Transcriptome Analysis of a Cotton Cultivar Provides Insights into the Differentially Expressed Genes Underlying Heightened Resistance to the Devastating Verticillium Wilt

Cotton is an important economic crop worldwide. Verticillium wilt (VW) caused by Verticillium dahliae (V. dahliae) is a serious disease in cotton, resulting in massive yield losses and decline of fiber quality. Breeding resistant cotton cultivars is an efficient but elaborate method to improve the resistance of cotton against V. dahliae infection. However, the functional mechanism of several excellent VW resistant cotton cultivars is poorly understood at present. In our current study, we carried out RNA-seq to discover the differentially expressed genes (DEGs) in the roots of susceptible cotton Gossypium hirsutum cultivar Junmian 1 (J1) and resistant cotton G.hirsutum cultivar Liaomian 38 (L38) upon Vd991 inoculation at two time points compared with the mock inoculated control plants. The potential function of DEGs uniquely expressed in J1 and L38 was also analyzed by GO enrichment and KEGG pathway associations. Most DEGs were assigned to resistance-related functions. In addition, resistance gene analogues (RGAs) were identified and analyzed for their role in the heightened resistance of the L38 cultivar against the devastating Vd991. In summary, we analyzed the regulatory network of genes in the resistant cotton cultivar L38 during V. dahliae infection, providing a novel and comprehensive insight into VW resistance in cotton.

Evaluation of Cotton Cultivar and At-Plant Nematicide Application on Seasonal Populations of Reniform Nematode

The reniform nematode, Rotylenchulus reniformis (Linford and Oliveira), remains a common, widespread nematode pest of cotton across the southern United States. Trials were conducted during 2017 at three non-irrigated locations: one location in Hamilton, MS, and two locations in Tchula, MS, in field settings with a history of cotton production and documented economically-damaging reniform nematode populations. Trials were designed to evaluate the response of two cotton cultivars to in-furrow nematicides consisting of aldicarb, 1,3-dichloropropene, and a non-treated control applied for nematode suppression. No significant interactions between cotton cultivar and nematicide were observed. However, treatment with 1,3-dichloropropene produced greater plant biomass, and plant height compared to aldicarb-treated cotton and the nontreated. Nematode densities were suppressed with the use of 1,3-dichloropropene compared to aldicarb and the non-treated control. The use of 1,3-dichloropropene resulted in positive early-season plant growth responses; however, these responses did not translate into greater yield.

An in vitro co-culture system for rapid differential response to Fusarium oxysporum f. sp. vasinfectum race 4 in three cotton cultivars

Fusarium oxysporum f. sp. vasinfectum race 4 (FOV4) is a devastating fungus pathogen that causes Fusarium wilt in both domesticated cotton species, Gossypium hirsutum (Upland) and G. barbadense (Pima). Greenhouse and field-based pathogenicity assays can be a challenge due to non-uniform inoculum levels, the presence of endophytes, and varying environmental factors. Therefore, an in vitro co-culture system was designed to support the growth of both domesticated cotton species and FOV4 using an inert polyphenolic foam substrate with a liquid medium. A Fusarium wilt-susceptible Pima cotton cultivar, G. barbadense ‘GB1031’, a highly resistant Pima cotton cultivar, G. barbadense ‘DP348RF’, and a susceptible Upland cotton cultivar, G. hirsutum ‘TM-1’, were evaluated for 30 days during co-culture with FOV4 in this foam-based system. Thirty days after inoculation, disease symptoms were more severe in both the susceptible cultivars, which displayed higher percentages of foliar damage, and greater plant mortality, than observed in ‘DP348RF’, the resistant Pima cotton cultivar. This foam-based in vitro system may be useful for screening cotton germplasm for resistance to a variety of fungus pathogens and to facilitate the study of biotic interactions in domesticated cotton species under controlled environmental conditions.

Flagellin FLiC Enhances Resistance of Upland Cotton to Verticillium dahliae

The mechanism by which flagellin induces an immune response in plants is still unclear. The purpose of this study is to reveal the effect and mechanism of flagellin in inducing plants to produce an immune response to increase the resistance to Verticillium dahliae (VD). The full-length flagellin gene C (FliC) was cloned from an endophytic bacteria (Pseudomonas) in the root of upland cotton cultivar Zhongmiansuo 41. The FliC protein purified in vitro has 47.50% and 32.42% biocontrol effects on resistant and susceptible cotton cultivars, respectively. FLiC can induce allergic reactions in tobacco leaf cells and immune responses in cotton plants. Smearing FLiC to cotton and performing RNA-seq analysis, it is significantly enriched in the activity of positive ion transporters such as potassium ions and calcium ions (Ca2+), diterpenoid biosynthesis, phenylpropane biosynthesis and other disease-resistant metabolic pathways. FLiC inhibits the expression of calcium antiporter activity gene (GhCAA) to accelerate intracellular Ca2+ influx and stimulate the increase of intracellular hydrogen peroxide (H2O2) and nitric oxide (NO) content. The coordinated regulation of Ca2+, H2O2 and NO enhances disease resistance. The resistance of transgenic FLiC gene Arabidopsis to VD was significantly improved. The FLiC gene can be used as an anti-VD gene and as a regulator to improve resistance to VD.

Response of Xanthomonas citri pv. malvacearum isolates to cotton differing in susceptibility to the bacterium and their predicted type III effectors.

From 2015 to 2020, 342 isolates of Xanthomonas citri pv. malvacearum (Xcm) were obtained from cotton fields in Texas, including 64 isolates collected from symptomatic cultivars that were thought to be resistant to race 18 strains of Xcm (the predominant race in the USA). Symptoms on highly resistant cultivars prompted concern that a new race of Xcm was present. The 342 isolates were inoculated on a race 18 susceptible (DP 1747NR B2XF) and race 18 resistant (S295) cotton cultivar and none of the isolates caused blight type symptoms (water soaking and chlorosis) on S295, indicating that the B12 resistant gene was still beneficial for disease management. Four cultivars, varying in their field response to bacterial blight, were inoculated with each of 17 isolates of Xcm and the incidence of plants exhibiting bacterial blight symptoms averaged 87% for DP 1747NR B2XF, 51% for partially susceptible NG 4936 B3XF, 16% for partially resistant DP 1646 B2XF, and 0% for S295. Xcm isolates from Texas (11), Georgia (1) and Oklahoma (1) were sequenced, and their type three effectors (T3Es) were predicted. All isolates (GA, OK, TX) had the same T3E proteins as previously identified Xcm race 18 isolates (tested for 25 genes), including XopJ. Race 1, 2, 3, and 12 of Xcm included in the comparisons were all missing the XopJ gene. Use of cultivars with the B12 gene is an effective strategy to manage bacterial blight of cotton.

Molecular cloning and characterization of GhERF105, a gene contributing to the regulation of gland formation in upland cotton (Gossypium hirsutum L.)

Background Gossypium hirsutum L. (cotton) is one of the most economically important crops in the world due to its significant source of fiber, feed, foodstuff, oil and biofuel products. However, the utilization of cottonseed was limited due to the presence of small and darkly pigmented glands that contain large amounts of gossypol, which is toxic to human beings and non-ruminant animals. To date, some progress has been made in the pigment gland formation, but the underlying molecular mechanism of its formation was still unclear. Results In this study, we identified an AP2/ERF transcription factor named GhERF105 (GH_A12G2166), which was involved in the regulation of gland pigmentation by the comparative transcriptome analysis of the leaf of glanded and glandless plants. It encoded an ERF protein containing a converved AP2 domain which was localized in the nucleus with transcriptional activity, and showed the high expression in glanded cotton accessions that contained much gossypol. Virus-induced gene silencing (VIGS) against GhERF105 caused the dramatic reduction in the number of glands and significantly lowered levels of gossypol in cotton leaves. GhERF105 showed the patterns of spatiotemporal and inducible expression in the glanded plants. Conclusions These results suggest that GhERF105 contributes to the pigment gland formation and gossypol biosynthesis in partial organs of glanded plant. It also provides a potential molecular basis to generate ‘glandless-seed’ and ‘glanded-plant’ cotton cultivar.

Molecular cloning and characterization of GhERF105, a gene contributing to the regulation of gland formation in upland cotton (Gossypium hirsutum L.)

Background: Gossypium hirsutum L. ( cotton) is one of the most economically important crops in the world due to its significant source of fiber, feed, foodstuff, oil and biofuel products. However, the utilization of cottonseed was limited due to the presence of small and darkly pigmented glands that contain large amounts of gossypol, which is toxic to human beings and non-ruminant animals. To date, some progress has been made in the pigment gland formation, but the underlying molecular mechanism of its formation was still unclear.Results: In this study, we identified an AP2/ERF transcription factor named GhERF105 (GH_A12G2166), which was involved in the regulation of gland pigmentation by the comparative transcriptome analysis of the leaf of glanded and glandless plants. It encoded an ERF protein containing a converved AP2 domain which was localized in the nucleus with transcriptional activity, and showed the high expression in glanded cotton accessions that contained much gossypol. Virus-induced gene silencing(VIGS) against GhERF105 caused the dramatic reduction in the number of glands and significantly lowered levels of gossypol in cotton leaves. GhERF105 showed the patterns of spatiotemporal and inducible expression in the glanded plants. Conclusions: These results suggest that GhERF105 contributes to the pigment gland formation and gossypol biosynthesis in partial organs of glanded plant. It also provides a potential molecular basis to generate ‘glandless-seed’ and ‘glanded-plant’ cotton cultivar.

Molecular cloning and characterization of GhERF105, a gene contributing to the regulation of gland formation in upland cotton (Gossypium hirsutum L.)

Background: Gossypium hirsutum L. ( cotton) is one of the most economically important crops in the world due to its significant source of fiber, feed, foodstuff, oil and biofuel products. However, the utilization of cottonseed was limited due to the presence of small and darkly pigmented glands that contain large amounts of gossypol, which is toxic to human beings and non-ruminant animals. To date, some progress has been made in the pigment gland formation, but the underlying molecular mechanism of its formation was still unclear.Results: In this study, we identified an AP2/ERF transcription factor named GhERF105 (GH_A12G2166), which was involved in the regulation of gland pigmentation by the comparative transcriptome analysis of the leaf of glanded and glandless plants. It encoded an ERF protein containing a converved AP2 domain which was localized in the nucleus with transcriptional activity, and showed the high expression in glanded cotton accessions that contained much gossypol. Virus-induced gene silencing(VIGS) against GhERF105 caused the dramatic reduction in the number of glands and significantly lowered levels of gossypol in cotton leaves. GhERF105 showed the patterns of spatiotemporal and inducible expression in the glanded plants. Conclusions: These results suggest that GhERF105 contributes to the pigment gland formation and gossypol biosynthesis in partial organs of glanded plant. It also provides a potential molecular basis to generate ‘glandless-seed’ and ‘glanded-plant’ cotton cultivar.