The Population of Fusarium oxysporum f. sp. cubense in Brazil Is Not Structured by Vegetative Compatibility Group or by Geographic Origin.

Fusarium wilt, caused by the soilborne fungus Fusarium oxysporum f. sp. cubense (Foc), is considered one of the most destructive diseases of bananas in Brazil. In this study, a collection of 194 monosporic isolates from several banana-producing regions located in different climatic zones along a south-to-north transect in Brazil was formed to assess the genetic structure of the population of Foc. The isolates underwent pathogenicity tests, PCR diagnosis for the detection of tropical race 4, and screening of SIX homolog genes that produce putative effector proteins. The vegetative compatibility group (VCG) of 119 isolates was determined by pairing against 17 internationally known VCG-tester strains. A group of 158 isolates was selected for simple sequence repeat (SSR) genotyping. There was moderate diversity of Foc in Brazil. Eight VCGs were identified: 0120, 0122, 0124, 0125, 0128, 01215, 01220, and 01222, of which 78% of isolates belong to a single VCG, whereas 22% of isolates are assigned to multiple VCGs, belonging to complexes of VCGs. The distribution of VCGs is uneven and independent of the banana genotype. The isolates of a VCG shared a similar profile of SIX homologs, but there was no association with geographic region. Four SSR loci were polymorphic, and, on average, 7.5 alleles were detected per locus. Thirty-five multilocus genotypes (MLGs) were identified. There was no association between VCG and MLGs, and no genetic structure of the population of Foc in Brazil was detected.

Pre-selection of banana somaclones resistant to Fusarium oxysporum f. sp. cubense, subtropical race 4.

- Fusarium wilt, caused by Fusarium oxysporum f. sp. cubense (Foc), is one of the most destructive diseases affecting banana crops worldwide. Therefore, the development of resistant cultivars is a promising alternative to mitigate the effects of the disease on banana plantations. The objectives of this study were to induce somaclonal variation in banana cultivars of the Silk and Cavendish types and to select somaclones resistant to subtropical race 4, thereby enabling the production of fruit in areas where this race is present. Shoot clump apexes of the Grand Naine and Maçã (Silk) cultivars were grown in MS medium. The cultures were subcultured four times. They were then challenged with fusaric acid (FA) in an experiment consisting of four treatments with different concentrations (0.1, 0.2, 0.3, and 0.4 mM) and five repetitions, each consisting of a Petri dish containing seven multiple shoot clumps in MS culture medium supplemented with 2.5 mg/L benzylamine purine. Multiple shoot clumps without the addition of FA were also used in the experiment, and were subcultured three times and maintained in a dark room. The multiple shoot clumps that survived the treatment with FA were transferred to MS medium and maintained in the growth chamber in the presence of light. The regenerated plants were later planted in tanks containing soil infested with an isolate classified as Foc subtropical race 4 (Foc STR4), and were evaluated for resistance to the pathogen at 90 days after inoculation (d.a.i.). Pathogen structures were confirmed by root clarification and root staining technique. All somaclones of the Maçã (Silk) cultivar were susceptible to Fusarium wilt and two somaclones of the Grand Naine cultivar were selected as resistant. The addition of FA as a selective agent was effective in the selection of somaclones among plants of the Grand Naine cultivar, as shown by the selection of two somaclones resistant to Foc STR4. The next step will consist of the agronomic and market potential validation of the selected somaclones, aiming to confirm their potential use by producers.

Molecular, Histological and Histochemical Responses of Banana Cultivars Challenged with Fusarium oxysporum f. sp. cubense with Different Levels of Virulence.

Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is the most limiting factor in the banana agribusiness worldwide. Therefore, studies regarding pathogen attack mechanisms, and especially host defense responses, in this pathosystem are of utmost importance for genetic breeding programs in the development of Foc-resistant banana cultivars. In this study, analysis at the molecular, histological and histochemical levels of the Musa spp. x Foc interaction was performed. Three Foc isolates representative of race 1 (R1), subtropical race 4 (ST4) and isolate 229A, which is a putative ST4, were inoculated in two Prata-type cultivars (Prata-Anã and BRS Platina) and one cultivar of the Cavendish type (Grand Naine). Of seven genes related to plant-pathogen interactions, five were overexpressed in 'BRS Platina' 12 h after inoculation (HAI) with Foc R1 and ST4 but had reduced or negative expression after inoculation with Foc 229A, according to RT-qPCR analyses. While hyphae, mycelia and spores of the Foc 229A isolate grow towards the central cylinder of the Grand Naine and Prata-Anã cultivars, culminating in the occlusion of the xylem vessels, the BRS Platina cultivar responds with increased presence of cellulose, phenolic compounds and calcium oxalate crystals, reducing colonization within 30 days after inoculation (DAI). In general, these data indicate that the cultivar BRS Platina has potential for use in banana-breeding programs focused on resistance to Foc tropical race 4 (TR4) and in aggregating information on the virulence relationships of the Foc pathogen and the defense responses of banana plants after infection.

Gene Expression, Histology and Histochemistry in the Interaction between Musa sp. and Pseudocercospora fijiensis.

Bananas are the main fruits responsible for feeding more than 500 million people in tropical and subtropical countries. Black Sigatoka, caused by the fungus Pseudocercospora fijiensis, is one of the most destructive disease for the crop. This fungus is mainly controlled with the use of fungicides; however, in addition to being harmful to human health, they are associated with a high cost. The development of resistant cultivars through crosses of susceptible commercial cultivars is one of the main focuses of banana breeding programs worldwide. Thus, the objective of the present study was to investigate the interaction between Musa sp. and P. fijiensis through the relative expression of candidate genes involved in the defence response to black Sigatoka in four contrasting genotypes (resistant: Calcutta 4 and Krasan Saichon; susceptible: Grand Naine and Akondro Mainty) using quantitative real-time PCR (RT-qPCR) in addition to histological and histochemical analyses to verify the defence mechanisms activated during the interaction. Differentially expressed genes (DEGs) related to the jasmonic acid and ethylene signalling pathway, GDSL-like lipases and pathogenesis-related proteins (PR-4), were identified. The number and distance between stomata were directly related to the resistance/susceptibility of each genotype. Histochemical tests showed the production of phenolic compounds and callosis as defence mechanisms activated by the resistant genotypes during the interaction process. Scanning electron microscopy (SEM) showed pathogenic structures on the leaf surface in addition to calcium oxalate crystals. The resistant genotype Krasan Saichon stood out in the analyses and has potential for use in breeding programs for resistance to black Sigatoka in banana and plantains.

Genetic Improvement for Resistance to Black Sigatoka in Bananas: A Systematic Review.

Bananas are an important staple food crop in tropical and subtropical regions in Asia, sub-Saharan Africa, and Central and South America. The plant is affected by numerous diseases, with the fungal leaf disease black Sigatoka, caused by Mycosphaerella fijiensis Morelet [anamorph: Pseudocercospora fijiensis (Morelet) Deighton], considered one of the most economically important phytosanitary problem. Although the development of resistant cultivars is recognized as most effective method for long term control of the disease, the majority of today's cultivars are susceptible. In order to gain insights into this pathosystem, this first systematic literature review on the topic is presented. Utilizing six databases (PubMed Central, Web of Science, Google Academic, Springer, CAPES and Scopus Journals) searches were performed using pre-established inclusion and exclusion criteria. From a total of 3,070 published studies examined, 24 were relevant with regard to the Musa-P. fijiensis pathosystem. Relevant papers highlighted that resistant and susceptible cultivars clearly respond differently to infection by this pathogen. M. acuminata wild diploids such as Calcutta 4 and other diploid cultivars can harbor sources of resistance genes, serving as parentals for the generation of improved diploids and subsequent gene introgression in new cultivars. From the sequenced reference genome of Musa acuminata, although the function of many genes in the genome still require validation, on the basis of transcriptome, proteome and biochemical data, numerous candidate genes and molecules have been identified for further evaluation through genetic transformation and gene editing approaches. Genes identified in the resistance response have included those associated with jasmonic acid and ethylene signaling, transcription factors, phenylpropanoid pathways, antioxidants and pathogenesis-related proteins. Papers in this study also revealed gene-derived markers in Musa applicable for downstream application in marker assisted selection. The information gathered in this review furthers understanding of the immune response in Musa to the pathogen P. fijiensis and is relevant for genetic improvement programs for bananas and plantains for control of black Sigatoka.

Improvements in the Resistance of the Banana Species to Fusarium Wilt: A Systematic Review of Methods and Perspectives.

The fungus Fusarium oxysporum f. sp. cubense (FOC), tropical race 4 (TR4), causes Fusarium wilt of banana, a pandemic that has threatened the cultivation and export trade of this fruit. This article presents the first systematic review of studies conducted in the last 10 years on the resistance of Musa spp. to Fusarium wilt. We evaluated articles deposited in different academic databases, using a standardized search string and predefined inclusion and exclusion criteria. We note that the information on the sequencing of the Musa sp. genome is certainly a source for obtaining resistant cultivars, mainly by evaluating the banana transcriptome data after infection with FOC. We also showed that there are sources of resistance to FOC race 1 (R1) and FOC TR4 in banana germplasms and that these data are the basis for obtaining resistant cultivars, although the published data are still scarce. In contrast, the transgenics approach has been adopted frequently. We propose harmonizing methods and protocols to facilitate the comparison of information obtained in different research centers and efforts based on global cooperation to cope with the disease. Thus, we offer here a contribution that may facilitate and direct research towards the production of banana resistant to FOC.

Genetic Diversity and Azole Fungicide Sensitivity in Pseudocercospora musae Field Populations in Brazil.

Pseudocercospora musae, causal agent of Sigatoka leaf spot, or yellow Sigatoka disease, is considered a major pathogen of banana (Musa spp.). Widely disseminated in Brazil, this study explored the genetic diversity in field populations of the pathogen from production areas in the Distrito Federal and the States of Bahia, Minas Gerais, and Rio Grande do Norte. Resistance to demethylation inhibitor (DMI) fungicides was also examined. For 162 isolates from 10 banana growing regions, analysis of mating type idiomorph frequency was conducted, together with estimation of genetic diversity at 15 microsatellite loci. A total of 149 haplotypes were identified across the examined populations, with an average genetic diversity of 4.06. In general, populations displayed 1:1 proportions of idiomorphs MAT1-1 and MAT1-2, providing evidence for sexual recombination. Multilocus linkage disequilibrium also indicated asexual reproduction contributing to the genetic structure of certain populations. AMOVA revealed that 86.3% of the genetic differentiation of the pathogen occurred among isolates within populations. Discriminant Analysis of Principal Components (DAPC) identified six most probable genetic groups, with no population structure associated with geographic origin or collection site. Although genetic similarity was observed among certain populations from different states, data revealed increasing genetic differentiation with increasing geographic distance, as validated by Mantel's test (r = 0.19, P < 0.001). On the basis of DMI fungicide sensitivity testing and CYP51 gene sequence polymorphism, isolates from the Distrito Federal separated into two main groups, one with generally higher EC50 values against eight DMI fungicides. A clear phenotype-to-genotype relationship was observed for isolates carrying the CYP51 alteration Y461N. Conventionally adopted fungicides for control of Sigatoka leaf spot are likely to be overcome by combined sexual and asexual reproduction mechanisms in P. musae driving genetic variability. Continued analysis of pathogen genetic diversity and monitoring of DMI sensitivity profiles of Brazilian field populations is essential for the development of integrated control strategies based on host resistance breeding and rational design of fungicide regimes.