The Stevia rebaudiana Germplasm Bank at the University of Cordoba, Colombia, plays a pivotal role in conserving and efficiently utilizing the genetic variability of this species. Despite safeguarding promising genotypes with valuable traits, such as late flowering or a significant diterpenoid glycoside content, there is a need for an efficient mass propagation protocol for elite genotypes. This study aims to develop efficient in vitro micropropagation protocols for three elite S. rebaudiana genotypes (L020, L102, and Morita II). The methods employed various combinations of cytokinins and auxins following organogenesis protocols. The results showed that optimal shoot multiplication (17.3 shoots per explant) for L020 was achieved when cultures were grown on a basal medium MS supplemented with 1 μM 6-benzylaminopurine (BAP). For L102, optimal shoot multiplication (18.5 shoots per explant) was achieved in MS supplemented with 1 μM BAP and 0.5 μM naphthalene acetic acid (NAA), while for Morita II, the best treatment was an MS supplemented with 2 μM BAP and 0.5 μM NAA, producing 16.4 shoots per explant. This study successfully achieved micropropagation for promising S. rebaudiana genotypes, highlighting the significant impact of genotype on tissue culture, particularly in shoot multiplication. Developing a successful micropropagation system is crucial for the conservation and improvement of S. rebaudiana, with significant implications for its future use and performance.

The Germplasm Bank of Stevia rebaudiana segregants of the University of Córdoba is a strategy for the use of genetic variability and efficient crop yield. There are genotypes with important characteristics such as: high tolerance to salt stress and climatic change (high CO₂ in the Earth surface), late flowering, erect architecture, and high contents of steviol glycosides. However, there is a lack of in-depth studies of morphophysiological and biochemical indicators such as gas exchange, chlorophyll a fluorescence, chloroplast pigments, and antioxidant systems, which allow us to develop early selection tools for elite genotypes. The genotypes (L020, Morita II, and L102) were found to have elite characteristics such as high efficiency in water use, excellent biomass production, and a more robust antioxidant system than the genotypes (L057 and L082). The L020 genotype presented the highest content of stevioside and rebaudioside A, followed by the Morita II genotype. We found a close correlation between the electron transport rate and the mechanisms that increase photosystem complexes. In this sense, non-photochemical cooling modulated by the release of heat by the leaves is a fact that is confirmed by the greater activity of the xanthine pool to protect the photochemical complexes in S. rebaudiana.

Limnospira maxima is a remarkable organism showing great potential as a versatile and sustainable food source, offering a powerful solution to address the pressing issues of malnutrition and undernourishment worldwide. L. maxima contains high amounts of proteins, vitamins, minerals, and essential fatty acids. It can be grown in both bioreactors and open systems; however, before considering industrial production, optimization studies of the cultivation must be conducted to obtain knowledge about the ideal environmental conditions. Additionally, for the molecular typing of L. maxima strains and their industrial scaling, high-quality and large quantity DNA extraction is required. Notwithstanding, DNA extraction from L. maxima can be challenging due to the low amount of DNA in cells and the presence of difficult-to-remove substances such as polysaccharides and polyphenols. In this study, the quality and quantity of DNA extracted from two types of L. maxima samples (Limnospira maxima strain SISCA accession GenBank: OR195505.1) were evaluated using three commercially available DNA extraction kits and two types of input biological material. The results showed that Pbact-P kit had the highest quantity and quality of DNA, while CTAB-P allowed for a higher quantity and quality of RNA, making them optimal protocols for nucleic acid extraction to improve PCR, rt-PCR, and genome sequencing of L. maxima compared with other extraction methods.

Stevia rebaudiana Bertoni is a strategically important perennial crop because it is the main botanical source of steviol glycosides, a group of high-intensity, non-caloric sweeteners increasingly demanded by the global food and beverage industry. Despite the rapid expansion of stevia cultivation, commercial production remains strongly dependent on a narrow genetic base, particularly on clonally propagated cultivars such as ‘Morita II’, which has long served as the industrial benchmark because of its favourable rebaudioside A profile and processing consistency. This dependence has raised concerns about limited adaptive capacity, genetic erosion and restricted long-term breeding progress. In this review, we provide an integrated and critical synthesis of current knowledge on the genetic diversity of S. rebaudiana, the biosynthetic and regulatory architecture of steviol glycosides, and the conventional and emerging strategies available for crop improvement. Unlike previous reviews, this article explicitly connects domestication-driven genetic bottlenecks, wild germplasm mobilisation, metabolic pathway regulation, advanced analytical phenotyping and precision breeding into a single systems-oriented framework. We examine the roles of wild germplasm, somaclonal variation, polyploidy, molecular markers, omics-assisted approaches and transgene-free genome editing as complementary tools to broaden the stevia breeding base while preserving industrial quality standards. We finally propose an integrative roadmap for the sustainable genetic improvement of stevia, positioning ‘Morita II’ not as an endpoint, but as a benchmark within a broader diversification strategy.

Stevia rebaudiana (Bert.) is a natural sweetener with diterpenic glycosides, offering 300 times more sweetness than sucrose, and boasting low-calorie content, along with medicinal and therapeutic properties. To address a limited genetic diversity, a germplasm bank comprising 115 segregated lines of stevia was established in Colombia. Notwithstanding, the fungus Athelia rolfsii adversely impacted key stevia lines, leading to wilting and plant mortality. For this reason our study aimed to explore the biocontrol potential of native strains of Trichoderma spp., isolated from productive lots in Colombia's Cordoba department, as a solution to manage A. rolfsii. Soil samples underwent serial dilution technique (103 conidia.mL-1) to obtain Trichoderma spp. isolates, which were thoroughly morphologically and molecularly characterized. A total five native strains were identified through sequencing the PCR products using universal ITS primers, and comparisons with the GENBANK database confirmed their identity (OK310695.1, ON237703.1, ON238106.1, ON237737.1, and ON237923.1). Their biocontrol efficacy against A. rolfsii was evaluated using the dual culture technique. Remarkably, all strains exhibited significant biocontrol capacity against the A. rolfsii isolate INVEPAR-05 (OK271308.1), except for Trichoderma viride INVEPAR-T10 (ON237923.1). These findings suggest that Trichoderma spp. could be a potential option to manage the fungus in stevia.

Intraspecific phenotypic diversity in clonally propagated crops is frequently constrained by narrow domestication histories and the widespread use of a limited number of elite cultivars. In Stevia rebaudiana, commercial production has largely centred on cv. ‘Morita II’, raising concerns about reduced diversity and adaptive potential. This study characterised and structured phenotypic diversity within a segregating population derived from ‘Morita II’ under tropical field conditions. Eighty-six progeny-derived genotypes (clonally propagated) plus the commercial control (87 genotypes total) were evaluated using 25 agromorphological descriptors (qualitative and quantitative). Quantitative traits showed broad variation, including plant height (28.26–119.50 cm) and dry yield rate (0.94–28.55 g plant⁻¹). Multivariate analyses of mixed descriptors (PCA and hierarchical clustering based on Gower distance) identified plant architecture, vegetative growth, and phenology as the main sources of differentiation. The first two principal components explained 19.65% and 12.58% of total phenotypic variance, respectively (32.23% cumulative). Hierarchical clustering (UPGMA; dissimilarity cut-off = 0.25) resolved four phenotypic groups (GI–GIV) with sizes n = 3, 1, 66, and 17, respectively, enabling the definition of contrasting ideotype candidates based on recurrent trait combinations. These results provide a quantitative baseline for phenotypic structuring, prioritization of materials for further evaluation, and management of clonal stevia collections in tropical production systems. These ideotype candidates should be considered preliminary until validated across environments and linked to chemical quality traits.