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甘蔗品种黑穗病抗性评价体系的建立
阙友雄, 许莉萍, 林剑伟, 陈天生, 陈如凯
, 李依龙 (福建农林大学
农业部甘蔗生理生态与遗传改良重点开放实验室,350002) 摘
要 为了建立甘蔗品种黑穗病抗性评价体系,选用9个引进品种,设计一个包括6个对照品种在内的田间试验。首次采用混合小种进行人工浸渍接种,通过整个新植蔗生长季病害进展曲线下的面积、茎感染率和株感染率3个病情指数,以及病害流行学参数潜伏侵染期和持续发病期的分析,对其抗病性进行评价,在分析以上参数相关程度的基础上,引进了系统聚类分析法进行进一步评价。结果显示:9个引进甘蔗品种中,ROC26属于感病品种,其余属于抗病或高抗品种;3个病情指数和持续发病期的两两相关均为显著正相关,潜伏侵染期与这些参数的相关为负相关,但未达到显著水平;在类间距离大于1.0的条件下,可将15个品种聚为6类,进一步明确了各品种抗黑穗病性的相似程度;抗性鉴定标准对照种NCo310、F134、NCo376和Ya71-374的应用,明确了接种源为小种1和小种2,通过一次接种试验,明确了供试品种对2个小种的抗性水平,标准对照种的抗性表现,还说明了本生长季发病条件基本是适宜的。试验结果表明,本文建立的甘蔗品种抗黑穗病评价体系是切实可行的。 关键词:甘蔗品种
黑穗病 抗性评价
相关系数 聚类分析 Establishment of
Evaluation System of Smut Resistance for Sugarcane Varieties
QUE
You-xiong,XU
Li-ping,LIN Jian-wei,CHEN Tian-sheng CHEN Ru-kai,LI
Yi-long (Key Lab of Sugarcane Eco-Physiology & Genetic Improvement, Ministry of Agriculture, Fujian Agriculture and Forestry University ,Fuzhou 350002) Abstract
In order to establish the evaluation system of
smut resistance for sugarcane varieties, nine introduced sugarcane
varieties were chosen and a field experiment including six other control
varieties was conducted. By means of manual dipping inoculation, smut
resistance for tested sugarcane varieties was evaluated on the basis of
analyses on three infected indexes of the area under disease progress
curve, infected rate of stalk, infected rate of plant and on two disease
epidemic parameters of latent infection period (LIP) and sustained
disease duration (SDD). Systemic cluster analyses based on correlation
coefficients of these parameters showed that ROC26 was sensitive to smut
infection and the other varieties were resistant in total of nine
introduced varieties. Positive significant correlation between three
infection indexes and SDD, and negative insignificant correlation
between infection indexes and LIP were found. Fifteen sugarcane
varieties could be clustered into 6 clusters on the basis of cluster
distance being more than 1.0. It resulted in more clear understanding of
smut resistance
in tested sugarcane varieties. It suggested that smut race 1 and race 2
were included in inocula and the condition for Ustilago
scitaminea infection was suitable in the crop season by setting up
the standard varieties of NCo310, F134, NCo376 and Ya71-374 for smut
identification.
In addition, the results of sugarcane varieties resistant to smut race 1 and race 2 were known by only a simple field trial. Form above, a feasible evaluation system for sugarcane smut resistance was established. Key words:
Sugarcane varieties;
Smut (Ustilago scitaminea Syd.);
Evaluation of resistance;
Correlation coefficient;
Cluster analysis Sugarcane smut incited by fungus of Ustilago scitaminea is a worldwide disease [1].
It infects sugarcane through the bud, then the mycelia permeate the bud
scale and twist the meristem zone until the elicitation of the smut whip
which contains thousands of chlamydospore. The chlamydospore then spread
into other buds and caused serious losses in yield and in sucrose
content. As one of the main disease in mainland China, several methods
are available to control smut disease, but resistance cultivars are the
most effective [2].
The accuracy and efficiency of resistance identification directly
influence the process of resistance breeding [3].
Although there are several methods being used in the evaluation of smut
resistance of sugarcane, such as structural characters of buds assay[4,5],
tissue invertase assay,chemical
assay[6], histological staining, and ultra-structural assay, only resistance
identification by artificial dip-inoculation and inspection of tested
material planted in field in full growth season is popular in the world[3].
Recently, marker-assisted selection (MAS) provides a new approach to
selection of crop target characters. But application of this technology
should be based on the establishment of technology platform containing
series of linkage markers, which we still haven’t established. So, up
to now, the measure of resistance identification of sugarcane varieties
is still the artificial inoculation and plant breeding [7].
In
this study, the resistance identification of the varieties introduced in
National 948 Project were performed so as to construct the evaluation
system of smut resistance for sugarcane varieties and at the same time
further provide the information about disease resistance of these tested
varieties. 1
MATERIALS AND METHODS 1.1
Tested materials 1.1.1
Sugarcane varieties Total
of 15 sugarcane(Saccharum
spp)varieties,
which contains five introduced in National 948 Project( HoCP91-555、HoCP93-746、HoCP93-750、CP85-1308、CP89-1509、RF93-244、
RB72-454、ROC25
and ROC26),four standard control varieties NCO376, NCO310, F173 and F134
,the national uniform control variety ROC10 and the local control
variety Mingtang70-611 were used as tested plant materials. 1.1.2
Tested inocula Teliospores of Ustilago scitaminea race 2 and race 1 both were
collected from Guangxi and Fujian, respectively, mingled together. They
were air-dried, sealed in plastic bags and stored at 4℃. Prior to use,
germination of the spores was checked [1, 3]. 1.2
METHODS 1.2.1
Inoculation and field design
Artificial inoculation and field breeding were conducted to identify the
resistance [3].
Teliospore suspension contained 5×106 spores/ml[3]
were prepared,
which was about 40 spores in the 60× microscope, note that in the
preparation process using some washing soda as surface activator which
could ruin the surface tension and prevent the spores from floating on
the surface. All the sugarcane stalks were cut into two-bud sets and
dip-inoculated by immersion in the prepared teliospore suspension for
ten minutes. Then the inoculated sets were incubated at 25℃ under
humid condition for 24 hours before being planted in field. Each one-row
plot was 3.5 m long with 1.2m space between plots within rows. Plots
were arranged in a randomized complete-block design with two
replications (plots). Thirty sets for every kind of material were
inoculated. There were twenty buds in every plot. 1.2.2
Data collection and statistical analysis After
the neat emergence of seedlings, check the emergence number of each plot
and begin the investigation of the infection situation when the first
whip-like sort appears. Data collections were made every 15 days until
whip-like sorts did not produce. The appearance date of the first whip,
the number of infected stalks (including the flank tiller bud), the
number of total stalks (including the flank tiller bud) and the number
of infected plants (clump) were recorded. In order to judge new whip,
every whip was labeled with cards on which the appearance date and the
sequence number of the plant and stalk were recorded. The final
cumulative infected stalks (Ymax) were used for the determination of the
resistance reaction type. After the standardization conversion of the
data in every index , systematical clustering of the tested materials
was performed by using UPGMA (unweighted pair
group method using arithmetic average).The five indexes used in
cluster were LIP(latent infection period),SDD(sustained disease
duration), IP(the cumulative percentage of infected plants),IS(the
cumulative percentage of infected stalks) and AUDPC(area under disease
progress curves)
[3].The
formula of AUDPC is as follows: AUDPC = (IS1 + IS2)/2
×(t2-t1), where IS1 and IS2 is
the adjacent investigated IS, respectively, while the t2 and
t1 are the adjacent investigation time. Both correlation and
cluster analyses were completed by Systemic Analysis Soft (SAS) 3.0.
Before cluster, standardization conversion of all data used was
conducted. The formula of range standardization value is as follows [3]:
1.2.3
Ratings of smut resistance According to the rating standard put forward by predecessor, the rating
of smut resistance is showed in Table 1. Table
1 Smut resistant ratings in sugarcane
2
RESULTS AND DISCUSSION 2.1
Correlation analysis of infection index Results
of correlation analysis among IP, IS and AUDPC were given in Table 2. It
showed that there existed significant positive correlations at 0.01
levels between every two of infection indexes of IP, IS and AUDPC, and
the same level of significant positive correlations between every
infection index and disease epidemic parameter of SDD was also found.
And the correlation coefficients between every two of AUDPC, IS, IP and
SDD were all positive. Although there existed negative correlations
between LIP and the infection index IS, IP and AUDPC. The correlation
coefficients were somewhat large, but they did not reach the significant
level of 0.05, so was the correlation coefficient between LIP and
disease epidemic parameters SDD. Table 2 Correlation coefficients between disease epidemic parameters and
infection indexes
Notes:
IP is the percentage of plant infected, IS is the percentage of stalk
infected, AUDPC is the area under the disease progress curves,
LIP is latent infection period, SDD sustained disease duration.
Significance at 0.05 and 0.01 levels is marked with * and **,
respectively. (r0.05=0.763,r0.01=0.840) 2.2
The
resistance reaction types of tested materials All
of the 15 sugarcane varieties tested were infected and seemed to have
different levels of infection. The corresponding results of infection
indexes and disease epidemic parameter were showed in Table 3. As
for infection indexes, the IP of Ya71-374 is maximal, while the maximal
value of IS and AUDPC existed in ROC26, so Ya71-374 and ROC26 were the
two most susceptible. According to the final cumulative IS, the
resistance reaction types of the tested 15 varieties were as follows: 5
high resistance, 4 resistance, 3 susceptibility and the surplus 3 are
middle resistance .There was no any tested variety exhibiting high
susceptibility. In the identification system of smut resistance, the
resistance reaction type of F134 and NCo310 was middlingly susceptible
and susceptible, respectively. The result accounted for that there
existed both Ustilago scitaminea race
1 and race 2 in the tested inocula.
Considering the conditions for Table
3 Performance of smut
resistance of tested 15 varieties
Ustilago
scitaminea infection
and the development of smut, the highly susceptible standard control
variety Ya71-374 only appeared to be middlingly susceptible, and the
susceptible standard variety F134 also only appeared to be middlingly
susceptible to race 2, the condition appeared not to be adequate
according to the resistance reaction type of the two tested varieties,
so the identification resistance reaction type should have been
overestimated. On the other hand, the resistance reaction type of
susceptible standard control variety NCo310 was accorded with
expectation, so the condition for infection and smut development was
adequate accordingly and the identification result was suitable.
According to the above consideration, the smut infection and development
condition of this growing season was basically adequate. 2.3
Cluster identification of smut resistance of tested materials When
using LIP、SDD、IP
and IS as cluster indexes, the tree map for UPGMA of 15 tested sugarcane
varieties was produced and showed in Figure 1.At the distance of 1.0,
the 15 tested varieties were clustered into six groups. According to the
cluster result, cluster 1 included 3 varieties,Ya71-374, NCo310and
ROC26. Cluster II included only one variety134 and cluster III included
the single variety ROC10.Cluster IV included 2 varieties Mingtan70-611
and CP89-1509.
Cluster V included 3 varieties, CP85-1308, HoCP93-746 and RB72-454, and
cluster VI included 5 varieties, NCo376, HoCP91-555, RF93-244, ROC25 and
HoCP93-750. The resistance of cluster I and cluster VI were the weakest
and strongest, respectively. The resistance character was gradually
increased from cluster I to cluster VI.
By
using LIP、SDD、IP
and AUDPC as cluster indexes, the cluster result of the 15 tested
sugarcane varieties was showed in Figure 2. Set the same classification
standard above, the 15 tested varieties were also clustered into six
groups. And the result is as follows: cluster 1 included 2
varieties,Ya71-374 and NCo310. Cluster II included 2 varieties ROC26 and
F134.Cluster III included only one variety ROC10.Cluster IV included 3
varieties Mingtan70-611 , CP89-1509
and RB72-454. Cluster V included 2 varieties, CP85-1308 and HoCP93-746.
Cluster VI included 5 varieties, NCo376, HoCP91-555, RF93-244, ROC25 and
HoCP93-750. The resistance of cluster I and cluster VI were the weakest
and strongest, respectively. From cluster I to cluster VI, the
resistance character was also gradually increased.
图2
15个品种的聚类树状图 3
DISCUSSION Generally,
the identification of resistance reaction types of sugarcane to Ustilago
scitaminea smut is based on the final cumulative IS all through the
growing season [3].
For two reasons, setting the standard control varieties is indispensable
to the artificial inoculation and field breeding. Firstly, because the
resistant expression maybe influence by external condition, the
identification results of standard control varieties are helpful to
determine whether the condition for smut infection and development is
adequate or not. Secondly, the existence of standard control varieties
is good for the verdict of Ustilago
scitaminea smut race. Nowadays, in mainland China,there exist race 1
and race 2 in the sugarcane planting area at the same time, so
application of only one smut race can’t assess completely the
resistance of sugarcane genotypes to the both races. For the first time,
the commingling races were used for inocula in this study to evaluate
the resistance of tested materials to both races at one time, and the
identification efficiency was improved. The
introduction target of cluster analysis is to better understand the
similarity of relevant resistance parameters and to demonstrate the
resistance dissimilarity of various varieties. The aim has been achieved
by clustering with four parameters LIP,SDD,IP and IS, which produced
basically similar results for using other four parameters LIP、SDD、IP
and AUDPC. AUDPC came from
the increasing value of IS, and was a dynamic parameter that involved
the whole process of disease infection and development. Moreover, AUDPC
can reflect the total disease epidemic volume of the whole growing
season. Despite of that, the substitution of AUDPC for IS in cluster
reduced the homologous results. From
above, it was known that the condition for smut infection and
development was adequate in the present study, so the research results
objectively reflected the resistance levels of various genotypes. Due to
this point,ROC26
was sensitive to smut infection and the other varieties were resistant
in total of nine introduced varieties. ROC26 was introduced through folk
channel and recommended by predominant sugarcane breeding regions
because of its new code. Through the execution of National 948 project,
ROC26 was exchanged and resistance was identified. The resistance
identification result of ROC26 is far from optimistic, so high attention
should be paid for its further use in production.
In a word, with the establishment of the evaluation system of
smut resistance for sugarcane varieties, resistance identification for
smut will be more convenient and economical. References 1.
Que Y X(阙友雄), Xu L P(许莉萍),Zou T T(邹添堂) et al.Primary analysis of molecular diversity in populations of
the fungus Ustilago scitaminea
Syd..Journal of Agricultural
Biotechnology(农业生物技术学报),2004,12(6):685~689 2.
Xu L
P(许莉萍),Chen
R K(陈如凯).
Current status and prospects of smut and smut resistance breeding in
sugarcane.Fujian J.of Agri.Sci.(福建农业学报),
2000, 15 (2): 26~31 3.
Xu L
P(许莉萍),Chen
R K(陈如凯),Chen
P H(陈平华).Analysis
on infection index of smut caused by Ustilago scitaminea in sugarcane
segregated population.Chinese Jounal of Tropical Crops(热带作物学报),2004,25(3):33~36. 4.
Gong
D M(龚得明),Lin
Y Q(林彦铨),Chen
R K(陈如凯).
Study on the breeding techniques for smut resistance in sugarcaneⅢ.Relation
between bud characteristics and resistance to smut.Acta Agronomica Sinica(作物学报),1996,22(3):362~264 5.
Padmanaban
P, Alexander K C, Shanmugam N. Studies on certain characters associated
with smut resistance. Indian Phytopath. 1988, 41:594~598. 6.
Lloyd
H L, Naidoo G. Chemical assay potentially suitable for determination of
smut resistance of sugarcane cultivars. Plant Dis., 1983, 67:1103~1105 7.
Chao
C P, Hoy J W, Saxton A M and Martin F A
Heritability of resistance
and repeatability of clone reactions to sugarcane smut in Louisiana[J].
Phytopath , 1990, 80 (7): 622~626 |