Method for isolation and identification of bacillus circulans

A technique for the identification of Bacillus circulans, which is applied in the field of isolation and identification of Bacillus circulans, achieving the effect of high screening accuracy

Active Publication Date: 2019-11-19
NANJING INST OF PROD QUALITY INSPECTION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In order to ensure the safety of food packaging paper, the rapid and efficient isolation and identification of Bacillus circulans has strong application value and social significance. At present, there is no simple and rapid method for the isolation and identification of Bacillus circulans from environmental bacteria.

Method used

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  • Method for isolation and identification of bacillus circulans
  • Method for isolation and identification of bacillus circulans
  • Method for isolation and identification of bacillus circulans

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Embodiment 1: Establishment of optimal culture conditions

[0060] Generally, the choice of selective enrichment medium is to add nutrients or antibiotic components that are sensitive to other bacteria and insensitive to the target bacteria on the basis of the basic medium. The basal medium of this example is improved tryptic soybean broth, and the enrichment medium of Bacillus is not selected in order to reduce the dominant proliferation of other Bacillus with high nutritional requirements.

[0061] Sample preparation: The screening subject of this experiment is the food contact paper purchased in the market, routine supervision and random inspection, and inspection by enterprises.

[0062] Control substance: Take 25g of samples without any growth of miscellaneous bacteria verified by the experiment, add positive strains respectively: Bacillus circulans, Bacillus lysine, Bacillus subtilis, Streptococcus hemolyticus, and obtain 1~9×10 4Bacterial solution at a concentra...

Embodiment 2

[0080] Embodiment 2: Antibiotic and bacteriostatic compound selection

[0081] Due to the complex composition of environmental microorganisms, too many bacteria in the enrichment process will have more interference in the later identification process. The screening of Bacillus, according to the sensitivity of different microorganisms to different antibiotics, inhibits the growth of miscellaneous bacteria and ensures the dominant growth of Bacillus circulans. The following antibiotics and bacteriostatic compounds were selected for experimentation:

[0082] (1) Cephalosporin antibiotics have a broad antibacterial spectrum and are effective against both Gram-positive and Gram-negative bacteria. Sensitive bacteria are Pasteurella multocida, Pasteurella hemolyticus, Salmonella, Escherichia coli, Streptococcus, Staphylococcus, etc. Some Pseudomonas aeruginosa and Enterococcus are drug-resistant.

[0083] (2) Lincomycin hydrochloride is a bacteriostatic agent, and sensitive bacteri...

Embodiment 3

[0110] Example 3: Separation and Identification of Columbia Blood Agar Plates

[0111] According to the experimental results of Examples 1 and 2, the growth status of different positive bacterial strains in antibiotics was determined, and the Columbia blood agar plate added with novobiocin and nalidixic acid sodium antibiotics was determined as the isolation and identification medium.

[0112] Columbia Blood Agar

[0113] Tryptone 12.0g

[0114] Animal tissue protein digestion solution 5.0g

[0115] Yeast Extract 3.0g

[0116] Beef Extract 3.0g

[0117] Cornstarch 1.0g

[0118] Sodium chloride 5.0g

[0119] Agar 13.5g

[0120] Distilled water 1000.0mL

[0121] Dissolve each component in distilled water, heat to dissolve, sterilize at 121°C for 15 minutes, add sterile defibrinated sheep blood 5mL / 100mL and antibiotics, shake well and distribute for use.

[0122] Add antibiotics:

[0123] Novobiocin 0.01g

[0124] Nalidixic acid sodium 0.01g

[0125] Sample: Take 25g ...

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Abstract

The invention provides a method for isolation and identification of bacillus circulans, wherein the method comprises the following steps: (1) environmental bacteria containing bacillus circulans is subjected to selective enrichment culture by a selective enrichment culture medium, the proliferation of non-bacillus circulans is reduced, and the bacillus circulans is highly efficiently proliferates,wherein the selective enrichment culture medium is a modified tryptone soybean broth culture medium containing cephalosporin antibiotics and benzylpenicillin potassium; and (2) the bacterial solutionobtained in the step (1) is cultured in a Columbia blood agar plate containing antibiotics, and the strains having hemolytic ring appearing are isolated and are identified according to the colony morphology. The method can further use the molecular biological method to identify the strains isolated in the step (2). The method can simply and quickly isolate and identify bacillus circulans from theenvironmental bacteria, and has high screening precision.

Description

technical field [0001] The invention belongs to the field of microorganism detection, and in particular relates to a method for isolating and identifying circulus bacillus. Background technique [0002] Bacillus (Bacillus) is a Gram-positive, aerobic or facultative anaerobic endospore rod-shaped bacteria, due to the production of stress-resistant endospores, Bacillus microorganisms can resist various extreme environments, Such as high temperature, extreme acid, extreme salt, and resistance to various fungicides. Bacillus includes nearly 50 species such as Bacillus subtilis, Bacillus licheniformis, Bacillus pumilus, Bacillus circulus, Bacillus mycoides, Bacillus polymyxa, Bacillus anthracis, and Bacillus cereus. [0003] The conventional detection method for Bacillus is to judge according to the characteristics of colonies and bacteria combined with identification materials of strains, but it is easy to misjudgment when facing samples with inconspicuous phenotypic characteri...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/02C12Q1/04C12R1/07
CPCC12N1/20C12N1/02C12Q1/045
Inventor 陈碧霄周帆张驰杨淼乔玲韩冰闻博芹徐宁杨洋王迪冯诗旸王灿李园园王晓丽祝晨辰刘艳容杨婷婷杜亚琼刘秀吕伟孙建娜张辰刘虹杜鹃
Owner NANJING INST OF PROD QUALITY INSPECTION
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