Methods For Rapid Identification Of Pathogens In Humans And Animals

Abstract

The present invention provides methods of: identifying pathogens in biological samples from humans and animals, resolving a plurality of etiologic agents present in samples obtained from humans and animals, determining detailed genetic information about such pathogens or etiologic agents, and rapid detection and identification of bioagents from environmental, clinical or other samples.

1 - 22 . (canceled) 
     
     
         23 . A method of identifying a bioagent comprising the steps of: selecting at least one pair of oligonucleotide primers, wherein one member of said pair of primers hybridizes to a first conserved region of nucleic acid from said bioagent and the other member of said pair of primers hybridizes to a second conserved region of nucleic acid from said bioagent, wherein said first and second conserved regions flank a variable nucleic acid region that when amplified creates a unique base composition signature among at least five bioagents; amplifying nucleic acid from at least one said bioagent with said pair of oligonucleotide primers to produce an amplification product; determining the molecular mass of said amplification product by mass spectrometry; calculating the base composition of said amplification product from said molecular mass; comparing said base composition of at least one said bioagent to five or more calculated or measured base compositions of amplification products of known bioagents produced by using said at least one pair of oligonucleotide primers, thereby identifying said bioagent. 
     
     
         24 . The method of  claim 23 , wherein said comparing comprises comparing said base composition of at least one said bioagent to nineteen or more calculated or measured base compositions of amplification products of known bioagents produced by using said at least one pair of oligonucleotide primers. 
     
     
         25 . The method of  claim 23 , wherein said bioagent is a bacterium, virus, mold, fungus or parasite. 
     
     
         26 . A method of identifying a bioagent comprising the steps of: selecting at least one pair of oligonucleotide primers, wherein one member of said pair of primers hybridizes to a first conserved region of a nucleic acid encoding a protein that participates in translation, replication, recombination, repair, transcription, nucleotide metabolism, amino acid metabolism, lipid metabolism, uptake, secretion, antibiotic resistance, virulence, or pathogenicity, and the other member of said pair of primers hybridizes to a second conserved region of said nucleic acid encoding a protein that participates in translation, replication, recombination, repair, transcription, nucleotide metabolism, amino acid metabolism, lipid metabolism, uptake, secretion, antibiotic resistance, virulence, or pathogenicity, wherein said first and second conserved regions flank a variable nucleic acid region that when amplified creates unique base composition signatures among at least five bioagents; amplifying nucleic acid from at least one said bioagent with said pair of oligonucleotide primers to produce an amplification product; determining the molecular mass of said amplification product by mass spectrometry; calculating the base composition of said amplification product from said molecular mass; comparing said base composition of at least one said bioagent to five or more calculated or measured base compositions of amplification products of known bioagents produced by using said at least one pair of oligonucleotide primers, thereby identifying said bioagent. 
     
     
         27 . The method of  claim 26 , wherein said molecular masses of said amplification products are determined by ES1-TOF mass spectrometry. 
     
     
         28 . The method of  claim 26 , wherein said bioagent is a bacterium, virus, mold, fungus or parasite. 
     
     
         29 . The method of  claim 26 , wherein said comparing comprises comparing said base composition of at least one said bioagent to nineteen or more calculated or measured base compositions of amplification products of known bioagents produced by using said at least one pair of oligonucleotide primers.