CN101491199B - Tobacco black shank resistance disease nursery identification method - Google Patents

Abstract

The invention relates to a method for identifying tobacco black shank resistance in disease nursery. A continuous cropping tobacco field in a historical high occurrence area of the tobacco black shank disease is selected as a disease nursery; plants are inducted to develop the disease through the improvement of the field humidity, so that the complex operation of strain separated storage and culture is reduced; the strain is sourced from the field; the quantity and variety of the microspecies for inoculation are theoretically unlimited; and the method has better diversity and representativeness, and the inoculation conditions are under the control of nature, so the identification results reflect the local practical situation of the day more correctly. A one-off uprooting investigation method after the collection of the flue-cured tobacco is adopted so as to greatly reduce the workload, save the cost and bring about a correct and reliable result. The method is characterized by low costand simple and feasible operation. The method is an economical and practical method and has bright market and application prospect.

Description

A kind of disease nursery identification method of black shank resistance

Technical field

The invention belongs to disease resistance of plant authenticate technology field, be specifically related to the evaluation of black shank resistance, particularly relate to a kind of method of the black shank resistance being carried out the evaluation of disease garden.

Background technology

Black shank is to endanger one of severe diseases in the World tobacco, also is the main disease of China tobacco simultaneously, can endanger all cultivation tobaccos such as flue-cured tobacco, air-curing of tobacco leaves, suncured tabacco, burley tobaccos, Turkish tobaccos.North Carolina, USA accounts for 1%～2% of gross yield because of the tobacco leaf that black shank loses every year, and the loss that causes because of balck shank in the period of nineteen ninety-five and 1996 two just reaches 3,610 ten thousand dollars, accounts for 35.13% of disease total losses, occupies first of each disease.This disease is at China's widely dispersed, and except that the Heilongjiang Province did not find as yet, all the other respectively plant cigarette provinces and regions (containing Taiwan Province) all generation.Average attack rate is 5%～12%, and the serious field piece incidence of disease is up to 75%, even causes total crop failure.According to incompletely statistics, China's black shank every year on average onset area up to 76373hm ², production loss 2869.26 kilograms, the output value is lost above 1.23 hundred million yuans.Production practices show, prevent and treat that balck shank is most economical, effective, the measure of environmental protection is exactly a breeding resistant variety, and resistance identifies that nature becomes an important ring in the work such as anti-source digging utilization, breeding for disease resistance, resistance monitoring.

The crop disease-resistant breeding is different from the selection of other economical characters, and the phenotype of disease resistance is the performance that crop and pathogen are done under certain environmental condition mutually.Compare with the phenotype of economical character, crop disease-resistant is the mutual work of two kinds of biologies, and therefore, the performance of disease resistance then exists for prerequisite with pathogen, and environmental condition must satisfy two kinds of biological growth and developments again simultaneously.The screening and assessment in black shank breeding and anti-source all must seed selection or check in the holocoenotic selection of balck shank is pressed, otherwise, carry out anti-balck shank breeding and be blindly, poor efficiency.

Existing tobacco resistance authentication method mainly contains the greenhouse and identifies inoculation method and sick garden live body bacterial strain artificial infection survey method seedling stage.Seedling stage, survey method adopted zoospore or mycelium suspended liquid to soak the root inoculation usually, but owing to generation zoospore or mycelial quantity variance between each strains tested are bigger, the bacterium that connects of these methods is measured wayward, the test operation complexity, grade scale is difficult for grasping, and is subject to the influence of the factors such as walk time, vitality and infiltration capability of zoospore or mycelial concentration and zoospore.Result and the field resistance difference of utilizing this method to identify are big, can not accurately reflect the disease resistance of kind.The live body bacterial strain artificial infection of sick garden identifies it is authentication method commonly used at present.At present the shortcoming of existing authentication method be inoculation with collection, preservation and the choose reasonable of bacterial strain is the work long-term, continuous, that operating technology is higher, cost is bigger, and the diversity of inoculating microspecies quantity, the kind of usefulness to compare with the field trial method be limited all the time.And disease survey method in the past according to disease resistance grade scale natural occurrence in disease send out initial stage, Sheng phase and not each investigation of phase once, if then each investigation in the 20th day, 40 days, 60 days is once in the inoculation back in artificial infection.All should investigate the whole plant of each kind at every turn.Because the black shank site of pathological change mainly at basal part of stem, investigates some difficulty.

Summary of the invention

The objective of the invention is to deficiency at the prior art existence, a kind of disease nursery identification method of black shank resistance is provided, adopt disease over the years district occurred frequently continuous cropping vega malingering garden, provide necessary condition by increasing field humidity for the plant morbidity, reduce bacterial strain and separated the complex operations of preserving and cultivating, allowing needs seed selection or the check in the holocoenotic selection of balck shank is pressed of the kind of evaluation, the disposable investigation methods of uprooting after adopting flue-cured tobacco to gather, qualification result is accurate, and can accurately reflect the actual conditions of there and then, cost is low, easy to operation.

Purpose of the present invention is achieved through the following technical solutions: a kind of disease nursery identification method of black shank resistance comprises following sequential steps:

One, the selection in sick garden

(1), select that black shank is popular throughout the year in history, take place seriously, the incidence of disease reaches 80% above plot;

(2), local 5～July is hot and humid, average annual rainfall is big;

(3), continuous cropping, natural irrigation conditions is good, the tobacco crop-planting is consistent with managerial skills, operates routinely and manages.

Two, field management

Other farming operation management routinely.During the tobacco crop-planting, fill the gaps with seedlings, apply fertilizer after the cigarette transplantation of seedlings.Guaranteeing that the cigarette seedling all survives behind the 15d, pours water in the beginning field, guarantees in the ditch of field open fire as seen, and the later stage needs fertile situation supplementary fertilizer according to each kind.

Three, disease survey and evaluation

The investigation of uprooting after flue-cured tobacco is gathered is carried out classification and evaluation of resistance by national standard to identification of species.

The present invention has taken the lead in proposing a kind of black shank nursery identification method with practical value, has realized without artificial infection, effective under field conditions (factors) identification of species resistance.Take place by providing advantage to bring out disease, reduce numerous and diverse bacterial classification and preserve work such as cultivation, inoculum formulation and inoculation, its bacterium source is in the land for growing field crops, microspecies quantity, the kind of inoculation usefulness are unlimited in theory, diversity, better representative, inoculation condition also is nature control, so qualification result can accurately reflect the actual conditions of there and then; The disposable investigation methods of uprooting after adopting flue-cured tobacco to gather, significantly reduced workload, provide cost savings, qualification result accurately and reliably, and can accurately reflect the actual conditions of there and then, having cost characteristics low, easy to operation, is a kind of comparatively economical and practical method, has better market prospect.

Embodiment

Embodiment 1:

1.1 the addressing and the foundation in the sick garden of field spontaneous induction

The stable geographical ecotope that helps falling ill, falling ill is the basis in disease garden.Germ survives the winter in the residual body of diseased plant, soil, muck with chlamydospore and resting mycelium body, can survive about 3 years.The soil that carries disease germs, class fertilizer, irrigation water and sick seedling are the main primary source of infection of black shank.Germ borrows wind and rain, irrigation water, farming operation etc. to repeat to infect again, makes disease more and more heavier, and then climing and full field.It is movable in the soil layer of 0～5cm down that tobacco black shank bacterium mainly concentrates on apart from soil table, and by wound or directly intrusion, infecting the position mainly is basal part of stem, and the susceptible stage is mainly before buddingging.The hot and humid disease that helps takes place, and humidity is popular key factor.24～32 ℃ of temperature, relative moisture is 3～5d more than 80%, and an onset peak will appear in the field, and humidity is big more, and morbidity is faster heavier.

1.1.1 geographical and ecological requirement the in sick garden:

According to the suitableeest ecological condition and other factors that balck shank takes place, establish following sick garden target: balck shank is popular throughout the year in history, takes place serious; 5～July is hot and humid, and average annual rainfall is big; Sick garden area about 2～3 mu, continuous cropping, natural irrigation conditions is good, crop-planting is consistent with managerial skills; Have a good transport service.

1.1.2 addressing process

1.1.2.1 selected area balck shank history and the investigation of main meteorological data

According to the addressing target, establish Kunming, Yuxi as selected purpose district.

1.1.2.2 site inspection

In May, 2005 to July is on the occasion of the black shank onset peak period, ground such as Kunming grassy marshland, Shidian, Kunming, Yuxi Hongta District, Yuxi river and mountain, Chengjiang, Yuxi, Yuxi Huaning have successively been investigated, understood local incidence over the years, gathered the balck shank standard specimen in the disease district occurred frequently, and the record place name, taken photo.

1.1.2.3 the preliminary of addressing determined

According to the addressing target, to establish impregnable pass town, Yuxi Jiangchuan County, institute of Yan He town, Yuxi Hongta District cigarette section proving ground, Kunming Xun Dian county sci-tech popularization and stand that 3 plot are arranged is balck shank Chang Faqu over the years in the proving ground, the incidence of disease reaches more than 80%, is continuous cropping field.

The geographical position and the meteorological data of selected area are as follows:

Geographical position, impregnable pass town, Yuxi Jiangchuan County: 1932 meters of height above sea level, 24 ° 20 ' of north latitude, 30 ° of north latitude, 16.7 ℃ of average annual temperature, 749.2 millimeters of annual rainfalls; Geographical position, experimental field, Yan He town, Yuxi Hongta District cigarette section institute proving ground: N:24 ° 14.382 ', and E:102 ° 30.024 ', height above sea level 1635m, average temperature of the whole year 16-17 ℃, 880～900 millimeters of annual rainfalls; Xun Dian county, Kunming sci-tech popularization experimental field, the proving ground geographical position N:25 ° 30.759 ' that stands, E:103 ° 15.910 ', height above sea level 1878m, 14.7 ℃ of average annual temperature, 1352.2 millimeters of annual rainfalls.

1.1.3 the check in sick garden and foundation, verification experimental verification:

1 materials and methods

1.1 experimental cultivar

Yuxi is ground and with Yuxi river and mountain 18 kinds is arranged, it is respectively L8, NC1071, N.nesophila, N.nudicaudis, N.plumbaginifolia, G28, K326, NC95, MD609, little gold 1025, No. three, innovation, Venus 6007, red big, N.stocktonii, land Huang, Barley37, Barley21, build ripple No. 1.

Xun Dian point in Kunming has 13 kinds, and it is respectively L8, NC1071, N.nesophila, N.nudicaudis, N.Plumbaginifolia, G28, K326, NC95, MD609,1025, No. three, innovation, Venus 6007, red big.

With Yunnan kind K326 is high anti-check variety, red greatly susceptible check variety.

1.2 test determination method

Yuxi grind and: grind with the proving ground in Yunnan Provine Tobacco Science Inst. and to carry out test determination.Utilize ill continuous cropping vega malingering garden, lure its natural occurrence.The sub-district randomized arrangement repeats for three times, and every kind is every to repeat to plant delegation, every row 35 strains.

The field layout drawing

Kunming Xun Dian point: carry out test determination in the Kunming Xun Dian sci-tech popularization proving ground of standing, N:25 ° 30.759 ', and E:103 ° 15.910 ', height above sea level: 1878m.Utilize ill continuous cropping vega malingering garden, lure its natural occurrence.The sub-district randomized arrangement repeats for three times, and every kind is every to repeat to plant delegation, every row 20 strains.

The field layout drawing

Yuxi river and mountain: utilize ill continuous cropping vega malingering garden, lure its natural occurrence.The sub-district randomized arrangement repeats for three times, and every kind is every to repeat to plant delegation, every row 25 strains.

The field layout drawing

The conventional floating seedlings of cultivation management, of the right age transplanting, the cigarette seedling survives the field, back and waters more, guarantees field humidity, and fertilising is management routinely.The plant strain growth phase is not all used bactericide, and the insect pest management is identical with field management.

The classification of disease severity should meet the regulation of YC/T 39.Natural occurrence in disease send out initial stage, Sheng phase and not each investigation of phase once all should investigate the whole plant of each kind at every turn, and calculate diseased plant rate, the disease index of each kind, be main then with the disease index according to the resistance against diseases of evaluating each kind.

The classification of disease severity is by the regulation of the tobacco business standard YC/T39 of the People's Republic of China (PRC).The evaluation of resistance standard is undertaken by the method for the tobacco business standard YC/T41-1996 of the People's Republic of China (PRC).

Disease grade scale (is unit with whole strain)

0 grade: complete stool is anosis;

1 grade: stem's scab is no more than stem girth 1/2, or half is slightly wilting with lower blade, or scab appears in bottom minority blade;

2 grades: stem's scab surpasses 1/2 of stem girth, or blade more than half accent withers;

3 grades: stem's scab is around stem girth, or 2/3 is wilting with blade;

4 grades: the whole blades of diseased plant are wilting or withered.

Disease index calculates

Disease refers to=∑ (diseased plant numbers at different levels * this disease level value)/(investigating total strain number * superlative degree value) * 100

1.3 statistical analysis

Disease is referred to do variance analysis, measure the sick difference that refers between each expert evidence.

2 results and analysis

Table 2-3 grinds with field disease disease resistance and significantly analyzes

More than field, 3 different locations nature qualification result show the red large and small gold 1025 of susceptible variety and disease-resistant variety K326, G28 and to reform No. three differences extremely remarkable.Red big grind in Yuxi and disease that point, Yuxi river and mountain point, Kunming Xun Dian orders refers to be respectively 60.95,35.67,27.5 of susceptible contrast, disease-resistant contrast K326 disease refers to be respectively 0,1.67,1.67, and height is felt the highest disease of germplasm and is referred to be respectively 99.05,94.00,79.17.Yuxi grind and occurring degree higher.

2-3 sees that 3 tests all can more as one man reflect the disease resistance of each experimental cultivar to black shank from table, and anti-susceptible difference is obvious.This explanation is set up the black shank natural resistance and is identified that the garden is feasible in above-mentioned place, its result can pass judgment on the disease resistance difference of experimental cultivar substantially.In view of Yuxi river and mountain point and the red big susceptible check variety disease of Kunming Xun Dian point refer on the low side, Yunnan tobacco black shank bacterium biological strain is formed single, 3 plot measurement result basically identicals have only kept Yuxi and have ground and malingering garden, plot, proceed the screening study of disease-resistant germ plasm resource.

Grind from Yuxi and the qualification result in 2006～2008 years, this disease garden occurring degree height, red serious disease referred to reach 61.43 in 2007, the K326 disease refers to be 24.52, red serious disease referred to reach 76.85 in 2008, and the K326 disease refers to be 7.87, and anti-susceptible difference is extremely remarkable, annual intercurrent disease difference is less, test data is accurate, complete, continuous, and crop-planting is consistent with managerial skills, away from peasant's vega, explanation is set up the sick garden of spontaneous induction at this, is used to identify that the germ plasm resource disease resistance is feasible.After build up in sick garden, save its ecosystem environment from damage, the people who avoids subenvironment for a change as far as possible.

1.1.3.22006～2008 annual main meteorological data (table 2-4)

Table 2-42006～2008 year 5-9 month main meteorological data

Year	Month	Ten days samming (℃)	Temperature depature (℃)	The temperature model year	Rainfall (mm)	Rainfall anomaly (%)	The rainfall model year	Sunshine (hr)	Sunshine anomaly %	Percentage of sunshine %	Sunshine model year	The weather model year
													2006	5	18.5	-1.7	Extremely low	125	44	Slightly many	200	-4	50	Normally	Low form
	6	21.2	0.1	Normally	55	-60	Special few	159	17	39	Slightly many	Type short of rain
														7	21.2	0.4	Slightly high	196	10	Slightly many	109	-8	26	Normally	Normal type
	8	20.5	0.1	Normally	66	-64	Seldom	99	-27	24	On the low side	The few photograph short of rain
														9	18.5	-0.8	On the low side	95	-12	Less slightly	105	-12	28	Less slightly	Low form
2007	5	18.5	-1.7	Extremely low	142	64	On the high side	160	-23	39	On the low side	Low temperature overcast and rainy day is few to be shone
														6	21.8	0.7	Higher	64	-39	Special few	233	72	57	Extremely many	High temperature and little rainfall are light how
	7	20.9	0.1	Normally	175	-1	Normally	70	-41	16	On the low side	Few according to type

	8	20.3	-0.1	Normally	246	17	Slightly many	103	-24	26	On the low side	Few according to type
														9	18.1	-1.1	Special low	71	-32	On the low side	116	-3	31	Normally	Low temperature is short of rain
2008	5	19.6	-0.6	Lower slightly	75	-13	Less slightly	195	-6	48	Normally	Normal type
														6	20.4	-0.7	On the low side	178	29	Slightly many	160	18	39	Slightly many	Low form
	7	20.1	-0.7	Special low	264	49	On the high side	125	5	30	Normally	Low temperature is rainy
														8	20.1	-0.3	Lower slightly	139	-25	Less slightly	130	-4	32	Normally	Normal type
	9	19.3	0	Normally	69	-36	On the low side	99	-17	27	Less slightly	Type short of rain

1.1.3.3 auxiliary equipment

Based on the test in sick garden and the check of major parameter, Yan He town, addressing Yuxi Hongta District institute of cigarette section proving ground meets the requirement in disease garden fully.The agricultural garden farming construction in sick garden comprises sprinkling irrigation system, ridge hardening of cement and advances draining and enclose the clear stasis of blood of ditch etc. the unified reconstruction in base is arranged.

1.1.4 the characteristics in sick garden and function

1.1.4.1 geography and climate

Be positioned at institute of Yan He town, Yuxi Hongta District cigarette section proving ground, N:24 ° 14.382 ', and E:102 ° 30.024 ', height above sea level 1635m, average temperature of the whole year 16-17 ℃, 880～900 millimeters of annual rainfalls.

1.1.4.2 geographic isolation is with favourable conditions

Isolate with the farmland.

1.1.4.3 evaluation scale

Can be used for 2.3 mu of disease garden areas, one time performance is identified 60 parts of germplasm materials simultaneously.

1.1.4.4 traffic and irrigation are convenient.

Traffic convenience, irrigation convenience, about 17 kilometers apart from Yuxi, the beautiful first highway of approach.Irrigate and to satisfy the demand throughout the year.

Embodiment 2:

The anti-balck shank of tobacco germplasm is identified

Black shank is the crushing fungal disease of a kind of worldwide tobacco that is caused by Phytophthora nicotianae [Phytophthora parasitica var.nicotianae (Breda deHean) Tucker].Cause of disease Pseudomonas Mastigomycotina Phytophthora is the hot and humid facultative parasite of happiness, can endanger all tobacco types, can both infect root, stem, leaf in any stage of tobacco growing, causes wilting, stem damage and plant death.In recent years, because the growth of continuous cropping vega enlarged areas and the continuous cropping time limit, black shank has and increases the weight of popular trend.For alleviating the loss that this disease causes, people are once with multiple distinct methods control such as cultural control, chemical control, breeding for disease resistance black shank, but because the zoospore of this germ formation speed is fast and quantity is big, infect short, rapid onset of incubation period after the cigarette strain, can take place repeatedly to infect again a season of growth, should disease also mix generation sometimes simultaneously, be difficult to control in case disease is popular with other disease.If the loss that prophylactico-therapeutic measures balck shank not in place causes can reach 100%.At the serious harm of present black shank, the research disease-resistant varieties has become the emphasis of current control black shank.The application utilizes ill continuous cropping vega malingering garden, lures its natural occurrence, sets up natural resistance and identifies the garden, carries out the disease resistance of germ plasm resource and identifies, be intended to for effective control black shank generation, reduce economic loss theoretical foundation be provided.

1 materials and methods

1.1 experimental condition

This research field trial was ground with the proving ground in Yunnan Provine Tobacco Science Inst. in 2006～2008 years and is carried out, and belonged to subtropics land plateau type weather, 15.5～23.7 ℃ of average temperatures of the whole year, average annual rainfall 700～900mm.Cultivating soil is a red soil, and root is made layer 35～45cm.Preceding crop is a mortise pin.

1.2 material

Had 18 kinds in 06 year, be respectively L8, NC1071, N.nesophila, N.nudicaudis, N.plumbaginifolia, G28, K326, NC95, MD609, little gold 1025, No. three, innovation, Venus 6007, red big, N.stocktonii, land Huang, Barley37, Barley21, build ripple No. 1.

Experimental cultivar was 21 in 07 year, be respectively NC2514, build ripple No. 1, Islangold, CV78-5, heavy foliage point, Venus 6007, big Red Star, Shanxi too 7615, CV78-4, No. 1, the Wujiang River, CV16-2, big willow leaf 2014, land Huang, Daris Special, Q1236:Hicks Q46, long gold-tinted plate, No. 7, Xu Jin, willow leaf point and Black Shank Resistant.

Experimental cultivar was 39 in 08 year, was respectively CV16-2, Islangold, CV78-5, Shanxi too 7615, No. 1, the Wujiang River, build ripple No. 1, CV78-4, Daris Special, land Huang, NC2514, little gold 1025, MD609, reform No. three, N.plumbaginifolia, G28, NC95, NC1071, L8, N.nudicaulis, Coker Tobacco, H80A432, peace is selected 79-1, Pan Yuanhuang, NC86, ミ ィ ニ Le ト, NC-22-NF, BL-921, Japan's flue-cured tobacco 1, Longyan C2, draw No. 1 in Tianjin, the COM cigarette, OX414, RG12, K358, RG17, D101, OX2028.

With K326 is disease-resistant check variety, and the big gold dollar of safflower is susceptible check variety.

1.3 experimental scheme

06 year: grind with the proving ground in Yunnan Provine Tobacco Science Inst. and to carry out test determination.Utilize ill continuous cropping vega malingering garden, lure its natural occurrence.The sub-district randomized arrangement, three repetitions, each kind plantation delegation, distance between rows and hills is 1.2m * 0.5m, every row 35 strains.

Adopted the block design of single factor completely random in 07 year, each kind (being) is 1 processing, 3 repetitions, each kind plantation 1 row, every row 35 strains.Distance between rows and hills is 1.2m * 0.5m, beer bottle location reason moisture in the soil.

08 year: adopt the block design of single factor completely random, each kind (being) was a processing, three repetitions, and every kind is planted 3 moisture in the soils, every moisture in the soil 18 strains, distance between rows and hills is 120cm * 50cm, the experimental field area is 2.3 mu.

Test adopts seedling-cultivating tray live, floating seedlings, and every germplasm sowing two is coiled, and last week carries out hardening in transplanting, three leaf-cuttings, seedling age 67 days, May 6 transplanted, and had reached the standard in strong sprout during transplanting.The spraying pesticide on the same day of transplanting (time 200mm/ mu) pest control.Each kind fertilization type is consistent with total amount, N: P2O5: K2O=1: 1: 2.5, the base manure of nitrogenous fertilizer, potash fertilizer and phosphate fertilizer and proportion of additional fertilizer were 3: 1.Burley tobaccos L8 suitably increases fertilizer.The tobacco composite fertilizer special that the fertilizer that use at every turn in the field is all produced as Yunnan Prov. Tobacco Leaf Corp..

Transplant back group's phase and carry out the height ridging, whole field period sprays the insect protected agricultural chemicals 4 times altogether, does not spray the anti-microbial type agricultural chemicals.Other cultivation management measures all require to carry out according to the production technology scheme of Yunnan Provine Tobacco Science Inst..

1.1.4 disease resistance is measured

Utilize ill continuous cropping vega malingering garden, carry out the field and pour water, keep humidity, lure its natural occurrence.The classification of disease severity should meet the regulation of YC/T39.After the natural occurrence, carry out repeatedly disease classification investigation, all should investigate whole plant of each kind at every turn, and calculate each the kind incidence of disease and disease index, as the resistance index,, estimate the resistance against diseases of respectively measuring kind according to the resistance criteria for classifying with disease index.And average disease referred to do variance analysis, measure the difference of anti-sense degree between each expert evidence.The disease resistance evaluation of germ plasm resource finishes according to last tobacco leaf picking, and the investigation of uprooting is calculated disease index and judged.

1.1.5 field occurrence regularity research

Utilize ill continuous cropping vega malingering garden, adopt the field to pour water, lure its natural occurrence.The classification of disease severity should meet the regulation of YC/T39.Carry out disease classification investigation every 10d after the natural occurrence, all should investigate whole plant of each kind at every turn, and calculate the incidence of disease and the disease index of each kind, draw the field its growth, judge the Sheng of disease and send out the phase.According to the resistance criteria for classifying, estimate the resistance against diseases of respectively measuring kind.And average disease referred to do variance analysis, measure difference disease-resistant between each expert evidence.

The disease survey grade scale:

0 grade: complete stool is anosis;

1 grade: stem's scab is no more than 1/2nd of stem girth, or half is slightly wilting with lower blade, or scab appears in bottom minority blade.

2 grades: stem's scab surpasses 1/2nd of stem girth, or half is wilting with lower blade.

3 grades: stem's scab is around stem girth, or 2/3rds is wilting with blade.

4 grades: the whole blades of diseased plant are wilting or withered.

According to the diseased plant survey data, be calculated as follows diseased plant rate (generally rate) and disease index (severity).

* 100 is counted in diseased plant rate (%)=morbidity strain number/total strain of investigation;

Disease index=∑ (diseased plants at different levels count * should disease level)/(investigate total strain and count the highest sick level of *) * 100.

(2) evaluation of resistance standard

High anti-: disease index 0～5, represent with HR; Disease-resistant: disease index is 5.01～25, represents with R; In anti-: disease index 25.01～50, represent with MR; Middle sense: 50.01～75, represent with MS; Susceptible: disease index is represented with S more than 75.

1.1.6 data are handled

Test data is analyzed with practical statistical analysis and DPS data handling system thereof that the Tang Qi of Zhejiang University justice, Feng Mingguang compile.

2 results and analysis

2.1 the resistance difference between expert evidence

According to the last investigation result in field, calculate disease index, according to evaluation of resistance standard evaluation germ plasm resource disease resistance.From table 2-5 as can be known: 18 kinds for examination had 5 high anti-kinds in 2006, be respectively NC1071, NC95, land Huang, G28, K326,5 disease-resistant varieties are arranged, be respectively N.nesophila, N.plumbaginifolia, MD609, No. three, innovation, build ripple No. 1, anti-kind Venus 6007 and N.stocktoii in 2 are arranged, the red big and little gold 1025 of sense kind in 2 is arranged, 4 susceptible variety Barley 21, Barley 37, L8 and N.nudicaulis are arranged.Barley 21, Barley 37, L8 and N.nudicaulis performance are susceptible, and its susceptible degree overweights the red big and little gold 1025 of check variety, and difference is extremely remarkable.NC1071, NC95, the Huang Gaokang that lands are not remarkable with contrast disease-resistant variety G28, K326, No. three differences of innovation.Disease-resistant variety N.plumbaginifolia, MD609 performance is disease-resistant, and is not remarkable with contrast disease-resistant variety G28, K326, No. three differences of innovation.All the other varietal resistances are placed in the middle.

The evaluation of resistance of table 2-5 2006 tobacco germplasm

Annotate: * is (SSR) result of middle and later periods hurdle multiple ratio." ± " front digitized representation disease index mean value; Back numeral standard deviation.Capitalization shows the significance of difference on P＜0.01 level, and lowercase shows the significance of difference on P＜0.05 level.Down together.

After tobacco leaf picking finished in 2007, the investigation of uprooting was calculated disease index, according to the disease resistance of evaluation of resistance standard evaluation germ plasm resource.Significance test of difference result between the average disease of each kind referred in 07 year shows: the resistance of different tobacco kind confrontation balck shanks has notable difference, and the disease index of tobacco bred willow leaf point is the highest, is 95.00, and K326's is minimum, is 24.52.The disease that No. 7, susceptible variety willow leaf point, Black Shank Resistant and Xu Jin refers to be higher than extremely significantly the big gold dollar of check variety safflower; Kind Q1236:Hicks Q46, length do not have significant difference to gold-tinted plate and susceptible contrast, but kind is long the disease of gold-tinted plate are referred to be lower than susceptible contrast.The otherness result of each disease-resistant variety and check variety K326 shows: the disease of kind heavy foliage point, big willow leaf 2014 refers to be higher than extremely significantly contrast; The big Red Star of kind, Venus 6007, CV16-2, Islangold, CV78-5, disease refer to be significantly higher than contrast; The kind Shanxi too 7615, No. 1, the Wujiang River, build ripple No. 1, CV78-4, Daris Special, land Huang, NC2514 and contrast and do not have significant difference, but disease refers to all be higher than contrast.NC2514 and K326 disease index are minimum, can recommend breeding for disease resistance to select for use.

The significance of difference between table 2-6 2007 each expert evidence disease refer to

The significance of difference between table 2-7 2008 each expert evidence disease refer to

After tobacco leaf picking finished, the investigation of uprooting was calculated disease index, according to the disease resistance of evaluation of resistance standard evaluation germ plasm resource.By 39 parts of anti-balck shanks of tobacco germplasm are identified (table 2-7), the average disease of disease-resistant contrast K326 refers to 7.87, and evaluation of resistance is in disease-resistant scope, and the red big average disease of susceptible contrast refers to be 76.85, and evaluation of resistance is in susceptible scope.Tobacco bred BL-921 morbidity is the heaviest, and disease index is the highest, reaches 99.07, be higher than red serious disease and refer to, remarkable with the red big differences of susceptible contrast, all the other kind morbidities are lighter than red big, wherein Pan Yuan Huang, Shanxi too 7615 with red big differences not remarkable, all the other kinds and red big-difference are remarkable.The disease index of RG17 is minimum, is 0.93.High anti-kind N.plumbaginifolia, G28, NC95, NC86, Japanese flue-cured tobacco 1, COM cigarette, OX414, K358, RG17 are not remarkable with disease-resistant check variety K326 difference, all disease-resistant variety CV16-2, CV78-4, Daris Special, land Huang, NC25 14, NC1071, L8, N.nudicaulis, Coker Tobacco, H80 A432, pacify select 79-1, ミ ィ ニ Le ト, NC-22-NF, Longyan C2, Tianjin draw No. 1, RG12, D101, OX2028 and K326 difference also not remarkable.Susceptible variety have BL-921, red big, Pan Yuanhuang, Shanxi too 7615, CV78-5, Islangold, MD609, No. 1, the Wujiang River, No. 1, little gold 1025 are extremely remarkable with K326 difference to build ripple.

2.2 tobacco germplasm resistance qualification result

Supplied 18 kinds (table 2-8) of examination that 5 high anti-kinds are arranged in 2006, be respectively NC1071, NC95, land Huang, G28, K326,5 disease-resistant varieties are arranged, be respectively N.nesophila, N.plumbaginifolia, MD609, No. three, innovation, build ripple No. 1, anti-kind Venus 6007 and N.stocktoii in 2 are arranged, the red big and little gold 1025 of sense kind in 2 is arranged, 4 susceptible variety Barley 21, Barley 37, L8 and N.nudicaulis are arranged.

Can be divided into 4 classes substantially according to resistance to the disease resistance qualification result of 21 expert evidences (table 2-8) in 2007 to tobacco black shank bacterium: disease-resistant variety, in anti-kind, middle sense kind and susceptible variety.Wherein disease-resistant variety has: K326 and NC2514 account for 9.52% of total expert evidence; In anti-kind have: build ripple No. 1, Islangold, CV78-5, heavy foliage point, Venus 6007, big Red Star, Shanxi too 7615, CV78-4, No. 1, the Wujiang River, CV16-2, big willow leaf 2014, land Huang and Daris Special, totally 13, account for 61.90%.Middle sense kind has: Q1236:Hicks Q46, long gold-tinted plate and red big, accounts for 14.29% by totally 3; Susceptible variety has Xu Jin No. 7, willow leaf point and Black Shank Resistant accounts for 14.29%.

39 expert evidence disease resistances were identified in 2008 and tentatively find out, can fall into 5 types substantially the resistance level of tobacco black shank bacterium: high anti-kind, disease-resistant variety, in anti-kind, middle sense kind and susceptible variety.Wherein high anti-kind has N.plumbaginifolia, G28, NC95, NC86, Japanese flue-cured tobacco 1, COM cigarette, OX414, K358, RG17, totally 9, accounts for 23.08% of expert evidence sum; Disease-resistant variety has CV16-2, CV78-4, Daris Special, land Huang, NC2514, K326, NC1071, L8, N.nudicaulis, Coker Tobacco, H80 A432, peace selects 79-1, ミ ィ ニ Le ト, NC-22-NF, Longyan C2, Tianjin to draw totally 19 of No. 1, RG12, D101, OX2028, accounts for 48.72% of expert evidence sum; In the Wujiang River is arranged No. 1, No. three, No. 1, MD609, innovation of building ripple of anti-kind, totally 4, account for 10.26% of expert evidence sum; In the sense kind have Islangold, CV78-5, Shanxi too 7615, little gold 1025, totally 4, account for 10.26% of expert evidence sum; Susceptible variety has red big, Pan Yuanhuang, BL-921, accounts for 7.69% of expert evidence sum.Wherein the disease-resistant variety proportion is the highest, and the susceptible variety proportion is minimum.Qualification result sees Table 2-8.

The anti-balck shank result's statistics of table 2-8 tobacco germplasm

Table 2-9 varietal resistance gathers

(inconsistent several breed differences are little between year, adopt mean value to judge the disease resistance level)

2.3 the growth and decline of the tobacco Germplasm Identification material state of an illness are dynamic

Passed through in 2007 continuous 7 morbidity surveys of each identification of species, it is dynamic to analyze each variety source field morbidity growth and decline.The result shows the growth with the field growing phase, and disease presents overall ascendant trend, each germplasm morbidity Sheng phase focus mostly on late May to late June.21 parts of confessions are planted experimentally matter and can be divided 3 types because of the asynchronism(-nization) of onset peak place: the 1st type is that susceptible variety Black Shank Resistant, No. 7, Xu Jin, willow leaf point onset peak concentrate on Time of Day section in May 25 to June 5, fall ill to have slightly June 5 to June 16 and slow down, stable disease after mid-June, this veriety disease time concentrates on late May to mid-June.The 2nd type for long gold-tinted plate, big willow leaf 2014, heavy foliage point, big Red Star, Venus 6007, CV16-2, Q1236:Hicks Q46, red greatly, the Shanxi too 7615, build No. 1 onset peak of ripple in Time of Day section in June 5 to June 16, this veriety disease time concentrates on late May to late June; The 3rd type for big willow leaf 2014, CV78-5, No. 1, the Wujiang River, CV78-4, Daris Special, Huang, NC2514, the K326 morbidity Sheng phase of landing June 16 to June 27, be mid or late June, these kind disease times concentrate on early June to early July.Anti-or disease-resistant variety during the 3rd veriety is all, except that big willow leaf 2014 morbidities early, all the other germplasms late resistance height of all falling ill.First kind kind is all susceptible variety, and morbidity early and resistance is low.The disease resistance of finding out germ plasm resource thus is sooner or later relevant with morbidity, this may with the later stage plant thick, cuticula thickens, xylem forms and makes germ be difficult for invading relevant.

Claims (2)

1. the disease nursery identification method of a black shank resistance comprises following sequential steps:

One, the selection in sick garden

(1) select that black shank is popular throughout the year in history, take place seriously, the incidence of disease reaches 80% above plot;

(2) locality is hot and humid 5～July, and average annual rainfall is big;

(3) continuous cropping, natural irrigation conditions is good, and tobacco crop-planting and field management are operated routinely and are managed;

Two, the investigation of uprooting after the tobacco crop is gathered is carried out classification and evaluation of resistance by national standard to identification of species.

2. the disease nursery identification method of black shank resistance according to claim 1, it is characterized in that: during the tobacco crop-planting, fill the gaps with seedlings and apply fertilizer after the cigarette transplantation of seedlings, guarantee that the cigarette seedling all survives behind the 15d, pour water in the beginning field, guarantee the interior open fire of field ditch as seen, the later stage needs fertile situation supplementary fertilizer according to each kind.