Method for detecting chikungunya virus

Abstract

Compositions, methods and kits for detecting Chikungunya viral nucleic acids. Particularly described are methods for detecting very low levels of the viral nucleic acids using nucleic acid amplification.

What is claimed is: 
     
         1 . A kit for amplifying and detecting a Chikungunya virus (CHIKV) nucleic acid sequence, comprising:
 (a) a first primer up to 100 bases long and comprising a target-complementary 3′ terminal sequence consisting of 15-44 contiguous bases of SEQ ID NO:46, said target-complementary 3′ terminal sequence of said first primer being fully contained within the sequence of SEQ ID NO:46, and said first primer optionally comprising a first primer 5′ sequence that is not complementary to CHIKV nucleic acids;   (b) a second primer up to 100 bases long and comprising a target-complementary 3′ terminal sequence consisting of 15-40 contiguous bases of SEQ ID NO:65, said target-complementary 3′ terminal sequence of said second primer being fully contained within the sequence of SEQ ID NO:65, and said second primer optionally comprising a second primer 5′ sequence that is not complementary to CHIKV nucleic acids;   (c) a third primer up to 100 bases long and comprising a target-complementary 3′ terminal sequence consisting of 15-43 contiguous bases of SEQ ID NO:66, said target-complementary 3′ terminal sequence of said third primer being fully contained within the sequence of SEQ ID NO:66, and said third primer optionally comprising a third primer 5′ sequence that is not complementary to CHIKV nucleic acids; and   (d) a hybridization probe for detecting a nucleic acid amplification product synthesized using said primers,
 wherein said primers and said hybridization probe are in packaged combination with each other. 
   
     
     
         2 . The kit of  claim 1 , wherein the target-complementary 3′ terminal sequence of said first primer is SEQ ID NO:106. 
     
     
         3 . The kit of  claim 1 , wherein the target-complementary 3′ terminal sequence of said second primer is SEQ ID NO:145. 
     
     
         4 . The kit of  claim 1 , wherein the target-complementary 3′ terminal sequence of said third primer is SEQ ID NO:146. 
     
     
         5 . The kit of  claim 1 , wherein said hybridization probe is up to 39 bases in length and comprises 15-39 contiguous bases of SEQ ID NO:82. 
     
     
         6 . The kit of  claim 5 , wherein said hybridization probe comprises SEQ ID NO:162. 
     
     
         7 . A method for detecting whether a Chikungunya virus (CHIKV) nucleic acid sequence is present in a test sample comprising nucleic acids, said method comprising the steps of:
 (a) contacting nucleic acids of the test sample with a set of amplification oligonucleotides comprising,
 (i) a first primer up to 100 bases long and comprising a target-complementary 3′ terminal sequence consisting of 15-44 contiguous bases of SEQ ID NO:46, said target-complementary 3′ terminal sequence of said first primer being fully contained within the sequence of SEQ ID NO:46, and said first primer optionally comprising a first primer 5′ sequence that is not complementary to CHIKV nucleic acids, 
 (ii) a second primer up to 100 bases long and comprising a target-complementary 3′ terminal sequence consisting of 15-40 contiguous bases of SEQ ID NO:65, said target-complementary 3′ terminal sequence of said second primer being fully contained within the sequence of SEQ ID NO:65, and said second primer optionally comprising a second primer 5′ sequence that is not complementary to CHIKV nucleic acids, and 
 (iii) a third primer up to 100 bases long and comprising a target-complementary 3′ terminal sequence consisting of 15-43 contiguous bases of SEQ ID NO:66, said target-complementary 3′ terminal sequence of said third primer being fully contained within the sequence of SEQ ID NO:66, and said third primer optionally comprising a third primer 5′ sequence that is not complementary to CHIKV nucleic acids; 
   (b) performing an in vitro nucleic acid amplification reaction using nucleic acids of the test sample as templates together with said set of amplification oligonucleotides, whereby, if said test sample comprises said CHIKV nucleic acid sequence, there is produced an amplification product; and   (c) detecting with a hybridization probe any of said amplification product that may have been produced in the in vitro nucleic acid amplification reaction,
 wherein detecting said amplification product in an amount greater than a cutoff value indicates that the CHIKV nucleic acid sequence is present in the test sample, and 
 wherein detecting said amplification product in an amount less than the cutoff value indicates that the CHIKV nucleic acid sequence is absent from the test sample.