| Giardia and Cryptosporidium are important human and animal communicable pathogens,and have a wide range of infection hosts.Their clinical symptoms are mainly diarrhea after infection,causing huge economic losses to animal husbandry and aquaculture.Cryptosporidiosis not only causes severe diarrhea in mammals and respiratory diseases in poultry,but also causes severe diarrhea in people and secondary infection of AIDS.It is one of the leading causes of death in these patients.Cryptosporidium has been listed as one of the most harmful microorganisms among more than 150 water-borne pathogens;it is classified as a Class B biological warfare agent by the CDC.C.parvum is a microscopic animal-animal communicable parasite that is widely found in many vertebrates and is parasitic to humans and most mammals.The disease caused is cryptosporidiosis,mainly diarrhea.Mainly,it can be life-threatening when it is serious.C.andersonisoni causes mainly digestive tract and respiratory diseases in young animals,leading to decreased productivity or death.G.duodenalis is an important zoonic parasite that inhabits the gallbladder and small intestine of the human body.It mainly inhabits the duodenum and causes giardiasis.Clinical manifestations Mainly for abdominal pain,diarrhea,abdominal distension,vomiting,fever and anorexia.Giardia has experienced epidemic outbreaks in many countries in Europe and America,and the disease is also widespread in China.According to reports,the total infection rate of Giardia is 2.52%in China,and the Giardia infection rate in Jilin Province is as high as 25.20%.The quality standards of urban water supply and hygiene standards for drinking water published in China all make detection of Cryptosporidium and Giardia indispensable indicators.The infections of G.duodenalis,C.parvum,and C.andersonisoni were mostly caused by contamination of the water source by their oocysts,and the common symptoms of infection were mainly diarrhea.The detection of G.duodenalis,C.parvum,and C.andersoni is particularly important in public safety.When cattle and sheep are infected by these three parasites,their immunity declines,and the feed utilization,milk production,and slaughter rate are greatly reduced,and cattle and sheep are extremely important as important experimental animals in scientific research work.The influence of the health status of the animal used as an experimental animal directly affects the experiment data and results analysis of the experimental personnel.The three parasites belong to the zoonosis,and there is a great potential safety hazard for both the farmer and the experimenter.Therefore,this experiment established a method for the detection of sheep,cattle Giardia,and Cryptosporidium by multiplex PCR,and conducted an epidemiological investigation.In this experiment,the establishment of double PCR for C.parvum and G.duodenum was used to select the highly conserved 18S rRNA gene of Cryptosporidium and Giardia TPI gene to design primers.Through the optimization of various conditions,the annealing temperature was determined.At 57°C,the extension time was 30s and the sensitivity was 5×10~2/g feces.Using the established double PCR method of Cryptosporidium parvum and G.duodenalis,193 sheep fecal samples were collected from Changchun and Baicheng areas for detection,according to established system operation and optimized reaction conditions.A double PCR amplification was performed,and 48.7%(94/193)samples of G.duodenalis positive were detected by double PCR,and the genotype was assemblage A.And 2.6%(5/193)samples were positive for Cryptosporidium parvum,and the genotype was C.parvum IIa.Experimental G.duodenalis,C.parvum,and C.andersoni triple PCR was used to select the highly conserved Cryptosporidium 18S rRNA gene and the Giardia TPI gene to design primers.Through the optimization of various conditions,the annealing temperature was determined.At 58°C,the extension time was 60s and the sensitivity was 1×10~3/g feces.Using established triple-PCR methods for the detection of cows with G.duodenalis,C.parvum,and C.andersonisoni,the fecal samples from 156 cows in Changchun were collected and tested.Triple-PCR amplifications were performed to optimize the reaction conditions.There were77.6%(121/156)positive samples of G.duodenalis,and the genotype was assemblage A;And 3.2%(5/156)positive samples of C.parvum,and the genotype were C.parvum genotype II、Human C.parvum genotype HFL5。... |
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