beta-lactamase

The β-lactamase refers to an inactivating enzyme capable of catalyzing the hydrolysis of an amide bond in a β-lactam ring in a biomolecule. Bacterial production of β-lactamase is the main mechanism of bacterial resistance to β-lactam antibiotics. Β-lactamase antibiotics including penicillins, cephalosporins, atypical β-lactams, etc., are the most widely used, the most widely studied, and the most widely used drugs. Basic Information Specialist classification: Infectious disease inspection and classification: biochemical examination Applicable gender: whether men and women apply fasting: not fasting Tips: According to the nutritional status of the whole body, milk, eggs, fruits, soy milk, etc. are given in the meal. Normal value The type and proportion of the flora in the body are normal, and the human body is in a dynamic balance. Clinical significance The mechanism of bacterial resistance: 1. Inactivated enzymes - mainly beta-lactamases (more than 200 types), which render antibiotics ineffective. 2, target changes - the target changes with antibiotics, so that the role of antibiotics decreased. 3, the change of bacterial membrane permeability - so that antibiotics can not or rarely enter the bacteria to reach the target. The development of bacterial resistance from hospital strains (such as Enterobacteriaceae, Staphylococcus aureus) to out-of-hospital strains (such as Streptococcus pneumoniae, Streptococcus pyogenes, Neisseria gonorrhoeae), resistance to resistance to a single drug The drug develops to multi-drug resistance, such as Staphylococcus aureus resistance to penicillin; ESBL strain is resistant to a variety of commonly used antibiotics. Abnormal results Enterobacteriaceae, Staphylococcus aureus, Streptococcus pneumoniae, Streptococcus pyogenes, Neisseria gonorrhoeae and other infections, but the use of antibiotics is not effective or the effect is not obvious, bacteria produce drug resistance. The people in need of examination have patients with the above conditions. Precautions Inappropriate crowd: No. Forbidden before examination: Pay attention to normal eating habits and pay attention to personal hygiene. Requirements for inspection: Actively cooperate with the doctor. Inspection process Detection of β-lactamase In clinical specimens, Staphylococcus aureus, Haemophilus influenzae, Neisseria gonorrhoeae, and a few strains of Enterococcus were recently found to be resistant to penicillin and ampicillin. Since the above strain produces β-lactamase, the aforementioned antibiotic is hydrolyzed. The following methods are used to detect the enzyme: 1. Microbial activity disappearance method This method is an indicator strain of Bacillus subtilis that is highly sensitive to penicillin. If the strain to be tested produces β-lactamase and destroys penicillin, there is Bacillus subtilis growth on both sides of the line, otherwise the indicator is still very large. Antibacterial ring. 2, starch-iodine determination The beta lactamase destroys the beta lactam ring, and the iodine combines with the opened beta lactam ring to convert the blue starch-iodine complex to colorless. METHODS: A suspension of penicillin 6000 μg/ml was prepared with pH 6.0 phosphate buffer solution. 0.1 ml of the solution was placed in a well of a microdilution plate. Several colonies of the test strain were selected and suspended therein, shaken for 30 min, and allowed to stand at room temperature for 1 h. Add 2 drops of starch solution, add 1 drop of iodine solution, mix and observe the color change immediately. When the positive person adds iodine solution, it will appear blue immediately, such as disappearing blue within 10min, suggesting that the bacteria produce β-inner Amidase. 3. Acid measurement Penicillin is hydrolyzed by β-endase to penicillin acid, the pH value drops below 6.8, with phenol red indicator, from red (purple) color (stock solution: citrate buffer pH 8.5) → yellow (pH 6.8 the following). 4, cephalosporin thiophene color reaction The β-lactam ring of Nitrocefin is opened by β-lactamase and the matrix changes from yellow to red. Liquid method: 10 mg of cefa nitrothiophene was dissolved in 1 ml of dimethyl sulfoxide, and then diluted 1:20 with 0.1 mol/L PBS (pH 7.0), and finally the concentration was 500 μg/m1, and the solution was yellow or pale orange. Store 4 to 10 ° C for several weeks. Take 0.05ml of the solution in the well of the micro-dilution plate or in the small test tube, pick the colony of the test, make a thick suspension, mix with the matrix liquid in the concave hole, and change the cephalosporin nitrothiophene from yellow to room temperature at room temperature for 10 to 30 minutes. Red is positive. If the color is not obvious, the observation can be extended for 6 hours. It has been reported that the results of the paper method for the detection of β-lactamase by Haemophilus influenzae and Neisseria gonorrhoeae are accurate, but for some strains, a small amount of β-lactamase, which is tightly bound to the bacterial cells, such as saprophytic Staphylococcus, etc., better by liquid method. Quality control is applied to any of the above detection methods. Not suitable for the crowd Generally no taboos. Adverse reactions and risks Generally no complications and harm.