OBJECTIVE: To construct a modABC gene knockout strain of Proteus mirabilis and explore the effect of modABC gene deletion on biological characteristics of Proteus mirabilis. METHODS: Fusion PCR was used to obtain the fusion gene of modABC and the kanamycin-resistant gene Kn, which was ligated with the suicide vector pCVD442 and transduced into Proteus mirabilis. The modABC gene knockout strain of Proteus mirabilis was obtained after homologous recombination with the suicide vector. PCR and Sanger sequencing were used to identify genomic deletion of modABC gene in the genetically modified strain. The concentration of molybdate in the wild-type and gene knockout strains was determined using inductively coupled plasma mass spectrometry (ICP-MS), and their survival ability in LB medium was compared under both aerobic and anaerobic conditions. RESULTS: PCR and sanger sequencing confirmed genomic deletion of modABC gene in the obtained Proteus mirabilis strain. The concentration of intracellular molybdenum in the modABC gene knockout strain was 1.22 mg/kg, significantly lower than that in the wild-type strain (1.46 mg/kg, P < 0.001). Under the aerobic condition, the modABC gene knockout strain grown in LB medium showed no significant changes in survival ability compared with the wild-type strain, but its proliferation rate decreased significantly under the anaerobic condition and also when cultured in nitrate-containing LB medium under anaerobic condition. CONCLUSION Homologous recombination with the suicide vector can be used for modABC gene knockout in Proteus mirabilis. modABC gene participates in molybdate uptake and is associated with anaerobic growth of Proteus mirabilis in the presence of nitrate.
Humans ; Gene Deletion ; Nitrates ; Proteus mirabilis/genetics* ; Gene Knockout Techniques

OBJECTIVE: A core genome multilocus sequence typing (cgMLST) scheme to genotype and identify potential risk clonal groups (CGs) in Proteus mirabilis. METHODS: In this work, we propose a publicly available cgMLST scheme for P. mirabilis using chewBBACA. In total 72 complete P. mirabilis genomes, representing the diversity of this species, were used to set up a cgMLST scheme targeting 1,842 genes, 635 unfinished (contig, chromosome, and scaffold) genomes were used for its validation. RESULTS: We identified a total of 205 CGs from 695 P. mirabilis strains with regional distribution characteristics. Of these, 159 unique CGs were distributed in 16 countries. CG20 and CG3 carried large numbers of shared and unique antibiotic resistance genes. Nine virulence genes ( papC, papD, papE, papF, papG, papH, papI, papJ, and papK) related to the P fimbrial operon that cause severe urinary tract infections were only found in CG20. These CGs require attention due to potential risks. CONCLUSION This research innovatively performs high-resolution molecular typing of P. mirabilis using whole-genome sequencing technology combined with a bioinformatics pipeline (chewBBACA). We found that the CGs of P. mirabilis showed regional distribution differences. We expect that our research will contribute to the establishment of cgMLST for P. mirabilis.
Genome, Bacterial ; Proteus mirabilis/genetics* ; Multilocus Sequence Typing ; Molecular Epidemiology ; Genotype

Proteus mirabilis lipase (PML) features tolerance to organic solvents and great potential for biodiesel synthesis. However, the thermal stability of the enzyme needs to be improved before it can be used industrially. Various computational design strategies are emerging methods for the modification of enzyme thermal stability. In this paper, the complementary algorithm-based ABACUS, PROSS, and FoldX were employed for positive selection of PML mutations, and their pairwise intersections were further subjected to negative selection by PSSM and G_REMLIN to narrow the mutation library. Thereby, 18 potential single-point mutants were screened out. According to experimental verification, 7 mutants had melting temperature (T_m) improved, and the ΔT_m of K208G and G206D was the highest, which was 3.75 ℃ and 3.21 ℃, respectively. Five mutants with activity higher than the wild type (WT) were selected for combination by greedy accumulation. Finally, the T_m of the five-point combination mutant M10 increased by 10.63 ℃, and the relative activity was 140% that of the WT. K208G and G206D exhibited certain epistasis during the combination, which made a major contribution to the improvement of the thermal stability of M10. Molecular dynamics simulation indicated that new forces were generated at and around the mutation sites, and the rearrangement of forces near G206D/K208G might stabilize the Ca²⁺ binding site which played a key role in the stabilization of PML. This study provides an efficient and user-friendly computational design scheme for the thermal stability modification of natural enzymes and lays a foundation for the modification of PML and the expansion of its industrial applications.
Enzyme Stability ; Lipase/chemistry* ; Molecular Dynamics Simulation ; Proteus mirabilis/metabolism* ; Solvents/chemistry*

Tyrosine is an important aromatic amino acid. Besides its nutritional value, tyrosine is also an important precursor for the synthesis of coumarins and flavonoids. Previously, our laboratory constructed a Saccharomyces cerevisiae strain LTH0 (ARO4K229L, ARO7G141S, Δaro10, Δzwf1, Δura3) where tyrosine feedback inhibition was released. In the present study, heterologous expression of betaxanthins synthesis genes DOD (from Mirabilis jalapa) and CYP76AD1 (from sugar beet B. vulgaris) in strain LTH0 enabled production of yellow fluorescence. The engineered strain LTH0-DOD-CYP76AD1 was subjected to UV combined with ARTP mutagenesis, followed by flow cytometry screening. Among the mutants screened, the fluorescence intensity of the mutant strain LTH2-5-DOD-CYP76AD1 at the excitation wavelength of 485 nm and emission wavelength of 505 nm was (5 941±435) AU/OD, which was 8.37 times higher than that of strain LTH0-DOD-CYP76AD1. Fourteen mutant strains were subjected to fermentation to evaluate their tyrosine producing ability. The highest extracellular tyrosine titer reached 26.8 mg/L, which was 3.96 times higher than that of strain LTH0-DOD-CYP76AD1. Heterologous expression of the tyrosine ammonia lyase FjTAL derived from Flavobacterium johnsoniae further increased the titer of coumaric acid to 119.8 mg/L, which was 1.02 times higher than that of the original strain LTH0-FjTAL.
Flavobacterium ; High-Throughput Screening Assays ; Mirabilis ; Saccharomyces cerevisiae/genetics* ; Tyrosine

High cholesterol is one of the important factors inducing cardiovascular and cerebrovascular diseases. Drug therapy is the main method for reducing cholesterol, but has the disadvantages such as high cost and side effects. Studies have shown that intestinal bacteria play important roles in cholesterol metabolism. However, there are few reports on the screening and functional evaluation of cholesterol-lowering intestinal bacteria. In this study, 36 bile-tolerant bacteria were screened from healthy people stool through culturomics using bovine bile acid or artificial mixed bile acids as substrates. Taking Lactobacillus rhamnosus GG (LGG) as a positive control, three bile acid concentration groups (0 g/L, 0.3 g/L, 3 g/L) were set up to evaluate the cholesterol-lowering ability of bile-tolerant bacteria in vitro. Ten bacteria (including Proteus mirabilis, Providencia stuartii, Proteus vulgaris et al) were identified as the dominant cholesterol-lowering bacteria. Six of the above bacteria, Proteus mirabilis, Providencia stuartii, Proteus vulgaris, Proteus penneri, Wohlfahrtiimonas chitiniclastica, Providencia rettger, were evaluated for their ability to reduce triglycerides in vitro and tolerance to artificial gastric juice. Comparing with strain LGG, the six bacteria showed better triglyceride-lowering ability in vitro. With the decrease of pH value of artificial gastric juice and the increase of treatment time, the survival rate of six bacteria decreased. The above screening experiments and functional evaluation provide a basis for further development of potential cholesterol-lowering bacterial products.
Animals ; Cattle ; Cholesterol ; Gammaproteobacteria ; Humans ; Proteus mirabilis ; Providencia

The purpose of this study is to investigate the correlation of cell surface hydrophobicity (CSH) and biofilm formation or adhesion in Candida albicans (C. albicans) and several pathogenic bacteria. All of C. albicans (n=82) and 7 bacterial species (Escherichia coli, n=25; Klebsiella pneumoniae, n=33; Morganella morganii, n=21; Proteus mirabilis, n=33; Proteus vulgaris, n=12; Pseudomonas aeruginosa, n=31; Staphylococcus aureus, n=31) were isolated clinically. CSH was quantified with microbial adhesion to hydrocarbons. Biofilm formation was determined by tetrazolium salt reduction assay. Adhesion assay was performed by counting colonies after culture the microbes adhered to HeLa cells. Although high CSH-expressing bacterial species showed greater adherence to HeLa cells and larger amounts of biofilm formation on polystyrene, the significant relationships within same species were not shown. In C. albicans, however, strong positive correlations were observed between CSH and biofilm formation (r =0.708; p < 0.05) or cell adhesion (r =0.509; p < 0.05). These results suggest that hydrophobic force of bacteria may play a minor role in adhesion and biofilm formation, but CSH of C. albicans may be an important factor for adherence on surface and biofilm forming process.
Bacteria ; Biofilms* ; Candida albicans* ; Candida* ; Cell Adhesion ; HeLa Cells ; Humans ; Hydrocarbons ; Hydrophobic and Hydrophilic Interactions* ; Klebsiella pneumoniae ; Morganella morganii ; Polystyrenes ; Proteus mirabilis ; Proteus vulgaris ; Pseudomonas aeruginosa ; Staphylococcus aureus

Proteus mirabilis (P. mirabilis) meningitis in a neonate is rare, but its recognition is important because the disease progresses rapidly and has poor prognosis. A 4-day-old premature female infant born at 36 weeks and 5 days of gestation presented with symptoms of fever and icteric skin. Initial cerebrospinal fluid findings suggested bacterial meningitis, and treatment with antibiotics was started. On the third day, P. mirabilis growth was found in both blood and cerebrospinal fluid cultures and brain computed tomography revealed normal findings. The patient showed improved clinical symptoms, but brain magnetic resonance imaging on hospital day 18 revealed a brain abscess measuring 4.5×3.1×3.1 cm in the right frontal lobe. Cyst drainage was performed immediately and a catheter was inserted. Follow-up computed tomography revealed a tiny abscess remaining in the right frontal lobe, and follow-up magnetic resonance imaging later demonstrated marked interval regression in the size of the abscess. The patient was discharged on day 57 of hospitalization in good condition. Serial brain imaging should be considered in neonatal cases of P. mirabilis meningitis.
Abscess ; Anti-Bacterial Agents ; Brain Abscess* ; Brain* ; Catheters ; Cerebrospinal Fluid ; Drainage ; Female ; Fever ; Follow-Up Studies ; Frontal Lobe ; Hospitalization ; Humans ; Infant ; Infant, Newborn ; Magnetic Resonance Imaging ; Meningitis* ; Meningitis, Bacterial ; Mirabilis ; Neuroimaging ; Pregnancy ; Prognosis ; Proteus mirabilis* ; Proteus* ; Skin

Pet turtles are well-known to harbor an array of bacterial pathogens which can cause zoonotic infections in humans as well as opportunistic infections in the turtles itself. Essential oils are the natural plant extracts which have been traditionally used for disease treatment. In the present study, the essential oil of lavender (EOL) was examined for its antibacterial activity against thirty-eight strains of turtle-borne pathogenic bacteria belonging to seven species; Aeromonas hydrophila, A. caviae, A. dhakensis, Citrobacter freundii, Proteus mirabilis, Salmonella enterica and Pseudomonas aeruginosa. Antibacterial activity of EOL was tested by means of disk diffusion, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) tests. In addition, the antimicrobial susceptibility pattern of 11 commonly used antimicrobials was examined and the multiple antibiotic resistance (MAR) index was calculated. The results revealed that EOL was active against all tested turtle-borne pathogenic bacteria except P. aeruginosa. The range of MIC and MBC values of EOL against isolates except P. aeruginosa were recorded as 0.5-1% (V/V) and 0.5-2% (V/V), respectively. The MBC/MIC ratio was detected as <4, revealing that the tested EOL was bactericidal. Besides, most of the isolates were resistant to different antimicrobials in antimicrobial disk diffusion test. MAR index values of the tested strains were ranging from 0.27 to 0.91. The outcomes indicate that EOL has a potential to be used as an antibacterial agent against pathogenic bacteria isolated from pet turtles.
Aeromonas hydrophila ; Animals ; Bacteria* ; Citrobacter freundii ; Diffusion ; Drug Resistance, Microbial ; Guinea Pigs ; Humans ; Lavandula* ; Microbial Sensitivity Tests ; Oils, Volatile ; Opportunistic Infections ; Plant Extracts ; Proteus mirabilis ; Pseudomonas aeruginosa ; Salmonella enterica ; Turtles ; Zoonoses

The usage of essential oils as antimicrobial agents is gaining attention. Besides, pet turtles were known to harbor a range of pathogenic bacteria while the turtle keeping is a growing trend worldwide.The current study examined the antimicrobial activity of lemon grass oil (LGO) against seven species of Gram negative bacteria namely; Aeromonas hydrophila, A. caviae, Citrobacter freundii, Salmonella enterica, Edwardsiella tarda, Pseudomonas aeruginosa, and Proteus mirabilis isolated from three popular species of pet turtles. Along with the results of disc diffusion, minimum inhibitory and minimum bactericidal concentration (MIC and MBC) tests, LGO was detected as effective against 6 species of bacteria excluding P. aeruginosa. MIC of LGO for the strains except P. aeruginosa ranged from 0.016 to 0.5% (V/V). The lowest MIC recorded in the E. tarda strain followed by A. hydrophilla, C. freundii, P. mirabilis, and S. enterica. Interestingly, all the bacterial species except E. tarda were showing high multiple antimicrobial resistance (MAR) index values ranging from 0.36 to 0.91 upon the 11 antibiotics tested although they were sensitive to LGO.
Aeromonas hydrophila ; Animals ; Anti-Bacterial Agents ; Anti-Infective Agents ; Bacteria* ; Citrobacter freundii ; Cymbopogon* ; Diffusion ; Edwardsiella tarda ; Gram-Negative Bacteria ; Guinea Pigs ; Mirabilis ; Oils, Volatile ; Proteus mirabilis ; Pseudomonas aeruginosa ; Salmonella enterica ; Turtles*

PURPOSE: To evaluate the change in pathogens in cultured Jones tubes used in lacrimal bypass surgery according to postoperative period and to provide basic data related to preventive antibiotics or functional lacrimal stent development. METHODS: Fifty patients who underwent Jones tubes removal were enrolled in this study. Removed Jones tubes were cultured to identify bacteria and were tested for antibiotic sensitivity. The results were further analyzed according to the period between lacrimal bypass surgery and tube removal. RESULTS: Among 50 cases, 24 (48%) showed cultured bacteria of Staphylococcus aureus, 5 (10%) were Pseudomonas, and another 5 (10%) were Gram-positive bacilli. Although Staphylococcus aureus was the most frequently cultured organism, Proteus mirabilis was the most common cultured organism in patients who underwent tube removal more than 10 years after lacrimal bypass surgery. There was no significant correlation between cultured organism and the period between lacrimal bypass surgery and tube removal. Eighty four percent of cultured Staphylococcus aureus showed resistance to penicillin, and 53% of cultured Staphylococcus aureus showed resistance to methicillin. CONCLUSIONS: Staphylococcus aureus was the most frequently cultured organism according to Jones tube-related lacrimal bypass surgery. A large proportion of cultured Staphylococcus aureus showed resistance to penicillin and methicillin. Proteus mirabilis should be considered the most common pathogen in patients more than 10 years after lacrimal bypass surgery.
Anti-Bacterial Agents ; Bacteria ; Humans ; Methicillin ; Penicillins ; Postoperative Period* ; Proteus mirabilis ; Pseudomonas ; Staphylococcus aureus ; Stents