Ehrlichia canis DIVA (differentiate infected from vaccinated animals)

A method for determining the presence or absence of an antibody or fragment thereof, in a test sample, wherein the antibody or fragment thereof specifically binds to one or more purified polypeptides consisting of the amino acid sequence set forth in SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:22 or combinations thereof comprising:

• contacting the test sample with one or more purified polypeptides consisting of the amino acid sequence set forth in SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:22, or combinations thereof, wherein the one or more purified polypeptides are optionally linked to a heterologous protein, an indicator reagent, an amino acid spacer, a signal sequence, a stop transfer sequence, a transmembrane domain, a protein purification ligand, or a combination thereof, under conditions suitable for specific binding of the one or more purified polypeptides to the antibody or fragment thereof;
• and
• detecting the presence or absence of specific binding;
• wherein the presence of specific binding indicates the presence of the antibody or fragment thereof, and wherein the absence of specific binding indicates the absence the antibody or fragment thereof.

A method of distinguishing between an animal that has been infected with E. canis from an animal that has not been infected with E. canis, the method comprising:

• (a) contacting a biological sample from an animal with one or more purified polypeptides consisting of the amino acid sequence set forth in SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:22, or combinations thereof, wherein the one or more purified polypeptides are optionally linked to a heterologous protein, an indicator reagent, an amino acid spacer, a signal sequence, a stop transfer sequence, a transmembrane domain, a protein purification ligand, or a combination thereof;
• and
• (b) detecting whether antibodies in the sample specifically bind to the one or more purified polypeptides;
• wherein if antibodies in the sample specifically bind to the one or more purified polypeptides, then the animal has been infected with E. canis.

A method of distinguishing between animals that (1) have been infected with Ehrlichia canis and animals that (2) have not been infected with E. canis regardless of whether the animal has been vaccinated with an E. canis vaccine, wherein the E. canis vaccine does not elicit antibodies that specifically bind to an antigen consisting of SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, or SEQ ID NO:22, the method comprising:

• (a) contacting a biological sample from an animal with one or more purified E. canis polypeptides that do not specifically bind to antibodies that are a component of the animal's immune response to an E. canis vaccine;
• wherein the one or more purified E. canis polypeptides consist of the amino acid sequence set forth in SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:22, or combinations thereof and wherein, optionally, the one or more purified E. canis polypeptides are linked to a heterologous protein, an indicator reagent, an amino acid spacer, a signal sequence, a stop transfer sequence, a transmembrane domain, a protein purification ligand, or a combination thereof;
• (b) detecting whether antibodies in the sample specifically bind to the one or more purified E. canis polypeptides;
• and
• (c) wherein if antibodies in the sample specifically bind to the one or more purified E. canis polypeptides, then the animal has been infected with E. canis.