Epidemiological Investigation And Prevention And Control Measures Of Porcine Reproductive And Respiratory Syndrome In Guangdong

Porcine reproductive and respiratory syndrome(PRRS)is caused by porcine reproductive and respiratory syndrome virus(PRRSV),mainly characterized by reproductive failure,respiratory syndrome of piglets and high mortality in weaning and nursery pigs.The disease was first discovered in the United States in 1987 and was first confirmed in China in 1996.Due to the high mutation of PRRSV,a highly pathogenic PRRS appeared in 2006,causing huge economic losses to the pig industry in China.In order to better control the occurrence of PRRS,this study investigated the epidemiology of PRRS in Guangdong,and on this basis,a PRRSV strain GD-KP with the characteristics of recombination of classic strains and variant strains was isolated and further research on molecular bioinformatics,pathogenicity and immune effects of PRRSV strain GD-KP.Based on this,a comprehensive prevention and control program for PRRS was proposed.In this study,3080 suspect PRRS disease samples from different regions of Guangdong were collected from 2011 to 2015,and PRRSV nucleic acid detection was performed by RT-PCR.The prevalence of PRRS at different times and in different regions was studied.During the five years from 2011 to 2015,the positive rate of PRRSV samples in Guangdong was 49.3%,38.3%,34.8%,42.9% and 18.8%,respectively.Among them,the positive rates of 658 PRRSV nucleic acids in eastern Guangdong were 42%,67%,48.1%,51% and 11.9% respectively.The positive rates of 1249 PRRSV nucleic acids in western Guangdong were 53%,16%,32.2%,36.2% and 15.5%,respectively.The positive rates of537 PRRSV nucleic acids in central Guangdong were 40%,46%,35.5%,51.6%,and 15%,respectively;the positive rates of 636 PRRSV nucleic acids in northern Guangdong were62%,24%,23.2%,29.9%,and 32.9 %respectively.In summary,the PRRS epidemic in theeastern part of Guangdong is the most serious,and the detection rate of PRRSV in northern Guangdong is the lowest,but it is on the rise.14 PRRSV isolates were isolated from suspected PRRS pathogens and identified by IFA and RT-PCR.The sequences of NSP2 and ORF5 genes of the obtained PRRSV isolates were determined.The obtained PRRSV NSP2 and ORF5 gene sequence was compared with that of PRRSV reference strain.The results showed that PRRSV isolates and PRRSV reference strains can be divided into 4 subgroups,and 14 strains of PRRSV isolates are divided into 2 subgroups.In addition to PRRSV GD-KP strains,the remaining 13 isolates and PRRSV JAX1 belong to the same subgroup.The PRRSV GD-KP,QY2010,QYYZ and the US isolate NADC30 belong to a new subgroup.The neutralization epitope analysis of the obtained PRRSV isolate and PRRSV reference strain GP5 protein indicated that compared with the position 39 th amino acid leucine L39(Leu)of subgroup 1,subgroup 2,subgroup 3 and subgroup 4 was mutated to phenylalanine F39 alanine(Phe),isoleucine I39(Ile),and serine S39(Ser),respectively.Subgroup 1,subgroup 2 and subgroup 3 strains(including subgroup 4 NADC30 strain)at amino acid position 38 is histidine H38(His),but subgroup 4 of GD-KP,QY2010 and QYYZ(2011)at amino acid position 38 was mutated to tyrosine Y38(Tyr).The results of the analysis of 10 PRRSV representative strain GP5 protein antigen sites showed that compared with the classic strain(represented by VR-2332),the mutant strain(represented by JXA1 strain)have two more potential135~140 and 180~185 amino acids,and less than one antigenic sites in the 190~198.The GP5 protein of PRRSV GD-KP strain has both the 190-198 antigenic sites of the classical strain and 135~140 antigenic sites of the mutant strain,suggesting that the GD-KP strain may originate from the recombination of classic and mutant strains.To learn more about the molecular characteristics of PRRSV GD-KP,bioinformatics methods were used to analyze the physicochemical properties,solubility,signal peptide,transmembrane region,hydrophilicity and hydrophobicity,post-translational modification sites,secondary structure and antigenic epitopes of GP5 protein of PRRSV GD-KP strain,and through SWISS The-Model program predicts the three-dimensional structure of the GP5 protein.The results showed that GP5 protein of PRRSV GD-KP strain found a common region of 30~40 amino acids(SANGNSSSYSQ)through the principles ofintegrated ?-turn,hydrophilicity,flexibility,surface probability region,and B-lymphocyte antigen epitope prediction.This region is shown to be a very important antigenic site of the GP5 protein of the PRRSV GD-KP strain.Glycosylation site prediction analysis revealed multiple glycosylation sites in the GP5 protein,located at amino acids 34~37,44~47,and51~54,respectively.Signal peptide of GP5 protein was predicted by the online software SignalP tool: the signal peptide was located between amino acids 1~31.The PRRSV GD-KP strain has three transmembrane helix sequences on the GP5 protein sequence,which are located at amino acids 15~34,66~88,103~125,respectively.The amino acids 1~14 and89~102 were located outside the cell membrane,35~65 and 126~200 amino acids were located in the cell membrane.In order to understand the pathogenic characteristics of PRRSV,the pathogenicity of PRRSV GD-KP strains in piglets was studied.The results showed that after 60-day-old piglets were infected with the PRRSV GD-KP strain,the loss of appetite and apathy Showed in the first 4 days after infection,and the feed intake and mental status returned to normal on the 5th day.On the eleventh day of culling and necropsy,the moderate lesions were seen in the lungs,the edema of the hilar lymph nodes,Spleen slightly swollen,Inguinal lymphadenopathy,bleeding.Histopathological examination revealed that there were a large number of necrotic macrophages in the alveoli and necrosis in the central lymph nodes.The results showed that PRRSV GD-KP had pathogenicity for piglets,but not cause piglets to die.In the prevention and control of PRRS,it is difficult for a single vaccine to achieve effective prevention and control of PRRS.For this reason,four different combinations of PRRSV vaccines were used in this study.Group 1: 14-day-old Immunized with PRRS attenuated vaccine and inactivated vaccine.Group 2: 14-day-old immunized PRRS attenuated vaccines and 35-day-old immunized inactivated vaccine.Group 3:14-day-old immunized PRRS attenuated vaccines and 35-day immunized attenuated vaccines.Group 4:14-day-old immunized PRRS attenuated vaccines.The Immune effects of the PRRS vaccine against PRRSV GD-KP strains were evaluated by antibody titer,viremia,and production parameters after immunization.The results showed that the 14-day-old immunized attenuated vaccine and the 35-Day immunized inactivated vaccine group had the bestresults.Finally,the infection status of PRRSV in pigs was evaluated by clinical symptoms,production parameters and laboratory tests.A corresponding prevention and control plan was formulated for different PRRSV infection status,and its plan was implemented and evaluated.In summary,the prevention and control program for porcine reproductive and respiratory syndrome(PRRS)is proposed,that is evaluate the status of infections-formulate prevention and control plans and implement-effectiveness assessments.