N Protein Muants of Porcine Reproductive and Respiratory Syndrome Virus

Abstract

The present invention provides a genetically modified PRRS virus, methods to make it and related polypeptides, polynucleotides and various components. Vaccines comprising the genetically modified virus and polynucleotides are also provided.

1 . A composition comprising a PRRS infectious agent selected from the group consisting of:
 a.) a genetically modified PRRS virus comprising an N protein which has been modified in at least one conserved region selected from the group consisting of the NLS-2 region, NoLS region and the NES region such that the conserved region has been eliminated, and wherein the genetically modified PRRS virus is attenuated;   b.) an infectious RNA molecule encoding the genetically modified PRRS virus of a.); and   c.) an isolated polynucleotide molecule comprising a DNA sequence encoding the infectious RNA molecule of b.).   
     
     
         2 . The composition of  claim 1  which has been further modified to result in the elimination of the conserved NLS-1 region. 
     
     
         3 . The composition of  claim 1  wherein the conserved region has been eliminated by the introduction of a non-conservative amino acid substitution. 
     
     
         4 . The composition of  claim 1  or  2  wherein the conserved region has been at least partially deleted. 
     
     
         5 . The composition of  claim 1  wherein the conserved region is the NoLS region. 
     
     
         6 . The composition of  claim 1  wherein the conserved region is the NLS-2 region. 
     
     
         7 . The composition of  claim 1  wherein the conserved region is the NES region. 
     
     
         8 . The composition of  claim 1  wherein the PRRS virus is a North American PRRS virus. 
     
     
         9 . The composition of  claim 1  wherein the PRRS virus is a European PRRS virus. 
     
     
         10 . The composition of  claim 8  wherein the conserved region is the NLS-2 region. 
     
     
         11 . The composition of  claim 10  wherein residues 42 and 43 of the N protein are glycines. 
     
     
         12 . The composition of  claim 10  wherein residues 42 and 43 of the N protein are glycine and residue 44 is an asparagine. 
     
     
         13 . The composition of  claim 10  wherein the NLS-2 region has been at least partially deleted. 
     
     
         14 . The composition of  claim 13  wherein at least one of residues 43 through 48 of the N protein have been deleted. 
     
     
         15 . The composition of  claim 14  wherein both residues 43 and 44 of the N protein have been deleted. 
     
     
         16 . The composition of  claim 14  wherein residues 43, 44, and 46 of the N protein have been deleted. The composition of  claim 14  wherein residues 44, 46, and 47 of the N protein have been deleted. 
     
     
         17 . The composition of  claim 14  wherein residues 46, 47, and 48 of the N protein have been deleted. 
     
     
         18 . The composition of  claim 1  that contains additional nucleotide mutation, substitutions and/or deletions, designed to minimize the probability of reversion. 
     
     
         19 . A vaccine for protecting a porcine animal from infection by a PRRS virus comprising the composition of  claims 1  in an amount effective to produce immunoprotection against infection by a PRRS virus; and a carrier acceptable for veterinary use. 
     
     
         20 . A method for protecting a porcine animal from infection by a PRRS virus, which comprises vaccinating the animal with an amount of the vaccine of  claim 19  that is effective to produce immunoprotection against infection by a PRRS virus. 
     
     
         21 . A transfected host cell comprising a composition according to  claim 1 . 
     
     
         22 . A method for making a genetically modified and attenuated PRRS virus, which method comprises:
 a.) mutating a DNA sequence encoding an infectious RNA molecule which encodes a PRRS virus, to produce a genetically modified PRRS virus comprising an N protein which has been modified in an at least one conserved region selected from the group consisting of the NLS-2 region, NoLS region and the NES region such that the conserved region has been eliminated;   b.) introducing the genetically modified PRRS virus into a host cell capable of supporting PRRS replication.   
     
     
         23 . A method of  claim 22  wherein the genetically modified PRRS virus is a North American PRRS virus. 
     
     
         24 . A method of  claim 22  wherein the genetically modified PRRS virus is a European PRRS virus. 
     
     
         25 . The method of  claim 22  wherein the host cell capable of supporting PRRS replication is a MARC-145 cell. 
     
     
         26 . The method of  claim 22  wherein the host cell capable of supporting PRRS replication is comprised within a live porcine animal.