CN101832889B - Sheet washing method for rapid pine wood nematode separation technology - Google Patents
Sheet washing method for rapid pine wood nematode separation technology Download PDFInfo
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- CN101832889B CN101832889B CN2010101668074A CN201010166807A CN101832889B CN 101832889 B CN101832889 B CN 101832889B CN 2010101668074 A CN2010101668074 A CN 2010101668074A CN 201010166807 A CN201010166807 A CN 201010166807A CN 101832889 B CN101832889 B CN 101832889B
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Abstract
The invention relates to a sheet washing method for rapid pine wood nematode separation technology. The method comprises the following steps of: preparing a wood veneer by sampling; soaking the wood veneer in water and flushing and filtering the wood veneer; and observing the treated wood veneer by using an anatomical lens and verifying the microscopic morphology of nematodes to make correct verification. Due to the adoption of the method, the separation time of pine wood nematodes is shortened to between 1 and 5 minutes and the separation speed of the pine wood nematodes is increased by hundreds of times. The method is simple and practical, has less investment, can be implemented by using a conventional nematode separating tool and contributes to improving the detectable rate of the nematodes and the working efficiency of inspection and quarantine and greatly increasing the passing speeds of wood products and wood packaged goods.
Description
[technical field]
The invention belongs to insect pest preventing and controlling quarantine technical field, relate to a kind of sheet washing method of rapid pine wood nematode separation technology.
[background technology]
Pine wood nematode is the universally acknowledged No.1 killer of pine forest, it is the international forest disease worm of quarantining, pine wood nematode enters xylem by the wound that Monochamus alternatus supplements the nutrients, colonize in the resin canal, move simultaneously prolific, spread all over complete stool gradually, and cause resin canal parenchyma cell and epithelial destruction and death.In a single day pine tree infects pine wood nematode, and the fastest more than 40 days promptly withered, and pine forest only needs the 3-5 year from falling ill to destroying.So pine wood nematode has the No.1 killer's of pine forest title again, owing to still do not have the effectively preventing method at present both at home and abroad, so pine nematode claims " pine cancer again.Pine wood nematode in the propagation of China, spread the impetus, caused the Party Central Committee, the leader's of State Council great attention, the General Office of the State Council in 2002 sends " about further reinforcement pine nematode prevention and wipe out and treat insect pests and plant diseases the notice of work " for this reason specially; But in recent ten years, pine wood nematode is fast, troubling in propagation, the rate of propagation of China! According to State Administration of Forestry's report, pine nematode has spread to 123 counties of 12 provinces at present, causes nearly 3,000,000,000 yuan of direct economic loss, nearly hundred billion yuan of indirect economic loss, the safety of more than 500,000,000 mu of pine forest of serious threat China.This situation will constitute a serious threat to the ecological safety of China if continue development, and consequence is hardly imaginable! A host of facts prove that pine that catches an illness and goods thereof (Wooden packing container, wooden block dish etc.) are the carriers of propagating pine wood nematode; Preventing one of measure of pine wood nematode long-distance communications, is the quarantine of getting twice the result with half the effort.China's inspection and quarantine department, forestry quarantine departments, the time of existing separation pine wood nematode, generally more than 6-12 hour, tripping device and detection method as China 03103072.6 related pine wood nematode, though mask work can be able within 20～30 minutes, finish, but the separation of dozens of minutes is oversize proving time, and quarantine has influenced the efficient of work for inspection and the card speed of being open to the custom of Wood products and wooden packing goods, simultaneously costing an arm and a leg of equipment set and be difficult to popularize.The speed of nematode is slow, the time is long because prior art is separated, quarantine departments can not draw assay fast, import can not in time be handled, the transhipment and the use of goods have been influenced, increased the cost of corporate operation, because the import wooden packing need rest on the port for a long time and wait the quarantine result, also can offer an opportunity simultaneously for the anthropochory of pine wood nematode.Therefore, explore the quick isolation technics of pine wood nematode, strengthen quarantine, the long-distance communications of containment pine wood nematode, very urgent.
[summary of the invention]
In order to overcome the above-mentioned shortcoming of prior art, the invention provides a kind of disengaging time of pine wood nematode that makes and foreshorten in the 1-5min, the sheet washing method of the rapid pine wood nematode separation technology that have that simple and practical, small investment, inspection cost are low, the efficient of the recall rate height of nematode and quarantine and examination work is high.
The technical solution adopted for the present invention to solve the technical problems is: a kind of sheet washing method of rapid pine wood nematode separation technology the steps include:
The first step is sampling, chooses one of the wood sample of long 8-12cm, wide 6-8cm, thick 2cm, heavy 20-100g, and the rarest 20-50 gram of wooden unit weight detects its moisture;
Second step was to produce veneer, with cutter wooden unit was split into the thick veneer of 0.1-1cm, and the optimum thickness of veneer is 0.1-0.3cm;
The 3rd step was soaked and flushing, at normal temperatures, veneer was put into container, added water cover and crossed veneer, constantly stirred and washed in soaked process, and the time is controlled in the 1-5min, extracts washing fluid then;
The 4th step was to filter, and poured washing fluid into two-layer nematode sieve and filtered, and the upper strata sieve aperture of two-layer nematode sieve is that 120-300 order, lower floor's sieve aperture are 500 orders, then liquid at the bottom of the lower screen is moved to double dish, and liquid accounts for 1/3 of double dish volume at the bottom of the lower screen;
The 5th step was to observe in the double dish with anatomical lens whether nematode is arranged, and when in observing double dish nematode being arranged, with pipettor nematode was moved on on the microslide, and covered is carried out microscopic morphology and identified; In the time can not making accurate evaluation to nematode, then nematode is sucked plastics finger bottle with pipettor, send expert appraisal.
Described wooden unit is when being split into veneer, and every veneer has at least the surface more than 1/4 not to be subjected to the destruction of cutter.Described coolant-temperature gage is 25-38 ℃.
The invention has the beneficial effects as follows: utilize the present invention to make the disengaging time of pine wood nematode foreshorten to 1-5min, velocity of separation improves hundreds of times, simple and practical, the small investment of the present invention, its inspection cost is low, do not need specialized equipment, utilize the existing instrument that separates nematode just can carry out the work, improved the recall rate of nematode and the efficient of quarantine and examination work, accelerated the circulation of Wood products and the card speed of being open to the custom of wooden packing goods greatly.
[embodiment]
The present invention is further described below in conjunction with embodiment:
A kind of pine wood nematode fast separating process is got one of the wood sample of long 8-12cm, wide 6-8cm, thick 2cm, heavy 30-100g, with cutter wooden unit is split into 0.1-0.3cm or the thick veneer of 0.4-0.6cm; At normal temperatures, veneer is put into container, add water cover and cross veneer, the optimum water temperature degree is 25-38 ℃, stirs and wash 1-5min in soaked process; Pour washing fluid into two-layer nematode sieve and filter, the upper strata sieve aperture of two-layer nematode sieve is that 120-300 order, lower floor's sieve aperture are 500 orders, then liquid at the bottom of the lower screen is moved in the double dish, and liquid accounts for 1/3 of double dish volume at the bottom of the lower screen; Observe with anatomical lens whether nematode is arranged, with pipettor nematode is moved on to microslide then, when in observing double dish nematode being arranged, covered is carried out microscopic morphology and is identified; If can not make accurate evaluation, with pipettor nematode is sucked plastics finger bottle, send expert appraisal.
By wooden unit is split into veneer, make to colonize in fat road parenchyma cell and epithelial nematode and expose surface at veneer.Wooden unit is when being split into veneer, every veneer has at least the surface more than 1/4 not to be subjected to the destruction of cutter, the lip-deep nematode of this part wood chip form that will be kept perfectly depends on or exposes surface at veneer, cooperate and to be soaked in water, to wash and technical finesse such as stirring, the nematode on veneer surface will come off in the free water, reach the effect of separating nematode fast from wood chip.
Test one: adopt different-thickness wood chip sample (0.1-0.3cm and 0.4-0.6cm) washing by soaking to separate nematode, 10 samples of duplicate test, observations.
Test two: adopt the water logging bubble sample (5 ℃, 10 ℃, 15 ℃, 20 ℃, 25 ℃, 30 ℃) of different temperatures, washing separates nematode, duplicate test 3 times, observations.
Test three: adopt different time (5,30,60,180,360,720,1440min) to separate nematode, separating resulting is observed in duplicate test 2 times.
Test four: contrast nematode spill-out in the different sample 1min.
Test five: same sample respectively with electronic sawing sheet with manually use the chopper film-making, soak, washing separates nematode, duplicate test 5 times, observation contrasts the nematode spill-out.
Test six: handle (do not do technical finesse or implement stirring, flushing processing immediately) 1min with distinct methods respectively after the sample immersion of the same race, separate nematode, observe the nematode spill-out.
Above-mentioned each sample is all got 50g, adopts the present invention that 300 wood samples are carried out the nematode separation test, and test findings shows:
Sample can be after the technical finesse of 1-5min, that isolates nematode accounts for 99%, 150 wood samples that contain pine wood nematode are carried out the quick separation test of nematode, and 100% wood sample can both be cutting out partial nematode after the technical finesse of 1-5min (accounting for more than 25% of wood sample nematode total amount) as a result.
Test findings is analyzed: 1, dried thickness sees Table 1 to the influence of nematode spill-out, as can be seen from Table 1, dried thickness influences the spill-out of nematode, and 0.1-0.3cm dried thickness sample is Duoed more than 40% than the spill-out of 0.4-0.6cm dried thickness sample nematode.
Table 1 different-thickness wood chip nematode spill-out relatively
2 water temperatures to the influence of nematode spill-out as can be seen from Table 2, water temperature is very little to the influence of nematode spill-out, test shows that 3 ℃-38 ℃ water temperature can both be isolated nematode.
Table 2 different temperatures nematode spill-out relatively
The influence of 3 soaked time length as can be seen from Table 3, the soaked time length of sample is influential to the nematode spill-out, the trend that the nematode spill-out has the prolongation with the soaked time to increase.
The different soaked time nematode spill-outs of table 3 relatively
4 as can be seen from Table 4 the wood chip of 10 samples can both after 1 minute technical finesse, isolate nematode, the spill-out of 50 gram sample nematodes reaches 150 at most, the rarest 6.
Nematode spill-out in the different wooden sample 1min of table 4
The artificial as can be seen from Table 5 film-making sample separation nematode population of the influence of 5 flaking methods than the high 3-8 of power saws slice separation nematode population doubly.
Artificial film-making of table 5 and mechanical film-making nematode spill-out are relatively
The influence of 6 disposal routes can find out that from table 6 the nematode fractional dose of the 1min of the sample that enforcement is stirred immediately after the immersion, flushing is handled doubles above than the sample of not doing technical finesse.
The nematode spill-out of table 6 different disposal method relatively
Conclusion (of pressure testing)
1 adopts the present invention to test, and intercepts and captures 7 batches of pine wood nematodes from the wooden packing that enters the territory, and intends 4 batches of pine wood nematodes, and many batches of aphelenchoides detect the live body pine wood nematode in the wooden packing of express mail first at home, obtain remarkable social benefit.
2 sheet washing methods are simple to operate, utilize the existing instrument that separates nematode just can carry out, and the time is fast, can determine whether to have in the sample nematode at short notice, in immigration wooden packing quarantine, played the effect of pine wood nematode quarantine primary dcreening operation, also be applicable to that epidemic situation generaI investigation and timber allocation and transportation quarantine that forest department does pine wood nematode in the open air make a definite diagnosis the method whether pine wood nematode is arranged fast when coming to testify simultaneously.
3 present inspection and quarantine systems separate the time of the hiding nematode of timber generally more than 6-12 hour.Utilize sheet washing method to carry out nematode and separate, the disengaging time more than original 6-12 hour is foreshortened to 1-5min, velocity of separation is nearlyer more than 300 times than original raising; Compare with the achievement in research that is on the leading domestic level at present (20-30min finishes the nematode detachment process, more than 10,000 yuan of complete nematode separation equipment price), following advantage is arranged: the first, velocity of separation improves nearly 20 times; The second, simple and practical, the small investment of sheet washing method utilizes the existing equipment that separates nematode just can carry out the work.
4 experiment showed, that wood moisture content is less to the spill-out influence of nematode, and the timber water cut can both be isolated nematode from 3%-38%; But the nematode state that the direct decision of the height of wood moisture content is separated, general inward wooden packing moisture lower (below 15%).Therefore, lessly directly be separated to the live body nematode, the nematode overwhelming majority who separates is the worm attitude (dead worm, dormancy worm attitude) that does not have vigor; The nematode that moisture is separated at the wood sample more than 25% is the live body nematode basically.
5 because the wooden packing moisture that enters the territory is lower, most samples, and the nematode of adopting this method to separate is dead worm or dormancy worm attitude basically; But the nematode complete form of overflowing; Can carry out kind and identify, the part nematode progressively shows vigor after absorbing moisture through the short time (5-10min or longer time); Or wood sample continuing soaked more than 24 hours, 95% above sample can both be separated to the live body nematode as a result.
6 wood samples that take off from the pine tree that catches an illness, because of the moisture height, the nematode overwhelming majority who separates is the worm that lives, so sheet washing method also is adapted to the pine wilt disease generaI investigation of woodland.
If 7 sample dried thicknesses are 0.1-0.3cm, check sample weight is the 30-60 gram; Dried thickness is 0.3-0.6cm, and then example weight should be the 60-100 gram.Can suitably increase timber weight for increasing the nematode spill-out, but wood chip 0.1-0.3cm is an optimum thickness.
The inapplicable the present invention of 8 machine-processed veneers.
The nematode that 9 the present invention adapt to all timber and goods thereof separates.
Claims (7)
1. the sheet washing method of a rapid pine wood nematode separation technology is characterized in that step is:
The first step is sampling, chooses one of the wood sample of long 8-12cm, wide 6-8cm, thick 2cm, heavy 20-100g, detects its moisture;
Second step was to produce veneer, adopted the mode of manually producing, and with cutter wooden unit was split into the thick veneer of 0.1-1cm;
The 3rd step was soaked and flushing, and veneer is put into container, added water cover and crossed veneer, constantly stirred and washed in soaked process, and the time is controlled in the 1-5min, extracts washing fluid then;
The 4th step was to filter, and poured washing fluid into two-layer nematode sieve and filtered, and the upper strata sieve aperture of two-layer nematode sieve is that 120-300 order, lower floor's sieve aperture are 500 orders, then liquid at the bottom of the lower screen is moved in the double dish;
The 5th step was to observe in the double dish with anatomical lens whether nematode is arranged, and observed in the double dish with anatomical lens whether nematode is arranged earlier, when in observing double dish nematode being arranged, with pipettor nematode was moved on on the microslide, and covered is carried out microscopic morphology and identified.
2. the sheet washing method of rapid pine wood nematode separation technology as claimed in claim 1 is characterized in that: in the first step, wooden unit weight is the 20-50 gram.
3. the sheet washing method of rapid pine wood nematode separation technology as claimed in claim 1 is characterized in that: the thickness of veneer is 0.1-0.3cm in second step.
4. the sheet washing method of rapid pine wood nematode separation technology as claimed in claim 1 is characterized in that: in the 3rd step, water temperature is controlled between 25-38 ℃.
5. the sheet washing method of rapid pine wood nematode separation technology as claimed in claim 1 is characterized in that: in the 4th step, liquid accounts for 1/3 of double dish volume at the bottom of the described lower screen.
6. the sheet washing method of rapid pine wood nematode separation technology as claimed in claim 1 is characterized in that: in the 5th step, in the time can not making accurate evaluation to nematode, with pipettor nematode is sucked in the plastics finger bottle, send expert appraisal.
7. as the sheet washing method of claim 1 or 3 described rapid pine wood nematode separation technologies, it is characterized in that: described wooden unit is when being split into veneer, and every veneer has at least the surface more than 1/4 not to be subjected to the destruction of cutter.
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| Application Number | Priority Date | Filing Date | Title |
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| CN2010101668074A CN101832889B (en) | 2010-04-30 | 2010-04-30 | Sheet washing method for rapid pine wood nematode separation technology |
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| CN2010101668074A CN101832889B (en) | 2010-04-30 | 2010-04-30 | Sheet washing method for rapid pine wood nematode separation technology |
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| CN101832889B true CN101832889B (en) | 2011-11-23 |
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| CN105432566B (en) * | 2015-10-30 | 2018-10-26 | 泸县农林局 | A kind of method of quick detection Bursaphelenchus xylophilus |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1430061A (en) * | 2003-01-21 | 2003-07-16 | 厦门大学 | Fast separation device of pine material nematode and its detection method |
| CN1645135A (en) * | 2005-01-21 | 2005-07-27 | 江西出入境检验检疫局检验检疫综合技术中心 | Rapid inspection for nematode larva of dormant pine wood |
| CN1724991A (en) * | 2005-07-21 | 2006-01-25 | 云南农业大学 | Portable fast separation device of pine material nematode and its separation method |
| CN1731130A (en) * | 2005-08-01 | 2006-02-08 | 厦门大学 | Apparatus for quick separation of pine wood nematode and detection method therefor |
| CN101153334A (en) * | 2007-09-29 | 2008-04-02 | 重庆大学 | Rapid sampling method for pine wood nematode molecule detection |
| WO2009136456A1 (en) * | 2008-05-07 | 2009-11-12 | 独立行政法人森林総合研究所 | Dna extraction method for bursaphelenchus xylophilus from wood chips, lamp primer set for bursaphelenchus xylophilus, and detection method for bursaphelenchus xylophilus from wood chips |
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- 2010-04-30 CN CN2010101668074A patent/CN101832889B/en not_active Expired - Fee Related
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1430061A (en) * | 2003-01-21 | 2003-07-16 | 厦门大学 | Fast separation device of pine material nematode and its detection method |
| CN1645135A (en) * | 2005-01-21 | 2005-07-27 | 江西出入境检验检疫局检验检疫综合技术中心 | Rapid inspection for nematode larva of dormant pine wood |
| CN1724991A (en) * | 2005-07-21 | 2006-01-25 | 云南农业大学 | Portable fast separation device of pine material nematode and its separation method |
| CN1731130A (en) * | 2005-08-01 | 2006-02-08 | 厦门大学 | Apparatus for quick separation of pine wood nematode and detection method therefor |
| CN101153334A (en) * | 2007-09-29 | 2008-04-02 | 重庆大学 | Rapid sampling method for pine wood nematode molecule detection |
| WO2009136456A1 (en) * | 2008-05-07 | 2009-11-12 | 独立行政法人森林総合研究所 | Dna extraction method for bursaphelenchus xylophilus from wood chips, lamp primer set for bursaphelenchus xylophilus, and detection method for bursaphelenchus xylophilus from wood chips |
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