Prevalence of Class 1, 2, and 3 Integrons and Biofilm Formation in Pseudomonas aeruginosa and Acinetobacter baumannii among ICU and nonICU Patients

Background: Infections caused by Pseudomonas aeruginosa or Acinetobacter baumannii are of greatest concern for hospitalized patients, particularly those in intensive care units (ICUs). The aims of this study were to investigate the prevalence of integrons and biofilm formation among P. aeruginosa and A. baumannii isolates collected from ICU and non-ICU inpatients. Materials and Methods: A total of 90 P. aeruginosa and 90 A. baumannii isolates were recovered from patients admitted into diverse units of Shahid Mohammadi hospital in Bandar Abbas from January to December 2014. Bacterial identification was carried out by phenotypic methods and PCR. Antibiotic susceptibility was measured by disk diffusion assay. The presence of Class 1, 2, and 3 integrons were evaluated by multiplex-PCR. Biofilm quantification was done by microtiter method. Results: The highest number of isolates (48%) were recovered from ICU patients. 81% of P. aeruginosa isolates were sensitive to piperacillin/tazobactam and ticarcillin, while 60% were resistant to third generation of cephalosporins. In case of A. baumannii, all the isolates were sensitive to colistin, but 98% were resistant to other antibiotics (p≤0.05). Susceptibility to ceftazidime, ticarcillin, imipenem, and piperacillin/tazobactam were higher among isolates obtained from non-ICU patients. Class 1 integron was detected in 13.3% of the P. aeruginosa and 40% of the A. baumannii isolates, while Class 2 integron was harbored by 7 and 6.6% of the isolates, respectively. Furthermore, 23% of the A. baumannii and 12% of the P. aeruginosa isolates showed strong biofilm activity. Conclusion: Class 1 integron-positive isolates were resistant to three classes of antibiotics and predominantly observed in specimens collected from ICU patients showing strong biofilm

Biosynthesis and recovery of selenium nanoparticles and the effects on matrix metalloproteinase-2 expression

Today, green synthesis of nanoparticles is attracting increasing attention. In the present study, the Bacillus sp. MSh-1 was isolated from the Caspian Sea (located in the northern part of Iran) and identified by various identification tests and 16S ribosomal DNA analysis. The reduction time course study of selenium ion (Se4+) reduction by using this test strain was performed in a liquid culture broth. Then, the intracellular NPs (nanoparticles) were released by the liquid nitrogen disruption method and thoroughly purified using an n-octyl alcohol water extraction system. Characterization of the separated NPs on features such as particle shape, size and purity was carried out with different devices. The energy dispersive X-ray and X-ray diffraction patterns showed that the purified NPs consisted of only selenium and are amorphous respectively. In addition, the transmission electron micrograph showed that the separated NPs were spherical and 80–220 nm in size. Furthermore, the cytotoxicity effect of these extracted biogenic selenium (Se) NPs on the fibrosarcoma cell line (HT-1080) proliferation and the inhibitory effect of the Se NPs on MMP-2 (matrix metalloproteinase-2) expression were studied using the MTT [3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay and gelatin zymography. Biogenic Se NPs showed a moderately inhibitory effect on MMP-2 expression

Acute and subacute toxicity of novel biogenic selenium nanoparticles in mice

Context: In the present investigation, acute and subacute toxicity of the biogenic Se nanoparticles (Se NPs) has been reported. Objective: To characterize the Se NPs produced by a bacterium species and to evaluate their toxicity and impact on clinical chemistry and hematological parameters of NMRI mice. Materials and methods: The Se NPs were prepared by Bacillus sp. MSh-1 in a culture medium containing SeO2 (1.26 mM) and their physiochemical properties investigated using TEM, XRD and FT-IR. The LD50 of Se NPs and SeO2 were determined and the subacute toxicity evaluated by orally administration of 0, 2.5, 5, 10 and 20 mg kg−1 of Se NPs to male mice for 14 consecutive days. Parameters of blood cells, AST, ALT, ALP, creatinine, BUN, cholesterol, bilirubin, triglyceride and CPK were experimentally measured. Results: The XRD and TEM analyses showed that the spherical NPs were amorphous, in the size range of 80–220 nm. The toxicological evaluation showed that the LD50 values of SeO2 and Se NPs were 7.3 and 198.1 mg kg−1, respectively. No biochemical changes were observed from the administration of 2.5, 5 and 10 mg kg−1 of Se NPs, but a dose of 20 mg kg−1 was accompanied with signs of toxicity including lower body weight and changes in clinical chemistry and hematological parameters. Conclusion: The biogenic Se NPs were less toxic than synthetic Se NPs and much less (26-fold) toxic than the SeO2, which demonstrates the important role of Bacillus sp. MSh-1 in conversion of a highly toxic Se compound to the less toxic Se NPs. Keywords: L

Iron limitation enhances acyl homoserine lactone (AHL) production and biofilm formation in clinical isolates of Acinetobacter baumannii

Acinetobacter baumannii is an important source of infections in intensive care units (ICUs) of our hospitals in Kerman, Iran and the most frequently isolated strains produce biofilm. There is a little information about role of iron (Fe) levels on acyl homoserine lactone (AHL) production and biofilm formation in this microorganism. In the present study, we investigated the influence of iron-III limitation on AHL, siderophore, catechol and virulence factors in the biofilm forming clinical strains of A. baumannii. A total of 65 non-duplicated multidrug resistance (MDR) strains of A. baumannii were isolated from patients in ICUs of 2 hospitals in Kerman, Iran. Antibiotic susceptibility, siderophore and other iron chelators, hemolysis, cell twitching motility, capsule, gelatinase and DNase were studied. Presence of quorum sensing, LuxI and LuxR genes was detected by multiplex-PCR. AHL activity quantified by colorimetric method and the functional groups were determined by Fourier Transform Infra-Red Spectroscopy (FT-IR). Biofilm formation was detected by microtiter plate technique. All of the isolates were resistant to third generation of cephalosporins, ciprofloxacin, levofloxacin, tetracycline, whereas, 78% and 81% were resistant to amikacin and carbapenems, respectively. The siderophore activity was highest at 20 mM Fe3C (70%); however, it decreased to 45% as concentration of Fe3C increased to 80 mM. Furthermore, screening of the isolates for LuxI and LuxR genes showed that presence of both genes required in the isolates with high AHL activity. FT-IR analysis indicated CDO bond of the lactone ring and primary amides. Significantly, a higher amount of AHL (70%) was detected in the presence of low concentration of iron-III (20 mM); as iron concentration increased to 80 mM, the AHL activity was reduced to 40% (P � 0.05). All the isolates exhibited twitching motility and had a capsule. No any gelatinase or DNase activity was detected. Quantification of the biofilm formation introduced 23 isolates with efficient attachment to microplate wells and strong biofilm. We found that both the AHL production and biofilm formation were regulated by iron concentration in a dose dependent manner. These findings provide evidence that iron limitation plays an important regulatory role in AHL and siderophore production resulting in strong or weak biofilm, thereby helping the organism to persist in less available micronutrient environment

Effect of selenium supplementation with sodium selenite and selenium nanoparticles on iron homeostasis and transferrin gene expression in sheep: A preliminary study

The present research aimed at evaluating the effects of sodium selenite and selenium nanoparticles (Se NPs) on iron homeostasis and the expression of transferrin and its receptor-binding protein genes. Twenty one Lori–Bakhtiary sheep were randomly allocated into 3 groups. Groups 1 and 2 orally received Se NPs and sodium selenite (1 mg kg�1) for 10 consecutive days, respectively. Group 3 served as the control. Blood and sternal bone marrow samples were collected at different supplementation intervals. Various factors such as serum iron concentration, total iron binding capacity (TIBC), and transferrin saturation percent were determined. The expression of transferrin and transferrin binding receptor genes was also studied. Results showed a decreasing trend in serum iron concentration particularly during the early and middle stages of supplementation (0–20 days) with Se NPs or selenium ions. Conversely, the TIBC level increased in sera especially during these periods (0–20 days) in animals that received selenium NPs or selenium ions. Our results also showed that expression of transferrin and its receptor genes was considerably increased during supplementation of the animals by both selenium compounds for 10 or 20 days. After this period, the expression of the mentioned genes significantly decreased, especially in animals that received selenium ions

Molecular Analysis and Expression of bap Gene in Biofilm-Forming Multi-Drug-Resistant Acinetobacter baumannii

Background: Acinetobacter baumanniiis commonly resistant to nearly all antibiotics due to presence of antibiotic resistance genes and biofilm formation. In this study we determined the presence of certain antibiotic-resistance genes associated with biofilm production and the influence of low iron concentration on expression of the biofilmassociated protein gene (bap) in development of biofilm among multi-drug-resistant A. baumannii (MDRAB). Methods: Sixty-five MDRAB isolates from clinical samples were collected. Molecular typing was carried out by random amplified polymorphism DNA polymerase chain reaction (RAPD-PCR). Biofilm formation was assayed by the microtiter method. Results: The sequence of bap was determined and deposited in the GenBank database (accession no. KR080550.1). Expression of bap in the presence of low iron was analyzed by relative quantitative real time PCR (rqRT-PCR). Nearly half of the isolates belonged to RAPD-types A and B remaining were either small clusters or singleton. The results of biofilm formation revealed that 23 (35.4%), 18 (27.7%), 13 (20%), and 11 (16.9%) of the isolates had strong, moderate, weak, and no biofilm activities, respectively. ompA and csuE genes were detected in all, while bap and blaPER-1 were detected in 43 (66%) and 42 (64%) of the isolates that showed strong and moderate biofilm activities (p ≤ 0.05), respectively. Analysis ofbapexpression by rqRT-PCR revealed five isolates with fourfold bap overexpression in the presence of low iron concentration (20 µM). Conclusion: The results suggest thatbapoverexpression may influence biofilm formation in presence of low iron concentration

Biosynthesis and recovery of rod-shaped tellurium nanoparticles and their bactericidal activities

In this study, a tellurium-transforming Bacillus sp. BZ was isolated from the Caspian Sea in northern Iran. The isolate was identified by various tests and 16S rDNA analysis, and then used to prepare elemental tellurium nanoparticles. The isolate was subsequently used for the intracellular biosynthesis of elemental tellurium nanoparticles. The biogenic nanoparticles were released by liquid nitrogen and purified by an n-octyl alcohol water extraction system. The shape, size, and composition of the extracted nanoparticles were characterized. The transmission electron micrograph showed rod-shaped nanoparticles with dimensions of about 20 nm � 180 nm. The energy dispersive X-ray and X-ray diffraction spectra respectively demonstrated that the extracted nanoparticles consisted of only tellurium and have a hexagonal crystal structure. This is the first study to demonstrate a biological method for synthesizing rod-shaped elemental tellurium by a Bacillus sp., its extraction and its antibacterial activity against different clinical isolates

Efficacy of biogenic selenium nanoparticles against Leishmania major: In vitro andin vivo studies

Project: This study investigated the in vitro and in vivo effectiveness of biogenic selenium nanoparticles(Se NPs), biosynthesized by Bacillus sp. MSh-1, against Leishmania major (MRHO/IR/75/ER). Procedure:The 3-(4,5-dimethylthiozol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay was used to evaluatethe cytotoxicity effects of the biogenic Se NPs against both promastigote and amastigote forms of L.major. In a separate in vivo experiment, we also determined the preventive and therapeutic effects ofbiogenic Se NPs in BALB/c mice following subcutaneous infected with L. major. Results: The MTT assaysshowed that the highest toxicity occurred after 72 h against both promastigote and amastigote formsof L. major. The cytotoxicity of Se NPs was higher at all incubation times (24, 48, and 72 h) against thepromastigote than the amastigote form (p < 0.05). The 50% inhibitory concentrations (IC50) of the Se NPswere 1.62 ± 0.6 and 4.4 ± 0.6 �g ml−1against the promastigote and amastigote forms, respectively, aftera 72-h incubation period. Apoptosis assays showed DNA fragmentation in promastigotes treated with SeNPs. In an animal challenge, prophylactic doses of biogenic Se NPs delayed the development of localizedcutaneous lesions. Moreover, daily administration of Se NPs (5 or 10 mg kg−1day−1) in similarly infectedBALB/c mice that had not received prophylactic doses of Se NPs also abolished the localized lesions after14 days. Conclusion: Based on these in vitro and in vivo studies, biogenic Se NPs can be considered as anovel therapeutic agent for treatment of the localized lesions typical of cutaneous leishmaniasis

Green synthesis of gold nanoparticles by the marine microalga Tetraselmis suecica

The application of green-synthesis principles is one of the most impressive research fields for the production of nanoparticles. Different kinds of biological systems have been used for this purpose. In the present study, AuNPs (gold nanoparticles) were prepared within a short time period using a fresh cell extract of the marine microalga Tetraselmis suecica as a reducing agent of HAuCl4 (chloroauric acid) solution. The UV–visible spectrum of the aqueous medium containing AuNPs indicated a peak at 530 nm, corresponding to the surface plasmon absorbance of AuNPs. The X-ray diffraction pattern also showed a Bragg reflection related to AuNPs. Fourier-transform infrared spectroscopy was performed for analysis of surface functional groups of AuNPs. Transmission electron microscopy and particle-size-distribution patterns determined by the laser-light-scattering method confirmed the formation of well-dispersed AuNPs. The most frequent size of particles was 79 nm

The removal of ρ-chlorophenol in aqueous cultures with free and alginate-immobilized cells of the microalga Tetraselmis suecica

The present study aimed at evaluating the ability of some isolated cyanobacterial and microalgal strains for the removal of ρ-chlorophenol (ρ-CP), an environmentally harmful contaminant. To identify the most efficient species, a screening program was carried out using 15 microalgal and cyanobacterial strains. Among them, Tetraselmis suecica was able to remove 67 % of the ρ-chlorophenol at an initial concentration of 20 mg L−1 from the medium within a 10-day period. The efficacy of the process was dependent on the ρ-chlorophenol concentration. At concentrations above 60 mg L−1 of the pollutant, no removal was observed due to the inhibitory effect of ρ-chlorophenol on the T. suecica cells. The effect of cell immobilization in alginate beads on T. suecica removal capacity was also examined. Using this technique, the removal efficacy for the initial ρ-CP concentration of 20 mg L−1 increased up to 94 %