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Means and methods for distinguishing FECV and FIPV

A one-part, feline coronavirus technology, applied in biochemical equipment and methods, medical preparations containing active ingredients, pharmaceutical formulations, etc., can solve problems such as inability to distinguish other diseases, inability to qualify, etc.

Active Publication Date: 2012-11-21
乌得勒支大学控股有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Fourth, as fully discussed by Pedersen in his recent review, FECV79-1683 cannot qualify as a true FECV (Pedersen 2009)
Because there is no specific diagnostic test for FIPV, it is often not possible to distinguish FIP from other disorders that share some of the same symptoms

Method used

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  • Means and methods for distinguishing FECV and FIPV
  • Means and methods for distinguishing FECV and FIPV
  • Means and methods for distinguishing FECV and FIPV

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] In this example, 6 clinical samples (stools) were analyzed. RNA was extracted from clinical samples, RT-PCR was performed on the extracted RNA, and after the first PCR (1st run) and nested PCR (2nd run) it was run by agarose gel electrophoresis (see image 3 ) to analyze the product.

[0073] Materials and methods

[0074] Nested RT-PCR was used to amplify the region of the FCoV spike protein gene containing the targeted mutation. Genomic RNA was extracted from feces of 6 healthy cats using QIAamp Viral RNA Mini Kit and Qiagen RNeasy Mini Kit (Qiagen, Inc.) according to the manufacturer's instructions. cDNA was synthesized using M-MLV reverse transcriptase (RT), followed by polymerase chain reaction (PCR) amplification using Taq DNA polymerase. All enzymes were used according to the manufacturer's instructions (Promega Corp., Madison, WI). Both reactions were primed with specific primers (see Primer Table 1). Primers were designed using the FCoV genome sequences wi...

Embodiment 2

[0078] In this example, fecal or plasma samples from 47 healthy cats, clinical samples from 54 cats with proven FIP, and fecal samples from 14 cats with proven FIP were analyzed.

[0079] Materials and methods

[0080] Extraction of genomic RNA, synthesis, amplification and sequencing of cDNA were carried out according to the materials and methods in Example 1.

[0081] result

[0082] The nucleic acid sequence encoding methionine at amino acid position 1049 was detected in feces or plasma (47 / 47) from all healthy cats ( Figure 4A ). found out later Figure 4A 包含源自证实患有FIP的猫的5条序列(Q093501030_326B_4546.scf、Q093501032_327B_4546.scf、Q093501036_321S_4546.scf、Q093501038_321A_4546.scf和Q093501046_K11_019.ab1),这意味着在来自健康猫的42 / 42源自粪便或 Methionine exists at amino acid position 1049 in FCoV in plasma. 2 / 54 also amplified from cats with proven FIP derived from lesions ( Figure 4B ) RNA and 12 / 14 from feces ( Figure 4C This sequence is observed in the RNA of ). Importantly, it was dem...

Embodiment 3

[0084] We continue to collect samples and cats through Dutch veterinarians and owners. In this example, the following samples were analyzed:

[0085] Fecal samples from 352 healthy cats;

[0086] White blood cell samples from 89 healthy or non-FIP suspicious cats;

[0087] Plasma samples from 89 healthy or non-FIP suspicious cats;

[0088] Serum samples from 56 healthy or non-FIP suspicious cats;

[0089] FIP lesion samples (mesenteric lymph node (LN) and / or kidney and / or spleen and / or omentum and / or lung and / or LN and / or liver and / or ascites) from 97 cats with proven FIP;

[0090] WBC samples from 34 cats with proven FIP;

[0091] Plasma samples from 34 cats with proven FIP; and

[0092] Serum samples from 15 cats with proven FIP.

[0093] Materials and methods

[0094] Extraction of genomic RNA, synthesis, amplification and sequencing of cDNA were carried out according to the materials and methods in Example 1.

[0095] result

[0096] Samples from healthy cats

[...

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PUM

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Abstract

The invention provides methods and means for distinguishing FECV and FIPV, and methods and means for determining whether FIPV is present in a sample. Further provided are primers and probes for detecting FIPV specific nucleic acid sequences encoding a spike protein, antibodies for detecting a FIPV, and an immunogenic composition and use thereof for eliciting an immune response against a feline coronavirus, preferably a FIPV.

Description

technical field [0001] The present invention relates to the field of veterinary diagnostics, more particularly the present invention relates to the field of feline coronaviruses and their identification. Background technique [0002] Feline coronavirus (FCoV) is a virus in domestic and non-domestic cats (including but not limited to cats, lions, tigers, leopards, jaguars, lynxes, caracals, cheetahs, pumas, and servals) common pathogens. FCoV seropositivity can be as high as 90% in multiple cat households. FCoV is closely related to canine coronavirus (CCoV) and porcine transmissible gastroenteritis virus (TGEV). There are two serotypes (type I and type II) of FCoV, of which serotype I is the main one, accounting for 80-95% of FCoV infections. Type II FCoV may result from RNA recombination in animals co-infected with serotype I FCoV and CCoV, in which the spike gene of CCoV or part thereof is incorporated into the genome of FCoV (this apparently rarely occurs event). Fel...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70
CPCC12Q1/701C12Q2600/156C12Q1/04A61K39/00A61P31/12A61P37/04C12Q1/6876A61K39/215A61P31/14
Inventor 彼得鲁斯·约瑟夫斯·玛丽·罗提尔张惠雯赫尔曼·F·艾格伯林克
Owner 乌得勒支大学控股有限公司
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