Evaluation of antibacterial, antifungal and antibiofilm activities of A. baumannii-derived tannase and gallic acid against uropathogenic microorganisms.

One of the most prevalent infectious diseases and a key driver of antibiotic prescriptions in pediatrics is urinary tract infection (UTI). Due to the emergence of more resistant uropathogenic bacterial and fungal strains, current treatments are no longer effective, necessitating the urgent development of novel antibacterial and antifungal drugs. In this study, the antifungal, antibacterial, and anti-biofilm capabilities of compounds, such as tannase (TN) and gallic acid (GA), which were produced from a novel natural source, Acinetobacter baumannii (AB11) bacteria, were assessed for the inactivation of uropathogenic microorganisms (UMs). Ammonium sulphate precipitation, ion exchange, high-performance liquid chromatography, and gel filtration were used to purify TN and GA that were isolated from A. baumannii. A 43.08 % pure TN with 1221.2 U/mg specific activity and 10.51 mg/mL GA was obtained. The antibacterial, antifungal and anti-biofilm activities of TN and GA were evaluated against UMs and compared to those of commercially available antibiotics including sulfamethoxazole (SXT), levofloxacin (LEV), ciprofloxacin (CIP), amikacin (Ak), and nitrofurantoin (F). The results showed that TN and GA were superior to commercial antibiotics in their ability to inactivate UMs and considerably reduced biofilms formation. Additionally, the GA emerges as the top substitute for currently available medications, demonstrating superior antibacterial and antibiofilm properties against all UMs evaluated in this study. The results of this investigation showed that A. baumannii-derived TN and GA could be utilized as an alternative medication to treat UTIs.

Broad-Spectrum Bioactivity of Chitosan N-acetylglucosaminohydrolase (Chitosan NAGH) Extracted from Bacillus ligniniphilus.

Background: The genus Bacillus has species with strains that produce Chitosan N-acetylglucosaminohydrolase (NAGH), a hydrolytic enzyme. Objective: A novel bacterium, Bacillus ligniniphilus, was characterized as producing Chitosan NAGH. This study further examine its antibiofilm properties and its possible uses against biofilm-producing bacteria. Methods: Various sea soil samples were evaluated for the presence of Chitosan NAGH. The chosen isolate, Bacillus ligniniphilus 61, was then used to extract and purify Chitosan NAGH using precipitation in ammonium sulfate followed by polyethylene glycol-treated dialysis and gel-permeation chromatography. Biofilm inhibition and antimicrobial activity of Chitosan NAGH was estimated against different bacterial species. Both gene expression profiling of biofilm-related genes and an extracellular polymeric substance (EPS) inhibition assay were performed. Results: The BL61 strain was able to produce much more Chitosan activity than the other strains, as the latter only exhibited antimicrobial activity at low concentration levels; however, they did show as antibiofilm agents at varying proportions. Chitosan NAGH caused a uniform decrease in EPS formation in each isolate. Many biofilm-related genes, e.g., IcaABCD, decreased, but genes related to autoinducer synthetase were not affected by Chitosan NAGH. EPS, which is responsible for polysaccharide formation, was underexpressed at 3-fold down. Conclusions: The current study results allow future researchers to look for better and newer compounds with the antibiofilm property that inhibits the formation of biofilm created by a wide range of bacteria without affecting their growth.

A Novel Genetic Determination of a Lectin Gene in Iraqi Acinetobacter baumannii Isolates and Use of Purified Lectin as an Antibiofilm Agent.

BACKGROUND: Lectin was initially called hemagglutinin or agglutinin because of its capacity to agglutinate human as well as human erythrocytes. They are a heterogeneous group of proteins or glycoproteins of nonimmune origin. Because of their chemical properties, they have become a useful tool in several fields such as immunology, cell biology, molecular biology, membrane structure, pharmacology, cancer research, clinical chemistry, and genetic engineering. OBJECTIVE: The wide applications of lectins users urged the need to isolate lectins from a new strain of bacteria can produce new and high yield of lectin because the current production of lectin from Pseudomonas spp. is very expensive. The goal of this study was to screen the ability of Acinetobacter baumannii isolates to produce lectin and detection of its phenotypic and genotypic profiles and detection of lectin ability to inhibit ofbiofilm formation. METHODS: Fifty-one isolates from different sources were collected and detected genetically by using the recA gene. Phenotypic detection of lectin by using semi-quantitative analysis and quantitative analysis in microtiter plate. Genotypic detection of lectin by designed lec gene and used PCR technique. The lectin was extracted by using glass beads and purified by chromatographyic technique followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis for determination the molecular size of lectin and finally detection the spectrum of biofilm inhibition by the purified lectin toward biofilm producers. RESULTS: Of 51 A. baumannii isolates, 17 (33.3%) have been found to produce lectin. Ten of 17 were sequenced, of which 2 were submitted and tested by the gene bank National Center for Biotechnology Information (NCBI), and accession numbers (KX766405.1 and KX766406.1) were obtained. These 17 isolates were phenotypically and genotypically positive for lectin and showed different lec gene expression in semi-quantitative and quantitative analysis. The activities ranged between 4-128 U/mL. Lectin purified by ammonium sulfate precipitation was used to inhibit biofilm formation. We found reduction at three different types of bacteria ranging from 26% for Klebsiella pneumonia, 46.7% for P. stutzeri and 53% for A. baumannii. These results suggested that lectin has a promising application as an antibiofilm agent to combat the growing number of multidrug-resistant pathogen-associated infections. CONCLUSIONS: Lectin has been detected recently in A. baumannii, but the genetic property of this lectin has not yet been fully studied. In our study, we determined the presence of the lectin gene (lec gene) in A. baumannii by using PCR technique, and lec PCR products were identified with various source of isolation and sequenced to screening for epidemiology and submitted to the gene bank NCBI under accession number (KX766405.1 and KX766406.1). HIGHLIGHTS: A. baumannii has an ability to produce lectin protein; Lec gene was detected in A. baumannii, and the sequence was recorded under accession number KX766405.1 and KX766406.1.; Lectin was extracted by glass beads and purified by chromatographyic technique; Lectin had strong effect against biofilm formation.

Chitosan extracted from Aspergillus flavus shows synergistic effect, eases quorum sensing mediated virulence factors and biofilm against nosocomial pathogen Pseudomonas aeruginosa.

The biological methods for extraction of chitosan were used as alternative procedures for chemical methods In biological methods, the chitosan was extracted from A. flavus by using of Lactobacillus paracasei for demineralization and Bacillus subtilis for deproteinization. The yield of extracted chitosan reached to 53.8%, pH was 7.8 and complete solubility in 1% acitic acid. Purified chitosan had the ability to reduce the biofilm forming capacity of P. aeruginosa clearly visible in light microscopic staining and scanning electron microscopy. The QS dependent phenotype and QS regulated gene expression was significantly reduced in the influence of chitosan. A significant decrease in biofilm formation was seen in the presence of chitosan. The chitosan treatment showed a decrease in the expression of lasR and rhlR genes. Same time production of pyocyanin and proteases was also inhibited in dose dependent manner. Chitosan led to increasing antimicrobial activity of antibiotics and had synergism effect, thus chitosan may be a useful adjuvant agent for the treatment of many bacterial infections in combination with antibiotics.

Screening, nutritional optimization and purification for phytase produced by Enterobacter aerogenes and its role in enhancement of hydrocarbons degradation and biofilm inhibition.

In this study, a novel isolate of Enterobacter aerogenes isolated from contaminated soils with hydrocarbons had extracellular phytate-degrading activity. Enterobacter aerogenes isolates were identified by biochemical tests and confirmed by16S rRNA gene products (amplified size 211bp) for genotypic detection. The phytase activity was reached to maximum activity when this isolate was cultivated under the optimal conditions which consisted of using minimal salt medium containing 1%(w/v) rice bran as a sole source for carbon and 2% (w/v) yeast extract at pH 5.5 and temperature of 50°C for 48 h. The phytase had purified to homogeneity by 50% ammonium sulphate precipitation, ion exchange and gel filtration chromatography with 75.7 fold of purification and a yield of 30.35%. The purified phytase is a single peptide with approximate molecular mass of 42 kDa as assessed by SDS-PAGE. The highest degradative ability by Enterobacter aerogenes of black oil, white oil and used engine oil had observed after 72 h of incubation. Rapid degradation of black oil and used engine oil had also observed while slow degradation of white oilat all time of incubation. The purified phytase inhibited biofilm formation ability in a dose-dependent manner for all Gram-negative and Gram-positive biofilm-forming bacteria and a significant difference in cell surface hydrophobicity was observed after exposure of planktonic cells to phytase for hour. The hydrolyzing effect of phytase released by Enterobacter aerogenes for complex salts of phosphorus that are insoluble in the soil led to increase of phosphorus concentrations and enhanced the ability of Enterobacter aerogenes to degrade a specific hydrocarbon in contaminated soil so that the phytase has a promising application in bioremediation of contaminated soils with hydrocarbons.

Chitosanase purified from bacterial isolate Bacillus licheniformis of ruined vegetables displays broad spectrum biofilm inhibition.

A number of bacterial species produces chitosanases which has variety of applications because of its high biodegradability, non-toxicity and antimicrobial assets. In the present study chitosanase is purified from new bacterial species Bacillus licheniformis from spoiled vegetable. This novel strain of Bacillus licheniformis isolated from spoilt cucumber and pepper samples has the ability to produce the chitosanase enzyme when grown on chitosan substrate. Study also examined its antibiofilm properties against diverse bacterial species with biofilm forming ability. The purified chitosanase inhibited the biofilm formation ability for all Gram-negative and Gram-positive biofilm-forming bacteria [biofilm producers] tested in this study in congo red agar and microtiter plate's methods. Highly antibiofilm activity of chitosanase was recorded against Pseudomonas aeruginosa followed by Klebsiella pneumoniae with reduction of biofilm formation upto 22 and 29%, respectively compared with [100] % of control. Biofilm formation has multiple role including ability to enhance resistance and self-protection from external stress. This chitosanase has promising benefit as antibiofilm agent against biofilm forming pathogenic bacteria and has promising application as alternative antibiofilm agents to combat the growing number of multidrug resistant pathogen-associated infections, especially in situation where biofilms are involved.