Research Of CSFV Distribution Rule And Tropism Of Tissues In Acutely And Subacutely Infected Pigs

Swine Fever also known as Classical Swine Fever(CSF),caused by Classical Swine Fever Virus(CSFV) is a highly contagious and hemorrhagic disease of pigs and has significant economical losses in pig industry.For research the danamic distribution of tissues in acutely and subacutely infected pigs,firsty Acute and subacute animal infection models were established.Then adopting the optimization of visualization of CSFV RNA in situ hybridization(ISH) method and histopathological method to comparative research the viral distribution and tissue pathological changes in systematic of acute and subacute infection. Then provided experiment data for further illuminates the viral distribution in the infected tissues on the RNA molecule level.1. Research of CSFV distribution rule and tropism of tissues in acutely infected pigsIn this study, eight 30 ages health weaned piglets were inoculated with CSFV highly virulent(5×103.61TCID50/mL) by the means of neck muscle injection, one to two pigs were killed on days post infection(dpi)1?3?5?7, pig's temperature were measured and clinical observations were recorded daily. Tonsil?submandibular lymph nodes?duodenum?spleen?liver?lung?kindey and so on sixteen tissues were collected to recorded pathological changes.1dpi the body temperatue up to above 40?. 1dpi?7dpi the clinical score of infected pigs graduedly increased,7dpi it up to the maximum. After autopsy, lymph nodes enlarged obviously,splenic edges infarction,kidney hematoma and soon were see. Then acute animal infection models were established successfully. Laid important fundmental for further illuminates the viral distribution in the infected tissues on the RNA molecule level.Using optimizated RNA visualization ISH methods to locate viral RNA in infected tissues,results showed that, viral RNA could be detected in Tonsill?submandibular lymph nodes? duodenum?spleen?liver?lung?kindey on 1dpi after actuely infected. 1dpi?7dpi the amount of RNA gradually increased. viral RNA were mainly concentrated in tonsile?spleen?lymph nodes.Results of HE showed that:1dpi in addition to submandibular lymp node,other tissues appears to varying degress of pathological changes,the phenomenon such as interstitial hyperemia of lymphoid nodule of tonsile,spleen artery hemorrhage and inflammatory granulocyte infiltration,hyperemia of kidney?liver?lung.With the course of the disease,the pathological changes of each tissues obviously aggravating,the main phenomenon amog them were the necrotic,lymphoid tissue emptying,seriously destroying structure of lymphocytes in the immune organs.while in the whole process,histologic lessions in lung was slighter than others.At the same time,adopting CSFV and TCID50 methods to verify it.Results of fluorescent quantitation RT-PCR showed that:1dpi could detecte CSFV nucleic in 16 tissues,and the CT value is dreased gradually. TCID50 results showed that:from 1dpi to articulo mortis,the CSFV relatively infected density appear to total upwards trend in sixteen tissues?organs. Compared to 1dpi.CSFV tropism from high to low is pancreatic, mesenteric lymph nodes, spleen,submandibular lymph nodes, ileovcaecal valve, tonsil, liver, lung, kidney, ovarian (testes), caecum,stomach, heart, bladder, muscle, duodenum?With the course of disease prolonged,viral lode is more and more high in infected tissues,the amount of viral RNA is more and more.that means it has the positive correlation with the pathogenetic severity.2.Research of CSFV distribution rule and tropism of tissues in subacutely infected pigsIn this study, eighteen 30 ages health weaned piglets were inoculated with CSFV moderate virulent(5×103.66TCID50/mL) by the means of neck muscle injection, one to two pigs were killed on 1?3?6?10?12?13?18?20?24?28dpi, pig's temperature were measured and clinical observations were recorded daily. Tonsil?submandibular lymph nodes?duodenum?spleen?liver?lung?kindey and so on sixteen tissues were collected to recorded pathological changes.2dpi temperature rised to above 40?.After autopsy, splenic edges infarction, ileocecal valve ulcer, lymph nodes enlarged obviously and appear to typically"marble liked" lesion.The course of disease lasted 28d, then subacute animal infection model was established successfully. Provided important fundmental for further illuminates the viral distribution in the infected tissues on the RNA molecule level.Using optimizated RNA visualization ISH methods to locate viral RNA in infected tissues,results showed that,1dpi viral RNA could be detected in Tonsil?submandibular lymph nodes?duodenum?liver?kindey,3dpi viral RNA could be detected in spleen?lung.6dpi-28dpi the amount of RNA gradually increased,especially after 18dpi.HE results showed that:the more serious lessions occoured in immune organs,especially after 18dpi,each tissues appear seriously pathological changes,such as severe necrosis and empty of tonsil lymph nodes cells, massive hemorrhage in the eages of germinal center of submandibular lymph nodes,a larger number inflammatory cells infiltration,spleen appear diffuse hemorrhage,lymphocytes began to collapse,while lung has the opposite changes, it's pathological changes has the trend to alleviate on 20dpi-28dpi.At the same time,adopting CSFV and TCID50 methods to verify it.Results of fluorescent quantitation RT-PCR showed that:1dpi viral RNA could be detected in Tonsil?submandibular lymph nodes?duodenum?liver?kindey,3dpi it could be detected in spleen and lung,6dpi it could de detected in kinds of sixteen tissues.TCID50 results showed that:from 1dpi to articulo mortis,the CSFV relatively infected density appear to total upwards trend in sixteen tissues?organs. compared to 1dpi. CSFV tropism from high to low is Tonsil, ileocecal valve, spleen,submandibular lymph nodes, duodenum, kidney, cecum, mesenteric lymph nodes, liver, lung, stomach, ovary (testes), myocardial, bladder, pancreas, muscle.With the course of disease prolonged,viral lode is more and more high in infected tissues.the amount of viral RNA is more and more, that means it has the positive correlation with the pathogenetic severity. But the RNA ISH results of lung showed that the amount of viral RNA has the anticorrelation with the course of disease, and it's lesions alleviated in the later infection.Compare the difference of distribution rule and tropism of tissues between actuely and subacutely infected pigs.In the course of acute disease,CSFV RNA has the faster diffuse speed in spleen and submandibular nodes and higher viral load in duodenum,In this study,acute and subacute animal infection models were established successfully. Adopting the optimization visualization of CSFV RNA ISH method and regular detect methods to research the CSFV distribution rule and tropism after virulent and moderate virulent strains infected.Then provided important basis for study CSFV pathogenesis.