María Francisca Perera

Introduction: The growing demand for renewable energy together with the environmental impact of fossil fuels, have intensified global interest in sustainable bioethanol production. Saccharomyces cerevisiae is the preferred microorganism for industrial fermentation due to its productivity and stress tolerance, but cumulative stress during successive cycles reduces process efficiency. Therefore, selecting stress-tolerant strains capable of adapting to fluctuating conditions is crucial. Predictive microbiology (PM), which applies mathematical models to predict and quantify microbial responses to environmental factors, remains a valuable but still limited approach in yeast-based bioethanol production. Methods: In this study, diverse PM models were applied to evaluate the effects of temperature, pH, sucrose, and ethanol concentrations on S. cerevisiae strains isolated from industrial fermentations in Tucumán, Argentina. Thirteen native isolates were compared with a commercial reference strain (Calsa). Results: The primary and secondary models used achieved excellent fits in all cases ( R ² > 0.9), effectively describing and anticipating growth responses under stress conditions relevant to industrial fermentations of the evaluated strains, which displayed wide tolerance ranges suggesting potential suitability for cell recycling and high-density fermentation, pending validation under fermentation conditions. Strains T415, Le384, LF84, and SR350 emerged as promising candidates for further validation in industrial bioethanol fermentations to improve the stability and sustainability of industrial bioethanol production, matching or outperforming the control strain. Discussion: Overall, this study underscores the usefulness of PM tools for characterizing and selecting native yeast strains with enhanced stress tolerance in local bioethanol production, as they allow for anticipation of physiological behavior in growth-based assays that mimic key industrial stresses, reduction of experimental workload, and strengthen strain selection and evaluation criteria.

Understanding the genetic diversity and population structure of sugarcane germplasm banks is essential for generating progenies with maximum variability. In this study, 350 accessions from the EEAOC germplasm bank were genotyped using DArT-seq markers. Genetic diversity, population structure, and variability were assessed through Bayesian analysis, principal coordinate analysis (PCoA), and analysis of molecular variance (AMOVA). Additionally, different sizes of core collections were evaluated. After filtering, 74,969 high-quality SNPs were retained, and two outlier genotypes were excluded. The mean observed heterozygosity (HO) was 0.28, while the mean expected heterozygosity (HE) was 0.3. Polymorphic information content (PIC) values ranged from 0 to 0.38 (mean 0.22), and the mean discrimination power (Dj) was 0.28. Structure and PCoA analyses consistently revealed three genetic clusters. AMOVA indicated that most of the genetic variation was found within subpopulations, while 10.25% was attributable to differences among them (p < 0.0001), where ΦFST suggested moderate genetic differentiation. Core collection analysis showed that a subset of 35 genotypes (10%) captured nearly 96% of the total genetic diversity, while a 30% core captured over 98%. These results provide valuable information for the effective management and utilization of sugarcane genetic resources and support the design of breeding strategies to develop superior cultivars.

Sugarcane diseases can be controlled by an integrated management approach where biotechnological tools can successfully contribute. The Obispo Colombres Agroindustrial Experimental Station (EEAOC) in Tucumán (Argentina’s main sugarcane producer) has successfully implemented multiple strategies that greatly enhance the productivity of sugarcane fields. The local breeding program develops resistant varieties by applying molecular markers to reveal the presence of Bru1 gene for brown rust resistance throughout the EEAOC germplasm collection. In addition, SNP alleles linked to novel sources of resistance were identified following a selective genotyping strategy. Another strategy is the implementation of a seed cane sanitation project using hydrothermal therapy, an in vitro culture technique, molecular diagnosis of diseases, and bionanoparticles. As a result, the incidence of systemic diseases has significantly decreased in the production fields. More recently, the use of biological products has shown to be effective for disease control in EEAOC varieties. In summary, several biotechnological strategies including molecular markers associated with resistant sources, in vitro culture of apical meristems, molecular diagnostic techniques, and the use of bioproducts are being successfully used for the sustainable management of sugarcane diseases in Tucumán, Argentina.

Acidovorax avenae subsp. avenae (Aaa) is the causal agent of red stripe in sugarcane, a disease characterized by two forms: leaf stripe and top rot. Despite the importance of this disease, little is known about Aaa infection mechanisms, and studies of virulence factors (VFs) have not been reported. Previous analysis of Aaa isolates obtained from sugarcane fields in Tucumán, Argentina, suggested that pathogenicity was not related to genetic groups determined by rep‐PCR analysis. Therefore, the aims of this work were to analyse VFs produced by Aaa strains from different genetic groups and to evaluate the association among VFs, between VFs and Aaa pathogenicity and between VFs and Aaa genetic diversity. Bacterial motility (sliding, swimming and twitching), extracellular enzyme activities (proteases, amylases and endoglucanases), H ₂ O ₂ ‐mediated oxidative stress, cell adhesion and exopolysaccharide (EPS) production were assayed in 14 Aaa isolates from different genetic groups. Pearson's correlation analysis and principal component analysis (PCA) were performed. For each VF analysed, values were variable presenting quantitative traits among the strains. Significant correlations were observed among the VFs, though no clear trend was identified between them and the pathogenicity or genetic group. Results reinforce the idea that a single VF does not account for the infection outcome, but a more complex process occurs where the interplay among VFs must be considered. The results of this study are crucial because they provide insight into the infection mechanism of this bacterium.

Sugarcane breeding programs incorporate foreign material to broaden the genetic base, expanding the gene pool. In South America, the Inter-university Network for the Development of the Sugarcane Industry (RIDESA) and Estación Experimental Agroindustrial Obispo Colombres (EEAOC) sugarcane breeding programs from Brazil and Argentina, respectively, have never exchanged materials. In that sense, the knowledge of the genetic diversity and population structure among sugarcane genotypes of both germplasm banks, determined in a reliable way through their molecular profiles, will provide valuable information to select the best parental accessions for crossing aimed at the efficient introgression of desirable alleles. For that, the aim was to determine the genetic diversity and population structure of 96 Saccharum commercial hybrids from RIDESA and EEAOC sugarcane breeding programs by using TRAP, SSR and markers related to disease resistance (e.g. Bru1 and G1). Genetic structure was determined through genetic similarity analysis, analysis of molecular variance (AMOVA), Multidimensional scaling (MDS), and a Bayesian method. Average PIC values were 0.25 and 0.26, Ho values were 0.24 and 0.28, and He values were 0.25 and 0.28, for TRAP and SSR primers, respectively. Genetic similarity, MDS, and analysis of structure revealed that Brazilian and Argentinean genotypes clustered in two groups clearly differentiated, whereas AMOVA suggested that there is more variability within programs than between them. Regarding Bru1 markers, Brazilian genotypes showed high frequency of haplotype 1 (71.4%) whereas Argentinean genotypes showed high frequency of haplotype 4 (80.8%); haplotypes 1 and 4 are indicated for the presence and absence of the brown rust resistance gene (Bru1), respectively. Respecting the G1 marker, most of the evaluated genotypes (60.4%) showed the presence of the fragment, in a similar proportion for genotypes of both programs. In conclusion, the exchange of materials, at least the most diverse genotypes, between RIDESA and EEAOC breeding programs will allow extending the genetic base of their germplasm banks, and the knowledge of genetic diversity will help breeders to better manage crosses, increasing the probability of obtaining more productive varieties.

A significant increase in the occurrence of red stripe (caused by Acidovorax avenae subsp. avenae ) has been observed in the last decade in Argentina. Considering that no extensive sampling of the main sugarcane‐producing area in the country has been conducted to characterize the diversity and population structure of A . avenae subsp. avenae , molecular markers were employed to analyse 112 isolates from Tucumán. By using repetitive element polymorphism‐based polymerase chain reaction (rep‐PCR) almost all isolates were differentiated and grouped into 10 clusters, revealing a high genetic diversity. Using the amplified fragment length polymorphism (AFLP) technique, five pairs of isolates were discriminated that could not be distinguished with rep‐PCR. Cluster analysis showed no clear association between isolate clustering, sugarcane host genotype, crop age, type of tissue sampled, fertilization, or year of sampling. Linkage equilibrium analysis by using rep‐PCR data indicated that the population has some degree of clonality. Three housekeeping genes were also sequenced: ugpB and pilT sequences were highly similar to A . avenae subsp. avenae sequences from other Argentinian isolates, whereas the lepA sequence did not reveal significant similarity. An additional four housekeeping genes could not be amplified, suggesting the existence of differences in those regions. Subsequently, virulence of 14 A . avenae subsp. avenae isolates was evaluated under controlled conditions. Results showed a differential level of aggressiveness among the isolates on a resistant sugarcane variety. This study confirmed that rep‐PCR is an adequate tool for genetic analysis and population structure characterization in bacteria, and revealed both high genetic diversity and clonal population structure of A . avenae subsp. avenae in Tucumán, Argentina.

The red stripe disease caused by Acidovorax avenae subsp. avenae in sugarcane, has become a quite relevant issue in Argentina because of its high incidence in the sugarcane growing area. The resistance of host plants is the most promising method for controlling the disease. In that sense, the Estación Experimental Agroindustrial Obispo Colombres (EEAOC) has a Sugarcane Breeding Program, which generates new varieties with higher productivity and good sanitary behavior. The lack of an effective screening technique to select resistant sugarcane genotypes limits the cultivar selection process. To develop a practical and affordable method for achieving the expression of the red stripe disease, three available inoculation techniques were evaluated under controlled conditions over two sugarcane varieties, with a previously adjustment of soil composition and nutrition and relative humidity. They consisted in (i) scrubbing the leaf surface with a cotton ball soaked in the suspension of A. avenae subsp. avenae; and spraying inoculum under two conditions: (ii) leaves pre-treated with a refined sand scarification and (iii) leaves with no scarification. Fifteen plants were inoculated per cultivar and treatment according to a randomized protocol with three replicates and the severity of the disease was evaluated on a scale of 1- 9 according to the International Society of Sugarcane Technologists. The spray inoculation using a bacterial suspension of A. avenae subsp. avenae without abrasives was also field tested. These results contribute to sugarcane breeding programs, providing a tool to assess the resistance to red stripe of their materials, overcoming the lack of bacterial pressure or favorable conditions for the disease.