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1.
PeerJ ; 8: e9020, 2020.
Article in English | MEDLINE | ID: mdl-32523805

ABSTRACT

Acinetobacter baumannii, an important emerging pathogen of nosocomial infections, is known for its ability to form biofilms. Biofilm formation increases the survival rate of A. baumannii on dry surfaces and may contribute to its persistence in the hospital environment, which increases the probability of nosocomial infections and outbreaks. This study was undertaken to characterize the biofilm production of different strains of A. baumannii and the effects of chemical compounds, especially antibiotics, on biofilm formation. In this study, no statistically significant relationship was observed between the ability to form a biofilm and the antimicrobial susceptibility of the A. baumannii clinical isolates. Biofilm formation caused by A. baumannii ATCC 17978 after gene knockout of two-component regulatory system gene baeR, efflux pump genes emrA/emrB and outer membrane coding gene ompA revealed that all mutant strains had less biofilm formation than the wild-type strain, which was further supported by the images from scanning electron microscopy and confocal laser scanning microscopy. The addition of amikacin, colistin, LL-37 or tannic acid decreased the biofilm formation ability of A. baumannii. In contrast, the addition of lower subinhibitory concentration tigecycline increased the biofilm formation ability of A. baumannii. Minimum biofilm eradication concentrations of amikacin, imipenem, colistin, and tigecycline were increased obviously for both wild type and multidrug resistant clinical strain A. baumannii VGH2. In conclusion, the biofilm formation ability of A. baumannii varied in different strains, involved many genes and could be influenced by many chemical compounds.

2.
Microb Drug Resist ; 25(9): 1338-1346, 2019 Nov.
Article in English | MEDLINE | ID: mdl-31295061

ABSTRACT

Of all the Proteus spp., Proteus mirabilis is the most common species identified in clinical specimens and is a leading agent of complicated urinary tract infection. This study was undertaken to understand the antimicrobial susceptibility, prevalence of antibiotic resistance genes, and molecular typing of P. mirabilis isolates collected from three hospitals in northern Taiwan. The results showed that the collected isolates of P. mirabilis were susceptible to most antibiotics except cefazolin and tigecycline. Many resistance genes were detected in the collected isolates, of which TEM genes were the most common. Resistance to third- or fourth-generation cephalosporins was related to the presence of at least one of the tested extended-spectrum ß-lactamase (ESBL) or AmpC genes. The presence of the VEB-1 gene seemed to be a good predictor for both cefepime and ceftazidime resistance, which was further supported by quantitative polymerase chain reaction results. Of the four imipenem-resistant P. mirabilis isolates, three isolates could hydrolyze imipenem by mass spectrometry analysis. Molecular typing by pulsed-field gel electrophoresis showed that the pulsotyping of the selected P. mirabilis isolates was heterogeneous. By analyzing the relationship of antimicrobial resistance and the presence of resistance genes, revision of the Clinical and Laboratory Standards Institute cefepime and ceftazidime MIC breakpoints for Enterobacteriaceae to predict ESBL producers might possibly be needed.


Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Resistance, Bacterial/genetics , Proteus Infections/drug therapy , Proteus mirabilis/drug effects , Anti-Bacterial Agents/administration & dosage , Hospitals , Humans , Microbial Sensitivity Tests , Molecular Epidemiology , Polymerase Chain Reaction , Proteus Infections/epidemiology , Proteus Infections/microbiology , Proteus mirabilis/genetics , Proteus mirabilis/isolation & purification , Taiwan , beta-Lactamases/genetics
3.
J Microbiol Methods ; 160: 84-86, 2019 05.
Article in English | MEDLINE | ID: mdl-30928423

ABSTRACT

In this study, we developed a method to assess influence of different medical tubing on biofilm formation by A. baumannii. The results of biofilm quantification and scanning electron microscopy showed that the biofilm formation susceptibility of different tubing materials was rubber latex > polyvinyl chloride > silicone.


Subject(s)
Acinetobacter baumannii/physiology , Biofilms , Catheter-Related Infections/microbiology , Humans , Latex/chemistry , Polyvinyl Chloride/chemistry , Silicones/chemistry
4.
J Microbiol ; 55(2): 130-136, 2017 Feb.
Article in English | MEDLINE | ID: mdl-28120193

ABSTRACT

Efflux pumps play an important role in antimicrobial resistance for Acinetobacter baumannii. However, the function of the Emr pump system and the relationship between Emr and drug resistance has not been characterized in A. baumannii. In this study, four possible groups of emr-like genes were found by searching a genome database. Among them, A1S_1772 (emrB) and A1S_1773 (emrA) were demonstrated to be co-transcribed as a single operon. Moreover, during osmotic stress, A1S_1772 showed the largest change in gene expression compared to the other emrB-like genes, and deletion of A1S_1772 (AB ΔemrB) significantly slowed cell growth in 20% sucrose. Using a phenotypic microarray analysis, the AB ΔemrB mutant was more susceptible to colistin and nafcillin, paromomycin, spiramycin, and D,L-serine hydroxmate than the wild type. The spot assay, time kill assay and minimal inhibition concentration determination also indicated that the wild type could tolerate colistin better than the AB ΔemrB mutant. Finally, the increased expression levels of all emrB-like genes, including A1S_0775, A1S_0909, A1S_1772, and A1S_1799, in colistin resistance-induced A. baumannii further supported the possible involvement of the emrB genes in A. baumannii colistin resistance. Together, the Emr pump systems in A. baumannii contribute to adaptation to osmotic stress and resistance to colistin.


Subject(s)
Acinetobacter baumannii/genetics , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Colistin/pharmacology , Drug Resistance, Bacterial , Membrane Proteins/genetics , Membrane Transport Proteins/genetics , Acinetobacter baumannii/drug effects , Acinetobacter baumannii/growth & development , Acinetobacter baumannii/metabolism , Bacterial Proteins/metabolism , Drug Resistance, Bacterial/genetics , Membrane Proteins/metabolism , Membrane Transport Proteins/metabolism , Microbial Sensitivity Tests , Oligonucleotide Array Sequence Analysis , Operon , Osmotic Pressure , Phenotype , Reverse Transcriptase Polymerase Chain Reaction , Sequence Deletion , Sucrose/pharmacology , Transcription, Genetic
5.
J Microbiol Immunol Infect ; 50(2): 224-231, 2017 Apr.
Article in English | MEDLINE | ID: mdl-26055688

ABSTRACT

BACKGROUND/PURPOSE: Efflux pumps are one of the major mechanisms of antimicrobial resistance in Acinetobacter baumannii. This study aimed to understand the distribution of different types of pump genes in clinical isolates of multidrug-resistant A. baumannii (MDRAB) and to reveal the relationship between their presence and expression with antimicrobial resistance. METHODS: MDRAB isolates were collected from five hospitals in Taiwan. Different categories of pump genes, including adeB, adeJ, macB, abeM, abeS, emrA-like, emrB-like, and craA, were chosen, and their presence in the collected isolates was determined. Three induced resistant strains of A. baumannii ATCC 17978 to tigecycline, imipenem, and amikacin were also included. The expressions of the selected pump genes were determined using quantitative reverse transcription-polymerase chain reaction. RESULTS: Twenty-one MDRAB clinical isolates were obtained from five hospitals. All of the studied pump genes were present in the collected MDRAB isolates except one isolate that lacked the emrA-like gene. The gene expression of these efflux pumps was variable among the strains. The upregulation of the adeB, adeJ, and macB genes was responsible for tigecycline resistance, and the increased abeS expression was strongly related to amikacin resistance. Of all the antibiotics studied, tigecycline was the strongest inducer of gene expression for many efflux pumps in A. baumannii. CONCLUSION: Efflux pump genes are universally present in the collected clinical MDRAB isolates. The upregulation of the adeB, adeJ, macB and abeS genes is more related with antibiotic resistance.


Subject(s)
Acinetobacter Infections/microbiology , Acinetobacter baumannii/drug effects , Acinetobacter baumannii/genetics , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Membrane Transport Proteins/genetics , Acinetobacter baumannii/metabolism , Anti-Bacterial Agents/pharmacokinetics , Bacterial Proteins/metabolism , DNA, Bacterial/analysis , Drug Resistance, Multiple, Bacterial , Genes, Bacterial , Humans , Membrane Proteins/genetics , Membrane Transport Proteins/metabolism , Microbial Sensitivity Tests , RNA, Bacterial/analysis
6.
PLoS One ; 10(10): e0141107, 2015.
Article in English | MEDLINE | ID: mdl-26484669

ABSTRACT

Multidrug-resistant Acinetobacter baumannii has recently emerged as an important pathogen in nosocomial infection; thus, effective antimicrobial regimens are urgently needed. Human antimicrobial peptides (AMPs) exhibit multiple functions and antimicrobial activities against bacteria and fungi and are proposed to be potential adjuvant therapeutic agents. This study examined the effect of the human cathelicidin-derived AMP LL-37 on A. baumannii and revealed the underlying mode of action. We found that LL-37 killed A. baumannii efficiently and reduced cell motility and adhesion. The bacteria-killing effect of LL-37 on A. baumannii was more efficient compared to other AMPs, including human ß-defensin 3 (hBD3) and histatin 5 (Hst5). Both flow cytometric analysis and immunofluorescence staining showed that LL-37 bound to A. baumannii cells. Moreover, far-western analysis demonstrated that LL-37 could bind to the A. baumannii OmpA (AbOmpA) protein. An ELISA assay indicated that biotin-labelled LL-37 (BA-LL37) bound to the AbOmpA74-84 peptide in a dose-dependent manner. Using BA-LL37 as a probe, the ~38 kDa OmpA signal was detected in the wild type but the ompA deletion strain did not show the protein, thereby validating the interaction. Finally, we found that the ompA deletion mutant was more sensitive to LL-37 and decreased cell adhesion by 32% compared to the wild type. However, ompA deletion mutant showed a greatly reduced adhesion defect after LL-37 treatment compared to the wild strain. Taken together, this study provides evidence that LL-37 affects A. baumannii through OmpA binding.


Subject(s)
Acinetobacter Infections/drug therapy , Acinetobacter baumannii/drug effects , Anti-Bacterial Agents/pharmacology , Bacterial Outer Membrane Proteins/metabolism , Cathelicidins/pharmacology , Gene Expression Regulation, Bacterial , Acinetobacter Infections/metabolism , Acinetobacter Infections/microbiology , Amino Acid Sequence , Antimicrobial Cationic Peptides , Bacterial Adhesion/drug effects , Bacterial Outer Membrane Proteins/genetics , Blotting, Western , Cell Movement/drug effects , Flow Cytometry , Fluorescent Antibody Technique , Humans , Microbial Sensitivity Tests , Molecular Sequence Data , Mutation/genetics , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Sequence Homology, Amino Acid , beta-Defensins/pharmacology
7.
PLoS One ; 10(7): e0132843, 2015.
Article in English | MEDLINE | ID: mdl-26161744

ABSTRACT

Bacterial two-component regulatory systems (TCSs) facilitate changes in gene expression in response to environmental stimuli. TCS BaeR regulons influence tigecycline susceptibility in Acinetobacter baumannii through positively regulating the pump genes adeA and adeB. In this study, we demonstrate that an additional two transport systems, AdeIJK and MacAB-TolC, are also regulated by BaeSR. In the wild type and clinical tigecycline-resistant A. baumannii strains, gene expression of AdeIJK and MacAB-TolC increased after tigecycline induction, implicating their importance to tigecycline resistance in addition to AdeABC. Phenotypic microarray results showed that A. baumannii is vulnerable to certain chemicals, especially tannic acid, after deleting baeR, which was confirmed using the spot assay. The wild-type strain of A. baumannii also exhibited 1.6-fold and 4.4-fold increase in gene expression of adeJ and macB in the medium with 100 µg/mL tannic acid, but the increase was fully inhibited by baeR deletion. An electrophoretic motility shift assay based on an interaction between His-BaeR and the adeA, adeI and macA promoter regions did not demonstrate direct binding. In conclusion, A. baumannii can use the TCS BaeSR in disposing chemicals, such as tannic acid and tigecycline, through regulating the efflux pumps.


Subject(s)
Acinetobacter baumannii/metabolism , Bacterial Proteins/metabolism , Membrane Transport Proteins/metabolism , Minocycline/analogs & derivatives , Acinetobacter baumannii/drug effects , Acinetobacter baumannii/genetics , Acinetobacter baumannii/isolation & purification , Drug Resistance, Bacterial/drug effects , Drug Resistance, Bacterial/genetics , Electrophoretic Mobility Shift Assay , Gene Expression Regulation, Bacterial/drug effects , Genes, Bacterial , Microbial Sensitivity Tests , Minocycline/metabolism , Mutation/genetics , Phenotype , Tannins/pharmacology , Tigecycline
8.
BMC Microbiol ; 14: 119, 2014 May 09.
Article in English | MEDLINE | ID: mdl-24885279

ABSTRACT

BACKGROUND: Tigecycline resistance in Acinetobacter baumannii is primarily acquired through overexpression of the AdeABC efflux pump. Besides AdeRS, other two-component regulatory systems (TCSs) involving the regulation of this transporter have not been clarified. RESULTS: In this study, we found that the TCS genes baeR and baeS are co-transcribed and function as stress responders under high osmotic conditions. The baeSR and adeAB genes showed increased transcription in both the laboratory-induced and clinical tigecycline-resistant strains compared with the wild-type strain. The deletion of baeR in the ATCC 17978 strain led to 67-73% and 68% reduction in adeA and adeB expression, respectively, with a resultant 2-fold decrease in the tigecycline minimal inhibition concentration (MIC). In contrast, the overexpression of baeR resulted in a doubled tigecycline MIC, with a more than 2-fold increase in adeA and adeB expression. The influence of baeR knockout on adeAB gene expression can also be observed in the laboratory-induced tigecycline-resistant strain. A time-kill assay showed that the baeR deletion mutant showed an approximate 1-log10 reduction in colony forming units (CFUs) relative to the wild-type strain when the tigecycline concentration was 0.25 µg/mL throughout the assay period. The wild-type phenotype could be restored by trans-complementation with pWH1266-kanr-baeR. Increasing the tigecycline concentration to 0.5 µg/mL produced an even more marked 4.7-log10 reduction in CFUs of the baeR deletion mutant at 8 h, while only a 2.1-log10 reduction was observed for the wild-type strain. CONCLUSIONS: Taken together, these data show for the first time that the BaeSR TCS influences the tigecycline susceptibility of A. baumannii through the positive regulation of the resistance-nodulation-division efflux pump genes adeA and adeB.


Subject(s)
Acinetobacter baumannii/drug effects , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/metabolism , Membrane Transport Proteins/metabolism , Minocycline/analogs & derivatives , Protein Kinases/metabolism , Trans-Activators/metabolism , Acinetobacter baumannii/genetics , Acinetobacter baumannii/physiology , Drug Resistance, Bacterial , Gene Expression Regulation, Bacterial , Microbial Sensitivity Tests , Minocycline/pharmacology , Osmotic Pressure , Stress, Physiological , Tigecycline
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