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A Fusobacterium nucleatum subsp. animal strain isolated from human intestine and its application

A technology of Fusobacterium nucleatum and animals, applied in the direction of bacteria, microorganisms, microorganisms, etc., can solve the problems of inability to distinguish Fusobacterium nucleatum subspecies, lack of specificity, etc.

Active Publication Date: 2021-11-23
SHANGHAI TENTH PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The currently known primers and probes for detecting Fusobacterium nucleatum lack sufficient specificity, and they also have positive amplification for Fusobacterium hwasookii and Fusobacterium periodonticum, and they cannot be distinguished at the subspecies level of Fusobacterium nucleatum

Method used

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  • A Fusobacterium nucleatum subsp. animal strain isolated from human intestine and its application
  • A Fusobacterium nucleatum subsp. animal strain isolated from human intestine and its application
  • A Fusobacterium nucleatum subsp. animal strain isolated from human intestine and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] This example provides a Fusobacterium nucleatum subsp. animal strain THCT6b3 isolated from the intestine.

[0031] The strain has been preserved, named Fusobacterium nucleatumsubsp.animalis, its preservation number is CCTCC NO: M 2019367, and the preservation date is May 17, 2019. The preservation unit is the China Center for Type Culture Collection.

[0032] The 16SrRNA gene sequence of the strain is specifically shown in SEQ ID NO: 1, which is 99.80% consistent with the 16SrRNA gene sequence (NCBI accession number NR_113378.1) of the F.nucleatum subsp.animalis strain JCM 11025, and has 99.80% consistency with the F.nucleatum subsp.nucleatum The 16S rRNA gene sequence of strain ATCC 25586 (NCBI accession number NR_114702.1) has 98.38% identity.

[0033] The whole genome sequencing of this strain shows that this strain contains a circular chromosome, which is 2268559bp, GC content 27.32%, containing 2115 protein coding genes, 63 non-coding genes (45 tRNA genes, 5 sets o...

Embodiment 2

[0035] This example provides six specific DNA sequences for identifying the Fusobacterium nucleatum subsp. animal strain THCT6b3 in Example 1.

[0036] Through comparative genomics analysis, the six segments are used to identify the specific DNA sequences of the strain: THCT6b3_specific_region_1~THCT6b3_specific_region_6, the nucleotide sequences of which are shown in SEQ ID NO:8~SEQ ID NO:19 respectively.

[0037] The sequences of the amplification primers designed on the basis of the six specific DNA sequences used to identify the strain are shown in SEQ ID NO: 8 to SEQ ID NO: 19, respectively as follows (both 5' to 3') :

[0038]

[0039] The above six pairs of primers can be used for the specific recognition of THCT6b3 by amplifying the corresponding DNA fragments. Six pairs of primers can be applied to common PCR and real-time quantitative PCR. The above six sequences and corresponding primer pairs can be used alone or in combination.

Embodiment 3

[0041] This example provides specific DNA sequences that can be used to identify Fusobacterium nucleatum subspecies.

[0042] The specific method for obtaining the above-mentioned specific DNA sequence for identifying the subspecies of Fusobacterium nucleatum is: download the Fusobacterium nucleatum genome from NCBI, and combine the THCT6b3 genome; classify the subspecies according to the ANIb value; Through BLASTn analysis in the genome, find the regions that are conserved in all strains; perform BLASTn analysis on these conserved regions and the genomes of other subspecies, and keep the regions that do not exist in the genomes of other subspecies; thus obtain subspecies-specific DNA sequences.

[0043] Three specific DNA sequences: animalis_specific_region_1~animalis_specific_region_3, respectively shown in SEQ ID NO:20~SEQ ID NO:22. The subspecies-specific DNA sequence includes the sequence shown in SEQ ID NO:20~SEQ ID NO:22 or the sequence shown in SEQ ID NO:20~SEQ ID NO:2...

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Abstract

The present invention provides a Fusobacterium nucleatum subsp. animal strain THCT6b3 isolated from the intestinal tract. The 16SrRNA gene sequence of the strain is specifically shown in SEQ ID NO:1. The strain is classified as Fusobacterium nucleatum subsp.animalis, its preservation number is CCTCC NO: M 2019367, and the preservation date is May 17, 2019. The preservation unit is the China Center for Type Culture Collection. The invention also provides the application of the bacterial strain in the detection of Fusobacterium nucleatum in intestinal tract, and establishes a PCR-based molecular identification method of Fusobacterium nucleatum animal subspecies for the first time. The invention can promote the discovery of the composition and source of intestinal Fusobacterium nucleatum in patients with colorectal cancer, and will help to analyze the molecular epidemiological characteristics of Fusobacterium nucleatum.

Description

technical field [0001] The invention relates to the field of Fusobacterium nucleatum, in particular to a Fusobacterium nucleatum subsp. animal strain isolated from the intestinal tract and its application in the detection of the Fusobacterium nucleatum in the intestinal tract. Background technique [0002] Fusobacterium nucleatum (Fusobacterium nucleatum) is a human commensal bacterium, a Gram-negative obligate anaerobic bacterium that colonizes the oral cavity; it is also an opportunistic pathogenic bacterium that plays a leading role in periodontal disease, often in subgingival plaque obtained in isolation. Fusobacterium nucleatum can also cause infections in other parts of the body, including brain, lung, liver, blood, joints, chest, abdomen, intrauterine, etc. Fusobacterium nucleatum can currently be divided into four subspecies, including F.nucleatum subsp.nucleatum, F.nucleatum subsp.polymorphum, and F.nucleatum subsp.vincentii and animal subspecies (F. nucleatum sub...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12Q1/689C12N15/11C12R1/01
CPCC12Q1/689C12N1/205C12R2001/01
Inventor 秦环龙蔚青毕德玺李豪朱崟
Owner SHANGHAI TENTH PEOPLES HOSPITAL
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