Study On The Resistance Of Clinical Isolates Of Acinetobacter Baumaniii To Guanidine Disinfectant

[Background]Acinetobacter baumannii is an aerobic, Gram-negative bacillus. It belongs to Acinetobacter spp and can’t ferment glucose. It distributes widely in the natural environment, hospitals, and the surface of skin and mucous membrane of humans. As an opportunistic pathogen, Acinetobacter baumannii threaten critically ill patients with low immune function such as Intensive Care Unit patients and burn patients. It could cause pneumonia, meningitis, bacteremia, urinary tract infections, surgical wound infection, soft tissue infection, etc.In recent years, with widely used of broad-spectrum antibiotic and interventional procedures, the resistance of Acinetobacter baumannii to common antibiotics is tending to increase, and the proportion of multiple resistant strains also increase significantly. Some studies find that Acinetobacter baumannii has the resistant characteristic not only to antibiotics but also to disinfectants. Some Acinetobacter baumannii strains has higher disinfectant resistant than the standard strain. It brings a great difficulty for clinical treatment. It also brings a new challenge to hospital infection control.[Objectives]In order to get the resistance characteristic of Acinetobacter baumannii to antimicrobial agent and disinfectant, we collected46Acinetobacter baumannii strains from clinic in Shandong province, and tested them with26antimicrobial agents and guanidine disinfectants by MIC test and MBC test, PCR was used to test the disinfectant resistant genes. The main purposes are as follows:1. The current situation of Acinetobacter baumannii antimicrobial agent resistance isolated from clinic in Shandong province.2. The antimicrobial efficacy of multiple resistant Acinetobacter baumannii with different guanidine disinfectants by polyhexamethylene guanidine (PHMG) and polyhexamethylene biguanidine (PHMB).3. The relationship among disinfectant resistance gene, antibiotics characteristic and anti-disinfectant characteristic of Acinetobacter baumannii.4. To provide scientific basis for clinical reasonable disinfectants using, improving disinfect efficacy, preventing Acinetobacter baumannii nosocomial infection and controlling infectious disease prevalence.[Methods]Forty-six Acinetobacter baumannii were collected from hospitals of Jinan city and Linyi city of Shandong province. The antimicrobial agent resistance was tested by paper disk method and minimal inhibitory concentration (MIC) method. The content of polyhexamethylene was tested by acidic eosin colorimetric method. MIC and minimal bactericidal concentration (MBC) were used to test the resistance to guanidine disinfectants with Acinetobacter baumannii isolated from clinic, the standard strain Escherichia coli8099and the standard strain Acinetobacter baumannii (ATCC10501). PCR was used to test the disinfectant resistance gene of Acinetobacter baumannii.[Results]1. Among the46Acinetobacter baumannii strains,93.48%were multiple resistant bacteria. The resistance characteristic to antimicrobial agent has significant differences between areas and hospitals. 2. The mean, range and median of MIC of44Acinetobacter baumannii strains to PHMG were13.95(1.56-200) mg/L and6.25mg/L, respectively; for PHMB, they were18.82(3.12-200) mg/L and12.5mg/L, respectively.From the MIC results of44Acinetobacter baumannii strains from clinic, Acinetobacter baumannii ATCC10501and Escherichia coli8099to PHMG, we found there was no significant difference between clinical Acinetobacter baumannii strains and Acinetobacter baumannii ATCC10501(t=0.636,p=0.528) or Escherichia coli8099(t=0.636, p=0.528) on the resistance to PHMG.From the MIC results of44Acinetobacter baumannii strains from clinic, Acinetobacter baumannii ATCC10501and Escherichia coli8099to PHMB, we found that clinical Acinetobacter baumannii strains had a higher resistance than Escherichia coli8099(t=4.075, p<0.001), and no significant difference with Acinetobacter baumannii ATCC10501(t=-0.901,p=0.372).Comparing with the MIC results of clinical Acinetobacter baumannii strains to PHMG and PHMB, we found the clinical strains had a higher resistance to PHMB than to PHMG (t=-2.339,p=0.022).3. The mean, range and median of MBC of36Acinetobacter baumannii strains to PHMG were207.8(6.25-1600) mg/L and100mg/L, respectively; for PHMB, it were380.9(12.5-1600) mg/L and400mg/L, respectively.From the MIC results of36Acinetobacter baumannii strains from clinic, Acinetobacter baumannii ATCC10501and Escherichia coli8099to PHMG, we found that clinical Acinetobacter baumannii strains had a higher resistance than Escherichia coli8099(t=7.026, p<0.001), and no significant difference with Acinetobacter baumannii ATCC10501(t=0.281, p=0.780).From the MBC results of36Acinetobacter baumannii strains from clinic, Acinetobacter baumannii ATCC10501and Escherichia coli8099to PHMB, we found that clinical Acinetobacter baumannii strains had a lower resistance than Acinetobacter baumannii ATCC10501(t=-2.562,p=0.015), but had a higher resistance than Escherichia coli8099(t=5.460, p<0.001). Comparing with the MBC results of clinical Acinetobacter baumannii to PHMG and to PHMB, we found the clinical strains had a higher resistance to PHMB than to PHMG (t=-3.299,p=0.002).The results of MIC and MBC revealed PHMG had a higher ability in suppressing and killing bacteria than PHMB.4. Selected23clinical Acinetobacter baumannii strains to test the disinfectant resistance genes. None of them have qacA/B gene in chromosome or plasmid. The positive rate of carrying qacA/B is0%. We did not find qacA/B gene in clinical Acinetobacter baumannii. The standard strain Escherichia coli8099, clinical pseudomonas aeruginosa strain and clinical Escherichia coli strain have not qacA/B gene in chromosome or plasmid, either. Among the23clinical Acinetobacter baumannii strains,21of them have qacEAl-sull gene both in chromosome and plasmid,1strain only have qacEAl-sull gene in plasmid, and1strain is without qacEAl-sull gene both in chromosome and plasmid. The positive rate of qacEAl-sull gene in chromosome and plasmid were95.65%and91.30%, respectively. The pseudomonas aeruginosa strain and Escherichia coli strain isolated from clinic were found having qacEAl-sull gene both in chromosome and plasmid.[Conclusion]1. Acinetobacter baumannii isolated from Jinan and Linyi city of Shnadong province appeared multiple resistant tendency. The resistant of Acinetobacter baumannii isolated from different hospitals has significant difference.2. From the result of the MIC test we found that the resistant to PHMG of Shandong Province clinical Acinetobacter baumannii is no harder than Acinetobacter baumannii (ATCC10501) and Escherichia coli (8099). However, different results were shown in the MBC test:The resistant to PHMG of Shandong Province clinical Acinetobacter baumannii is significantly harder than Escherichia coli (8099) but weaker than Acinetobacter baumannii (ATCC10501).The results of this study showed that PHMB has better bactericidal efficacy on Acinetobacter baumannii isolated from clinic, than Acinetobacter baumannii (ATCC10501), and has better antimicrobial and bactericidal efficacy on Escherichia coli (8099) than Acinetobacter baumannii isolated from clinic, but no significant difference between Acinetobacter baumannii isolated from clinic and Acinetobacter baumannii (ATCC10501) in the resistance against the inhibition ability of PHMB.The resistant of Shandong Province clinical Acinetobacter baumannii to PHMB is harder than PHMG. It indicts that PHMG has better antimicrobial and bactericidal efficacy than PHMB.3.The positive rate of carrying qacA/B gene in chromosome or plasmid of Acinetobacter baumannii isolated from Shandong Province clinic is0%. It indicts that clinical Acinetobacter baumannii has a low positive rate of carrying qacA/B gene. The positive rate of carrying qacEAl-sull gene in chromosome and plasmid of Acinetobacter baumannii from Shandong province clinic are95.65%and91.30%, respectively. It indicts that clinical Acinetobacter baumannii carrying with qacE△1-sull gene would be a common phenomenon. The high rate carrying with qacE△1-sull gene in plasmid of Acinetobacter baumannii indicts that it would be easier to spread the resistance genes between strains or species. However, no association was found between qacEAl-sull gene carrying and the resistance phenotype from the result of MIC test and MBC test of Acinetobacter baumannii carrying with or without qacEAl-sull gene. Further study should be conducted to survey this relationship.4. Acinetobacter baumannii is a common opportunistic pathogen in nosocomial infections. Multiple resistances have become a popular trend. In hospital Acinetobacter baumannii disease prevention and control, blocking the transmission is an important means of measurements. Because of using low dose and single disinfectant for long period time, some bacteria become resistant to disinfectants. Since the multiple resistances Acinetobacter baumannii has the possibility with harder resistance to the standard strains, choose a proper disinfectant should be taken more attention in the clinic. Such as a disinfectant with single guanidine has better disinfection efficacy than a disinfectant containing biguanide.