Study On Pathogen Identification And Control Of Dry Rot In Newly Planted Apple Saplings

The widespread phenomenon of death in newly planted apple seedlings,especially in northern Shaanxi and eastern Gansu,severely limits the development of the apple industry.A stem disease similar to apple canker was found on the dead seedlings.By analyzing the symptoms and morphological characteristics of the suspected pathogenic bacteria,the disease was preliminarily identified as apple canker.To clarify the type of pathogen causing apple canker and further explore effective prevention and control measures,we conducted a systematic study on the pathogenic bacteria and made preliminary progress in prevention and control.The following research achievements were obtained:(1)The disease and pathogenic species causing death of newly planted apple saplings in Shaanxi region have been identified.A total of 168 suspected samples of cankered branches were collected from multiple areas in Shaanxi,and 126 fungi were isolated using tissue separation method.Based on morphology and ITS r DNA phylogenetic analysis,the isolated fungi belonged to five groups: Diaporthe sp.,Botryosphaeria sp.,Alternaria sp.,Fusarium sp.,Trichothecium sp.,and Diplodia sp.,accounting for 37.84%,21.62%,15.32%,18.02%,5.41%,and 1.80% of the isolates,respectively.Diaporthe sp.was the dominant strain.(2)Pathogenicity testing results showed that Alternaria sp.,Fusarium sp.,Trichothecium sp.,and Diplodia sp.could not infect apple branches,while Diaporthe eres and Diaporthe sojae of the Diaporthe genus,Botryosphaeria dothidea and Botryosphaeria kuwatsukai of the Botryosphaeria genus could infect both apple branches and fruit,and were confirmed as the pathogenic fungi of apple dry rot.(3)Multi-gene(ITS-EF1-α-HIS-TUB2-CAL)phylogenetic analysis of Diaporthe sp.and Botryosphaeria sp.showed that the isolated fungi were Diaporthe eres,Diaporthe sojae,Botryosphaeria dothidea,and Botryosphaeria kuwatsukai.(4)There are interspecies differences in the biological characteristics of four pathogenic bacteria.Among them,the D.eres strain has the fastest hyphal growth rate under dark conditions and the slowest under light conditions;the D.sojae strain has the fastest hyphal growth rate under 12 h light-dark alternation culture conditions and the slowest under dark conditions;the B.kuwatsukai strain has the fastest growth rate under light conditions and the slowest under dark conditions;and the B.dothidea strain has the fastest growth rate under 12 h light-dark alternation culture conditions and the slowest under dark conditions.PDA culture medium is the most suitable medium for the growth of the four pathogens,and the hyphal growth rate is slowest in apple bark decoction medium.The four pathogenic strains can grow at temperatures ranging from10℃ to 35℃,with D.eres and B.dothidea having the optimum growth temperature of25°C,and B.kuwatsukai and D.sojae having the optimum growth temperature of 30°C.The hyphae of the fungus Phlyctema vagabunda cannot grow normally above 35℃,while the hyphae of the grapevine trunk rot fungus can grow normally above 35℃.The lethal temperature for hyphae of D.eres,B.dothidea,and B.kuwatsukai is 55℃,while that of D.sojae is 50℃.D.eres,D.sojae,and B.dothidea have the fastest hyphal growth rate when yeast extract powder is used as a nitrogen source,while B.kuwatsukai has the fastest hyphal growth rate when sodium nitrate is used as a nitrogen source.The growth rate of the four pathogenic bacteria is slowest when urea is used as a nitrogen source.The four pathogenic bacteria have the fastest growth rate when glucose is used as a carbon source,while D.eres and D.sojae have the slowest growth rate when the carbon source is soluble starch,and B.dothidea and B.kuwatsukai have the slowest growth rate when the carbon source is lactose or maltose.(5)The sensitivity of four pathogens to several commonly used chemical agents was revealed,and the activity of four pesticides against apple dry rot pathogens was determined using a growth rate method indoors,and a toxicity regression equation was established.The EC50 values of the tested agents pyraclostrobin,azoxystrobin,fluazinam,and tebuconazole for D.eres were 0.4766 μg/m L,4.2304 μg/m L,5.6250μg/m L,and 4.8285 μg/m L,respectively;for D.sojae,the EC50 values were 1.1063μg/m L,8.9895 μg/m L,0.2721 μg/m L,and 7.5363 μg/m L,respectively;for B.kuwatsukai,the EC50 values were 0.9967 μg/m L,0.4763 μg/m L,0.8128 μg/m L,and7.6413 μg/m L,respectively;for B.dothidea,the EC50 values were 11.1022 μg/m L,1.8544 μg/m L,1.4526 μg/m L,and 12.7218 μg/m L,respectively.It can be seen that the fungus Diaporthe sp.is more sensitive to pyraclostrobin,while Botryosphaeria sp.is more sensitive to fluazinam and azoxystrobin.Therefore,these three agents should be considered a priority for controlling apple dry rot in field production.