Assay for Diagnosing Streptococcus Pneumoniae

Abstract

Assays for detecting anti-streptococcal antibodies in biological samples using one or more streptococcal antigens are described herein. Various combinations of antigens may be used in the assays. For example, one or more of Ply, PhtD, PhtE, LytB and PcpA may be utilized. Additional streptococcal antigens may also be used. The assays may also be used in combination with assays that detect streptococcal nucleic acids.

What is claimed is: 
     
         1 . The use of a PcpA antigen for detecting a past infection or active infection by  Streptococcus pneumoniae  in a subject, wherein said infection is determined by binding of antibodies in a sample obtained from a subject to said PcpA antigen. 
     
     
         2 . The use of a PcpA antigen and at least one additional antigen selected from the group consisting of PhtD, PhtE, LytB and Ply. 
     
     
         3 . The use of  claim 2  wherein a combination of antigens are selected from the group consisting of PcpA and Ply; PcpA and PhtD; PcpA and PhtE; PcpA and LytB; PcpA, Ply, and PhtD; PcpA, Ply, PhtD, and PhtE; Ply, PhtD, PhtE, and LytB; PcpA, PhtD, and PhtE; PcpA, PhtD, PhtE, and LytB; PcpA, PhtE, and LytB; PcpA, Ply, PhtE, and LytB; PcpA, PhtD, and LytB; PcpA, Ply, and LytB; PcpA, Ply, PhtD, and LytB; PcpA, Ply, and PhtE; and, PcpA, PhtD, and PhtE. 
     
     
         4 . A method of diagnosing pneumonia or an infection by  Streptococcus pneumoniae  in a subject comprising detecting in a biological sample from said subject antibodies against a PcpA antigen wherein the presence of said antibodies in the sample is indicative of infection. 
     
     
         5 . The method of  claim 4  additionally comprising detecting antibodies against at least one additional antigen selected from the group consisting of PhtD, PhtE, LytB and Ply, or one or more immunologically reactive fragments thereof. 
     
     
         6 . The method of  claim 5  wherein the antigens are selected from the group consisting of PcpA and Ply; PcpA and PhtD; PcpA and PhtE; PcpA and LytB; PcpA, Ply, and PhtD; PcpA, Ply, PhtD, and PhtE; Ply, PhtD, PhtE, and LytB; PcpA, PhtD, and PhtE; PcpA, PhtD, PhtE, and LytB; PcpA, PhtE, and LytB; PcpA, Ply, PhtE, and LytB; PcpA, PhtD, and LytB; PcpA, Ply, and LytB; PcpA, Ply, PhtD, and LytB; PcpA, Ply, and PhtE; and, PcpA, PhtD, and PhtE. 
     
     
         7 . The method of any one of  claims 4 - 6  comprising contacting a biological sample derived from the subject with the antigen for a time and under conditions sufficient for an antigen-antibody complex to form and then detecting the formation of an antigen-antibody complex. 
     
     
         8 . The method of  claim 7  wherein detecting the formation of an antigen-antibody complex comprises detecting human immunoglobulin in the antigen-antibody complex. 
     
     
         9 . The method of  claim 8  wherein detecting human immunoglobulin comprises contacting the antigen-antibody complex with a second antibody that binds to human immunoglobulin for a time and under conditions sufficient for said second antibody to bind to the human immunoglobulin in the complex and then detecting the bound anti-human immunoglobulin. 
     
     
         10 . The method of  claim 8  wherein the second antibody is labeled with a detectable marker or reporter molecule. 
     
     
         11 . A method for determining the response of a subject having pneumonia or an infection by  Streptococcus pneumoniae  to treatment with a therapeutic compound for said pneumonia or infection, said method comprising detecting antibodies against a PcpA antigen in a biological sample of the subject after treatment, wherein the amount of antibody detected is increased, unchanged, or decreased compared to the amount of antibody detectable in a biological sample of the subject obtained prior to treatment or that of a normal or healthy subject, wherein an unchanged or decreased amount of antibody after treatment indicates that the subject is not responding to treatment. 
     
     
         12 . The method of  claim 11  comprising detecting antibodies immunoreactive with at least one antigen selected from the group consisting of PhtD, PhtE, LytB and Ply. 
     
     
         13 . The method of any one of  claims 11  or  12  comprising contacting the biological sample with the antigen for a time and under conditions sufficient for an antigen-antibody complex to form and then detecting the formation of an antigen-antibody complex. 
     
     
         14 . The method of  claim 13  wherein detecting the formation of an antigen-antibody complex comprises detecting human immunoglobulin in the antigen-antibody complex. 
     
     
         15 . The method of  claim 14  wherein detecting human immunoglobulin comprises contacting the antigen-antibody complex with a second antibody that binds to human immunoglobulin for a time and under conditions sufficient for said second antibody to bind to the human immunoglobulin in the complex and then detecting the bound anti-human immunogobulin. 
     
     
         16 . The method of  claim 15  wherein the second antibody is labeled with a detectable marker or reporter molecule. 
     
     
         17 . The method of one of  claims 7  or  13  comprising performing an enzyme-linked immunosorbent assay (ELISA). 
     
     
         18 . The method of  claim 17  wherein the ELISA is a sandwich ELISA using a capture antibody and a detection antibody. 
     
     
         19 . A kit for detecting  Streptococcus pneumoniae  infection in a biological sample, the kit comprising: (i) the isolated and purified PcpA antigen; and, (ii) reagents for detecting the formation of an antigen-antibody complex. 
     
     
         20 . The kit of  claim 19  further comprising instructions for use. 
     
     
         21 . The method of any one of  claims 4 - 18  further comprising detecting  Streptococcus pneumoniae  nucleic acid in the biological sample. 
     
     
         22 . The method of  claim 21  wherein the nucleic acid corresponds to a nucleic acid encoding the PcpA protein. 
     
     
         23 . A solid matrix comprising an antigen selected from the group consisting of PcpA, Ply, PhtD, PhtE, and LytB adsorbed thereto. 
     
     
         24 . A solid matrix comprising a combination of isolated and purified antigens adsorbed thereto, said combination being selected from the group consisting of PcpA and Ply; PcpA and PhtD; PcpA and PhtE; PcpA and LytB; PcpA, Ply, and PhtD; PcpA, Ply, PhtD, and PhtE; Ply, PhtD, PhtE, and LytB; PcpA, PhtD, and PhtE; PcpA, PhtD, PhtE, and LytB; PcpA, PhtE, and LytB; PcpA, Ply, PhtE, and LytB; PcpA, PhtD, and LytB; PcpA, Ply, and LytB; PcpA, Ply, PhtD, and LytB; PepA, Ply, and PhtE; and, PcpA, PhtD, and PhtE. 
     
     
         25 . The solid matrix of  claim 23  or  24  wherein the solid matrix is selected from the group consisting of a microtiter plate or a microarray. 
     
     
         26 . A method of diagnosing infection by  Streptococcus pneumoniae  comprising contacting a biological sample with a solid matrix of any one of  claims 23 ,  24  or  25  under conditions suitable for an antibody reactive to one or more of the antigens to bind thereto, and detecting the binding of said antibody to at least one or none of said antigens.