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Abstract [Objectives] This study was conducted to determine in-vitro antibacterial effect of 58 Chinese herbs on Staphylococcus aureus， and screen single Chinese herbs with better in-vitro antibacterial effect for formulation of prescription. [Methods] Effective components in Chinese herbs were extracted with water， and inhibition zone diameters and minimal inhibitory concentrations of the 58 Chinese herbs against S. aureus were determined by agar plate punching method and micro two-fold method， respectively. [Results] Twenty two kinds of Chinese herbs including Terminalia chebula Retz.， Anemarrhena asphodeloides Bunge.， Prunella vulgaris L， Rheum palmatum L. and Mosla chinensis Maxim. had better antibacterial effect， among which T. chebula Retz.， A. asphodeloides Bunge.， P. vulgaris L， R. palmatum L. and C. sappan Linn showed the best antibacterial effect， with average inhibition zone over 20 mm and MIC lower than 15.6 mg/ml. [Conclusions] Traditional Chinese herbs have certain antibacterial effect， and this study will provide important theoretical reference for clinical medication and development of prescription.
Key words Staphylococcus aureus； Multi-drug resistance； Chinese herbal medicine； In-vitro anitbacterial experiment； Antibacterial effect
Staphylococcus aureus is a main pathogen of skin infection， which causes about a half of the total cases， with symptoms of cellulitis， crusted tetter， furuncle and serious scalded skin syndrome[1-2]. Malpractice might result in blood poisoning. At present， this disease is mainly treated with antibiotics， but the lack of standardization in application of antibiotics and the characteristics of S. aureus often give rise to drug tolerance in S. aureus[3-6]. In recent years， the multi-drug resistance of clinic S. aureus isolates has become severer and severer， and it is difficult to achieve anticipated treatment effect， greatly perplexing clinical medication.
Chinese herbal medicine has a long history with the characteristics of low toxicity and low price. It is widely applied in clinic， and scholars at home and abroad are paying more and more attention to how to screen medicine with antibacterial effect from traditional Chinese medicine[7-8]. Traditional Chinese medicine could enhance disease resistance in organisms， some Chinese herbal medicines have the effect of killing bacteria and viruses. Furthermore， some traditional Chinese medicines have been proved to have the effect of eliminating drug tolerance in bacteria. All these reveal that traditional Chinese medicine has a broad application prospect[9]. In this study， 58 Chinese herbs including Phellodendron chinense Schneid.， Anemarrhena asphodeloides Bunge. and Terminalia chebula Retz. were extracted with water， and the antibacterial ability of the 58 medicines on S. aureus was evaluated by in-vitro antibacterial experiment to screen Chinese herbal medicines with remarkable antibacterial effect. This study will provide important reference for further formulation of traditional Chinese medicine prescription. Materials and Methods
Materials and reagents
The tested strain was isolated， purified and identified by Clinical Laboratory， The Second Hospital of Qinhuangdao， and designated as JP-1. S. aureus ATCC25923 was provided by professor Zhu from Yangzhou University. 58 Chinese herbal medicines including Nepeta cataria L.， P. chinense， Saposhnikovia divaricata （Turcz.） Schischk. and Catsia tora Linn were all purchased from Qinhuangdao Minle Medicine Trade Co.， Ltd. Ordinary nutrient agar medium was purchased from Beijing Land Bridge Technology Co.， Ltd. Nutrient broth medium was purchased from Haibo Biotechnology Co.， Ltd. in Qingdao Hi-tech Park.
Instruments and equipment
G1040WS-vertial automatic pressure steam sterilizer （Zealway Instrument Inc.）； sterile operating platform （Beijing Yatai Kelong Instruments Co.， Ltd.）； refrigerator （Haier BioMedical）； water-jacket incubator （Shanghai Yiheng Scientific Instruments Co.， Ltd.）； conventional reagents and instruments， which were all provided by Preventive Veterinary Medicine Key Laboratory， Hebei Normal University of Science & Technology.
Methods
Preparation of bacterial liquid
The S. aureus isolate JP-1 cryopreserved at -80 ℃ was inoculated on ordinary nutrient agar medium and cultured at 37 ℃ overnight. The activated colony was picked and inoculated in 3 ml of liquid medium， and cultured at 37 ℃ under 180 r/min to logarithmic phase （OD600 nm was around 1.5）. The bacterial liquid was used for counting of bacteria and preserved at 4 ℃.
Preparation of medicine liquid from single Chinese herb
At first， the 58 Chinese herbs including Forsythia suspensa， P. chinense and C. tora were prepared to water extracts by water extraction method[10]. A certain amount of each Chinese herb was weighed， followed by removal of impurities， flushing， drying， and pulverization. For each medicine， 30 g of powder was weighed and poured into a beaker； 300 ml of water was added into the beaker， and the mixture was cooled with stirring twice； and after cooling and filtration， the mixture was centrifuged， and the supernatant was concentrated to 30 ml， obtaining the liquid with a finally concentration of 1 g/ml， which was subpackaged， sterilized at 121 ℃ for 10 min and preserved in a refrigerator at 4 ℃ for later use.
Determination of in-vitro inhibition zone diameter of Chinese herb （preliminary screening）
The test was carried out according to agar plate punching method reported in references[10-11]. The bacterial liquid in the logarithmic phase was diluted to 1×108 cfu/ml， and 100 μl of the bacterial liquid was uniformly coated on nutrient agar medium. Holes were formed on agar plates， which were baked with alcohol lamp， and 30 μl of prepared herbal liquid was added into each hole which was marked. The inoculated bacteria were cultured at 37 ℃ for 12 h， and the diameter of inhibition zone was measured with a vernier caliper. Each herb was set with three replicates， and the obtained values were averaged. The judgment was made according to following criterions： very sensitive： inhibition zone diameter≥20 mm， highly sensitive： 20 mm>inhibition zone diameter≥16 mm， moderately sensitive： 16 mm>inhibition zone diameter≥10 mm， lowly sensitive： inhibition zone diameter<10 mm. Determination of minimal inhibitory concentration （MIC）/minimal bactericidal concentration （MBC） of Chinese herb
According to antibacterial experiment in vitro， singe herbs with remarkable inhibitory effect on S. aureus （inhibition zone diameter of 15 mm） were selected for further determination of MIC and MBC by micro two-fold method. This test was carried out according to methods in references[12-13]. Each herbal liquid was diluted with 2×LB to 1， 0.5， 0.25 and 0.125 g/ml， different concentrations of herbal liquids were added into 96-hole bacterial culture plate， and bacterial liquid （1×106 cfu/ml） of the same volume was added into each hole. Positive control （LB+ bacterial liquid） and negative control （pure herbal medicine） were also set. After the addition of sample， the bacteria were cultured at 37 ℃ for 24 h. MIC is the corresponding lowest concentration at which growth of bacteria could not be observed visually. Culture medium without growing bacteria was coated on ordinary agar plate， followed by 24 h of culture， and the lowest concentration corresponding to the plate without bacterial growth was MBC.
Results and Analysis
Determination of in-vitro inhibition zone diameter of Chinese herbal medicine （preliminary screening）
The results of preliminary screening on the water extracts of the 58 traditional Chinese herbs by agar diffusion method （Table 1） showed that 34 water extracts among the 58 water extracts inhibited the S. aureus isolate and standard strain to different degrees. S. aureus was highly sensitive to 21 Chinese herbs （with inhibition zone diameters≥15 mm）， moderately sensitive to 12 kinds（ with inhibition zone diameters≥20 mm）， and lowly or not sensitive to other Chinese herbs.
Determination MIC and MBC of Chinese herb
MIC and MBC of herbal medicines （inhibition zone≥16 mm） highly sensitive to S. aureus isolate and standard strain were determined. The results （Table 2） showed that the water extracts of 10 traditional Chinese medicines including T. chebula and A. asphodeloides had significant antibacterial effect， with MIC lower than 31.25 mg/ml and MBC lower than 62.5 mg/ml. Other herbal liquids had certain antibacterial effect.
Discussion
Compared with other clinical departments， skin S. aureus isolate exhibited severer multi-drug resistance. Yang et al. detected 1 416 cases of patients with skin diseases from 13 children hospital， and found 1 043 cases were infected with S. aureus， and the isolate was resistant to several topical antibiotics （erythrocin 97.3%， penicillin 96.7%， clindamycin 89.0%）[14]. Other literatures also indicate that skin S. aureus has an increasing trend of resistance against ciprofloxacin （60.6% in 2006）， tetracycline （reaching 30%-40%） and aminoglycoside antibiotics （gentamicin and neomycin）. At present， skin S. aureus infection is mainly treated with some topical ointment and liniment， with the assistance of oral and injected antibiotics when serious[15]. Common topical skin antibiotics are more limited， such as chlorotetracycline ointment， erythromycin ointment， neomycin fluocinolone acetonide， ciprofloxacin emulsifiable paste and ofloxacin emulsifiable paste[16]. With the multi-drug resistance becoming severer， traditional topical skin antibiotics could not satisfy current clinical requirement， and it is urgent to develop new safe antibacterial agents. Traditional Chinese medicine has the advantages of natural raw material， comprehensive efficiency， lower side effect and lower drug residue， and many single traditional Chinese herbs have been proved to have antibacterial activity， and are anticipated to replace antibiotics in future. Researches show that the action speed and effect of partial traditional Chinese herbs are not weaker than western medicine， and have broad application value， and partial traditional Chinese medicine preparations have been widely applied in clinic， such as Sanhuang Tablet and berberine[17]. This study also showed that quite a lot of traditional Chinese herbs had remarkable inhibitory effect on S. aureus perplexing in clinic. S. baicalensis and T. chebula as a kind of traditional Chinese medicine have wide antibacterial spectrum， and are proved to have certain inhibitory effect on Escherichia coli and Pseudomonas aeruginosa by some studies[18]. The antibacterial property of these two traditional Chinese herbs was also verified in this study. M. officinalis is mostly used for treating bacterial diseases and inflammations， and widely applied in traditional medicine and Chinese medicine in Japan[19]. It also has very remarkable inhibitory effect on S. aureus. F. multiflora is often used in traditional medicine， and has been proved to have a significant effect on various intestinal bacteria. It also showed very strong inhibitory effect on S. aureus in this study. Furthermore， the results of this study also showed that P. cuspidatum， A. paniculata， S. officinalis， R. palmatum and C. chinensis all have an inhibition effect on S. aureus to different degrees， laying a foundation for further development of prescription. In this study， with drug-resistant skin S. aureus common in clinic as a main object， traditional Chinese herbs suitable for clinical application were screened through antibacterial test of single traditional Chinese herbs， and it is anticipated to develop traditional Chinese medicine prescription suitable for topical application of skin on this basis. There have been many literatures about the screening of single traditional Chinese herbs with inhibitory effect on S. aureus， but the screening was conducted in a smaller scale and thus has certain limitations. In order to develop topical skin prescription， 58 Chinese herbal medicines were widely screened. Certainly， there are certain inadequacy， for instance， the preparation of traditional Chinese medicine liquid should be prepared according to the classification of different medicines， and more repeated trials of clinical S. aureus isolate should be done. In-vitro antibacterial tests were carried out on S. aureus common for skin infection in this study， with an attempt to screen traditional herbal medicines with stronger inhibitory effect. This study lays a foundation for the topical preparation for skin infection of S. aureus.
References
[1] GAGNAIRE J， VERHOEVEN PO， GRATTARD F， et al. Epidemiology and clinical relevance of Staphylococcus aureus intestinal carriage： a systematic review and meta-analysis[J]. Expert Rev Anti Infect Ther. 2017， 15（8）： 767-785.
[2] DENG J， WANG Q， WANG Y， et al. Detection and drug resistance analysis of common skin lesion Staphylococcus aureus in lesions of atopic dermatitis[J]. China Journal of Leprosy and Skin Diseases， 2012，28（10）： 712-715. （in Chinese）
[3] MA L， LIU Y. Selection of antibacterial drugs for skin Staphylococcus aureus infection[J]. The Chinese Journal of Dermatovenereology， 2013， 27（3）： 225-227. （in Chinese）
[4] RAO X， LYU XJ， LIU YB， et al. Clinical analysis of 55 cases of Staphylococcus aureus infective septicemia[J]. Journal of Sichuan University： Engineering Science Edition， 2009，40（2）： 347-349， 360. （in Chinese）
[5] LIU XQ. Clinical infection distribution and drug resistance analysis of 896 cases of Staphylococcus aureus in a certain area[J]. Experimental and Laboratory Medicine， 2015（1）： 80-81. （in Chinese）
[6] SUN RR. Clinical survey of anti-infectious agent for skin Staphylococcus aureus infection[J]. Capital Medicine， 2007（20）： 41. （in Chinese）
[7] LI YN， TAO QC. Research status and thought of antibacterial effect of traditional Chinese medicine[J]. International Journal of Laboratory Medicine， 2014（2）： 198-200. （in Chinese）
[8] ZHANG XP， ZHU SS， LIU J， et al. Research progress in evaluation on Chinese materia medica in vitro bacteriostasis methods[J]. Drug Evaluation Research， 2014，37（2）： 188-192. （in Chinese）
[9] PENG J， XIE LL， XIE J. Research methods for antibacterial mechanism of traditional Chinese medicine[J]. Curr Biotech， 2015（1）： 66-69. （in Chinese）
[10] ZUO GY， YU W， XU GL， et al. Screen of anti-MRSA activity from 18 Chinese herbage medicine extracts[J]. China Pharmacist， 2005，8（7）： 78-80. （in Chinese）
[11] YAN XJ， ZHANG JL， YANG JF， et al. Antibacteral effect of six traditional Chinese medicines against methicillin-resistant Staphylococcus aureu[J]. China Tropical Medicine， 2009， 9（2）： 219-220. （in Chinese）
[12] ZUO GY， MENG FY， HAO XY， et al. Antibacterial alkaloids from Chelidonium majus linn （papaveraceae） against clinical isolates of methicillin-resistant Staphylococcus aureus[J]. J Pharm Pharm Sci， 2008， 11（4）： 90-94. [13] WANG F， ZUO GY， HAN J， et al. In vitro screen of anti-Staphylococcus aureus activity of 20 kinds of heat-clearing and toxicity-removing Chinese herbal medicines[J]. Chinese Journal of Infection Control ， 2013， 12（5）： 321-325. （in Chinese）
[14] YANG ZHOU， LIU Y， SUN J， et al. Antibiotic susceptibility of Staphylococcus aureus isolated from children with infectious skin diseases in China[J]. Journal of Microbes and Infections， 2012（1）： 37-43.
[15] GAO CL， BAI XW， LI H， et al. Staphylococcus aureus septicemia： Clinical analysis and treatment[J]. Chinese Journal of Nosocomiology， 2003， 13（9）： 82-84. （in Chinese）
[16] HU JP. Investigation of utilization conditions of topical preparations in dermatology department of a certain center[J]. Chinese Journal of Pharmacoepidemiology， 2013， 22（5）： 259-261. （in Chinese）
[17] LI YN， TAO QC. Research status and thought of antibacterial effect of traditional Chinese medicine[J]. International Journal of Laboratory Medicine， 2014（2）： 198-200. （in Chinese）
[18] LIU W， TANG JH， ZHOU LW， et al. Antibacterial effect and ultrastructure of traditional Chinese medicine of Terminalia chebula Retz.[J]. Jiangsu Agricultural Sciences， 2014（6）： 281-283. （in Chinese）
[19] WENG TI， WU HY， KUO CW， et al. Honokiol rescues sepsis-associated acute lung injury and lethality via the inhibition of oxidative stress and inflammation[J]. Intensive Care Med， 2011， 37（3）： 533-541. （in Chinese）
Key words Staphylococcus aureus； Multi-drug resistance； Chinese herbal medicine； In-vitro anitbacterial experiment； Antibacterial effect
Staphylococcus aureus is a main pathogen of skin infection， which causes about a half of the total cases， with symptoms of cellulitis， crusted tetter， furuncle and serious scalded skin syndrome[1-2]. Malpractice might result in blood poisoning. At present， this disease is mainly treated with antibiotics， but the lack of standardization in application of antibiotics and the characteristics of S. aureus often give rise to drug tolerance in S. aureus[3-6]. In recent years， the multi-drug resistance of clinic S. aureus isolates has become severer and severer， and it is difficult to achieve anticipated treatment effect， greatly perplexing clinical medication.
Chinese herbal medicine has a long history with the characteristics of low toxicity and low price. It is widely applied in clinic， and scholars at home and abroad are paying more and more attention to how to screen medicine with antibacterial effect from traditional Chinese medicine[7-8]. Traditional Chinese medicine could enhance disease resistance in organisms， some Chinese herbal medicines have the effect of killing bacteria and viruses. Furthermore， some traditional Chinese medicines have been proved to have the effect of eliminating drug tolerance in bacteria. All these reveal that traditional Chinese medicine has a broad application prospect[9]. In this study， 58 Chinese herbs including Phellodendron chinense Schneid.， Anemarrhena asphodeloides Bunge. and Terminalia chebula Retz. were extracted with water， and the antibacterial ability of the 58 medicines on S. aureus was evaluated by in-vitro antibacterial experiment to screen Chinese herbal medicines with remarkable antibacterial effect. This study will provide important reference for further formulation of traditional Chinese medicine prescription. Materials and Methods
Materials and reagents
The tested strain was isolated， purified and identified by Clinical Laboratory， The Second Hospital of Qinhuangdao， and designated as JP-1. S. aureus ATCC25923 was provided by professor Zhu from Yangzhou University. 58 Chinese herbal medicines including Nepeta cataria L.， P. chinense， Saposhnikovia divaricata （Turcz.） Schischk. and Catsia tora Linn were all purchased from Qinhuangdao Minle Medicine Trade Co.， Ltd. Ordinary nutrient agar medium was purchased from Beijing Land Bridge Technology Co.， Ltd. Nutrient broth medium was purchased from Haibo Biotechnology Co.， Ltd. in Qingdao Hi-tech Park.
Instruments and equipment
G1040WS-vertial automatic pressure steam sterilizer （Zealway Instrument Inc.）； sterile operating platform （Beijing Yatai Kelong Instruments Co.， Ltd.）； refrigerator （Haier BioMedical）； water-jacket incubator （Shanghai Yiheng Scientific Instruments Co.， Ltd.）； conventional reagents and instruments， which were all provided by Preventive Veterinary Medicine Key Laboratory， Hebei Normal University of Science & Technology.
Methods
Preparation of bacterial liquid
The S. aureus isolate JP-1 cryopreserved at -80 ℃ was inoculated on ordinary nutrient agar medium and cultured at 37 ℃ overnight. The activated colony was picked and inoculated in 3 ml of liquid medium， and cultured at 37 ℃ under 180 r/min to logarithmic phase （OD600 nm was around 1.5）. The bacterial liquid was used for counting of bacteria and preserved at 4 ℃.
Preparation of medicine liquid from single Chinese herb
At first， the 58 Chinese herbs including Forsythia suspensa， P. chinense and C. tora were prepared to water extracts by water extraction method[10]. A certain amount of each Chinese herb was weighed， followed by removal of impurities， flushing， drying， and pulverization. For each medicine， 30 g of powder was weighed and poured into a beaker； 300 ml of water was added into the beaker， and the mixture was cooled with stirring twice； and after cooling and filtration， the mixture was centrifuged， and the supernatant was concentrated to 30 ml， obtaining the liquid with a finally concentration of 1 g/ml， which was subpackaged， sterilized at 121 ℃ for 10 min and preserved in a refrigerator at 4 ℃ for later use.
Determination of in-vitro inhibition zone diameter of Chinese herb （preliminary screening）
The test was carried out according to agar plate punching method reported in references[10-11]. The bacterial liquid in the logarithmic phase was diluted to 1×108 cfu/ml， and 100 μl of the bacterial liquid was uniformly coated on nutrient agar medium. Holes were formed on agar plates， which were baked with alcohol lamp， and 30 μl of prepared herbal liquid was added into each hole which was marked. The inoculated bacteria were cultured at 37 ℃ for 12 h， and the diameter of inhibition zone was measured with a vernier caliper. Each herb was set with three replicates， and the obtained values were averaged. The judgment was made according to following criterions： very sensitive： inhibition zone diameter≥20 mm， highly sensitive： 20 mm>inhibition zone diameter≥16 mm， moderately sensitive： 16 mm>inhibition zone diameter≥10 mm， lowly sensitive： inhibition zone diameter<10 mm. Determination of minimal inhibitory concentration （MIC）/minimal bactericidal concentration （MBC） of Chinese herb
According to antibacterial experiment in vitro， singe herbs with remarkable inhibitory effect on S. aureus （inhibition zone diameter of 15 mm） were selected for further determination of MIC and MBC by micro two-fold method. This test was carried out according to methods in references[12-13]. Each herbal liquid was diluted with 2×LB to 1， 0.5， 0.25 and 0.125 g/ml， different concentrations of herbal liquids were added into 96-hole bacterial culture plate， and bacterial liquid （1×106 cfu/ml） of the same volume was added into each hole. Positive control （LB+ bacterial liquid） and negative control （pure herbal medicine） were also set. After the addition of sample， the bacteria were cultured at 37 ℃ for 24 h. MIC is the corresponding lowest concentration at which growth of bacteria could not be observed visually. Culture medium without growing bacteria was coated on ordinary agar plate， followed by 24 h of culture， and the lowest concentration corresponding to the plate without bacterial growth was MBC.
Results and Analysis
Determination of in-vitro inhibition zone diameter of Chinese herbal medicine （preliminary screening）
The results of preliminary screening on the water extracts of the 58 traditional Chinese herbs by agar diffusion method （Table 1） showed that 34 water extracts among the 58 water extracts inhibited the S. aureus isolate and standard strain to different degrees. S. aureus was highly sensitive to 21 Chinese herbs （with inhibition zone diameters≥15 mm）， moderately sensitive to 12 kinds（ with inhibition zone diameters≥20 mm）， and lowly or not sensitive to other Chinese herbs.
Determination MIC and MBC of Chinese herb
MIC and MBC of herbal medicines （inhibition zone≥16 mm） highly sensitive to S. aureus isolate and standard strain were determined. The results （Table 2） showed that the water extracts of 10 traditional Chinese medicines including T. chebula and A. asphodeloides had significant antibacterial effect， with MIC lower than 31.25 mg/ml and MBC lower than 62.5 mg/ml. Other herbal liquids had certain antibacterial effect.
Discussion
Compared with other clinical departments， skin S. aureus isolate exhibited severer multi-drug resistance. Yang et al. detected 1 416 cases of patients with skin diseases from 13 children hospital， and found 1 043 cases were infected with S. aureus， and the isolate was resistant to several topical antibiotics （erythrocin 97.3%， penicillin 96.7%， clindamycin 89.0%）[14]. Other literatures also indicate that skin S. aureus has an increasing trend of resistance against ciprofloxacin （60.6% in 2006）， tetracycline （reaching 30%-40%） and aminoglycoside antibiotics （gentamicin and neomycin）. At present， skin S. aureus infection is mainly treated with some topical ointment and liniment， with the assistance of oral and injected antibiotics when serious[15]. Common topical skin antibiotics are more limited， such as chlorotetracycline ointment， erythromycin ointment， neomycin fluocinolone acetonide， ciprofloxacin emulsifiable paste and ofloxacin emulsifiable paste[16]. With the multi-drug resistance becoming severer， traditional topical skin antibiotics could not satisfy current clinical requirement， and it is urgent to develop new safe antibacterial agents. Traditional Chinese medicine has the advantages of natural raw material， comprehensive efficiency， lower side effect and lower drug residue， and many single traditional Chinese herbs have been proved to have antibacterial activity， and are anticipated to replace antibiotics in future. Researches show that the action speed and effect of partial traditional Chinese herbs are not weaker than western medicine， and have broad application value， and partial traditional Chinese medicine preparations have been widely applied in clinic， such as Sanhuang Tablet and berberine[17]. This study also showed that quite a lot of traditional Chinese herbs had remarkable inhibitory effect on S. aureus perplexing in clinic. S. baicalensis and T. chebula as a kind of traditional Chinese medicine have wide antibacterial spectrum， and are proved to have certain inhibitory effect on Escherichia coli and Pseudomonas aeruginosa by some studies[18]. The antibacterial property of these two traditional Chinese herbs was also verified in this study. M. officinalis is mostly used for treating bacterial diseases and inflammations， and widely applied in traditional medicine and Chinese medicine in Japan[19]. It also has very remarkable inhibitory effect on S. aureus. F. multiflora is often used in traditional medicine， and has been proved to have a significant effect on various intestinal bacteria. It also showed very strong inhibitory effect on S. aureus in this study. Furthermore， the results of this study also showed that P. cuspidatum， A. paniculata， S. officinalis， R. palmatum and C. chinensis all have an inhibition effect on S. aureus to different degrees， laying a foundation for further development of prescription. In this study， with drug-resistant skin S. aureus common in clinic as a main object， traditional Chinese herbs suitable for clinical application were screened through antibacterial test of single traditional Chinese herbs， and it is anticipated to develop traditional Chinese medicine prescription suitable for topical application of skin on this basis. There have been many literatures about the screening of single traditional Chinese herbs with inhibitory effect on S. aureus， but the screening was conducted in a smaller scale and thus has certain limitations. In order to develop topical skin prescription， 58 Chinese herbal medicines were widely screened. Certainly， there are certain inadequacy， for instance， the preparation of traditional Chinese medicine liquid should be prepared according to the classification of different medicines， and more repeated trials of clinical S. aureus isolate should be done. In-vitro antibacterial tests were carried out on S. aureus common for skin infection in this study， with an attempt to screen traditional herbal medicines with stronger inhibitory effect. This study lays a foundation for the topical preparation for skin infection of S. aureus.
References
[1] GAGNAIRE J， VERHOEVEN PO， GRATTARD F， et al. Epidemiology and clinical relevance of Staphylococcus aureus intestinal carriage： a systematic review and meta-analysis[J]. Expert Rev Anti Infect Ther. 2017， 15（8）： 767-785.
[2] DENG J， WANG Q， WANG Y， et al. Detection and drug resistance analysis of common skin lesion Staphylococcus aureus in lesions of atopic dermatitis[J]. China Journal of Leprosy and Skin Diseases， 2012，28（10）： 712-715. （in Chinese）
[3] MA L， LIU Y. Selection of antibacterial drugs for skin Staphylococcus aureus infection[J]. The Chinese Journal of Dermatovenereology， 2013， 27（3）： 225-227. （in Chinese）
[4] RAO X， LYU XJ， LIU YB， et al. Clinical analysis of 55 cases of Staphylococcus aureus infective septicemia[J]. Journal of Sichuan University： Engineering Science Edition， 2009，40（2）： 347-349， 360. （in Chinese）
[5] LIU XQ. Clinical infection distribution and drug resistance analysis of 896 cases of Staphylococcus aureus in a certain area[J]. Experimental and Laboratory Medicine， 2015（1）： 80-81. （in Chinese）
[6] SUN RR. Clinical survey of anti-infectious agent for skin Staphylococcus aureus infection[J]. Capital Medicine， 2007（20）： 41. （in Chinese）
[7] LI YN， TAO QC. Research status and thought of antibacterial effect of traditional Chinese medicine[J]. International Journal of Laboratory Medicine， 2014（2）： 198-200. （in Chinese）
[8] ZHANG XP， ZHU SS， LIU J， et al. Research progress in evaluation on Chinese materia medica in vitro bacteriostasis methods[J]. Drug Evaluation Research， 2014，37（2）： 188-192. （in Chinese）
[9] PENG J， XIE LL， XIE J. Research methods for antibacterial mechanism of traditional Chinese medicine[J]. Curr Biotech， 2015（1）： 66-69. （in Chinese）
[10] ZUO GY， YU W， XU GL， et al. Screen of anti-MRSA activity from 18 Chinese herbage medicine extracts[J]. China Pharmacist， 2005，8（7）： 78-80. （in Chinese）
[11] YAN XJ， ZHANG JL， YANG JF， et al. Antibacteral effect of six traditional Chinese medicines against methicillin-resistant Staphylococcus aureu[J]. China Tropical Medicine， 2009， 9（2）： 219-220. （in Chinese）
[12] ZUO GY， MENG FY， HAO XY， et al. Antibacterial alkaloids from Chelidonium majus linn （papaveraceae） against clinical isolates of methicillin-resistant Staphylococcus aureus[J]. J Pharm Pharm Sci， 2008， 11（4）： 90-94. [13] WANG F， ZUO GY， HAN J， et al. In vitro screen of anti-Staphylococcus aureus activity of 20 kinds of heat-clearing and toxicity-removing Chinese herbal medicines[J]. Chinese Journal of Infection Control ， 2013， 12（5）： 321-325. （in Chinese）
[14] YANG ZHOU， LIU Y， SUN J， et al. Antibiotic susceptibility of Staphylococcus aureus isolated from children with infectious skin diseases in China[J]. Journal of Microbes and Infections， 2012（1）： 37-43.
[15] GAO CL， BAI XW， LI H， et al. Staphylococcus aureus septicemia： Clinical analysis and treatment[J]. Chinese Journal of Nosocomiology， 2003， 13（9）： 82-84. （in Chinese）
[16] HU JP. Investigation of utilization conditions of topical preparations in dermatology department of a certain center[J]. Chinese Journal of Pharmacoepidemiology， 2013， 22（5）： 259-261. （in Chinese）
[17] LI YN， TAO QC. Research status and thought of antibacterial effect of traditional Chinese medicine[J]. International Journal of Laboratory Medicine， 2014（2）： 198-200. （in Chinese）
[18] LIU W， TANG JH， ZHOU LW， et al. Antibacterial effect and ultrastructure of traditional Chinese medicine of Terminalia chebula Retz.[J]. Jiangsu Agricultural Sciences， 2014（6）： 281-283. （in Chinese）
[19] WENG TI， WU HY， KUO CW， et al. Honokiol rescues sepsis-associated acute lung injury and lethality via the inhibition of oxidative stress and inflammation[J]. Intensive Care Med， 2011， 37（3）： 533-541. （in Chinese）