intense to pale red (5R 4/14 (I), 7.5R 5/16 (K,L) and 5R 5/14 (J)). Photographs: René Monzalvo.

Figure 3. Impatiens mexicana with different spot mottling dispositions and spur directions. Dorsal views: (A) Intense, (B) Medium and (C) Diffuse. Lateral views of Hidalgo specimens with spot-lines, (D) Highly recurved, (E,F) Slightly recurved, (G) Recurved with a hook end. Lateral view of Veracruz specimens with spots, (H) Extremely recurved, (I) Reflexed, (J) Medium recurved, and (K) Perpendicular and recurved. Photographs René Monzalvo.

Figure 4. Impatiens mexicana. (A,B) Cleistogamous flowers, (C) Capsule, (D) Deforestation, (E) Flowering mostly in pairs, (F) Roots, (G) Adventitious roots, (H) Stem, (I) Fungus infected leaves, (J) Epiphyte nature. Photographs: René Monzalvo.

Figure 5. Impatiens mexicana. (A–E,G,J) Hidalgo specimens; (F,H,I,K,L) Veracruz specimens; (A) Axial and (E) Adaxial faces of leaves with reticulate venation; Spur and mottling variations; (H) Recurved with hook end and reflexed, (K) Extremely recurved, (G) Perpendicular, (I) Medium recurved, and (L) Slightly recurved; Frontal face of flowers; (B) Small mottling with lateral petals that are semi-united, (C) Big mottling with lateral petals united; (J) Roots. Illustrations drawn by Monserrat Monzalvo, using deposited materials from herbarium and organisms studied in the field.

Figure 6. Stereoscopic photographs of Impatiens mexicana. (A) Exploited capsule; (B) Immature flower; (C) Lower sepal, (D) Upper petal; (E) Seed; (F) Gynoecium; (G) Capsule and stipe; (H,I) United lateral petals; (J) Androecium; (K) Lateral sepals. Photographs by René Monzalvo.

Figure 7. Phylogenetic trees of representatives of Impatiens spp. using the Bayesian Inference (BI) and Maximum likelihood (ML) based on (A) ITS, (B) ITS + atpB-rbcL, and (C) ITS + atpB-rbcL + trnL-F. Posterior probability and bootstrap values are in a range of 0–1 and 0–100 respectively. Values of less than 0.60 or 60 are marked with “-”. The arrows point PP and BS values as well.

Table 2. Main morphological characteristics and geographic information reported and updated in I. mexicana. * New characteristics. Note: Measurements are given in millimeters, unless otherwise stated.

Morphological Attributes and Geographic Information	Rydberg 1910	Barringer 1991	Present Study
Total height	40 cm or more.	60 cm.	24–235 cm.
Stem	40 cm. Glabrous, striate, light green or tinged with purple, slender.	No information.	Glabrous; light or medium green; tinged, aqueous; oval bumps; occasionally purple pigmentation.
Leaves	Alternate, dentate with obliquely triangular, thin, and mucronate teeth; acute at both ends 2–5 L × 1–3 W cm; somewhat paler beneath.	Ovate, roughly dentate, apiculate teeth; the base is obtuse; rounded to 2.5–6 L × 1.5–3.2 W cm.	Alternate, ovate dentate with obliquely triangular mucronate teeth; apiculate or rarely truncate form at apex; dark green in the adaxial face and pale green in the abaxial face; reticulate venation. 6.07–69 L × 4–42 W.
Petiole	1–3 cm.	10–20 eglandular.	1–69 glabrous.
Stipe	No information.	1–2 cm.	2–47.
* Root	No information.	No information.	Primary root 150 L × 10 W with several adventitious roots.
* Cleistogamous flowers morphology	No information.	No information.	Flower is bright to pale yellow. Munsell’s color codes: (5Y 9/12, 5Y 9/8, 5Y 9/10) 13–16 L x 7–8 W. Reflexed spur 9–10, occasionally green in the distal region. Munsell’s color codes: (5GY 9/12, 5GY 9/10). Mottling patterns can be diffused or concentrated.
Distal lobes of the united lateral petals	No information.	Different.	Doblariform 6–17 L × 5–15.5 W.
Basal lobe of the united lateral petals	Somewhat shorter; boot-shaped; acute.	Entire and different.	Orbicular, oblong, and subequal; 1.5–6 L × 1–4.5 W.
* Flower color	No information.	No information.	Vivid to vivid-pale and strong to pale yellow closest to Munsell’s colors code: (7.5Y 9/12, 7.5Y 9/10, 5Y 9/12, 7.5Y 9/12).
Color and disposition of spots in the chasmogamous flower	Large purplish spots.	No information.	Intense to pale red spots. Munsell’s color codes: (5R 4/14, 5R 5/14 and 7.5 R 5/16) covering the dorsal surface (approximately ¾ of of the region) and lateral regions of the lower sepal, while in the spur the dots fade as they reach at the distal zone, in part of the lobes, spots do not cover all of them (internal face, ¾ of basal lobe and ½ or less of distal lobe) red spots-mark lines, individuals vary, presenting diffuse patterns or condensed and intense.
Lateral sepals	Obliquely ovate; abruptly acuminate; 5–6 L.	Free sepals.	Lateral sepals ovate with dichotomous to longitudinal venation; pale green-white, greener in the middle zone, sometimes with gray spots; 2.5–7 L × 2.5–7 W.
Lateral united petals	Oblong; truncate; 15 L.	United, bilobated, 0.8–1.0 cm L.	Orbicular, reniform or obcorded lobes that mostly do not overlap each other, and occasionally overlap themselves 10–24 L × 4–13 W.
Upper petal	Broadly ovate.	Spurred without crest 2–4 (medium-hump-spur).	Orbicular; yellow; occasionally has some red spots in the mid-posterior region; 4.5–10 L × 3.5–11.5 W; has a white or green large horn in the middle that finishes in a concave tip, and a horn with different lengths and orientations.
Spur	Reflexed.	No information.	Recurved (slightly, medium, extremely and may end in the shape of a hook); perpendicular; reflexed. The spur may present at distal tip of spur may present a slightly or accentuated bifurcation and, occasionally, two bulges at the extremes 4–16 L.
Capsule	Clavate, acute; 15–18 L × 3 W.	Glabrous and glaucous; fusiform; 1–1.5 cm L.	Glabrous and glaucous; usually elongated fusiform; rarely short fusiform; 5.14–26.5 L × 1.13–6 W.
* Seeds	No information.	No information.	Ellipsoid green seeds with a brown external tegument, wart series, and ornamentation in a polar region; a rectangular-square form; 2–5.5 L × 0.75–4.0 W.
Phenology	No information.	August.	April–October (May vary between populations).
Habitat	Cloud forests.	No information.	Tropical mountain cloud forests; conserved; usually near water bodies (flood lands, rivers, waterfalls, and slopes); secondary forests in open fields.
Altitude (masl)	No information.	2300.	1906–2657.

Table 3. Morphological, and geographic information (habitat and altitude): a comparison of I. mexicana with American native congeners.

Traits and Geographic Information	I. mexicana	I. aurella	I. capensis	I. ecornuta	I. noli-tangere	I. pallida	I. turrialbana
References	([24,43], Present study)	[3,24,25,31,32]	[3,24,32,68,69,70,71,72,73,75,76,77,78,79,86]	[3,24,29,31,32]	[1,21,30,32,33]	[24,30,71]	[22,23,24]
Leaves	Alternate; ovate; dentate with obliquely triangular mucronate teeth; apex reticulate and rarely truncate; apiculate at base. Reticulate green venation in adaxial and abaxial faces; 6.07–69 mm L × 4–42 mm W.	Alternate; ovate or oval; thin; bright green; a little paler beneath; 2–8 cm L × 1–4 cm W; coarsely dentate with obliquely triangular, finely mucronate teeth.	Alternate; glaucous or pale leaves that are ovate or elliptic and crenate; egg-shaped to elliptic; shallowly and remotely saw-toothed; 2–12 cm L, 1–6 cm W.	Alternate; ovate; elliptic; ovate-elliptic; elliptic-lanceolate; with obliquely triangular mucronate teeth; adaxial face light green and paler abaxial face; 2–10 cm L, 1–5 cm W.	Opposite becoming alternate; ovate-elliptic to ovate-lanceolate or oblong; margin serrate to subcordate; often glandular near base; 1.5–10 cm L × 1.5 cm W.	Alternate; thin; bright green; somewhat paler beneath; oval or ovate; 3–15 cm L; 2–9 cm W; crenate-dentate; with rounded or obliquely triangular mucronate teeth.	Leaves opposite; uppermost crowded to appear whorled; oblong-elliptic; 2–7.5 cm L, 1–3 cm W; remotely serrate above the middle, or often entirely; apiculate with a mucro. * Reticulate green venation in the adaxial face and green to reddish in the abaxial face.
Flower color (chasmogamous)	Vivid to vivid-pale and strong to pale yellow, the closest to Munsell’s color codes: 7.5Y 9/12, 7.5Y 9/10, 5Y 9/12, 7.5Y 9/12.	Orange color.	Wide color variation, including orange, orange-brown, lemon-yellow, pale and intense yellow and orange-red. Forma citrina: Perianth with a lemon-yellow color Forma albiflora: Flowers with a white or cream color.	Orange to pale yellow.	Intense yellow.	Light sulfur-yellow; citron yellow.	Scarlet color. * Yellow tones may be inside and outside the upper petal.
Lateral united petals	Orbicular; reniform or obcorded lobes that mostly do not overlap each other but occasionally overlap themselves; 10–24 mm L × 4–13 mm W.	Spatulate form.	Obovate; rounded-spatulate; 13.5 mm L × 7.4 mm W.	Orbicular or reniform; notched; oblanceolate or oblong; 12–14 mm L × 6 mm W.	Oblique and irregularly lobed; 22 mm L × 14.4 mm W.	Broadly obliquely spatulate; 23.1 mm L × 14.6 mm W.	Oblong-elliptic with two oblong or ovate lobes; 4 mm L × 3–4 mm W; emarginate at apex.
Geographical distribution	Central and southern Mexico (Hidalgo, Puebla, Veracruz, and Oaxaca).	Southwestern Canada: British Columbia; North-West United States; Washington, Idaho, and Montana [32].	Widely distributed in the northeastern United States, its native range includes all eastern states, most Canadian provinces, many mid-western states, and the Pacific northwest [75].	Southwestern Canada: British Columbia and North-West United States: Washington, Oregon, Idaho, and Montana [87].	America; southwestern to central Canada; EUA; Alaska, Washington, Oregon, and California; Temperate Eurasia [87].	Southeastern Canada; Mid-West, North-East, and South USA [87].	Costa Rica: the Alajuela, San José and Cartago provinces; Panamá: the Chiriquí and Bocas del Toro provinces [48].
Color and disposition of spots in the chasmogamous flower	Intense to pale red spots that can be constant or dispersed; dots can be alargated into lines, covering dorsal (approximately ¾ of the ventral region) and lateral regions of the lower sepal. In part of the lobes, spots do not cover everything (internal face, ¾ of basal lobe and ½ or more of distal lobe). Munsell’s color codes: 5R 4/14, 7.5R 5/16, and 5R 5/14.	Unspotted.	Posterior sepal orange and rarely pink; usually spotted (red or purple), but sometimes wholly unspotted. $ 1.-Yellow perianth with intense orange lateral petals. $ 2.-Yellow perianth with uniform light orange mottling completely covering the complete lateral petals and inner upper petal. $ 3.-Yellow perianth, with orange spots in the inner upper petal and spots that do not invade most of the 50% of lateral surface. $ 4.-Yellow perianth with dark orange uniform or dispersed spots that cover the lateral petal and inner upper petal.	Unspotted.	Unspotted or with small brownish/red spots.	Usually unspotted or with minute reddish or brownish dots.	Unspotted or * rarely dark spots in the middle region of the lower sepal and on the spur.
Spur	4–16 mm L, recurved: slightly (110°), medium (180°), and extremely (270°); reflexed (45°), and perpendicular (90°) spur whose distal tip may present a slightly or accentuated bifurcation; Rarely spotted.	8 mm L; recurved, abruptly recurved and S-curved; cylindric spur, curved or deflexed forward.	Recurved, reflexed and perpendicular (88° to 420°).	Spurless flowers.	Curved 6–12 mm L; rarely bent through 90°.	Recurved or bent at a right angle; very short (3–8 mm L), usually notched.	Incurved; inflated at the apex; 8 mm L; is * rarely slightly spotted.
Lateral sepals	Ovate-rounded; pale green-white; greener in the middle zone, sometimes with gray spots; 2.5–7 mm L × 2.5–7 mm W; Dichotomous to longitudinal venation; main vein in the middle is greener than the secondary venation.	Ovate; abruptly acuminate; 4–5 mm L; “Sepals pouched”.	Obovate; short acuminate; 5–6.3 mm L; “Sepals pouched”.	Oblique-elliptical; oval; abruptly acuminate at the apex; 6 mm L.	Flat and oblique; green.	Broadly ovate; acuminate; light green; 5–7 mm L.	Orbicular-ovate; apiculate; subcordate at the base; 8 mm L × 7 mm W; * Sepals yellow and yellow-red to red; Longitudinally ribbed venation; main vein in the middle is yellow and the secondary venation is red.
Habitat	Tropical mountain cloud forests (TMCF) with primary and secondary vegetation, which are near water bodies and, sometimes, valleys.	Moist streambanks and meadows in the steppe or lower montane zones; swamps and wet places.	Moist habitats; forests, in the lowland; steppe and lower montane zones; Lakes, pools and riverbanks; woodlands.	Moist forests in the mountain zone and wet soils.	Damp to wet; occasionally waterlogged soils, mainly in damp woods; on the edges of rivers, streams, and lakes.	Riverbanks and wet grounds; dry sites.	Valleys; near rivers.
Altitude (masl)	1906–2657	No information	39–1334	No information	700–2000	No information	1300–2850 [48]

$ Morphological variations reported in photographs are in [33]. * Personal observation [81].

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Monzalvo, R.; Escorcia-Guerrero, D.L.; García-Montes, M.A.; Rewicz, A.; Rewicz, T.; Manríquez-Morán, N.L. The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species. Diversity 2024, 16, 87. https://doi.org/10.3390/d16020087

AMA Style

Monzalvo R, Escorcia-Guerrero DL, García-Montes MA, Rewicz A, Rewicz T, Manríquez-Morán NL. The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species. Diversity. 2024; 16(2):87. https://doi.org/10.3390/d16020087

Chicago/Turabian Style

Monzalvo, René, Diana Lizbeth Escorcia-Guerrero, Mario Adolfo García-Montes, Agnieszka Rewicz, Tomasz Rewicz, and Norma L. Manríquez-Morán. 2024. "The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species" Diversity 16, no. 2: 87. https://doi.org/10.3390/d16020087

APA Style

Monzalvo, R., Escorcia-Guerrero, D. L., García-Montes, M. A., Rewicz, A., Rewicz, T., & Manríquez-Morán, N. L. (2024). The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species. Diversity, 16(2), 87. https://doi.org/10.3390/d16020087

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The Mexican Balsam, Impatiens mexicana Rydb: A Redescription Based on Morphological and Phylogenetic Studies, with an Update of the Current Geographical Range of the Species

Abstract

1. Introduction

2. Materials and Methods

2.1. Biological Material Sampling

2.2. Morphological Study

2.3. Phylogenetic and Genetic Distance Analysis

3. Results

3.1. Redescription

3.1.1. Distribution, Habitat, and Ecology

3.1.2. Phenology

3.1.3. Floral Visitors

3.1.4. Studied Localities

3.1.5. Specimens Examined and Deposited

3.1.6. Remarks and Differential Diagnosis

3.2. Phylogenetic and Distance Genetic Analyses

4. Discussion

4.1. A Morphological Discrimination of I. mexicana with American Balsam Congeners

4.2. Impatiens mexicana: Current Known Status

4.3. Molecular Studies: Phylogenetic Position and Genetic Distances of Impatiens mexicana

4.4. Limitations of the Study and Recommended Additional Research

4.5. Final Comments

Author Contributions

Funding

Institutional Review Board Statement

Data Availability Statement

Acknowledgments

Conflicts of Interest

Appendix A

References

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