| Soybeans As a high-yield and high-protein forage crop,alfalfa(Medicago sativa L.)is widely cultivated around the world.Alfalfa has strong adaptability and high nutritional value,so that it was known as "the king of forages".Alfalfa root rot is an important soil-borne disease with a wide variety of pathogens,which seriously affected the establishment of alfalfa,decreased of grass quality and yield,shorted the life of alfalfa grassland,and declined of grassland in advance that would caused severely losses to production.It is clear that the species and distribution of pathogen of alfalfa root rot in Heilongjiang Province were the premise and foundation for prevention and control of this disease.Therefore,this paper collected alfalfa root rot samples from the main alfalfa-growing areas in Heilongjiang Province,and their pathogens were isolated and identified.Based on these,sensitivity of fungicide and genetic diversity of the dominant strains causing alfalfa root rot were analyzed.The results will provide important theoretical basis for the control of alfalfa root rot and breeding disease-resistant varieties in Heilongjiang Province.Diseased plants of alfalfa root rot were collected from 6 locations of alfalfa cultivation area in Heilongjiang Province,included Xiangfang,Shuangcheng,Acheng,Lanxi,Daqing and Zhaodong District.The isolates isolated by tissue separation method,were identified by morphological characteristics,molecular biology identification and phylogenetic tree analysis.A total of 480 fungal strains were isolated from 6 areas in main alfalfa planting of Heilongjiang Province,and comprised Fusarium tricinctum 112 isolates(23.3%),F.oxysporum 67 isolates(14%),F.acuminatum 92 isolates(19.2%),F.solani 59 isolates(12.3%),and F.equiseti 45 isolates(9.4%),Phoma Medicaginis 36 isolates(7.5%),Plectosphaerella cucumerina 21 isolates(4.4%),Alternaria alternata 29 isolates(6.0%),and Chaetomium.qlobosum 19 isolates(3.9%),respectively.F.tricinctum causing alfafa root rot was dominant strains in Heilongjiang Province.On the premise of ascertaining dominant strains,the pathogenicity and sensitivity of dominant strains to carbendazim was evaluated.The results showed that 12/73 F.tricinctum isolates had strong pathogenicity,accounted for 16.4%;59/73 isolates had medium-pathogenicity,accounted for 80.8%;2/73 isolates had weak pathogenicity,accounted for 2.7%.In other words,the most of dominant population of F.tricinctum in Heilongjiang Province had moderate or higher pathogenicity with less weak pathogenic isolates.And there no sensitive strains were found at the effective dose of 1 ?g/ml and 10 ?g/ml of carbendazim,which indicated that the dominant pathogen populations of alfalfa in Heilongjiang Province had completely developed resistance to carbendazim,it was not suitable for the control of alfalfa root rot in this area.Amplified Fragment Length Polymorphism(AFLP)analysis was used to evaluate genetic diversity of 73 single spore isolates of F.tricinctum collected from 6 alfalfa growing areas of Heilongjiang Province.The results showed that 28 pairs of primers had stable amplification,good repeatability,high polymorphism and strong resolution.Fragment sizes ranged from 100 to 2000 bp.Using UPGMA cluster analysis,73 F.tricinctum isolates from different regions could be divided into 8 groups with similarity coefficient 0.66.Group I contained 2 isolates from Xiangfang population.There were 32 isolates in Group II,which were further divided into 3 subgroups,subgroup I contained 8 isolates from Xiangfang,subgroup II contained 11 isolates from Shuangcheng,and subgroup III contained 13 isolates from Acheng;Group III contained 7 isolates from Lanxi.Group IV contained 7 isolates from Daqing population.Group V contained 16 isolates and further divided into 4 subgroups,which subgroup I and subgroup II contained 7 isolates from Daqing population,subgroup III and IV contained 9 isolates from Zhaodong population.Group VI ang VII contained 6 isolates from Lanxi.Group VIII contained 3 isolates from Zhaodong.Analysis of molecular variance(AMOVA)indicated that there was significant correlation between the F.tricinctum population and geographical location or pathogenicity.There was moderate genetic differentiation among different population of geographical location or pathogenicity with 88.94% and 93.27% attributing to within the population,respectively.And there was greater genetic differentiation associated within the population.The results indicated that the geographical locations and pathogenicity of F.tricinctum population had a significant effect on the population differentiation of dominant strains of alfalfa root rot in Heilongjiang province. |
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