66 results about "Microsporidium" patented technology

The invention discloses a group of primers and application thereof in quick detection of nosema bombycis. The primers have the nucleotide sequences as shown in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3 and SEQ ID NO: 4. Based on the primers disclosed by the invention, related reagents applied to quick detection of nosema bombycis by an LAMP (Loop-Mediated Isothermal Amplification) method can be prepared by combining a conventional technology in the field, can detect the nosema bombycis by the LAMP method quickly and accurately, are suitable for operation on a small amount of sample and simple to operate, and have important significance in practical application.

A biologic insecticide contains insecticiding microbe chosen from fungus, bacteria, actinomycetes, virus, nematode, and microsporidiosis, the microfluid film for wrapping said microbe, and carrier.

The invention discloses an Antonospora locustae strain and application thereof in prevention and control of locust. The Antonospora locustae disclosed by the invention is named as AL2008L-04 strain, is obtained by isolating and screening from the field diseased Asiatic migrotory locust, and was collected in China General Microbiological Culture Collection Centre (CGMCC) in No. 3 of Beichen West Road No. 1 Court, Chaoyang District, Beijing on May 23, 2011 with the culture collection number of CGMCC No. 4901. Antonospora locustae AL2008L-04 strain CGMCC No. 4901 is abbreviated as AL2008L-04 strain. The locust prevention and control agent disclosed by the invention can be used for preventing and controlling grassland locust, migratory locust and other locusts. The product belongs to a biological pesticide, has no toxic or harmful effects on environment, human, livestock and poultry, is safe for natural enemy, and has broad application prospects. The Antonospora locustae strain provided by the invention has significance in improving the level of a prevention and control technique on locust in China.

The invention discloses a group of microsporidium molecule universal detection primers and a kit thereof. The universal detection primers include an upstream primer HMG1F and a downstream primer HMG1R; the nucleotide sequence of the upstream primer is as shown in SEQ ID NO.3, and the nucleotide sequence of the downstream primer is as shown in SEQ ID NO.4. The detection primers target at the gene HMG1 and can be applied to detecting a plurality of microsporidia simultaneously, and the detection results are reliable, easy to operate and high in sensitivity; especially, the universal detection primers have extremely important practical application significance to early detection of various microsporidia in a sample in practical quarantine work.

The invention discloses a multiplex PCR detection kit for newly-emerged intestinal protozoa, which is used for simultaneously detecting cryptosporidium, giardia intestinalis, cyclospora cayetanensis and microsporidia. The kit comprises the following primers: specific primers for cryptosporidium: shown in SEQ ID NO.1 and SEQ ID NO.2; specific primers for giardia intestinalis: shown in SEQ ID NO.3 and SEQ ID NO.4; specific primers for cyclospora cayetanensis: shown in SEQ ID NO.5 and SEQ ID NO.6; and specific primers for microsporidia: shown in SEQ ID NO.7 and SEQ ID NO.8. In addition, the invention also discloses a multiplex PCR detection method for cryptosporidium, giardia intestinalis, cyclospora cayetanensis and microsporidia. Various validations show that the multiplex PCR detection kit and detection method disclosed by the invention have the characteristics of rapidness, sensitivity, and strong specificity, and solve the cumbersome steps of multiplex PCR detection, and through one-time reaction, an effect of simultaneously detecting multiple genes and determining pathogens can be achieved, so that the workload is greatly reduced, the detection time is shortened, and the detection cost is reduced, therefore, the multiplex PCR detection kit and detection method disclosed by the invention have a greater development trend.

The invention discloses an ultra-low volume suspending agent containing nosema locustae and metarhizium anisopliae and used for grasshopper control as well as a preparation method of the suspending agent. The ultra-low volume suspending agent comprises raw materials in parts by weight as follows: 1-6 parts of metarhizium anisopliae spore powder, 1-10 parts of nosema locustae, 1-5 parts of a sulfonate anionic emulsifier, 0-5 parts of a non-ionic surfactant, 5-15 parts of vegetable oil, 50-90 parts of polyol and 0.001-0.1 parts of an ultraviolet protective agent. The ultra-low volume suspending agent combines nosema locustae and metarhizium anisopliae for use, so that respective defects of the nosema locustae and metarhizium anisopliae can be overcome, ideal control effect can be achieved, the flash point is high, volatilization is low, good liquidity is realized, requirements for ultra-low volume spray for the ground and an airplane can be met, phytotoxicity on plants is avoided, the environment is not polluted, the activity of spore and microsporidia can be better kept, the control effect is good, an aversion function is not produced, and antifeedant of pests is avoided.

The invention relates to a very important protein gene PTP2 related to propagation and infection of nosema bombycis nageli. A gene order CDS for encoding nosema bombycis nageli polar tube PTP2 is obtained according to 9-fold genome data of the nosema bombycis nageli and proven by a series of biotechnology clonings. The nosema bombycis nageli polar tube PTP2 obtained in the invention plays a very important role in studying a propagation and infection mechanism of microsporidian, thus further playing a role in productive practice and prevention and treatment of the microsporidian disease. Experiments and analyses such as RNAi intervention and the like indicate that after the polar tube protein PTP2 is intervened, the microsporidian loses the capacity of infecting a host, therefore, the polar tube protein PTP2 of the microsporidian and potential regulator genes thereof become new factors for treating microsporidian infection, in addition, the method of modifying genes can be adopted to express the polar tube protein PTP2 in an abundant way, thus killing part of pests such as locust and the like.

The present invention provides in situ hybridization probes which include a marker and a nucleic acid molecule able to hybridize exclusively with only one species of Encephalitozoon. The nucleic acid molecule may be, for example, complimentary to segment 878-896 of 16S rRNA of Encephalitozoon hellem spores. Specifically disclosed probes are those including the following nucleotides: (1) 5'-ACT CTC ACA CTC ACT TCA G-3' (Seq. I.D. No. 1) which is species specific for Encephalitozoon hellem, (2) 5'-CAG ACC ACT ATC TGC A-3' (Seq. I.D. No. 2) which is species specific for Encephalitozoon cuniculi and (3) 5'-GTT CTC CTG CCC GCT TCA G-3' (Seq. I.D. No. 3) which is species specific for Encephalitozoon intestinalis. The assay of the present invention utilizes a sample such as surface, ground or drinking water, suspected of containing one of the aforementioned species as a target organism. The microorganisms contained in the sample are fixed in a conventional manner and the probe is then introduced wherein it specifically binds with the target microorganism, if present. The sample is then washed to remove the unbound probe and the bound probe is detected in a conventional manner appropriate for the marker molecule of the probe.

The invention discloses an Eriocheir sinensis Microsporidia in-situ hybridization detection probe and kit. The sequence of the probe is disclosed as SEQ ID NO.1. After the probe is labeled by digoxin and combined with Microsporidia 18S subunit ribosome DNA (18S SSU rDNA) by hybridization, the Microsporidia infection can be judged under a microscope after alkaline phosphatase detection system color development. The invention also discloses a kit containing the probe, which has favorable specific, is simple to operate and can intuitively combine the pathogen detection and pathological changes. On the premise of efficiently and accurately detecting the Spiroplasma, the kit can analyze the infection rate and infection intensity of the sample section. The hybridization between the probe and 18S SSU rDNA is specific, the signal is progressively amplified, and thus, the kit disclosed by the invention is more sensitive and accurate than the conventional dyeing observation detection. The detection result proves that the section subjected to hybridization color development can be stored for a long time.

The invention discloses an LAMP (loop-mediated isothermal amplification) detection primer group for microsporidia in silkworm eggs and an application thereof. The primer group comprises outside primers EB1-F3/EB1-B3 and inside primers EB1-FIP/EB1-BIP, wherein the sequences of the primers are shown in SEQIDNO:1-4. An LAMP detection method and kit for microsporidia in silkworm eggs are established by utilizing the primers. The kit comprises the primer group, 2*reaction buffer, positive control substances, negative control substances, a developing liquid (or a fluorescence staining liquid), Bst DNA (deoxyribonucleic acid) polymerase, a sealing liquid and sterile water. The results of detection carried out by adopting the method can be visually observed in natural light or be observed through agarose gel electrophoresis or be observed and determined via a real-time fluorescence curve. The method is easy to operate, has the advantages of short detection time, easiness in determination of results and strong specificity, and can be used for detecting the DNA, at a concentration of 5.0*10<-3>ng/mu L, of the eggs laid by the silkworms infected with nosema bombycis.

The invention relates to a very important protein gene PTP3 related to propagation and infection of nosema bombycis nageli. A gene order CDS for encoding nosema bombycis nageli polar tube PTP3 is obtained according to 9-fold genome data of the nosema bombycis nageli and proven by a series of biotechnology clonings. The polar tube PTP3 obtained in the invention plays a very important role in studying a propagation and infection mechanism of microsporidian, thus playing a role in productive practice and prevention and treatment of the microsporidian disease. Experiments and analyses such as RNAi intervention and the like indicate that after the polar tube protein PTP3 is intervened, the microsporidian loses the capacity of infecting a host, therefore, the polar tube protein PTP3 of the microsporidian and potential regulator genes thereof become new factors for treating microsporidian infection, in addition, the method of modifying genes can be adopted to express the polar tube protein PTP3 in an abundant way, thus killing part of pests such as locust and the like.

The invention discloses a method for constructing transgenic Nosema bombycis, characterized by injecting a transgenic vector wrapped by liposome in body cavity of the Bombyx mori infected with Nosema bombycis, after 2-4 days, adding silkworm hemolymph in Bombyx mori BmN culture cells to cultivate; when microsporidian presents green fluorescent light, using an insect medium to extremely dilute and adding into a cell culture plate, selecting a culture pore where only one culture cell presents green fluorescent light, adding normal culture cells to culture for 2-4 days to obtain infected culture cells; repeating for at least three times, taking the obtained infected culture cells, conducting homogenate and crushing the cells, conducting differential centrifugation, purifying microsporidian, infecting the Bombyx mori, after the Bombyx mori falls ill, purifying the infected tissues to obtain the transgenic Nosema bombycis. According to the invention, exogenous genes can be integrated into the Nosema bombycis genome, the modification of Nosema bombycis by gene engineering is realized, the adjustment of expression of the microsporidian genes is realized, and a method for microsporidian gene function research is provided.

The invention discloses application of rabbit encephalitozoon cuniculi spore wall protein SWP1 to preparation of a reagent for diagnosing or detecting rabbit encephalitozoon cuniculi infection. A specific primer is designed according to the gene sequence of the rabbit encephalitozoon cuniculi spore wall protein SWP1, published by a database of NCBI (National Center of Biotechnology Information), and is amplified in a fox kidney sample infected with encephalitozoon cuniculi to obtain the SWP1nucleotide sequence shown as SEQ ID NO.2. The fused-expressed SWP1-GST is used for the detection of the sample, the result shows that the fused-expressed SWP1-GST protein has good specificity on the detection of fox serum infected with rabbit encephalitozoo cuniculi, and has no cross reaction with positive serum infected with toxoplasma gondii, neospora caninum and cryptosporidium. In addition, the research result shows that the sensitivity of the SWP1 protein, which is taken as a detection antigen, is obviously higher than that of the PTP2 (Protein-tyrosine-phosphatase 2) protein as a detection antigen. Therefore, the method for diagnosing or detecting the rabbit encephalitozoonosis cuniculi is high in sensitivity, and the diagnosis and prevention of the rabbit encephalitozoonosis cuniculi are further promoted.

The invention discloses a prevention method for tetra hybrid bombyx mori pebrine germ vertical transmission. The method comprises the following steps: for silkworm eggs which do not need to be subjected to cold storage treatment, enabling the silkworm eggs to stand for 15 to 30h at a temperature of 20 to 30 DEG C after bombyx mori lays the eggs, then carrying out immersion treatment for 6 to 10min in water with a temperature of 47 to 48 DEG C, taking out the silkworm eggs, then eluting the silkworm eggs with clear water stage by stage until the silkworm eggs are odorless after are, and carrying out hatching hastening after drying; for silkworm eggs subjected to cold storage treatment, taking the silkworm eggs out of a cold storage, protecting for hours at a temperature of 10 to 20 DEG C, carrying out immersion treatment in water with a temperature of 47 to 52 DEG C for 6 to 10min, taking out the silkworm eggs, then eluting the silkworm eggs with clear water stage by stage until the silkworm eggs are odorless, drying, and then carrying out hatching hastening. The method disclosed by the invention can inactivate 97 to 99% of tetra hybrid bombyx mori sporozoan pathogeny, reaches a relative control effect of 90 to 97% on bombyx mori pebrine germ vertical transmission, can reach silkworm egg hatchability of over 93%, and has no harmful effect on growth development of bombyx mori.

The present invention relates to Lymantria xylina cell lines established from pupal tissues of L. xylina Swinhoe, including NTU-LY-1, NTU-LY-2, NTU-LY-3, and NTU-LY-4. These four cell lines were confirmed to be newly established cell lines derived from L. xylina by random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and isozyme analysis. The genotypes and characteristics of the abovementioned cell lines are totally different from other insect cell lines. In addition, these four L. xylina cell lines are susceptible to insect baculovirus of L. xylina multiple nucleopolyhedrovirus (LyxyMNPV), LdMNPV-like virus, PenuMNPV, as well as microsporidia of Nosema sp. (isolated fromEurema blanda) and N. bombycis and the like. Therefore the invention can be applied in multiplication of the abovementioned insect-pathogenic microorganisms to produce biopesticides in pest control. In addition, the cell lines can also be used as hosts for the expression vectors of said baculovirus to produce recombinant proteins.

The invention discloses a silkworm microsporidia HMG1 gene as well as a specific primer set used for rapidly detecting silkworm microsporidia and application thereof. The primer set comprises an upstream primer HMG1-s and a downstream primer HMG1-sR, wherein a nucleotide sequence of the upstream primer is shown in SEQ ID No.5, and a nucleotide sequence of the lower primer is shown in SEQ ID NO.6. A target gene of the detection primer is a high mobility protein gene HMG1 related to sex of the silkworm microsporidia, when the high mobility protein gene HMG1 is taken as the target gene for a silkworm tissue, especially silkworm egg microsporidia molecular detection, the characteristics of reliable detection result, easy operation, strong specificity and high sensitivity can be realized, and the high mobility protein gene HMG1 can be applied to high-sensitivity and rapid silkworm microsporidia molecular detection, especially early detection of the silkworm microsporidia, and has great significance in practical application.

The invention discloses a method for early detection of grouper intestinal tract microsporidia and application thereof. The method includes the following steps that 1, a fore intestine of grouper is selected, and intestine content is scrapped; 2, genomic DNA of the fore intestine content of grouper is extracted; 3, with the extracted genomic DNA as a template, a specific primer is adopted to conduct PCR amplification; 4, a PCR amplification product in the step 3 is taken and subjected to electrophoretic analysis, a result is judged according to the size of the amplification product, if 450 bp stripes can be amplified through specific amplification, it is proved that the sample is infected by the grouper intestinal tract microsporidia. The problem that diagnosis is difficult as a grouper intestinal tract microsporidia individual is small is solved, programming and standardization of diagnosis are achieved, detection specificity and sensitivity are high, and the method can be applied to diagnosis of grouper intestinal tract microsporidia disease, applied to investigation of the molecular epidemiology and used for assessing whether aquaculture water and bait are suitable for grouper aquaculture or not.

The invention relates to a real-time fluorescent PCR detection kit for bumblebee microsporidia and a detection method thereof, which are specially used for detecting bumblebee microsporidia. The kit comprises: (1) a positive standard; (2) negative control; (3) PCR reaction solution; (4) ExTaq enzyme; (5) sterile water. The invention has the following advantages: (1) good stability and specificity:detection of bumblebee microsporidia has high specificity, has no cross reaction with other microsporidia and bumblebee pathogens, and has good repeatability; (2) high sensitivity: the sensitivity can reach 20 copies/[mu] L; (3) easy and fast operation: the whole reaction can be completed within 2 hours. The kit can be used for detecting Nosema bombycis and differentiating and diagnosing the Nosema bombycis from other Nosema bombycis, and has important significance for early prevention and timely treatment of the disease. The kit can be used for detecting Nosema bombycis and differentiating and diagnosing the Nosema bombycis from other Nosema bombycis.

A composition for controlling or preventing microsporidia in a fish or a seafood is prepared by using, as an active ingredient, at least one compound selected from the group consisting of compounds represented by the following chemical formula (I):

(wherein, R², R⁴, R⁵, R⁶ and R⁷ each independently represent a specific substituent), pharmaceutically acceptable salts thereof, and compounds generating the compounds represented by the chemical formula (I) by metabolism in the body of the fish or the seafood. By using this composition for controlling or preventing microsporidia in a fish to or a seafood, a method for controlling or preventing microsporidia in a fish or a seafood is provided, which prevents microsporidian infection in the muscle or organs of the fish or the seafood, and/or suppresses the growth of microsporidia in the muscle or organs of the fish or the seafood, and/or highly effective in eliminating microsporidia from the body of the fish or the seafood and also excellent in safety.

The invention discloses silkworm microsporidia milRNAs and application thereof, research finds that milRNA-4, milRNA-8, milRNA-10 and milRNA-12 have a promoting effect on the proliferation of microsporidia, milRNA-1, milRNA-3 and milRNA-11 have an inhibiting effect on the proliferation of microsporidia, and the milRNA-8 has the most obvious promoting effect on the proliferation of microsporidia, so that the milRNA-8 can be used for regulating the proliferation of microsporidia. The research also finds that the target gene of miRNAs is BmPEX16, and the copy number of the overexpressed silkworm microsporidia and the protein expression quantity of the NbPTP2 are both reduced, which indicates that the proliferation of the silkworm microsporidia is inhibited after the overexpression of the BmPEX16; after the BmPEX16 is knocked out, the proliferation of the silkworm microsporidia is promoted, so that the proliferation of the silkworm microsporidia can be regulated by regulating the expression of the BmPEX16 gene, and a new thought is provided for analyzing the prevention and control of the silkworm microsporidia.

The invention relates to the technical field of insect biological control, in particular to a locust microsporidia AL200801 strain as well as a biological preparation and application of the locust microsporidia AL200801 strain. The invention provides a locust microsporidia AL200801 strain, and the preservation number of the locust microsporidia AL200801 strain is CCTCC No. V202137. The locust microsporidia AL200801 strain disclosed by the invention has the advantages of high timeliness, high lethality, high sporulation rate, no toxicity and strong obligate property.

The invention discloses an extraction method of microsporidium DNA based on a 3D printing functive, a reagent kit and an application of the extraction method, and belongs to the technical field of a nucleic acid extraction method and application. Samples are subjected to splitting treatment with lysate, DNA extraction is realized through holding a 3D printing special-shaped functive by hand or machine to quick transfer in lysate, cleaning liquid and eluent, time-consuming tedious steps of pipetting, centrifuging and the like are not needed, the manual labor density is low, the cost is low, theflux is high, DNA quality is high, requirements for equipment and operating space are small, and the extraction method has automated and high-flux downstream application prospects. The invention alsoprovides an application reagent kit of the nucleic acid extraction method, and an application of the nucleic acid extraction method to PCR rapid diagnosis for microsporidium pathogeny in animal and plant hosts. The problems that through a present microsporidium in vitro PCR diagnosing technique, target nucleic acid preparation is complex and time-consuming and the flux is low can be solved, the extraction method can be widely applied to fast detection of various microsporidium, and the detection flux, the operation convenience, the detection sensitivity and the detection accuracy are notablyincreased.

The invention provides application of andrographis paniculata aqueous extract in preparation of drugs for preventing and treating microsporidiosis of honeybees. The andrographis paniculata aqueous extract provided by the invention is prepared into an andrographis paniculata aqueous extract syrup mixture with a certain concentration, and is loaded into a feeding machine, so that honeybees which are infected by microsporidia can eat freely, and the andrographis paniculata aqueous extract in the mixture can act on the honeybees and the microsporidia, therefore, the effect of effectively preventing and treating the microsporidiosis of honeybees can be achieved. The andrographis paniculata aqueous extract is non-poisonous to honeybees, and has a good prevention and treatment effect on the microsporidiosis of honeybees. The andrographis paniculata aqueous extract is further a natural plant extract, wide in raw materials, low in cost and easy to prepare; no pollution to honeybee products and the environment is caused, the andrographis paniculata aqueous extract is a novel drug, which is healthy, environment-friendly, ecological and efficient, for preventing and treating microsporidiosis of honeybees.

The invention, which relates to the technical field of insect cytobiology and pest biological control, provides a locusta migratoria cell line QAU-Lm-E-17 as well as a preparation method and application thereof. According to the invention, the accession number of the locusta migratoria cell line QAU-Lm-E-17 is CCTCC No. C2019123. The locusta migratoria cell line QAU-Lm-E-17 is a locusta migratoriacell line which is established for the first time at home and abroad and can be subjected to continuous passage; the locusta migratoria cell line QAU-Lm-E-17 can support in-vitro proliferation of obligate parasitic locust microsporidia and a valuable experimental material is provided for the deep research on the locusta migratoria physiology and locust microsporidia.