Application of antibiotic substitute drug melatonin in resisting infection of meningitis escherichia coli pathogens on child patients

Abstract

The invention provides application of an antibiotic substitute drug melatonin in preventing the infection of meningitis escherichia coli pathogens on child patients. Meningitis-type escherichia coli is separated and identified from muscovy duck cerebrospinal fluid, a meningitis model of 3-week-old ICR mice is constructed for the first time by utilizing the strain, and the successful construction of the mouse meningitis model is evaluated from the aspects of neurological symptom scoring, mortality difference statistics, the damage degree of a blood-brain barrier, expression of meningitis-related inflammatory factors and the like respectively. With antibiotics as references, the mice are treated with the melatonin and the four combined antibiotics respectively, one week later, the disease onset conditions of the meningitis mice are detected, results show that through the melatonin and the combined use of the antibiotics, the neurological symptoms of the meningitis mice can both be reduced, the survival rate of the meningitis mice is increased, integrity keeping of the blood-brain barrier is promoted by the melatonin, the invasion of inflammatory cells into the central nervous systemis reduced, the disease onset of the meningitis in the mice is prevented through the function of the brain-gut axis, and a new antibiotic substitute and treatment strategy are provided for preventingneonatal meningitis diseases.

Description

technical field

[0001] The invention relates to the application of melatonin (melatonin), a medicine replacing antibiotics, to resist meningitis Escherichia coli pathogenic infection in children. The present invention further provides a drug for replacing antibiotics to prevent the onset of meningitis and its application characteristics, which is expected to provide a new strategy for preventing the onset of meningitis. Background technique

[0002] Since Vieusseux first reported bacterial meningitis (BM) in 1806, it has been considered as an important fatal disease, and it is an infection of the central nervous system that is of great clinical concern in humans and animals. There are many pathogenic bacteria that cause bacterial meningitis, among which the mortality rate of neonatal meningitis caused by Escherichia coli ranks among the highest in the world. Meningitis Escherichia coli mainly include neonatal meningitis Escherichia coli (NMEC) and avian meningitis Escherich...

Examples

Embodiment 1

[0044] 1. Main drugs and reagents

[0045] Melatonin (melatonin, MT) was purchased from Sigma, USA; formalin, PBS, DAB reagent, hematoxylin, eosin, distilled water. Tryptone (Tryptone) and yeast extract (Yeast extract) were purchased from Oxoid Company; sodium chloride (NaCl), diamine hydrogen citrate (C 6 h 14 N 2 o 7 ), sodium acetate (CH 3 COONa), dipotassium hydrogen phosphate (K 2 HPO 4 ), magnesium sulfate (MgSO 4 ), manganese sulfate (MnSO 4 ) and other analytical reagents were purchased from Sinopharm Chemical Reagent Co., Ltd.; MacConkey agar was purchased from Haibo Company; , glucose (Glucose) were purchased from Sigma Company; Faststart Universal SYBR GREEN Master (ROX) Fluorescence Quantitative Kit was purchased from Roche Company; Reagents are domestic analytical grade reagents.

[0046] 2. Main instruments

[0047] Ultra-clean workbench was purchased from Suzhou Purification Equipment Co., Ltd.; fluorescent quantitative PCR7500 was purchased from ABI ...

Embodiment 2

[0082] (1) Preparation of APEC TW-XM strain

[0083] First recover the bacteria from the seed preservation tube, line the four zones on the LB solid plate, and culture overnight in a 37°C incubator. The next day, pick a single colony and culture it in the liquid LB medium for 16 hours, and then follow the ratio of bacteria and medium 1:100 Transfer the bacterial solution to the liquid LB medium and cultivate to OD 600 When =1, collect the bacterial liquid, wash the bacterial suspension twice with PBS, resuspend it with normal saline, and dilute it to cfu=10 7 , for mouse infection.

[0084] (2) Experiment of melatonin preventing the onset of meningitis in mice

[0085] The mice were randomly divided into four groups, 6 in the (NS) group, 6 in the melatonin (MT) group, and 26 in the APEC TW-XM (A) group (10-bacteria counting, 6-pathological sections, 10 for Statistical survival rate), APEC TW-XM+melatonin (A+MT) group 26 (10-bacteria count, 6-pathological section, 10 for sta...

Embodiment 3

[0104] (1) Preparation of APEC TW-XM strain

[0105] With embodiment 2.

[0106] (2) Melatonin regulates intestinal flora to prevent meningitis

[0107] The mice were randomly divided into four groups, APEC TW-XM group: daily intraperitoneal injection of 100 μl of normal saline; daily replacement of sterile water; A+MT group: daily intraperitoneal injection of 100 μl of 30 mg / kg MT; daily replacement of sterile water; Group A+antibiotics: intraperitoneally inject 100 μl of normal saline every day; replace sterile water containing antibiotics every day; A+antibiotics+MT: inject 100 μl 30 mg / kg MT intraperitoneally every day; replace sterile water containing antibiotics every day; 8 days for challenge (cfu=10 7 ) to collect samples.

[0108] (3) Sample collection

[0109] Permeability detection (EB staining): Same as Example 1.

[0110] Detection of inflammatory factors: same as Example 1.

[0111] The making of pathological section: with embodiment 2.

[0112] Notes to t...

technical field

[0001] The invention relates to the application of melatonin (melatonin), a medicine replacing antibiotics, to resist meningitis Escherichia coli pathogenic infection in children. The present invention further provides a drug for replacing antibiotics to prevent the onset of meningitis and its application characteristics, which is expected to provide a new strategy for preventing the onset of meningitis. Background technique

[0002] Since Vieusseux first reported bacterial meningitis (BM) in 1806, it has been considered as an important fatal disease, and it is an infection of the central nervous system that is of great clinical concern in humans and animals. There are many pathogenic bacteria that cause bacterial meningitis, among which the mortality rate of neonatal meningitis caused by Escherichia coli ranks among the highest in the world. Meningitis Escherichia coli mainly include neonatal meningitis Escherichia coli (NMEC) and avian meningitis Escherich...

Examples

Embodiment 1

[0044] 1. Main drugs and reagents

[0045] Melatonin (melatonin, MT) was purchased from Sigma, USA; formalin, PBS, DAB reagent, hematoxylin, eosin, distilled water. Tryptone (Tryptone) and yeast extract (Yeast extract) were purchased from Oxoid Company; sodium chloride (NaCl), diamine hydrogen citrate (C 6 h 14 N 2 o 7 ), sodium acetate (CH 3 COONa), dipotassium hydrogen phosphate (K 2 HPO 4 ), magnesium sulfate (MgSO 4 ), manganese sulfate (MnSO 4 ) and other analytical reagents were purchased from Sinopharm Chemical Reagent Co., Ltd.; MacConkey agar was purchased from Haibo Company; , glucose (Glucose) were purchased from Sigma Company; Faststart Universal SYBR GREEN Master (ROX) Fluorescence Quantitative Kit was purchased from Roche Company; Reagents are domestic analytical grade reagents.

[0046] 2. Main instruments

[0047] Ultra-clean workbench was purchased from Suzhou Purification Equipment Co., Ltd.; fluorescent quantitative PCR7500 was purchased from ABI ...

Embodiment 2

[0082] (1) Preparation of APEC TW-XM strain

[0083] First recover the bacteria from the seed preservation tube, line the four zones on the LB solid plate, and culture overnight in a 37°C incubator. The next day, pick a single colony and culture it in the liquid LB medium for 16 hours, and then follow the ratio of bacteria and medium 1:100 Transfer the bacterial solution to the liquid LB medium and cultivate to OD 600 When =1, collect the bacterial liquid, wash the bacterial suspension twice with PBS, resuspend it with normal saline, and dilute it to cfu=10 7 , for mouse infection.

[0084] (2) Experiment of melatonin preventing the onset of meningitis in mice

[0085] The mice were randomly divided into four groups, 6 in the (NS) group, 6 in the melatonin (MT) group, and 26 in the APEC TW-XM (A) group (10-bacteria counting, 6-pathological sections, 10 for Statistical survival rate), APEC TW-XM+melatonin (A+MT) group 26 (10-bacteria count, 6-pathological section, 10 for sta...

Embodiment 3

[0104] (1) Preparation of APEC TW-XM strain

[0105] With embodiment 2.

[0106] (2) Melatonin regulates intestinal flora to prevent meningitis

[0107] The mice were randomly divided into four groups, APEC TW-XM group: daily intraperitoneal injection of 100 μl of normal saline; daily replacement of sterile water; A+MT group: daily intraperitoneal injection of 100 μl of 30 mg / kg MT; daily replacement of sterile water; Group A+antibiotics: intraperitoneally inject 100 μl of normal saline every day; replace sterile water containing antibiotics every day; A+antibiotics+MT: inject 100 μl 30 mg / kg MT intraperitoneally every day; replace sterile water containing antibiotics every day; 8 days for challenge (cfu=10 7 ) to collect samples.

[0108] (3) Sample collection

[0109] Permeability detection (EB staining): Same as Example 1.

[0110] Detection of inflammatory factors: same as Example 1.

[0111] The making of pathological section: with embodiment 2.

[0112] Notes to t...

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