Method for detecting microorganisms

Abstract

Described herein are methods of detecting a wound infection and for detecting the presence or absence of microorganisms, for example, wound pathogens in a sample, by contacting a sample with an enzyme produced and/or secreted by the bacteria, and detecting modification or the absence of modification of the substrate, as an indicator of the presence or absence of the enzyme in the sample. The present invention also features a biosensor for detecting the presence or absence of bacteria in a sample.

1 . A method for detecting the presence or absence of a pathogenic bacterium in a sample of wound fluid obtained from a subject, comprising the steps of:
 a) contacting the sample with a detectably labeled substrate for an enzyme produced and/or secreted by each of  Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pyogenes, Pseudomonas aeruginosa, Enterococcus faecalis  and  Escherichia coli , wherein said enzyme is selected from the group consisting of a lysin, an exotoxin, a cell wall enzyme, a matrix binding enzyme, a protease, a hydrolase, a virulence factor enzyme, and a metabolic enzyme, wherein said enzyme is not produced and/or secreted by a nonwound-pathogenic bacterium under conditions that result in modification of said substrate by said enzyme; and   b) detecting the modification or the absence of the modification of said substrate, wherein modification of said substrate indicates the presence of said pathogenic bacterium in said sample, and wherein the absence of modification of said substrate indicates the absence of said pathogenic bacterium in said sample.   
     
     
         2 . The method of  claim 1 , wherein said pathogenic bacterium is  Serratia marcescens.    
     
     
         3 . The method of  claim 1 , wherein said pathogenic bacterium is  Enterococcus faecalis.    
     
     
         4 . The method of  claim 1 , wherein said hydrolase is a lipase. 
     
     
         5 . The method of  claim 1 , wherein said lysin is an autolysin. 
     
     
         6 . The method of  claim 1 , wherein said metabolic enzyme is beta-galactosidase. 
     
     
         7 . A method for detecting the presence or absence of a wound infection in a subject, comprising the steps of:
 a) contacting a sample of wound fluid from a subject with a detectably labeled substrate for an enzyme produced and/or secreted by each of  Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pyogenes, Pseudomonas aeruginosa, Enterococcus faecalis  and  Escherichia coli , wherein said enzyme is selected from the group consisting of a lysin, an exotoxin, a cell wall enzyme, a matrix binding enzyme, a protease, a hydrolase, a virulence factor enzyme, and a metabolic enzyme, wherein said enzyme is not produced and/or secreted by a non-wound-pathogenic bacterium, under conditions that result in modification of said substrate by said enzyme; and   b) detecting the modification or the absence of the modification of said substrate, wherein modification of said substrate indicates the presence of a wound infection in said subject, and wherein the absence of modification of said substrate indicates the absence of a wound infection in said subject.   
     
     
         8 . The method of  claim 7 , wherein said hydrolase is a lipase. 
     
     
         9 . The method of  claim 7 , wherein said lysin is an autolysin. 
     
     
         10 . The method of  claim 7 , wherein said metabolic enzyme is beta-galactosidase. 
     
     
         11 . A biosensor for detecting the presence or absence of a microorganism in a sample, said biosensor comprising a solid support and a detectably labeled substrate specific for an enzyme produced and/or secreted by said microorganism, said substrate attached to said solid support. 
     
     
         12 . The biosensor of  claim 11 , wherein the solid support comprises a material required to be free of microbial contaminants. 
     
     
         13 . The biosensor of  claim 11 , wherein said solid support is a selected from the group consisting of a wound dressing, a container for holding body fluids, a disk, a scope, a filter, a lens, foam, cloth, paper, a suture, and a swab. 
     
     
         14 . The biosensor of  claim 13 , wherein said container for holding body fluids is selected from the group consisting of a urine collection bag, a blood collection bag, a plasma collection bag, a test tube, a catheter, and a well of a microplate. 
     
     
         15 . The biosensor of  claim 11 , wherein said microorganism is a wound-specific bacterium. 
     
     
         16 . The biosensor of  claim 15 , wherein said bacterium is selected from the group consisting of  Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pyogenes, Pseudomonas aeruginosa, Enterococcus faecalis, Proteus mirabilis, Serratia marcescens, Enterobacter cloacae, Acetinobacter anitratus, Klebsiella pneumonia , and  Escherichia coli.    
     
     
         17 . The biosensor of  claim 15 , wherein said wound-specific bacterium is  Serratia marcescens.    
     
     
         18 . The biosensor of  claim 15 , wherein said wound-specific bacterium is  Pseudomonas aeruginosa.    
     
     
         19 . The biosensor of  claim 15 , wherein said wound-specific bacterium is  Staphylococcus aureus.    
     
     
         20 . The biosensor of  claim 15 , wherein said wound-specific bacterium is  Enterococcus faecalis.    
     
     
         21 . The biosensor of  claim 15 , wherein said wound-specific bacterium is  Staphylococcus epidermidis.    
     
     
         22 . The biosensor of  claim 11 , wherein said biosensor directly contacts said wound. 
     
     
         23 . A kit for detecting a wound infection, comprising a biosensor for detecting the presence or absence of a microorganism in a sample, said biosensor comprising a solid support and a detectably labeled substrate specific for an enzyme produced and/or secreted by said microorganism, wherein said substrate is attached to said solid support, and one or more reagents for detecting the enzyme produced and/or secreted by a microorganism causing said wound infection.