It is an established fact that multiple cyclic episodes of whole-genome doubling or polyploidization have led to evolution and speciation in flowering plants (Wendel, 2015). The wondrous cycles of genome doubling are thought to be correlated with periods of extinction or global climate change over the geological time scale, while polyploids often thrive in harsh or disturbed environments (Van de Peer et al., 2021), and could colonize new habitats (Moraes et al., 2022). There are contrasting viewpoints about polyploidy as an evolutionary force, spanning from “evolutionary dead end” to “major player in evolution” (Lavania, 2020), but an established ecological force (Van de Peer et al., 2021). Initial models of polyploid evolution based on studies spanned over 70 years, considered autopolyploidy as an evolutionary dead end (Stebbins, 1999), but subsequent studies led to believe that polyploidy per se both auto- and allo- are the source for evolutionary innovation and species diversification (Van de Peer et al., 2021). Evidence accumulated on ecological niche vis-a-vis polyploid establishment underpin that polyploids evince broader adaptability and vast ecological tolerance (teBeest et al., 2012) and owe higher invasive potential compared to their diploid relatives (Pandit et al., 2011), although the distribution of such polyploid lineages may have restricted range distribution in preferred environment (Villa et al., 2022). It is also surmised that there is differential response to both biotic interactions and abiotic stress on polyploids vs. non-polyploids (Van de Peer et al., 2021). Polyploid organisms are thought to be more resilient to extreme environments owing to increased genetic variation, bufferring effect and adaptive potential of duplicated genes (Van de Peer et al., 2017; Doyle and Coate, 2019). As such, stress response in general is an important factor in the establishment of polyploidy (Van de Peer et al., 2021).

Identification of several critical genome replication/duplication events during the periods of major environmental and climate change (Alix et al., 2017), have led to propose that environmental constraints could elicit polyploidization/genome duplication as an escape to overcome the vagaries of harsh environment as adaptive speciation and survival strategy (Levin, 2019).

While discussing the likely patterns of speciation in next 500 years, Levin (2019) opines that if the global climate undergoes major changes, then this will lead to an increase in the number of plant chromosomes, and thereby an increase in the current proportion of polyploids in angiosperms to 35–50%, and an overall proportion up to 50% of the Earth’s plant species as polyploids. He further argues that such polyploidy incidences would be more pronounced in short-statured herbaceous plants. This is consistent with our earlier observations (Lavania and Srivastava, 1988, 1990), where it is observed that the subcultures of the diploid vs. autoteraploid calli when grown over a passage of monthly subcultures under stressful environment in vitro, exhibit differential effect to polyploidization, whereby the diploids turn into polyploids but the tetraploids retain the original ploidy status.

The polyploidy/genome doubling is considered a natural consequence to overcome abiotic stress. It is known to bring about changes in transpiration, water use efficiency, photosynthetic rate, phenology, antioxidant response, and morphology etc. that confer success to polyploids (Maherali et al., 2009; Deng et al., 2012; Soltis and Soltis, 2014). A lot has been discussed about the significance of both auto- and allo- polyploidy in conferring novelty and value addition to plants (Levin, 1983, 2002; Lavania and Vimala, 2022), especially where the plant biomass is the source of economic product, and their active metabolite components are valued in industrial applications (Lavania, 2005). It was therefore planned to explore whether polyploidy could lead to changes that confer morphological robustness from cultivation perspective, targeting a species that is cultivated through vegetative tillers under commercial cultivation, and the sexual system is deficient.

The genus Cymbopogon Spengel comprises a group of aromatic grasses that are either densely or loosely tufted. Although, almost all the species produce essential oil in the secretory cells present in the vegetative tissues of shoot, leaf and inflorescence, but only six species are majorly used for commercial cultivation, and the C. khasianus is one of them (Kumar et al., 2000; Lavania et al., 2012; Yogendra et al., 2021). Soenarko (1977) has provided detailed information on morphological, anatomical, geographical and ecological data on 55 species from taxonomic perspective, pinpointing that most of the species are perennial, where lateral branches/tillers are appressed to the main axis (culm). The perennial growth habit makes the growing tiller prone to lodging, and more so in C. khasianus because it becomes taller than the other species in the fast-growing season during warm and humid conditions.

The present study was undertaken to explore ploidy mediated approach to help realize genetic enhancement of an elite clone of an industrially important aromatic grass that suffers from lodging, and examine histo-morphologocal, qualitative and yield contributing characters from breeding perspective.

Both the diploid and its corresponding autopolyploid grown under similar conditions were examined for recording the data on morphological, histological and metric traits and productivity analysis. A general account of observations recorded for morphologial and histological features is provided in Table 1, and the key features related to realization of sturdiness are given in Table 2. The data related to yield contributing characters and breeding potential of the developed polyploid clone are provided in Table 3. All the qualitative features related to the theme of the study are depicted in Figures 1–5.

It is observed that there is an overall increase in body size of the developed polyploid. This is consummated through increased cell size at all the organizational levels, including tissues, organs, and micromorphological constituents, and their associated developmental changes in cell wall thickening, and metabolic changes related to photosynthetic efficiency and secretion of secondary metabolites in the form of the essential oil. Whereas, tissue thickening enables plant sturdiness, an increased cell size adds to biomass and physiological efficiency for enhanced essential oil concentration, leading to overall lodging tolerance and enhanced productivity of the economic product. Specific details based on these observations are dealt in discussion section.

Vast data accumulated on the biology of polyploids for over more than 100 years suggest that polyploids are sturdier than diploids owing to the thicker stem and leaves and associated corresponding changes (Ramsey and Ramsey, 2014), sporting enhanced efficiency in stressed environment (Pandit et al., 2011; teBeest et al., 2012; Levin, 2019), as also offering the advantage of evolutionary novelty (Levin, 2002), and profound amenability in diverse habitats, albeit having restricted distribution (Villa et al., 2022). It is with this background that the present study was undertaken on a supposedly paleo-hexaploid (2n = 60) species of the Cymbopogon species complex (base number × = 10, Soenarko, 1977; Lavania, 1988) to further enhance its biological potential by ploidy elevation for wider adaptability and enhanced productivity.

The results presented here demonstrate successful realization of genetically stable clonal autopolyploids in a commercially important aromatic grass valued for its citral rich essential oil used for its multifarious applications in aroma industry. Whereas, the target species used in this study is known to produce high biomass compared to other species of the genus, but the high biomass producing canopy per se is fraught with danger of lodging when challenged by intense air velocity and rains encountered in the fast-growing season. The instant induced polyploid reported here has been found to overcome such lodging challenge on account of stout stem, thicker and broader leaves realized in the polyploid owing to enhanced sclerenchyma in the vascular and epidermal regions and waxy coating/siliceous phytolith in the cuticular surface. At the same time the induced polyploid offers commercial advantage of enhanced biomass and increased concentration of essential oil, thus a dual advantage from cultivation perspective.

Comparative assay of micromorphological characters in diploids and autotetraploids reveal a clear-cut increase in the cell size reflected at all levels, including epidermal cells, vascular tissue, stomatal guard cells, essential oil secretory cells, phytolith size, cuticle thickness, and even epicuticular waxy deposition, albeit there is decrease in frequency of all such cells in per unit area. Nevertheless, there is an overall enhancement in the area occupied by the cells constituting larger tissues and organs. In particular, for the tissues contributing to economically important part, i.e., essential oil secreting cells in the source biomass, there is an overall increase in the area occupied by the essential oil secreting cells by 26.7%, and biomass yield by 20.3%, essential oil concentration by 26.9%, enabling an overall increase in essential oil productivity by 34%. At the same time the instant polyploid clone could withstand lodging pressure challenged by strong air currents and heavy rains encountered during the growing season (Figure 1D). Further, when compared with an elite clone “Krishna” of a related species Cymbopogon flexuosus—taken as “Check” (Figure 1C), the polyploid clone of C. khasaianus is superior by 237% in terms of the productivity of the economic product, i.e., citral rich essential oil.

The exhaustive data presented amply suggest that there is an overall increase in body size of the developed polyploid consummated through increased cell size at organizational levels, including tissues, organs and micromorphological constituents, and their associated developmental and metabolic changes. The major constituent components that deserve particular attention are highlighted below:-

Artificial polyploidy is known to increase cell size in plants; axiomatically it is likely to enhance cell size of organs and tissues involved in metabolite production. However, effect of such polyploidy mediated changes could be species specific that could be inferred from micro-morphological examination of key components. Cymbopogon is a perennial C4 aromatic grass with numerous stiff stems arising from a short, rhizomatous root stock, indigenous to tropical and sub-tropical parts of the world. In these aromatic grasses the essential oil is stored in secretory channels that run parallel to the leaf mesophyll on the abaxial side embedded in between the vascular cylinders. The essential oil synthesis is at its maximum secretion with the onset of blooming. However, the fast growing tillers reaching the blooming stage are prone to lodging when challenged by strong winds and heavy showers. Therefore, realizing plant hardiness is considered an important requirement for optimum harvest. The present study on the development of lodging tolerant plants is an important step that could be achieved through induced polyploidy, vis-a-vis adding to enhanced productivity without causing any adverse effect (related to seed-based cultivation posed on account of autopolyploidy) on cultivation since the cultivation of this grass is done through vegetative plantation as the standard practice.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

YV conceptualized and reviewed the manuscript. UL conceived the experimentation, performed the field evaluation, and drafted the manuscript. MS and SL performed micromorphological analysis and histological studies. SS and SB jointly did cytological screening. All authors contributed to the article and approved the submitted version.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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