Bouteloua is the sole genus within the subtribe Boutelouinae and comprises 60 described species, with 52 to 55 of them present in Mexico and 25 to 27 species endemic to the country. The most recent phylogenetic study using plastid and nuclear markers proposed a new infrageneric classification for Bouteloua. Section Buchloë includes eleven species, one of which is B. diversispicula. This species exhibits a broad geographic distribution, ranging from the southern United States through Mexico to Central America, and displays considerable morphological variation. In this study, we aimed to investigate the phylogenetic placement of individuals of B. diversispicula previously known as Cathestucum brevifoliumvar. ramosum y C. brevifolium var. hirsutum. Phylogenetic hypothesis using maximum likelihood analysis was conducted on 110 Bouteloua taxa, including seven new from Churumuco, Michoacán, using two plastid markers (rpl32-trnL and rps16-trnK) and nuclear ITS (internal transcribed spacer). Six of the Bouteloua samples from Churumuco were nested within the clade of sect. Buchloë. Based on our molecular results and morphological evidence, we describe two new species of Bouteloua.

Study Site—The study site was located at Tierra Caliente, Michoacan. This region belongs to the Balsas Basin province (Rzedowski 1978; Mor- rone et al. 2017). According to Ibarra-Manrıquez et al. (2021), it contains two distinct plant communities, tropical deciduous and subdeciduous for- ests sensu Rzedowski (1978). The mean monthly temperature is 29.8C, and the mean annual precipitation is 564 mm. Specimens from other regions including the Mexican states of Jalisco, Guerrero, Oaxaca, and Chiapas, as well as Guatemala were also examined.

Taxon Sampling—Samples of fresh leaves were collected in December 2023 near Churumuco de Morelos, Michoacan. The samples were kept in silica gel until processing in the laboratory. Specimens for herbarium vou- chers were collected and deposited at the IBUG herbarium (University of Guadalajara; Thiers 2024).

DNA Extraction and Sequencing—DNA extraction was conducted using the CTAB method (Doyle and Doyle 1987), with modifications as described by Cota-Sanchez et al. (2006), from 100 mg of dry leaf tissue pre- served in silica gel. The plant material was ground using a mortar and pestle. Subsequently, the extracted DNA was dissolved in 100 mL of Tris EDTA buffer solution (TE). The concentration and purity of the DNA were determined using a NanoDrop 2000TM Spectrophotometer (Thermo Fisher Scientific, Wilmington, DE, USA). For amplification, we employed the same primers utilized by Peterson et al. (2015) (rpl32-trnL, rps16-trnK, and nuclear ITS). Polymerase chain reaction (PCR) was carried out using an AERISTM thermal cycler (Esco Healthcare, Singapore) under the fol- lowing conditions: initial denaturation at 95C for 4 min, followed by 35 cycles of denaturation at 94C for 30 sec, annealing at 45–50C for 30 sec, and extension at 72C for 1.5 min, with a final extension step at 72C for 10 min. The PCR products were visualized on a 1% agarose gel stained with GelRed and examined under a UV transilluminator (UVP). Subse- quently, the amplified products were analyzed on a SeqStudio-232000826 (Applied BiosystemsVR ) at the Laboratorio Nacional de Identificacion y Caracterizacion Vegetal (LaniVeg, University of Guadalajara). The plastid and nuclear DNA sequences were edited using Sequencher v. 4.1.2 (Gene Codes Corporation, Ann Arbor, MI, USA), and aligned using Phyde (Mu€ller et al. 2005). GenBank accession numbers; ITS: PQ329508–PQ329514; rps16-trnK: PQ381973–PQ381975; rpl32-trnL: PQ381976–PQ381978. All sequence alignments are available on the Dryad Digital Repository (Ruiz-Sanchez and Perez-Garcia 2025). Genomic DNA extraction, product purification, amplification, and sequencing were per- formed at the National Laboratory of Plant Identification and Characteri- zation (LaniVeg), University of Guadalajara.

Phylogenetic Analysis—Our sampling included 107 Bouteloua sam- ples; seven of them were newly generated, and 100 were used previously in the phylogenetic analysis of Bouteloua (Peterson et al. 2015; Appendix S1). Muhlenbergia ramulosa (Kunth) Swallen, Monanthochloe littoralis Engelm., and Scleropogon brevifolius Phil. were used as outgroups. Mulen- bergia ramulosa was used as the functional root. First, we used ModelTest- NG (Darriba et al. 2020) to identify the molecular evolution model that best fits our three different matrices. Next, we performed maximum likeli- hood (ML) analysis using RAxML v. 8 (Stamatakis 2014) and implemen- ted it through raxmlGUI v. 2.0.10 (Edler et al. 2021). Since our DNA matrices were incomplete (Appendix S1), we used ML to assess how well the method reconstructed the topology with incomplete data (Wiens and Morrill 2011; Jiang et al. 2014). Node support was estimated with a para- metric bootstrap with 1000 replicates. Because most of the Bouteloua sequences came from the Peterson et al. (2015) study, we followed their infrageneric classification of the genus.

Morphological Analysis—Additionally, we reviewed specimens deposited in the herbaria IBUG, IEB, MEXU, NY, and US (Thiers 2024). We used a ruler with a centimeter scale to take measurements, but for measurements of micromorphological characters, we used a stereoscopic microscope with a millimeter grid. We followed the terminology of Pierce (1979) and Peterson et al. (2015) for the species descriptions.