Resistance To Pokkah Boeng Disease In New And Main Cultivated Sugarcane Varieties

Pokkah boeng is an epidemic fungal disease that seriously affects development of the sugarcane industry in China. Resistance to pokkah boeng disease differs among sugarcane varieties. Breeding and planting resistant cultivars is the most economic and effective measure for controlling this disease. In this study, new sugarcane varieties bred by the China Sugarcane System and the main varieties cultivated in various sugarcane areas, were screened to �nd new elite pokkah boeng resistant varieties for application in sugarcane production. The natural �eld resistance to pokkah boeng disease of 60 new varieties was evaluated in regionalized experiment. Meanwhile, the �eld resistance of the 31 main cultivated varieties was analyzed in Lincang city, Puer city, Yuxi city, Yunnan Province, and in Yizhou district, Guangxi Province, where the incidence of pokkah boeng is particularly high. Field surveys showed that 35 (58.33%) of the 60 new sugarcane varieties were highly resistant to moderately resistant, and 25 (41.67%) were susceptible to highly susceptible. Of the 31 main cultivated varieties, 15 (48.39%) were highly resistant to moderately resistant, and 16 (51.61%) were susceptible to highly susceptible. The results suggested that, in wetter and rainy sugarcane areas with high incidence of pokkah boeng disease, it is necessary to increase efforts to eliminate the susceptible main cultivated varieties and promote the application of resistant new varieties. This will help to achieve a reasonable distribution of varieties, fundamentally control the outbreak of pokkah boeng disease, and facilitate the high-quality development of the sugarcane industry in China.


Introduction
Pokkah boeng is an important fungal disease that affects sugarcane (Saccharum o cinarum) worldwide.F. verticillioides and F. proliferatum are two major Fusarium species that cause sugarcane pokkah boeng disease in China, with F. verticillioides accounting for over 90% of the recorded disease (Lin et al. 2014).Many cultivated plants act as middle hosts of the pathogen, including rice, sorghum, maize, banana, and pumpkin (Huang et al. 2018;Rott et al. 2000).The main symptoms of pokkah boeng disease were yellowing, shriveling and twisting of young leaves.Seriously infected plants exhibited growth cessation, and the top becomes very small, ultimately resulting in death by growing tip rot (Figure 1) (Huang et al. 2018).Pokkah boeng disease was rst found in Java in 1890 (Rott et al. 2000).In 1921, with the breeding and popularization of the sugarcane variety POJ2878, pokkah boeng disease became epidemic and caused varying degrees of economic losses worldwide (Mohamed et al. 2008; Raid and Rott 2015; Singh et al. 2006;Vishwakarma et al., 2013).
In China, this disease has been reported in Fujian, Taiwan, Guangdong, Guangxi, Yunnan, Sichuan, and Hainan (Huang 1993; Huang and Li 2016;Liu et al. 1991;Lu 2007).In the 1980s, a severe outbreak of pokkah boeng occurred in Guangxi.The diseased plant rate reached 52.4% in Guitang 11, and the cane yield and brix were reduced by 14% and 7%, respectively (Liu et al. 1991).In 1989, the disease occurred suddenly in the Pearl River Delta, where Yuetang 57-423 and Yuetang 54-176 were highly susceptible varieties.The infected area covered over 400 hm 2 , and the incidence rate was 30-50%.In some areas the incidence rate reached 80%, which had a signi cant impact on local sugarcane production (Huang et al. 1990).Since 2000, with the promotion of susceptible varieties ROC 1, ROC 10, ROC 16, ROC 22, ROC 25, and Yuetang 93-159 throughout the sugarcane regions of China, pokkah boeng disease occurred frequently.This has led to an increasingly serious situation and pokkah boeng is becoming an important disease in the growing stage (Huang and Li 2016;Huang et al. 2018).
In recent years, there has been more rainfall and high moisture, and the susceptible varieties Yuetang 93-159, ROC 25, ROC 1, and Chuantang 79-15 which have high yield and high sugar-content have been planted across large areas.This has led to the outbreak of pokkah boeng disease in Lincang, Puer, Yuxi, Honghe, and Banna, and has caused severe reductions in sugar and yield.Therefore, sugarcane production is increasing under threat (Li et al. 2017a).Investigations have shown that in susceptible varieties the disease is serious and causes the death of a large number of stalks.The diseased plant rate was found to be 81.1% on average and 100% in seriously infected elds.The measured yield loss was 38.42% on average, reaching 48.5%.The sugar content was reduced by 3.14% on average, with reductions as great as 4.21% (Shan et al. 2018).
At present, the research on pokkah boeng disease is mainly focused on the taxonomy, biological characteristics, phylogenetic relationship, and metabolites of the pathogen (Guo et  Resistance to pokkah boeng disease differs among sugarcane varieties.Screening and planting resistant varieties is the most economic and effective method to control the disease.In recent years, under support from the China Sugarcane System, several new elite sugarcane varieties have been bred via a national joint research effort.However, these new varieties have not yet been evaluated their resistance to pokkah boeng disease.The results could provide a scienti c basis for the breeding and selection of elite resistant varieties for commercial cultivation.This will bene t the control of pokkah boeng disease in sugarcane production via the rational distribution of varieties.
This study combined the regionalization experiment of new sugarcane varieties with disease investigation in sugarcane planting areas.A eld resistance screening was conducted to identify the resistance of pokkah boeng disease in 60 new elite sugarcane varieties and 31 main cultivated varieties from 2016 and 2019.The determined resistance levels of the different varieties will provide a scienti c basis for the selection of varieties for sugarcane production.

Material And Methods
Tested varieties A total of 60 new elite sugarcane varieties bred in China in recent years (Table 1) and 31 of the main varieties cultivated in Yunnan and Guangxi sugarcane elds (Table 2) were tested in this study.ROC 22 was used as the control varieties of the regional experiment.Cultivar ROC 20 was used as a resistant control and ROC 25 as a susceptible control.

Disease occurrence and eld resistance evaluation of new varieties under natural condition
The 60 new varieties was planted in in two successive batches at regionalization experimental station in Kaiyuan and Lincang, Yunnan Province, China in March 2016-2017 and 2018-2019 respectively.. ROC 22 was used as a control varieties of the regionalization experiment, and ROC 20 and ROC 25 were used as the pokkah boeng-resistant and pokkah boeng-susceptible control varieties, respectively.A randomized complete block design with three repeats was used.Each plot consisted of ve rows (6 m in length and 5 m in wide; total area of 30 m 2 ) with 1 m spacing between rows.The varieties were infected by naturally occurring populations of the pathogen of pokkah boeng disease in the eld.To maximize infection, cultivar ROC 25, which is highly susceptible to pokkah boeng disease, was planted along the borders surrounding the trial elds.Two rows of ROC 25 were also planted repeatedly between every two test varieties.
When pokkah boeng incidence peaked in the susceptible control variety (October to November each year), pokkah boeng incidence of each new variety in the newly-planted and ratoon eld was analyzed to evaluate the eld resistance.For each variety, the number of diseased plants were recorded in three replicates by observe the main symptoms refer to Figure 1, and 100 successive millable stalks were investigated per replicate.Therefore, a total of 300 plants/stalks were surveyed per variety.The total number of surveyed plants and the number of diseased plants were recorded to calculate the natural diseased incidence.The eld disease incidence (%) = (the number of diseased plants / total number of surveyed plants) × 100%.Referring to the classi cation method of Laboratory of Diseases and Insect Pests of the Technology System of National Sugar Industry (2012), and making modi cations, the eld resistance to pokkah boeng disease was graded 1 to 5 by the natural diseased incidence in each variety.Grade 1, 2, 3, 4, and 5 represented highly resistant, resistant, moderately resistant, susceptible, and highly susceptible, respectively and the corresponding ranges of disease incidence were 0.0%, 0.1-10.0%,10.1-20.0%,20.1-40.0%,and 40.1-100.0%

Disease occurrence and eld resistance evaluation of main cultivated varieties under natural condition
From 2017 to 2019, from October to November, ineach year, pokkah boeng incidence of each new variety in the newly-planted and ratoon eld was analyzed to evaluate the eld resistance.The main cultivated varieties were from Lincang city, Puer city, and Yuxi city in Yunnan Province, and Yizhou district in Guangxi Province.These sugarcane varieties were infected by natural pathogen population of pokkah boeng disease in the eld.For each main cultivated variety, a three points sampling method was used in the elds of newly-planted and ratoon cane.The diseased plant was recorded by three professional sugarcane pathologists with visual inspection of main symptoms.100 successive plants were assessed at each sampling points in.Therefore, a total of 300 plants were surveyed per variety.The total number of surveyed plants and the number of diseased plants were recorded to calculate the eld diseased incidence.The eld resistance of the main cultivated varieties to pokkah boeng disease was graded 1 to 5 and evaluated as described in section 2.2.Date analysis DPS 9.01 data processing system was used for statistical analysis, and Duncan's new multiple range method was used for difference signi cance analysis (P < 0.05).

Field resistance of new varieties
The resistance control variety ROC 20 was highly resistant to pokkah boeng disease, and the susceptible control variety ROC 25 was highly susceptible in the three replicate samples of newly-planted and ratoon cane grown in the eld in Kaiyuan and Lincang.The pokkah boeng resistance of these controls was stable and consistent.The pokkah boeng resistance of each of 60 new varieties was stable and relatively consistent, with 35 (58.33%) found to be highly resistant to moderately resistant, and 25 (41.67%) to be susceptible to highly susceptible.Among them, ve new varieties (8.33%), Yuegan 49, Funong 11-2907, Mintang 11-610, Mintang 12-1404, and Guitang 11-1076 were found to be highly resistant.Furthermore, 15

Discussion
Pokkah boeng disease is an important epidemic fungal disease that is prone to outbreaks.Screening and planting resistant varieties is the most economic and effective method that can be used to control this disease.As pokkah boeng disease has become an important disease that seriously affects the development of the sugarcane industry in China, disease-resistant sugarcane breeding strategies should be implemented in the future.Pokkah boeng disease resistance should be regarded as one of the main economic indices in variety breeding programs.The evaluation and screening of sugarcane varieties that are resistant to pokkah boeng disease should be strengthened.In addition, disease-resistant resources should be explored and utilized, the breeding of intermediate material should be improved, and parental combinations should be rationally matched.This will aid in the selection and breeding of stable resistant new varieties for popularization and application in sugarcane production.
Yunnan Province is an important distribution center of wild sugarcane resources in China and one of the global origins of wild sugarcane (Chen et al. 2001;Fan et al. 2001).Wild sugarcane resources are an importance source of resistance genes in modern sugarcane breeding.Some studies have shown that the wild germplasm resources preserved in the National Nursery of Sugarcane Germplasm Resources in China contain excellent diseaseresistance genes, which is promising for the breeding of disease-resistant sugarcane varieties (Huang et al. 2012;Li et al. 2013Li et al. , 2015Li et al. , 2017b;;Xu et al. 2014).The new and main cultivated varieties that displayed resistance to pokkah boeng in this study may be used for breeding resistant sugarcane varieties.These varieties can be used as a disease-resistance parent to hybridize with wild germplasm resources.The heritability of the disease resistance should then be analyzed and a disease-resistant germplasm gene bank may be established.This will allow further selection of new sugarcane varieties with pokkah boeng resistance for application in the sugarcane industry.
The main factors that induce pokkah boeng disease outbreaks are planting susceptible varieties in large areas, and high rainfall and humidity.The al. 2016, 2019; Hilton et al. 2017; Lin et al. 2014, 2015; Liu et al. 2019; Viswanathan et al. 2011; Vishwakarma et al. 2016).There have also some investigations and studies on the occurrence of the disease, variety resistance, and its control techniques (Feng 2000; Goswami et al. 2013; Luo et al. 2017; Long et al. 2019; Wang et al. 2017a, 2017b).Outbreaks of pokkah boeng disease are caused by several main factors including the planting of susceptible varieties across large areas, high temperatures, a wet climate, the accumulation of pathogens, and improper cultivation and management measures (Shan et al. 2018).
most economic and effective control measure against pokkah boeng disease is to select and plant resistance varieties(Huang et  al. 2018; Shan et al.

Figures Figure 1
Figures

Table 1
). Evaluation of eld resistance to pokkah boeng disease in new sugarcane varieties under natural infection conditions a The values in the table are the mean±SE.Different lowercase letters indicate signi cant differences among disease incidence of different varieties at 0.05 level.bHR:Highlyresistant; R: Resistant; MR: Moderately resistant; S: Susceptible; HS: Highly susceptibleField resistance of main cultivated varietiesThe resistance of the newly-planted and ratoon cane of each of the 31 main cultivated varieties, which from different sugarcane areas in the eld, was stable and relatively consistent.Of the 31 main cultivated varieties, 15 (48.39%) were highly resistant to moderately resistant and 16 (51.61%)were susceptible to highly susceptible.There were ve (16.13%) highly resistant main cultivated varieties, namely ROC 20, Yuetang 83-88, Chuantang 61-408, Yunyin 10, and Guitang 21.There were also ve (16.13%) resistant main cultivated varieties: Yuetang 79-177, Yunzhe

Table 2
Evaluation of eld resistance to pokkah boeng disease in main cultivated sugarcane varieties under natural infection conditions a The values in the table are the mean±SE.Different lowercase letters indicate signi cant differences among disease incidence of different varieties at 0.05 level.b HR: Highly resistant; R: Resistant; MR: Moderately resistant; S: Susceptible; HS: Highly susceptible