The Study On The Pathogens And Control Of Jujube Fruit Shrink Disease

In this study, the symptom, initial infection period and onset of the jujube fruit shrink disease, the most serious diseases of jujube fruit, were investigated in Hebei Province. Molecular detection of Erwinia carotovora in the tissue of jujube fruit shrink disease, isolation of pathogen, diversity analysis of microbial community from the fruit of jujube fruit shrink disease by PCR-DGGE and molecular cloning, the analysis of bitterness compounds of fruit shrink disease by high pressure liquid chromatography (HPLC) and gas chromatography-mass chromatography(GC/MC), and the control of jujube fruit shrink disease were studied as well. The main results of this research were showed as follows:1 Survey of the symptom, infection period and onset of the jujube fruit shrink disease in the main jujube fields of Hebei ProvinceDuring 2007-2008, the symptom, infection period and development of symptoms of the jujube fruit shrink disease were observed at Fuping, Tang, Zanhuang and Xingtang counties in Hebei Province. The typical symptom of the jujube fruit shrink disease is that firstly the yellow spots appeared the near the parts of jujube fruit pedicel or the middle of the jujube fruit; subsequently, the spots expanded into the irregular sunken and light yellow spots. Over time, sarcocarp of jujube has become yellowish brown, soft and shrink, and the jujube fruit pedicel became dark yellow. A large number of jujube fruits will fall off after rain or heavy fog. The spots became light black, wrinkled and skinny in the late, and the diseased tissues were sponge-like necrosis, vascular browning. Jujube fruit losed the economic value because of the bitterness. In the course of the investigation, we foundind that the beginning of symptom period of jujube fruit shrink disease is almost the same but slightly different in different areas of Hebei Province. There are very few diseased fruits with early symptoms at mafanggou in Fuping county on 20th Aug 2007, in the next few days the development of diseased fruit was slow. On 30th Aug 2007 after two rainy days, we observed the rot development rate increased rapidly, and 80% of the diseased fruit with typical symptoms fell to the groud on 1th Sep 2007, occupying almost 50% of the total amount of fruit. In 2007, compared with the beginning to show symptoms period in Fuping county, jujube fruit shrink disease occurred 5 days earlier in Tang county. Parts of the diseased fruits were found on 20th August, and the diseased rate of fruit showed a gradually increasing trend. In 2008, the diseased fruit were found in mid and late July, and on 10th August, we collected the diseased fruit with late symptom. This study proved that the invasion period of fruit shrink disease pathogen is the mid-July to mid-August. 2 Molecular detection of Erwinia carotovora in the tissue of fruit shrink of jujubeThe DNA of diseased and healthy Z. jujuba var. henanhuizao, Z. jujuba var. fupingdazao, Z. jujuba var. qingyuanhongzao and Z. jujuba var. tangxiandazao, was extracted. Erwinia carotovora was detected by PCR with specific primers SR3F and SR1cR. The results show that, E. carotovora broadly exist in the diseased and healthy fruit of Chinese jujube, indicating that E. carotovora may not be the pathogen of jujube fruit shrink disease.3 Isolation and identification of pathogeny of jujube fruit shrink diseaseIsolation and identification of the pathogeny of jujube fruit shrink disease were carried in Z. jujuba var. fupingdazao, Z jujuba var.tangxianpozao, Z. jujuba var. zanhuangdazao, Z. jujuba var. xingtangpozao and Z. jujuba var. henanhuizao in 2006 and 2009. The results showed that the isolation rate of the fungal microbes was only up to 14.0%, and most of its recovering parts were the epiderm of jujube. Alternaria spp. separation rate is the highest, followed by Phoma spp. and Dothiorella gregaria Sacc.. The lowest isolation rates were Penicillium, Colletotrichum and several unidentified fungal and bacterial strains. Each selected species of Alternaria spp., Phoma spp. and Dothiorella gregaria Sacc, separately, was inoculated on jujube fruit in the field. The symptoms of jujube fruit inoculated with Alternaria spp., Dothiorella gregaria Sacc. and Phoma spp. by stabing were soft rot, and the symptoms were soft rot. The reisolation rares of the three inoculated fungi were 26.7%,13.3% and 6.7%, respectively.4 Diversity analysis of microbial community from the fruit of jujube fruit shrink disease by PCR-DGGEThe V3 variable fragment for 16S rDNA of bacteria and the variable fragment for 18S rDNA of fungi were amplified from the genome DNA by PCR. The DNA extracted from the healthy fruit of Z. jujuba var. fupingdazao and the fruit of jujube fruit shrink disease. The diversity of bacterial communities of Z. jujuba var. fupingdazao was studied by PCR-DGGE. The results showed that the PCR-DGGE technology was a feasible method for diversity analysis of Z. jujuba var. fupingdazao. The population of endophytic bacteria and endophytic fungi from the diseased friuts was fewer than the ones from the healthy friut, and there was little change in their predominant bacterial floras. However, the predominant fungal floras in diseased friuts were fewer than the ones in healthy friuts.5 The study on bacterial and fungal species diversity in fruit of jujube fruit shrink disease by the molecular cloningThe bacterial and fungal community structures of the healthy and diseased fruits of Z. jujuba var. fupingdazao and Z. jujuba var. henanhuizao were mainly studied during the ocurrence of jujube fruit shrink disease using molecular biology methods-16S rRNA gene clone library construction technology. The bacterial 16S rDNA and fungal 18S rDNA gene fragment length polymorphism were analysed in both fruit types with cloning and sequencing method. After sequence 10 tested positive clones randomly picked from eath sample, the results showed that two fungal and bacterial genes sequence existed in diseased fruit of Z. jujuba var. henanhuizao, which indicated that there were few micobial community in jujube fruit.6 The analysis of bitterness of fruit shrink disease by high pressure liquid chromatography (HPLC) and gas chromatography-mass chromatography (GC/MC)The results showed that water, ethyl acetate and butanol were unsuitable for extracting of bitter substances, and ethanol and hexane could extract the material with obvious bitterness from the diseased tissue. The bitterness compounds were analysed by high pressure liquid chromatography, and the peaks were not obviously different between diseased and healthy jujube samples.The hexane extract of Z. jujuba var. tangxianpozao, Z. jujuba var. zanhuangdazao, Z. jujuba var. fupingdazao, Z. jujuba var. henanhuizao and Z. jujuba var. xingtangpozao was analysed by GC/MS, finding different materials in the diseased samples compared with the healthy ones. The extract was compared with hexade-canenitrile, oleanitrile, hexadecanamide and 9-octadecenamide in NIST05.L Mass Gallery, and the result showed that their similarities were reached to 91%,95%,96% and 95%, respectively. And oleanitrile group "=N" with bitter taste exsisted in the chemical structure of hexadecanenitrile and oleanitrile.7 The control of jujube fruit shrink diseaseAccording to data and the isolation results of the jujube fruit shrink diseases pathogen from the initial experiment, nine fungicides were tested in the field. The results showed that the 0.5% NaHCO3+ 0.25% vegetable oils+ family using detergent and the lipid membrane with 200 times dilution+ streptomycin with 140 units/mL+40% omethoate diluted 1000 times have the best control effect, the second was the lipid membranes with 200 times dilution+12.5% Trapp v1 (WP) with 3000 times dilution+cypermethrin with 3000 times dilution and the filtrate of liquid culture medium of Streptomyces rimosus+film forming material with 1000 times dilution (peach gum sodium carboxymethyl cellulose with ration of 3 to 1), film-forming material 200 times dilutoin and carbendazim diluted 800 times had certain control effect.