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Sugarcane white leaf (SCWL) is a phytoplasma disease that has spread rapidly in Yunnan province, China, in recent years. It is now the main barrier to the development of the sugarcane industry in this region. Herein, we explored the insect vector and their dominant population of SCWL phytoplasma at Gengma in Lincang city in 2019, where the occurrence of SCWL is the most serious in Yunnan Province. Two leafhopper species, Tettigoniella viridis (Linnaeus) and Clovia conifer Walker, were tested positive for SCWL phytoplasma by using nested PCR, indicating that they are maybe the potential insect vectors of SCWL phytoplasma. Additionally, nymphs of T. viridis were tested strongly positive, and they were preliminarily identified as the dominant population. Since the main transmission medias of SCWL phytoplasma appears to be SCWL-infected seedcane and the leafhopper insect vectors, establishing a disease-free and healthy seedcane breeding base and eliminating the leafhopper insect vectors in sugarcane planting areas may prevent or control the spread of SCWL and reduce the natural transmission rate in the field. The results expand our theoretical and technical knowledge of sugarcane phytoplasma disease and provide a reference for formulating comprehensive control of SCWL.

Brown stripe is a major fungal disease damaging the leaves of sugarcane. Breeding and cultivating resistant varieties is the most economical and effective method of controlling the disease. The paper reports on the evaluation of sugarcane brown stripe resistance in novel varieties bred by the China Sugarcane System and major varieties under cultivation in the sugarcane cultivating region. Field resistance against brown stripe was identified in 60 novel varieties and in 31 major varieties under cultivation in Lincang, Puer, and Yuxi, in Yunnan Province, and in Yizhou, Guangxi Province, where the incidence and severity of brown stripe is particularly high. Field surveys showed that 32 (53.3%) of the 60 novel varieties were highly resistant to moderately resistant, and 28 (46.7%) were susceptible to highly susceptible. In addition 21 (67.7%) out of the 31 major cultivated varieties were highly resistant to moderately resistant, and 10 (32.3%) were susceptible to highly susceptible. We recommend the intensifying of efforts to eliminate the major varieties that are susceptible, and promotion of the adoption of elite resistant varieties in wet and rainy sugarcane cultivating regions with high incidence and severity of sugarcane brown stripe disease, to effectively manage the brown stripe epidemic.

Sugarcane brown rust, caused by Puccinia melanocephala, is one of the main diseases of sugarcane in China. The identification and discovery of new resistance genes have important theoretical and practical significance for preventing outbreaks of brown rust and ensuring the sustainable production of sugarcane. To screen for polymorphic simple-sequence repeat (SSR) molecular markers for localization of brown rust resistance genes, we used two populations that are suitable for genetic linkage map construction and mapping of new resistance genes to construct resistant and susceptible genetic pools. We then screened 449 pairs of primers to identify polymorphic SSR markers in the parental lines and the resistant/susceptible genetic pools. The results showed that 25 pairs of primers directed amplification of polymorphic DNA fragments between the parents of the cross combination ‘Yuetang 03-393’ × ‘ROC 24’, and 16 pairs of primers amplified polymorphic fragments between the parents of the cross combination ‘Liucheng 03-1137’ × ‘Dezhe 93-88’. Four pairs of primers (SMC236CG, SCESSR0928, SCESSR0636 and SCESSR2551) amplified polymorphic DNA fragments between the parental lines and the resistant/susceptible genetic pools in ‘Yuetang 03-393’ × ‘ROC 24’. The results of this study will establish a solid foundation for the mapping of new brown rust resistance genes, genetic linkage map construction and the development of closely-associated molecular markers in sugarcane.

The aim of this study was to assess brown rust resistance of new sugarcane varieties bred in China and main cultivated varieties in sugarcane planting area, and screen the elite new brown rust resistant varieties for popularization and application in production. In total, 60 new varieties in the national regional test of new sugarcane varieties were tested in Kaiyuan and Lincang, and 34 main cultivated varieties were studied in Lincang, Puer, and Yuxi, Yunnan province, and Yizhou, Guangxi province, China, where the incidence of brown rust was particularly high. The resistance of these sugarcane varieties to brown rust was investigated under natural inoculation and molecular marker-assisted identification was used to detect the brown rust resistance gene Bru1 . The results of field survey showed that 66 (70.21%) of the 94 new and main cultivated varieties were highly resistant to moderately resistant, and 28 (29.79%) were susceptible to highly susceptible. Molecular detection indicated that Bru1 gene was found among 54 (57.45%) of the 94 new and main cultivated varieties. Some main cultivated varieties that were currently planted across large areas such as Guitang 29, Guitang 44, Dezhe 03-83, Liucheng 03-1137, Yuetang 60, and Guitang 46 were highly susceptible to brown rust, and 31 new varieties such as Yuegan 48, Funong 09-2201, Guitang 08-120, Liucheng 09-15, Zhongzhe 1, Yunzhe 08-1609, Yunrui 10-187, and Zhongtang 1201 were resistant. Therefore, in the sugarcane planting areas with high incidence of brown rust and wet and rainy climates, more effort should be eliminated the main susceptible varieties and promoted the application of new resistant varieties. This will help to achieve a reasonable distribution of varieties, fundamentally control the outbreak of disease in sugarcane planting areas, and provide security for the high-quality development of sugarcane industry in China in the future.

The aim of this study was to assess brown rust resistance in 60 new sugarcane varieties bred by the China Sugarcane System in recent years and in 34 major varieties cultivated in sugarcane growing areas in China. The resistance of these sugarcane varieties to brown rust was investigated in the field, and molecular markers were used to detect the brown rust-resistance gene Bru1. The results of the field survey showed that 66 (70.21%) of the 94 sugarcane varieties were highly resistant to moderately resistant, and 28 (29.79%) were susceptible to highly susceptible. The Bru1 gene was detected in 54 (57.45%) of the 94 sugarcane varieties. Seven highly resistant varieties and five resistant varieties did not carry Bru1, suggesting that they carry other genes associated with brown rust resistance. This study provides a scientific basis and identifies disease-resistant germplasm for selection of plants for sugarcane production.

We assessed inheritance of resistance to sugarcane brown rust (Puccinia melanocephala) in selfing F1 populations of wild sugarcane germplasm Erianthus rockii ‘Yundian 95-19’ and E. rockii ‘Yundian 95-20’. We tested parent and selfing F1 individuals for the brown rust resistance gene, Bru1, that has been shown to confer resistance to brown rust in sugarcane. The Bru1 gene was not detected in E. rockii ‘Yundian 95-19’, E. rockii ‘Yundian 95-20’ or their selfing F1 individuals, and we found there was segregation of resistance in the two selfing F1 populations (segregation ratio: 3:1). The results confirmed resistance in E. rockii ‘Yundian 95-19’ and E. rockii ‘Yundian 95-20’ to sugarcane brown rust is controlled by a novel, single dominant gene.

Sugarcane leaf scald caused by Xanthomonas albilineans (Ashby) Dowson is an extremely destructive disease. In 2017, suspected symptoms of sugarcane leaf scald was observed, in Baoshan, Mengding and Jinping, Yunnan Province, China. The bacteria was isolated from the symptomatic stalks and was subsequently identified as the causal agent using Koch’s postulates. BLASTN analysis showed that the amplified nucleotide sequences of 16S rRNA, gyrB and rpoD genes from all isolates shared 100% sequence identity with the nucleotide sequences of corresponding genes of X. albilineans strain GPE PC73 (Acc no. FP565176). Based on symptom diagnosis in the field, colony morphology, validation of Koch’s postulates and molecular identification of 16S rRNA, gyrB and rpoD genes, the disease was confirmed as sugarcane leaf scald caused by X. albilineans. Partial 16S rRNA, gyrB and rpoD gene sequence analysis of 36 X. albilineans strains from Yunnan and Guangxi provinces, China, was determined. The results revealed that the gyrB gene was the most sensitive in differentiating X. albilineans strains from Yunnan and Guangxi provinces.

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 find new elite pokkah boeng resistant varieties for application in sugarcane production. The natural field resistance to pokkah boeng disease of 60 new varieties was evaluated in regionalized experiment. Meanwhile, the field 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.

Sugarcane (Saccharum officinarum L.) is the main sugar crop in China. Yunnan is the second largest sugarcane production province in China. In December 2018, leaf blight was first observed on almost every leaf of sugarcane on 'Huanan 54-11', 'Baimei' and 'Chongan' in Kaiyuan (103°27' E, 23°72' N), Yunnan. In October 2019, during our survey in the field in Lingcang (100°08' E, 23°88' N), Yunnan, this disease was also observed on 'ROC 25'. Symptoms of the disease initially appeared as wilted, which seemed to be cause by water stress. As the disease progressed, irregular straw-yellow and blighted lesion ran throughout the leaf lamina from leaf tip to entire leaf sheath, many small black conidia formed in the dead leaf tissue under humid conditions. Symptomatic leaf tissues were surface-sterilized with 70% ethanol for 30 s, 0.1% HgCl2 for 1 min, and rinsed with sterilized water three times, air dried on sterile filter paper, and plated on potato dextrose agar (PDA). Six isolates were obtained from six symptomatic leaf samples and were transferred onto potato carrot agar (PCA). Colonies on PDA were white with loose aerial hyphae at first, then turned to dark olive or dark. Colonies on PCA were grayish with sparse hyphae, then turned to dark gray. Conidiophores were brown, simple or branched, and produced numerous conidia in short chains. Conidia (n = 50) were obclavate to obpyriform or ellipsoid, brown to dark brown, with a cylindrical short beak at the tip (2.3 to 17.3 µm in length), and 15.3 to 46.6 μm × 4.2 to 17.9 μm, 2 to 7 transverse septa and 0 to 3 longitudinal septa. Morphologically, the isolates were identified as Alternaria tenuissima (Simmons 2007). Two representative isolates C4 and C5 were selected for molecular identification. The internal transcribed spacers (ITS), Histone 3 genes and plasma membrane ATPase were amplified with primer pairs ITS1/ITS4, H3-1a/H3-1b and ATPDF1/ATPDR1, respectively (Glass et al. 1995; Lawrence et al. 2013). The sequences were deposited in GenBank (ITS, MT679707-MT679708; Histone 3, MT710929-MT710930; ATPase, MT833928-MT833929). BLAST searches showed ≥99% nucleotide identity to the sequence of A. tenuissima (ITS, 100% to MN822571; Histone 3, 100% to MN481955; ATPase, 99% to JQ671875, 100% to MH492703, respectively). Thus, the fungus was identified as A. tenuissima based on morphological and molecular characteristics. For pathogenicity tests, five healthy 2-month-old potted sugarcane leaves were wounded with one sterile needle and inoculated with 20 μl of suspension of 106 conidia/ mL, and five plants were inoculated with distilled water as the controls. Plants were placed in a greenhouse at 25 to 35°C. After two months, the leaf wound inoculated with the putative pathogen displayed blighted as those observed in the field whereas the controls remained symptomless. The fungus was reisolated from symptomatic leaves with the same morphological and molecular traits as the original isolates. The fungus was not isolated from the control plants. Pathogenicity tests were repeated two times. A. tenuissima causing leaf blight on barley in China was reported in 2008 (Luo et al. 2008). Leaf spot disease of sugarcane caused by A. tenuis has been recorded in Maharashtra (Patil et al. 1974). To our knowledge, this is the first report on A. tenuissima affecting leaf blight on sugarcane in Yunnan Province, China. Identification of the causes of the disease is important to develop effective disease management strategies. The author(s) declare no conflict of interest. Funding: This research was supported by Sugar Crop Research System (CARS-170303), the Yunling Industry and Technology Leading Talent Training Program "Prevention and Control of Sugarcane Pests" (2018LJRC56), and the Yunnan Province Agriculture Research System. References: Glass, N. L., et al. 1995. Appl. Environ. Microbiol. 61:1323. Lawrence, D. P., et al. 2013. Mycologia 105:530. Luo, Z., et al. 2008. Acta Phytophy. Sin. 35(5): 469-470. Patil, A.O., et al. 1974. Res. J. Mahatma Phule Agric. Univ. 5(2): 122-123. Simmons, E. G. 2007. Alternaria: An Identification Manual. CBS Fungal Biodiversity Centre, Utrecht, The Netherlands. Caption for supplementary Figure 1 Supplementary Figure S1. Disease symptoms of sugarcane leaf blight disease and morphological characteristics of Alternaria tenuissima. (A) Typical straw-yellow and blighted lesions on naturally-infected leaves of sugarcane; (B) Infected symptoms on wounded leaves of sugarcane two months after artificial infection with A. tenuissima; (C) Colony of A. tenuissima on PDA; (D) Colony of A. tenuissima on PCA; and (E-F) Sporulation and conidia of A. tenuissima on PCA. (Scale bars = 100 μm; 20 μm).

Sugarcane white leaf (SCWL) is a devastating sugarcane (Saccharum officinarum) disease caused by a 16SrXI group phytoplasma, which is extremely harmful to sugarcane production. To determine the occurrence of SCWL in different varieties in 2018, we conducted a field survey and performed nested PCR detection of SCWL phytoplasma in cane-planting areas of Mangweng and Hepai in Gengma, Yunnan province, which are the areas most severely affected by SCWL in China. The results of the field survey showed that the symptomatic incidence of SCWL differed among varieties. The mean symptomatic incidence of SCWL on variety Yuetang60 was the highest (73.50%), and it was the lowest on Liucheng05-136 (13.67%). Using nested PCR, the SCWL phytoplasma was detected in symptomatic plants of all varieties more than 90% of the time; the SCWL phytoplasma was detected in 91 and 97% of symptomatic plants of Yingyu91-59 and Liucheng05-136 varieties, respectively. The SCWL phytoplasma was detected by PCR in 82% of the asymptomatic plant samples. The results of this study showed that field survey based on white leaf symptoms did not accurately reflect the actual occurrence of the SCWL phytoplasma.

Sugarcane mosaic disease is one of the most serious and prevalent viral diseases of sugarcane in China. Resistant varieties are the most economical and effective measures for controlling this disease. Resistance against Sugarcane streak mosaic virus (SCSMV) and Sorghum mosaic virus (SrMV) in 41 excellent sugarcane innovation germplasms and their parents (Saccharum officinarum L. “Ludashi” × Erianthus rockii Keng “Yundian 95-19”) was checked by using a stalk inoculation and RT-PCR detection during 2015 and 2016. Results indicated that among the 41 excellent sugarcane innovation germplasms and parents, 23 were highly (Grade 1) to moderately resistant (Grade 3) to SCSMV, meanwhile, 31 were highly (Grade 1) to moderately resistant (Grade 3) to SrMV. Ten germplasms were highly resistant (Grade 1) to resistant (Grade 2) to both SCSMV and SrMV. Among these, six germplasms (Yun 09-604, Yun 09-607, Yun 09-619, Yun 09-633, Yun 09-656, Yundian 95-19) were highly resistant (Grade 1) to both SCSMV and SrMV, accounting for 13.95% of the total germplasm materials. These results may provide resistance resource available for the breeding program of sugarcane cultivars against the major vrius associated with mosaic disease in China.

Field resistance against sugarcane thrips (Fulmekiola serratus) in 21 sugarcane varieties was tested following inoculation with a natural population. The results revealed differences in pest incidence levels in different varieties and between different leaf positions in the same variety. The tested varieties were subsequently categorized into four levels of resistance (Grades 1–4). The variety GT31 and LC05-136 showed the highest resistance (Grade 1), with a pest index of less than 0.35, while 12 varieties including FN91-21 and FN39 had a pest index of 0.35–0.45 (Grade 2). Five varieties including YT60 and YZ03-194 had a pest index of 0.45–0.55 (Grade 3), while FN38 and DZ03-83 had the lowest resistance (Grade 4), with a pest index of greater than 0.65. Correlation analysis further revealed that the number of thrips present on a single plant was related to the level of resistance; however, the correlation was relatively weak. These findings provide a preliminary evaluation of thrips resistance screening in sugarcane, laying the foundation for analysis of thrips-resistant genes, breeding of thrips-resistant varieties and resistance evaluation in new sugarcane germplasms.

Sugarcane streak mosaic virus (SCSMV) is one of the most important pathogens associated with sugarcane mosaic disease. In recent years, the rapid spread of SCSMV in Yunnan province, China, has raised concerns. In this study, we analyzed 110 sugarcane samples with mosaic symptoms taken from main growing areas in Yunnan province. RT-PCR detection results showed that 94% samples were positive for SCSMV. We subsequently analyzed the genetic diversity and population structure of SCSMV based on the coat protein (CP) sequences of 48 isolates. The overall mean value of nucleotide diversity for all isolates was 0.01127 ± 0.00082. Phylogenetic analysis based on CP gene sequences showed that all the Chinese isolates were clustered into one group, showing a distinct geographic feature. Analysis of population structure showed that the population from this study is not genetically different from that in China, but is genetically different from populations from other countries. Our results suggest that SCSMV has become the main pathogen of sugarcane mosaic disease in Yunnan province, China and that the recent rapid spread of SCSMV in Yunnan is not due to the invasion of new SCSMV populations.

Sugarcane mosaic disease is one of the most serious and prevalent viral diseases of sugarcane in China. The use of resistant varieties is the most economical and effective measures for controlling this disease. Sugarcane streak mosaic virus (SCSMV) and Sorghum mosaic virus (SrMV) are the two predominant pathogens of mosaic disease in the cane-growing regions of China. In 2015, 2016, double resistance to SCSMV and SrMV was identified once a year using a combination of artificial stalk-cutting inoculation and RT-PCR detection in 71 new elite sugarcane varieties/clones. Of the 71 new elite sugarcane varieties/clones, 24 were highly (Grade 1) to moderately resistant (Grade 3), and 47 were susceptible (Grade 4) to highly susceptible (Grade 5) to SCSMV. Furthermore, 27 were highly (Grade 1) to moderately resistant (Grade 3), and 44 were susceptible (Grade 4) to highly susceptible (Grade 5) to SrMV. Fifteen new elite sugarcane varieties/clones were highly resistant (Grade 1) to resistant (Grade 2) to SCSMV and SrMV. While, five new elite sugarcane varieties/clones (Yuegan 34, Yuetang 55, Yunzhe 03–258, Yunzhe 05–51, Yunzhe 06–80) were highly resistant (Grade 1) to SCSMV and SrMV. We defined the resistance of 71 new elite sugarcane varieties/clones to the two main pathogens of mosaic disease and identified 15 resistant to both SCSMV and SrMV. These results provide an elite resistance resource for effective prevention and control of mosaic disease and may serve as a reference for commercial varieties.

To confirm the phytoplasma association in 13 sugarcane white leaf (SCWL) symptomatic samples collected from Puer, China, nested PCR assays with phytoplasma universal primer pairs was performed. A ~ 1240 bp product was obtained from 10 out of 13 symptomatic samples, and 16S rRNA gene sequencing results showed that all sequences of SCWL isolates were 100% identical to each other. BLASTn analysis revealed 100% sequence similarity with the 16S rRNA genes from SCWL phytoplasma strains LC7 and LC9 from Lincang, Yunnan province, China belonging to 16SrXI-B subgroup. Virtual restriction fragment length polymorphism also confirmed that Puer SCWL strains were member of the 16SrXI-B subgroup.

Brown rust, caused by Puccinia melanocephala has become a serious problem for sugarcane production in China. Therefore, 50 new elite varieties bred by the Chinese Sugarcane System were screened for brown rust resistance and the presence of a major resistance gene, Bru 1. Identification of field resistance and molecular detection of the brown rust resistance gene Bru 1 in the 50 new elite varieties and 2 main cultivated varieties were carried out in 2014 and 2015 at regional experimental stations of the China Sugarcane System in Dehong and Baoshan, Yunnan Province, where the incidence of brown rust is particularly high. Resistance evaluations based on ratings of symptom severity resulting from field infection showed that 32 (64%) of the 50 new elite varieties were highly resistant (n = 13, 26%), resistant (n = 16, 32%) and moderately resistant (n = 3, 6%). Molecular detection identified Bru 1 in 27 (54%) of these resistant varieties suggesting that brown rust resistance in these new elite varieties is primarily controlled by Bru 1. The absence of Bru 1 in the remaining five resistant suggested the presence of additional brown rust resistance genes. Two main cultivated varieties were highly resistant and Bru 1 - positive varieties. Different series differed in the frequencies of resistant and Bru 1 - positive varieties. Yunzhe series varieties had the highest frequency of resistant varieties (87.5%) and the highest frequency of Bru 1-positive varieties (81.3%); In contrast, the Yuetang series varieties had the lowest frequency of resistant varieties (40%) and the lowest frequency of Bru 1-positive varieties (30%). In this study, we defined the field resistance of 50 new elite varieties to brown rust, and determined the distribution of brown rust resistant gene Bru 1 in different series varieties. These results will provide new options for commercial production with effective resistance to brown rust. Moreover, these new elite resistant varieties can be used for subsequent breeding and selection to produce agronomically superior varieties with effective brown rust resistance.

Brown rust caused by Puccinia melanocephala H. Sydow & P. Sydo is an important sugarcane disease that severely threatens sugarcane production in China. Breeding and planting resistant cultivars is the most cost-effective and efficient strategy for the control the disease. In addition, exploration, research, and utilization of resistant genes are the basis for resistance breeding. In the present study, four sugarcane varieties without the Bru1 gene and highly resistant to brown rust were used as male parents, and four varieties highly susceptible to brown rust were used as the female parents to configure hybrids. Six F1 hybrid populations were subjected to authenticity determination using SSR markers, brown rust resistance phenotype identification using artificial inoculation, and molecular detection of Bru1 gene. The results showed that the genetic segregation ratios in F1 hybrid populations of Yuetang 03-393 (HS) × ROC24 (HR) and Liucheng 03-1137 (HS) × Dezhe 93-88 (HR) were 3R:1S and 1R:3S, respectively, and Bru1 gene was not detected in their F1 hybrids. The results indicated that two dominant novel genes controlled brown rust resistance in Yuetang 03-393 (HS) × ROC24 (HR) and two recessive novel genes controlled brown rust resistance in Liucheng 03-1137 (HS) × Dezhe 93-88 (HR). Two genetically segregating populations that could be used for the localization of novel brown rust resistance genes were developed in the present study, and could lay a suitable foundation and provide theoretical reference for the analysis of the genetic mechanism of brown rust resistance, genetic map construction, mapping of novel resistance genes and the development of closely associated molecular markers in the future.

Leaf scald caused by Xanthomonas albilineans (Ashby) Dowson is a destructive disease of sugarcane, as it causes severe economic losses (Rott et al. 1995). X. albilineans has been reported in over 60 countries in Africa, Asia, and North and South Americas (Rott et al. 2000), but has so far not been reported in mainland China. X. albilineans is a quarantine pathogen for sugarcane germplasm imported into China. In June 2016, symptoms of leaf scald were observed on the newly-bred sugarcane cultivars ‘Guitang46’ and ‘Guitang06-2081’ in Beihai, Laibin and Baise, Guangxi province, China. The average disease incidence of cultivars ‘Guitang46’ and ‘Guitang06-2081’ was 44.6% and 50.1%, respectively. Typical symptoms of the disease were white stripes along veins. Severely infected plants also exhibited shorter stalk internodes, smaller leaves, and abnormal development of side shoots along the stalk. Leaves of side shoots had similar white stripes as those observed at the top of the stalk. A sudden wilting of all lea...