Methods, devices, kits and compositions for detecting roundworm, whipworm and hookworm
Methods, devices, kits and compositions for detecting the presence or absence of one or more helminthic coproantigens in a sample are disclosed herein. The methods, devices, kits and compositions of the present invention may be used to confirm the presence or absence of roundworm, whipworm and/or hookworm in a fecal sample from a mammal and may also be able to distinguish between one or more helminth infections. Confirmation of the presence or absence of roundworm, whipworm and/or hookworm in the mammal may be made, for example, for the purpose of selecting an optimal course of treating the mammal and/or for the purpose of determining whether the mammal has been rid of the infection after treatment has been initiated.
1. A method of diagnosing and treating a mammal infected with one or more helminthic worms, the method comprising:
(a) contacting a sample from the mammal with at least two antibodies selected from the group consisting of:
(i) a first antibody capable of specifically binding a roundworm coproantigen, but not a whipworm or hookworm coproantigen;
(ii) a second antibody capable of specifically binding a whipworm coproantigen, but not a roundworm or hookworm coproantigen; and
(iii) a third antibody capable of specifically binding a hookworm coproantigen, but not a whipworm or roundworm coproantigen;
(b) forming antibody-coproantigen complexes in the presence of the coproantigens, if any, in the sample;
(c) detecting the presence or absence of the antibody-coproantigen complexes, if any;
(d) diagnosing the mammal as having:
(i) a roundworm infection if the first antibody-coproantigen complex is present;
(ii) a whipworm infection if the second antibody-coproantigen complex is present; and
(iii) a hookworm infection if the third antibody-coproantigen complex is present; and
(e) administering an anti-helminthic agent to treat the mammal having the roundworm infection, the whipworm infection or the hookworm infection or combination thereof.
2. The method of claim 1 , wherein:
(a) the first antibody is capable of specifically binding a polypeptide having an amino acid sequence of SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 38, or Copro6728; or the first antibody was raised against an extract of whole roundworms, or extract of roundworm reproductive organs, or extract of roundworm intestines;
(b) the second antibody is capable of specifically binding a polypeptide having an amino acid sequence of SEQ ID NO:3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9; and
(c) the third antibody is capable of specifically binding a polypeptide having an amino acid sequence of SEQ ID NO:33 or SEQ ID NO: 34, or CoproASP5.
3. The method of claim 1 , wherein the first antibody is capable of specifically binding a polypeptide having an amino acid sequence of SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 38, or Copro6728; or the first antibody was raised against an extract of whole roundworms, or extract of roundworm reproductive organs, or extract of roundworm intestines.
4. The method of claim 1 , wherein the second antibody is capable of specifically binding a polypeptide having an amino acid sequence of SEQ ID NO:3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9.
5. The method of claim 1 , wherein the third antibody is capable of specifically binding a polypeptide having an amino acid sequence of SEQ ID NO:33 or SEQ ID NO: 34, or CoproASP5.
6. The method of claim 1 , wherein the mammal is a canine or a feline.
7. The method of claim 1 , wherein the roundworm is Toxocara canis or Toxocara cati, Toxocara vitulorum, Toxascaris leonina, Baylisascaris procyonis, Ascaridia galli, Parascaris equorum, Ascaris suum, Ascaris lumbricoides, Anisakis simplex , or Pseudoterranova decipiens.
8. The method of claim 1 , wherein the whipworm is Trichuris vulpis, Trichuris campanula, Trichuris serrata, Trichuris suis, Trichuris trichiura, Trichuris discolor or Trichocephalus trichiuris.
9. The method of claim 1 , wherein the hookworm is Ancylostoma caninum, Ancylostoma braziliense, Ancylostoma duodenal, Ancylostoma ceylanicum, Ancylostoma tubaeforme and Ancylostoma pluridentatum, Necator americanus , or Uncinaria stenocephala.
10. The method of claim 1 , wherein the step of detecting the presence or absence of the complexes further includes the step of providing at least one secondary antibody that binds to the one or more complexes.
11. The method of claim 10 , wherein the secondary antibody is labeled.
12. The method of claim 1 , wherein the first, second and third antibodies are immobilized on a solid support.
13. The method of claim 12 , wherein the solid support forms part of an enzyme-linked immunosorbent assay device.
14. The method of claim 13 , wherein the enzyme-linked immunosorbent assay device is a lateral flow immunoassay device.
15. The method of claim 1 , further comprising the step of contacting the sample with one or more reagents to detect one or more members of the group consisting of: one or more non-worm parasites, heartworm, one or more viruses, one or more fungi, and one or more bacteria.
16. The method of claim 15 , wherein the one or more reagents for the detection are one or more antibodies or one or more antigens recognized by antibodies specific for one or more non-worm parasites, heartworm, one or more viruses, one or more fungi or one or more bacteria.
17. The method of claim 1 , further comprising the step of determining the presence or absence of a nucleic acid from the roundworm, whipworm and hookworm.
18. The method of claim 17 , wherein the nucleic acid has a nucleotide sequence of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 38, Copro6728, or CoproASP5 or is a conservative variant of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 38, Copro6728, or CoproASP5.
19. The method of claim 17 , wherein the step of determining presence or absence of the nucleic acid is carried out by using a polymerase chain reaction (PCR)-based assay.
20. The method of claim 1 , wherein the first, second and third antibodies do not specifically bind any coproantigen derived from heartworm.
21. The method of claim 1 , wherein said anti-helminthic agent is selected from praziquantel/pyrantel pamoate and milbemycin oxime.
22. The method of claim 1 , wherein subsequent to step (e), further comprising repeating steps (a) to (c).