Arsenite tolerance and removal potential of the indigenous halophilic bacterium, Halomonas elongata SEK2.

The indigenous halophilic arsenite-resistant bacterium Halomonas elongata strain SEK2 isolated from the high saline soil of Malek Mohammad hole, Lut Desert, Iran, could tolerate high concentrations of arsenate (As5+) and arsenite (As3+) up to 800 and 40 mM in the SW-10 agar medium, respectively. The isolated strain was able to tolerate considerable concentrations of other toxic heavy metals and oxyanions, including Cadmium (Cd2+), Chromate (Cr6+), lead (Pb2+), and selenite (Se4+), regarding the high salinity of the culture media (with a total salt concentration of 10% (w/v)), the tolerance potential of the isolate SEK2 was unprecedented. The bioremoval potential of the isolate SEK2 was examined through the Silver diethyldithiocarbamate (SDDC) method and demonstrated that the strain SEK2 could remove 60% of arsenite from arsenite-containing growth medium after 48 h of incubation without converting it to arsenate. The arsenite adsorption or uptake by the halophilic bacterium was investigated and substantiated through Fourier-transform infrared spectroscopy (FTIR), Scanning Electron Microscope (SEM), and Energy Dispersive X-ray (EDX) analyses. Furthermore, Transmission electron microscope (TEM) analysis revealed ultra-structural alterations in the presence of arsenite that could be attributed to intracellular accumulation of arsenite by the bacterial cell. Genome sequencing analysis revealed the presence of arsenite resistance as well as other heavy metals/oxyanion resistance genes in the genome of this bacterial strain. Therefore, Halomonas elongata strain SEK2 was identified as an arsenite-resistant halophilic bacterium for the first time that could be used for arsenite bioremediation in saline arsenite-polluted environments.

Characterization of gut microbiota profile in Iranian patients with bipolar disorder compared to healthy controls.

Introduction: The human gut microbiota plays a crucial role in mental health through the gut-brain axis, impacting central nervous system functions, behavior, mood, and anxiety. Consequently, it is implicated in the development of neuropsychiatric disorders. This study aimed to assess and compare the gut microbiota profiles and populations of individuals with bipolar disorder and healthy individuals in Iran. Methods: Fecal samples were collected from 60 participants, including 30 bipolar patients (BPs) and 30 healthy controls (HCs), following rigorous entry criteria. Real-time quantitative PCR was utilized to evaluate the abundance of 10 bacterial genera/species and five bacterial phyla. Results: Notably, Actinobacteria and Lactobacillus exhibited the greatest fold change in BPs compared to HCs at the phylum and genus level, respectively, among the bacteria with significant population differences. Ruminococcus emerged as the most abundant genus in both groups, while Proteobacteria and Bacteroidetes showed the highest abundance in BPs and HCs, respectively, at the phylum level. Importantly, our investigation revealed a lower Firmicutes/Bacteroidetes ratio, potentially serving as a health indicator, in HCs compared to BPs. Conclusion: This study marks the first examination of an Iranian population and provides compelling evidence of significant differences in gut microbiota composition between BPs and HCs, suggesting a potential link between brain functions and the gut microbial profile and population.

Toxic heavy metal/oxyanion tolerance in haloarchaea from some saline and hypersaline ecosystems.

Toxic heavy metal/oxyanion contamination has increased severely through the last decades. In this study, 169 native haloarchaeal strains were isolated from different saline and hypersaline econiches of Iran. After providing pure culture and performing morphological, physiological, and biochemical tests, haloarchaea resistance toward arsenate, selenite, chromate, cadmium, zinc, lead, copper, and mercury were surveyed using an agar dilution method. On the basis of minimum inhibitory concentrations (MICs), the least toxicities were found with selenite and arsenate, while the haloarchaeal strains revealed the highest sensitivity for mercury. On the other hand, the majority of haloarchaeal strains exhibited similar responses to chromate and zinc, whereas the resistance level of the isolates to lead, cadmium, and copper was very heterogeneous. 16 S ribosomal RNA (rRNA) gene sequence analysis revealed that most haloarchaeal strains belong to the Halorubrum and Natrinema genera. The obtained results from this study showed that among the identified isolates, Halococcus morrhuae strain 498 had an exceptional resistance toward selenite and cadmium (64 and 16 mM, respectively). Also, Halovarius luteus strain DA5 exhibited a remarkable tolerance against copper (32 mM). Moreover, strain Salt5, identified as Haloarcula sp., was the only strain that could tolerate all eight tested heavy metals/oxyanions and had a significant tolerance of mercury (1.5 mM).

Isolation, Screening and Identification of Native and New Bacillus subtilis with Strong Antifungal Compound against Fusarium oxysporum.

The genus Fusarium causes a wide range of infections in human, animals and herbs. The purpose of this research was to investigate and identify the native strains of Bacillus subtilis playing an inhibitory role against Fusarium oxysporum by producing surfactin. B. subtilis was isolated from the soil of various parks in Tehran-Iran, and identified by biochemical tests. Growth inhibition zone, minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of B. subtilis were determined. After purification of surfactin, quantitative and qualitative analysis of surfactin conducted using high performance liquid chromatography (HPLC) . Finally, two selected native strains with the highest production rate of surfactin identified using PCR for 16S rRNA and phylogenetic tree was drawn. Sixty strains of B. subtilis were isolated from soil, after identification through phenotypical and biochemical tests, the antagonistic activity of 27 different strains against F. oxysporum by Agar well diffusion assay determined and the highest inhibition zone was 13.66 mm. Six strains showing the best inhibitory effect, were isolated and their metabolite were purified by methanol. MIC and MFC values of different strains were in the range of 0.5-1.6 and 1.6-2.6 mg/mL. Using HPLC, the purified surfactin content in B. subtilis was about 56.7 - 131.9 µg/mL. Based on the curves of the chromatogram, the preferred strains with the highest production of surfactin, by molecular identification, displayed high similarity to B. subtilis. We got a maximum amount of yellow and transparent surfactin from native strains. Furthermore, the selected bacteria can be good candidates for biological control of fungal pathogens.

Recombinant PBP2a/autolysin conjugate as PLGA-based nanovaccine induced humoral responses with opsonophagocytosis activity, and protection versus methicillin-resistant Staphylococcus aureus infection.

Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) reasons extreme infections, can resist various conventional antimicrobial agents, and cause morbidity and mortality worldwide. Vaccination seems to help modulate MRSA infections. Nanovaccine is considered a novel strategy in vaccine technology. The primary purpose of the present study was to develop a conjugate vaccine based on recombinant PBP2a and MRSA autolysin formulated in PLGA as a nanoparticle capable of enhancing protective responses against MRSA in the murine model. Materials and Methods: Recombinant PBP2a and autolysin have been expressed and purified by nickel-nitrilotriacetic acid (Ni-NTA) affinity column and characterized by SDS-PAGE and western blot. PLGA was bound to recombinant proteins by using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDAC) and adipic acid dihydrazide (ADH) as a linker and spacer, respectively. Conjugation of recombinant proteins to PLGA was confirmed by the AFM assay, zeta potential, and size distribution, and its efficacy was evaluated in mice. Total IgG, IgG1, IgG2a, IgG2b, and IgM titers were analyzed to assess immune responses. Lastly, the bioactivity of antibodies was tested by using the opsonophagocytosis assay. Results: Mice immunized with the r-PBP2a-r-autolysin-PLGA nanovaccine led to increased levels of opsonic antibodies and IgG1, IgG2a, IgG2b, and IgM when compared with other experimental groups. Our results confirmed that vaccination with nanovaccine could reduce the mortality rate against the sub-lethal dose of MRSA challenge. Furthermore, the nanovaccine could eliminate MRSA from the kidney of infected mice. Conclusion: This study may provide valuable insights into the protective power of the r-PBP2a-r-autolysin-PLGA conjugate vaccine against MRSA infection.

In silico prediction of B and T cell epitopes based on NDV fusion protein for vaccine development against Newcastle disease virus.

Newcastle disease (ND) is known as the most common diseases of economic importance worldwide. Vaccination against virulent strains of Newcastle disease virus (NDV) has failed during some outbreaks. Here, we aimed to assess the epitopes of NDV fusion protein as targets for a peptide-based vaccine. To explore the most antigenic epitopes on the F protein, we retrieved virulent strains of genotype VII from National Center for Biotechnology Information (NCBI). Linear and conformational B-cell epitopes were identified. Moreover, T-cell epitopes with high and moderate binding affinities to human major histocompatibility complex (MHC) class I and class II alleles were predicted using bioinformatics tools. Subsequently, the overlapped epitopes of B-cell and MHC class I and MHC class II were determined. To validate our predictions, the best epitopes were docked, to chicken MHC class I (B-F) alleles using the HADDOCK flexible docking server. Seven 'high ranked epitopes' were identified. Among them, 'LYCTRIVTF' and 'MRATYLETL' showed the highest scores. The other five epitopes including LSGEFDATY, LTTPPYMALK, LYLTELTTV, DCIKITQQV and SIAATNEAV obtained very encouraging results as well. SIAATNEAV had been recognized as a neutralizing epitope of F protein using monoclonal antibodies before. Taken together, our results demonstrated that the identified epitopes needed to be tested by in vitro and in vivo experiments.

Development of Escherichia coli asparaginase II for the Treatment of Acute Lymphocytic Leukemia: In Silico Reduction of asparaginase II Side Effects by a Novel Mutant (V27F).

Acute lymphoblastic leukemia (ALL) is a common blood disease in children that is accountable for many deaths. Due to major improvements in treatment procedures in the past 50 years, the survivability of this disease has risen dramatically to about 90 percent today. L-asparaginase (ASNase) has been used to treat ALL. The glutaminase (GLNase) activity of this enzyme causes some side effects and is unnecessary for anticancer activity. This study investigated mutagenesis in Escherichia coli ASNase II to find a mutant with lower GLNase activity via molecular dynamics (MD) simulation. Residues with low binding energy to asparagine (Asn) and high binding energy to glutamine (Gln) were chosen for mutagenesis. A mutant with low free binding energy to Gln was then selected for molecular docking and MD studies. The results showed that V27F is a good candidate for reducing GLNase activity and that it has little effect on enzyme ASNase activity. A simulation analysis showed that the V27F mutant was more stable than the WT ASNase and that mutagenesis was quite successful.

Bio-removal of Heavy Metalsusing Iron-oxidizing Bacteria: A Novel Approach in Environmental Biotechnology.

The pharmaceutical and hygienic productivity of wastewater containing pollutants, especially heavy metals such as nickel, andmercury are brought into the nature. Recently, bio-removal of heavy metals has attracted significant attention as an eco-friendly approach for the research departments of the pharmaceutical companies. In the current study, removal of heavy metals including mercury and nickel was assessed using isolatediron-oxidizing bacteria from different sources. To this end, bacterial populations were isolated from a variety of aquatic ecosystems; including Mahallat Pond, mountainous rivers, iron industry wastewater, and treated industrial wastewater. The bacteria were cultured and purified in iron-oxidizing media after which the removal of mercury and nickel was measured through culturing the isolated bacteria in 3 different media of Luria-Bertani, PHGII, and iron-oxidizing media containing the heavy metals (2 ppm). The results proved LB as a suitable medium for all the isolated bacteria in removing the heavy metals.It was shown that approximately 100% of the mercury was removed through the bacterial cultured in LB medium. The removal of nickel also reached its maximum of 30% by bacterial culture in LB medium. Then, the phylogenetic study according to 16S rDNA gene sequences showed thatthe isolated bacteria from iron industry wastewater was Bacillus velezensis CR-502 (T).In summary, this study demonstrated the impressive ability of these bacteria for mercury removal and theeffects of different mediaon the removal of mercury and nickel.

The effect of Faecalibacterium prausnitzii and its extracellular vesicles on the permeability of intestinal epithelial cells and expression of PPARs and ANGPTL4 in the Caco-2 cell culture model.

BACKGROUND AND OBJECTIVES: Gut microbiota such as Faecalibacterium prausnitzii play a major role in the regulation of gut barrier, inflammation and metabolic functions. Microbiota-derived extracellular vehicles (EVs) have been recently introduced as functional units mediating the eukaryotic and prokaryotic cell-microbiota interactions. In this paper, the effect of F. prausnitzii and its EVs on mRNA expression levels of tight junction genes (ZO1 and OCLN) as well as PPARs and ANGPTL4 genes in the human epithelial colorectal adenocarcinoma (Caco-2) cell line was evaluated. METHODS: F. prausnitzii was cultured on the Brain Heart Infusion (BHI) broth medium under anaerobic conditions, and its EVs were extracted by ultracentrifugation. This bacterium and its EVs were treated on the Caco-2 cells. After 24 h, the expression of the genes encoding TJ proteins such as ZO1 and OCLN, PPARs and ANGPTL4 was evaluated by quantitative real-time PCR. RESULTS: Unlike F. prausnitzii, its EVs significantly increased the expression of ZO1 and OCLN genes, and PPARα, PPARγ and PPARß/δ genes (except at a concentration of 100 µg/ml) as well as ANGPTL4 gene. CONCLUSIONS: The results of this study demonstrated that F. prausnitzii-derived EVs increased the intestinal barrier permeability via TJs (ZO1 and OCLN) as well as PPAR-α, PPAR-γ and PPAR ß/δ genes and their targeted gene (ANGPTL4) in the Caco-2 cell line. Accordingly, it is suggested that F. prausnitzii-derived EVs can be considered as a new bacterial postbiotic to cure dysbiosis-associated diseases including obesity and its related metabolic dysfunctions, according to the leaky gut hypothesis.

La amplia distribución de un microorganismo termoacidófilo en el mineral de una mina cuprífera a temperatura ambiente y en condición ácida, y su importancia en la biolixiviación del concentrado de calcopirita / The wide distribution of an extremely thermoacidophilic microorganism in the copper mine at ambient temperature and under acidic condition and its significance in bioleaching of a chalcopyrite concentrate

Thermoacidophiles can exist in a state of dormancy both in moderate temperatures and even in cold conditions in heap leaching. Sulphide mineral ores such as chalcopyrite produce sulfuric acid when exposed to the air and water. The produced sulfuric acid leads to the decrease of pH and exothermic reactions in heap leaching causing the temperature to increase up to 55 °C and the activation of thermoacidophilic microorganisms. The aim of the present study was to isolate indigenous extreme thermoacidophilic microorganisms at ambient temperature from Sarcheshmeh Copper Complex, to adapt them to the high pulp density of a chalcopyrite concentrate, and to determine their efficiency in chalcopyrite bioleaching in order to recover copper. In this study samples were collected at ambient temperature from Sarcheshmeh Copper Complex in Iran. Mixed samples were inoculated into the culture medium for enrichment of the microorganisms. Pure cultures from these enrichments were obtained by subculture of liquid culture to solid media. Morphological observation was performed under the scanning electron microscope. Isolates were adapted to 30% (w/v) pulp density. For the bioleaching test, the experiments were designed with DX7 software. Bioleaching experiments were carried out in Erlenmeyer flasks and a stirred tank reactor. The highest copper recovery in Erlenmeyer flasks was 39.46% with pulp 15%, inoculums 20%, size particle 90 pm and 160 rpm. The lowest recovery was 3.81% with pulp 20%, inoculums 20%, size particle 40 pm and 140 rpm after 28 days. In the reactor, copper recovery was 32.38%. Bioleaching residues were analyzed by the X-ray diffraction (XRD) method. The results showed no jarosite (KFe3(SO4)2(OH)6) had formed in the bioleaching experiments. It seems that the antagonistic reactions among various species and a great number of planktonic cells in Erlenmeyer flasks and the stirred tank reactor are the reasons for the low recovery of copper in our study.

Los microorganismos termoacidófilos pueden estar en estado latente tanto a temperatura moderada como baja, en lixiviación en pilas. Los minerales sulfurosos, como la calcopirita, producen ácido sulfúrico cuando se exponen al aire y al agua. El ácido sulfúrico producido conduce a la disminución del pH y a reacciones exotérmicas durante la lixiviación en pilas, lo que hace que la temperatura aumente hasta 55 °C y se activen los microorganismos termoacidófilos. El objetivo del presente estudio fue aislar del complejo de cobre Sarchesh-meh (Irán) microorganismos termoacidófilos extremos que proliferan a temperatura ambiente e investigar su adaptación a la alta densidad de pulpa del concentrado de calcopirita, así como su eficiencia para biolixiviarese mineral, con el objeto de recuperar el cobre. Se recogieron muestras a temperatura ambiente del citado complejo, y luego muestras mixtas se inocularon en un medio de cultivo de enriquecimiento. A partir de estos enriquecimientos, mediante el subcultivo del cultivo líquido a medio sólido, se obtuvieron cultivos puros. La observación morfológica se realizó bajo microscopio electrónico de barrido. Los aislados estaban adaptados al 30% p/v de densidad de pulpa. Para la prueba de biolixiviación, los experimentos fueron diseñados con el software DX7. Los experimentos de biolixiviación se llevaron a cabo en Erlenmeyers y en un reactor tanque con agitación. La mayor recuperación de cobre en los Erlenmeyers fue del 39,46% y se obtuvo con la pulpa al 15%, un inóculo del 20%, un tamaño de partícula de 90 µm y una agitación de 160 rpm. La menor recuperación fue del 3,81% y se obtuvo con la pulpa al 20%, un inóculo del 20%, un tamaño de partícula de 40 µm y una agitación de 140 rpm, después 28 días. En el reactor, la recuperación del cobre fue del 32,38%. El análisis de difracción de rayos X (XRD) no mostró que se formara jarosita (KFe3-#91;SO4-#93;2-#91;OH-#93;6) en los experimentos de biolixiviación. Dicha técnica sirve para determinar la estructura cristalina de una sustancia desconocida. Al parecer, las reacciones antagónicas entre las diversas especies y el mayor número de células planctónicas en los Erlenmeyers y en el reactor fueron las causas de la baja recuperación de cobre observada en este estudio.

The wide distribution of an extremely thermoacidophilic microorganism in the copper mine at ambient temperature and under acidic condition and its significance in bioleaching of a chalcopyrite concentrate.

Thermoacidophiles can exist in a state of dormancy both in moderate temperatures and even in cold conditions in heap leaching. Sulphide mineral ores such as chalcopyrite produce sulfuric acid when exposed to the air and water. The produced sulfuric acid leads to the decrease of pH and exothermic reactions in heap leaching causing the temperature to increase up to 55°C and the activation of thermoacidophilic microorganisms. The aim of the present study was to isolate indigenous extreme thermoacidophilic microorganisms at ambient temperature from Sarcheshmeh Copper Complex, to adapt them to the high pulp density of a chalcopyrite concentrate, and to determine their efficiency in chalcopyrite bioleaching in order to recover copper. In this study samples were collected at ambient temperature from Sarcheshmeh Copper Complex in Iran. Mixed samples were inoculated into the culture medium for enrichment of the microorganisms. Pure cultures from these enrichments were obtained by subculture of liquid culture to solid media. Morphological observation was performed under the scanning electron microscope. Isolates were adapted to 30% (w/v) pulp density. For the bioleaching test, the experiments were designed with DX7 software. Bioleaching experiments were carried out in Erlenmeyer flasks and a stirred tank reactor. The highest copper recovery in Erlenmeyer flasks was 39.46% with pulp 15%, inoculums 20%, size particle 90µm and 160rpm. The lowest recovery was 3.81% with pulp 20%, inoculums 20%, size particle 40µm and 140rpm after 28 days. In the reactor, copper recovery was 32.38%. Bioleaching residues were analyzed by the X-ray diffraction (XRD) method. The results showed no jarosite (KFe3(SO4)2(OH)6) had formed in the bioleaching experiments. It seems that the antagonistic reactions among various species and a great number of planktonic cells in Erlenmeyer flasks and the stirred tank reactor are the reasons for the low recovery of copper in our study.

Applying simple linear combination, multiple logistic and factor analysis methods for candidate fecal bacteria as novel biomarkers for early detection of adenomatous polyps and colon cancer.

BACKGROUND: Colorectal cancer (CRC) is the third leading cause of cancer, and presents a considerable disease burden, worldwide. Recently, the gut microbiota has been proposed as a potential risk factor for CRC, and even adenomatous polyps (AP). Here, the aim of this study was to investigate the role of selected gut bacteria as fecal bacterial biomarkers, in early detection of CRC and AP. MATERIAL AND METHODS: Fecal samples (n = 93) were collected from Taleghani Hospital, Tehran, Iran, between 2015 and 2017, from normal controls (NC), AP cases and CRC stage I patients, who were undergoing screening for colonoscopy. Absolute quantitative real time PCR (qPCR) assays were established for the quantification of bacterial marker candidates, in all cases and control groups. In order to evaluate the diagnostic value of bacterial candidates in distinguishing CRC from a polyp, receiver operating characteristic curve (ROC) was performed. Multiple logistic regressions were used to find the best combinations of the bacterial candidates, then, combinations were analyzed based on three methods, including linear combination, multiple logistic and factor analysis models. RESULTS: According to the logistic model, combination of Fusobacterium nucleatum, Enterococcus feacalis, Streptococcus bovis, Enterotoxigenic Bacteroides fragilis (ETBF) and Porphyromonas spp. showed improved diagnostic performance, compared to each bacterium alone, as area under the receiver operating characteristic (AUROC) increases to 0.97, with 95% confidence interval. It was found that a simple linear combination was an appropriate model for discriminating AP and CRC cases, compared to the NC, with a sensitivity of 91.4% and specificity of 93.5%. CONCLUSION: Our results indicated that based on fecal bacterial candidates, statistical simple linear combination model and ROC curve analysis, early detection of AP and CRC might be possible.

The association between fecal microbiota and different types of colorectal polyp as precursors of colorectal cancer.

BACKGROUND: Colon cancer (CRC) is a heterogeneous disorder, arising from precursors-adenoma and serrated polyp. Previous studies have demonstrated the relationship between the human gut microbiota and CRC; however, its correlation to the different early precursors of CRC is not properly understood. Here, we studied the relationship between targeted gut bacteria and different colorectal polyp types, location, size and grade of dysplasia. MATERIAL AND METHODS: In the present case-control descriptive study, selected fecal bacteria were assessed in 118 patients, referred for standard screening colonoscopy, including 31 normal controls, 21 hyperplastic polyp (HP), 16 sessile serrated polyp (SSA), 29 tubular adenoma (TA) and 21 villous/tubuvillous polyp (VP/TVP) cases, between 2015 and 2017, by absolute quantitative real time PCR technique (q PCR) in different ethnicity of Iranian population. The panel of bacteria was including Streptococcus bovis/gallolyticus, Enterococcus faecalis, Enterotoxigenic Bacteroides fragilis (ETBF), Fusobacterium nucleatum, Porphyromonas spp., Lactobacillus spp., Roseburia spp., and Bifidobacterium spp. RESULTS: Higher numbers of F. nucleatum, E. feacalis, S. bovis, ETBF and Porphyromonas spp. were detected in AP cases, consisting TA and especially VP/TVP, in contrast to samples from the normal, HP and SSA groups (P < 0.001). On the contrary, lower number of Lactobacillus spp., Roseburia spp. and Bifidobacterium spp. were detected in AP, compared to the normal, HP and SSA. Surprisingly, a significant correlation was found among selected gut bacterial quantity, the size, location and grade of dysplasia of polyp cases. DISCUSSION: These findings suggest that gut bacteria might contribute in early stages of colorectal carcinogenesis through the development of AP, but not SSA. In fact, AP and SSA are also different in terms of molecular pathways and tendencies to present in specific colorectal location. Overall, these findings may lead to development of CRC prevention therapies, targeting early protagonist bacteria of colorectal carcinogenesis from AP.

Distribution of non-tuberculosis mycobacteria strains from suspected tuberculosis patients by heat shock protein 65 PCR-RFLP.

The genus Mycobacterium contains more than 150 species. Non-tuberculosis mycobacteria (NTM) often cause extrapulmonary and pulmonary disease. Mycobacteria detection at species level is necessary and provides useful information on epidemiology and facilitates successful treatment of patients. This retrospective study aimed to determine the incidence of the NTM isolates and Mycobacterium tuberculosis (Mtb) in clinical specimens collected from Iranian patients during February 2011-December 2013, by PCR-restriction fragment length polymorphism analysis (PRA) of the hsp65 gene. We applied conventional biochemical test and hsp65-PRA identification assay to identify species of mycobacteria in specimens from patients suspected of having mycobacterial isolates. This method was a sensitive, specific and effective assay for detecting mycobacterial species and had a 100% sensitivity and specificity for Mtb and Mycobacterium avium complex (MAC) species. Using PRA for 380 mycobacterial selected isolates, including 317 Mtb, four Mycobacterium bovis and of the 59 clinical isolates, the most commonly identified organism was Mycobacterium kansasii (35.6%), followed by Mycobacterium simiae (16.9%), Mycobacterium gordonae (16.9%), Mycobacterium fortuitum (5.1%), Mycobacterium intracellulare (5.1%), Mycobacterium avium (5.1%), Mycobacterium scrofulaceum (3.4%), Mycobacterium gastri (3.4%), Mycobacterium flavescens (3.4%), Mycobacterium chelonae (3.4%) and Mycobacterium nonchromogenicum (1.7%). PRA method, in comparison with classical methods, is rapid, useful and sensitive for the phylogenetic analysis and species detection of mycobacterial strains. Mycobacterium kansasii is the most common cause of infection by NTM in patients with non-HIV and HIV which demonstrated a high outbreak and diversity of NTM strains in our laboratory.

Different Approaches to Detect "Nanobacteria" in Patients with Kidney Stones: an Infectious Cause or a Subset of Life?

PURPOSE: This research focused on the detection of nanobacteria in kidney stones of 30 Iranian patients without adding fetal bovine serum (FBS) to the culture media. MATERIALS AND METHODS: Nanobacteria were isolated from a nephro-ureterolithiasis extract of the urinary tract and kidney of patients and were cultured in the laboratory. The growth of nanobacteria was monitored using a spectrophotometer, and with inverted microscopy technique, their crystallization was analyzed after two days. The images from atomic force microscopy (AFM), transmission electron microscopy (TEM) and scanning electron microscopy (SEM) indicated the morphology and demonstrated the size of the cultured nanobacteria which is between 60 and 160 nm. Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) were used to study the chemical composition, surface functional groups and crystal structure of the igloo-like nanobacteria shell. FTIR spectra in theregion of 1000 to 1200 cm-1 and the XRD peaks provided evidence that the main components of the nanobacteria shell were apatite-based compounds. RESULTS: Nanobacteria infected all the 27 patients with apatite kidney stone, and none of the three patients who had uric acid kidney stone were infected as confirmed by the cultivation of the stones samples. The results showed that nanobacteria might play a fundamental role in the formation of apatite-based kidney stones. CONCLUSION: The biomineralization ability of nanobacteria may lead to calcification of the soft tissues, which in turn may result in other diseases. It is also suggested that nanobacteria may be a factor in calcification-related diseases and disorders with poorly characterized etiologies. This research with its different approaches, clarified significant doubts that nanobacteria act as contaminant, warranting continued investigation of its role in other diseases.

Recombinant PBP2a as a vaccine candidate against methicillin-resistant Staphylococcus aureus: Immunogenicity and protectivity.

Methicillin-resistant Staphylococcus aureus infections are focal and development of an effective vaccine can help to control this infection. Here, recombinant PBP2a was studied in mouse model. Following the preparation of recombinant PBP2a, Balb/c mice were injected subcutaneously with 20 µg of r-PBP2a formulated in Freund's adjuvant three times with three weeks intervals with proper control group. Total and specific isotype antibodies were evaluated on sera by ELISA. Opsonophagocytic activity was also investigated on the sera samples. Intraperitonealchallenge with a sub-lethal dose of MRSA (5 × 108 CFU) was done in experimental mice. Following that, the number of bacteria from kidneys of experimental mice were determined. Survival rate was recorded for 60 days. Significant increase of antibody with high level of IgG1, IgG2a and IgG2b isotypes was demonstrated in vaccinated mice versus the control group (P < 0.005). The bacterial load in the kidneys from immunized mice was 1000 times less thancontrol group (PBS) and opsonophagocytic activity of immunized mice sera significantly increased (P < 0.0001). Finally the life span of immunized mice after bacterial challenge was extended versus control mice. These results may indicate the capacity of PBP2a as a candidate vaccine to control the MRSA infections.

DBT desulfurization by decorating Rhodococcus erythropolis IGTS8 using magnetic Fe3O4 nanoparticles in a bioreactor.

Today, crude oil is an important source of energy and environmental contamination due to the continued use of petroleum products is a matter or urgent concern. In this work, two technological platforms, namely, the use of a robust desulfurizing bacteria and the use of nanotechnology to decorate the surface of the bacteria with nanoparticles (NP), were combined to enhance biodesulfurization (BDS). BDS is an ecologically friendly method for desulfurizing petroleum products while avoiding damage to the hydrocarbons due to the high temperatures normally associated with physical desulfurization methods. First, a bacterium known to be a good organism for desulfurization (Rhodococcus erythropolis IGTS8) was employed in cell culture to remove a recalcitrant sulfur molecule from a common sulfur-containing compound found in crude petroleum products (dibenzothiophene). 2-Hydroxybiphenyl (2-HBP) produced as a consequence of the BDS of dibenzothiophene was determined using Gibbs' assay. The synthesized NP were characterized by field emission scanning electron microscope, transmission electron microscopy, Fourier transform infrared spectroscopy, X-ray diffraction spectroscopy, and vibrating sample magnetometer. The field emission scanning electron microscope and transmission electron microscopy images showed the size of the NP is 7-8 nm. The decorated cells had a long lag phase, but the growth continued until 148 h (at OD600 = 3.408) while the noncoated bacteria grow until 96 h before entering the stationary phase at OD600 = 2.547. Gibbs' assay results showed that production of 2-HBP by decorated cells was 0.210 mM at t = 148 h, while 2-HBP production by nondecorated cells was 0.182 mM at t = 96 h. Finally, the experiments were repeated in a fermenter.

Evaluation of Ciprofloxacin (gyrA, parC Genes) and Tetracycline (tetB Gene) Resistance in Nosocomial Acinetobacter baumannii Infections.

BACKGROUND: Acinetobacter baumannii plays an important role in some types of nosocomial infections as an opportunist microorganism which increases levels of resistance to antibacterial drugs and disinfectants. OBJECTIVES: The aim of this study was to determine the resistance and sensitivity of A. baumannii to different antibiotics and evaluate the minimal inhibitory concentration (MIC) for Ciprofloxacin and Tetracycline; in addition to Surfanios, Citron and Aniosyme DD1 disinfectants, and also to detect the presence of gyrA, parC and tetB gene bands in the isolates. MATERIALS AND METHODS: In this study, 65 A. baumannii isolates were collected from the hospitalized patients in NIOC hospital (National Iranian Oil Company hospital) of Tehran, Iran during 2010-2011. The pattern of sensitivity to antibiotics was determined using CSLI disk diffusion and MIC methods. Furthermore, resistance of isolates to the common disinfectants (Surfanios Citron and Aniosyme DD1) was determined in different hospital wards. Presence of gyrA, parC and tetB gene bands was also detected by PCR method. RESULTS: Frequency of Acinetobacter resistance to Amikacin, Ciprofloxacin, co-Trimoxazole, Ceftazidime and Ceftriaxone was 100% in the isolates reviewed in this study. The frequency of resistance to Gentamicin and Tetracycline were 86.1% in the isolates. The MIC of Ciprofloxacin in all (100%) of isolates was 32-64 µg/mL which showed the resistance to Ciprofloxacin In 86.1% of cases the Gentamicin and Tetracycline MIC were ≥ 16 µg/mL and in 13.9% of isolates the Gentamicin and Tetracycline MIC were 4 µg/mL, these results showed the resistance and sensitivity to the Gentamicin and Tetracycline, respectively. Additionally, all (100%) of the A. baumannii isolates were resistant to disinfectant concentrations, which were used with the methods recommended by manufacturers (0.5%). In 100% of the isolates parC and gyrA genes bands were detected, and tetB gene was also detected in 86.1% of Tetracycline resistant isolates. CONCLUSIONS: Due to the high resistance of A. baumannii isolates to most antibiotics in our study and also its high resistance to the common disinfectants usually used in hospitals, it seems that more attentions should be paid for applying disinfectants. Since most of the isolates were collected from tracheal and sputum samples (46%), it seems that respiratory tract is the most t prevalent site of infection among Acinetobacter infections. Therefore, disinfecting the respiratory tract related equipment and instruments by using proper disinfectants seems to be an appropriate way to prevent these infections.

Pyocyanine Biosynthetic Genes in Clinical and Environmental Isolates of Pseudomonas aeruginosa and Detection of Pyocyanine's Antimicrobial Effects with or without Colloidal Silver Nanoparticles.

OBJECTIVE: Pyocyanine plays an important role in the pathogenesis of Pseudomonas aeruginosa, (P. aeruginosa) and is known to have inhibitory and bactericidal effects. This study has aimed to detect the phenazine biosynthetic operon (phz ABCDEFG) and two phenazine modifying genes (phzM and phzS) by polymerase chain reaction (PCR) and detection of its possible protein bands by sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE). The antimicrobial effects of pyocyanine alone and mixed with colloidal silver nanoparticles were studied. MATERIALS AND METHODS: In this descriptive study, clinical and environmental species of P. aeruginosa were isolated by thioglycollate medium culture and cetrimide agar, respectively. The existence of a phenazine biosynthetic operon and two phenazine modifying genes as well as their protein products were confirmed by PCR and SDS-PAGE, respectively. Pyocyanine was extracted with chloroform and its antimicrobial effects against bacteria such as; Escherichia coli (E. coli), P. aeruginosaand Staphylococcus aureus (S. aureus) bacteria and yeast Candida albicans (C. albicans) were tested using well, spot and disk diffusion methods. RESULTS: In this study, 3 out of 48 clinical strains were unable to produce pyocyanine on cetrimide and Mueller Hinton (MH) agar. Two strains did not have phenazine modifying gene bands. Another strain did not have the possible protein band of the phzM gene. Pyocyanine had antimicrobial effects against the microbial strains, which increased in the presence of silver nanoparticles. CONCLUSION: According to the results of the present study, some P. aeruginosa strains are unable to produce pyocyanine due to the absence of the phzM or phzS genes. Therefore, these genes have an important role in pyocyanine production in P. aeruginosa. Pyocyanine shows synergistic antimicrobial effects in the presence of silver nanoparticles against microbial strains.