Bacillus Subtilis Var Niger Research Articles

Effects of CeO2 Nanomaterial on Pseudomonas Aeruginosa Mediated Bronchiectasis Inflammation Response

There are many researches on the preparation of rare earth metals (REM) materials and bactericidal materials, but there are few researches on the preparation of REM oxides based on the laser evaporation (LE) method in the field of bactericidal. In this research, ZnO–CeO2 nanoparticles (NPs) were obtained by the LE method with 100 W single-mode CW CO2 laser as the light source and cerous carbonate and zinc acetate as the target materials. While physical characterization of the ZnO–CeO2 NPs, Staphylococcus aureus (SA), Bacillus subtilis var niger (BSVN), and pseudomonas aeruginosa (PAE) were undertaken as test objects to verify the bactericidal performance of the nanomaterial. A PAE mediated inflammation model of bronchiectasis rats was established, and 40 SD rats were screened and rolled into 4 groups: a control (Ctrl) group, a sham group, a model group without nano solution injection (Model group), and a model group injected with nano solution (Model +NPs group). The effects of nanomaterials on inflammation were analyzed. The results suggested that ZnO–CeO2 NPs were scattered rod particles with uniform morphological distribution. XRD tests revealed that with the increase of ZnO content, its characteristic peaks in the NPs also increased. The preparation of NPs exhibited good bactericidal performance against SA and BSVN, and increasing the concentration of NPs greatly inhibited the activity of PA. By comparing with the model group, the upregulated IL-17 and IL-6 were inhibited (P <0.01), while the downregulated IL-10 and TGF-β were reversed (P <0.01). It meant that the NPs alleviated the inflammation of bronchiectasis mediated by PA, and thus improved the lung function damaged by inflammation.

Integration of high volume portable aerosol-to-hydrosol sampling and qPCR in monitoring bioaerosols

In this study, the integration of a high volume, portable aerosol-to-hydrosol sampling technique and quantitative polymerase chain reaction (qPCR) was investigated for bioaerosol monitoring by adapting the RCS High Flow to sample air with mineral-oil-strips. Bacillus subtilis var niger and Pseudomonas fluorescens were aerosolized and collected by the RCS High Flow loaded with mineral-oil-strips for 1, 2 and 5 min. In addition, the adapted aerosol-to-hydrosol sampler was also tested for sampling environmental bacterial aerosols in four different environments (a back yard, a student dorm, a dining hall, and a play ground). The performances of the RCS High Flow with mineral-oil-strips were compared with the use of agar strips under similar conditions in all experiments. Air samples collected by the RCS High Flow were cultured, and in addition those collected with mineral-oil-strips were also quantified using qPCR. When sampling B. subtilis var niger aerosols, the use of mineral-oil-strips was shown to report significantly higher culturable concentrations than those obtained by agar strips regardless of the sampling time tested (p-value = 0.04). In contrast, the differences between the two methods when sampling P. fluorescens aerosols were not statistically significant (p-value = 0.5). When coupled with qPCR, the RCS High Flow loaded with mineral-oil-strips obtained significantly higher bacterial aerosol concentrations than those detected by the culturing method. The sampling time was observed to have negligible effects on the efficiency of the technology developed here. When sampling in different environments, the use of mineral-oil-strip was observed to yield significantly higher, about 4-12 times, culturable bacterial aerosol concentration levels compared to the use of agar. This study demonstrated a high volume (100 L min⁻¹) portable aerosol-to-hydrosol sampling technique, holding broad promise in monitoring airborne biological threats when coupled with qPCR technology. Yet, caution should be taken in relating the bioaerosol concentrations to health risks as qPCR detects both culturable and non-culturable cells including inactivated ones.

A novel method for measuring the charge distribution of airborne microbes

A novel method for measuring bioaerosol charge distribution was investigated using electrostatic sampling and quantitative polymerase chain reaction (qPCR). Here, Bacillus subtilis var niger and Pseudomonas fluorescens were aerosolized and precipitated into different regions of two 96-well plates placed inside an electrostatic sampler. The concentrations of negatively charged bacteria in the air samples collected were quantified using qPCR, and the relevant bioaerosol charge levels were calculated using an equation developed in this study. Experimental results showed that B. subtilis var niger bioaerosols had wider charge distribution than P. fluorescens. For B. subtilis var niger, 93% of them carried a charge level of 13–42 units, 2% carried a charge level of 8–13 or lower, and 5% carried a charge level of 42–195 or higher. While for P. fluorescens, 99% of P. fluorescens aerosols carried a charge level of 14–34 elementary units. The indicative charge levels of bioaerosols collected at a certain point in the electrostatic sampling line largely depends on the particle size, bacterial species, electrostatic field strength, and the sampling flow rates for a given electrostatic sampler. In general, the trends for the charge levels here were similar to those obtained in previous studies. Use of qPCR technology in this study can distinguish between biological and non-biological particles collected. Thus, this avoided counting the non-biological particles in traditional bioaerosol charge measurements in which an optical particle counter is used. The developed technique here can be also used to analyze the charge levels for other types of bioaerosols.

Inactivation of bacteria and fungus aerosols using microwave irradiation

This study investigated the survivals of both lab-generated and environmental bioaerosols when exposed to the microwave irradiation (2450 MHz) for ∼2 min at different output powers (700, 385 and 119 W). Control and exposed bioaerosols were collected by a BioSampler or a mixed cellulose ester (MCE) filter, and the air samples were further cultured. As a comparison, liquid-borne exposure for one species was also conducted. Environmental scanning electron microscope (ESEM) and transmission electron microscope (TEM) were used to study the membrane surface morphologies and intracellular components of the microwave-treated and untreated microbes. The survival rates of airborne Bacillus subtilis var niger spores were shown to be about 35%, 44% and 35% when exposed to the microwave irradiation for 1.5 min with high, medium and low power applied, respectively ( p-value=0.37). In contrast, the airborne Pseudomonas fluorescens were shown to have lower survival rates of 5.8%, 12.2% and 21% ( p-value=0.0045). Similar patterns but higher survival rates at respective powers were observed for airborne Aspergillus versicolor exposure ( p-value<.0001). For environmental bacterial and fungal bioaerosols, 30–40% of them were shown to survive the high power microwave irradiation for 1.7 min. Outdoor bioaerosols were shown to have stronger survival than the indoor bioaerosols when exposed to the microwave irradiation. ESEM and TEM images showed visible damages to the microwave-irradiated microbes. The results obtained here can be used to develop microwave-based air sterilization technologies especially targeted for biological aerosols.

Use of carbon nanotube filter in removing bioaerosols

In this study, the carbon nanotube (CNT) filters were created by depositing both single-walled and multi-walled CNT on polypropylene and polyamide membrane supports. The physical efficiencies of the CNT filters with different CNT loadings when removing the aerosolized Bacillus subtilis var niger, Pseudomonas fluorescens and NaCl were studied using an optical particle counter together with a button aerosol sampler. The removal efficiencies of CNT filters prepared on polypropylene membrane supports of 10 μm pore size when collecting B. subtilis var niger aerosols were shown to range from 10% to 95% in average for the CNT loadings of 0.2–1.6 μg/cm 2. When collecting aerosolized P. fluorescens, the efficiencies were shown to range from 5% to 60% given similar CNT loadings used. The results revealed that particle types and CNT loading had greater effects on the efficiencies than the membrane types and pore sizes tested. When collecting aerosolized NaCl, the efficiencies were observed relatively lower, i.e., below 50% for loadings of 0.3–1.5 μg/cm 2. Acid-rinsing was shown to exhibit statistically significant effect on the removal efficiency of single-walled CNT filters when collecting aerosolized NaCl. Multi-walled CNT filters were shown to perform better than single-walled CNT filters. The performances of CNT filters prepared were observed to vary greatly, which was likely due to the CNT loading efficiency, membrane support filtration variance and the CNT uniformity on the support. The information obtained here can be used to develop future indoor air pollution control measures.

A methodological study on testing and evaluating of filtration efficiency of canister against microbial aerosol

To establish a testing and evaluating method for filtration efficiency of the canister against microbial aerosol. Serratia marcescens aerosol served as model of bacterial aerosol, Bacillus subtilis var niger aerosol as model of spores aerosol, bacteriophage f(2) aerosol as model of viral aerosol. Employing the microbial aerosol testing platform was established in lab, models of microbial aerosol generated artificially were sampled quantitatively by air samplers before and after filtrating by canisters, respectively. Filtration efficiency was determined by the concentration of microbial aerosol in the air sample before and after filtrating. The four canisters of 1-1, 1-2, 1-3, 1-4 were tested for the filtration efficiency against Serratia marcescens, Bacillus subtilis var niger and phage f(2) aerosol. The two canisters of 543 and 544 canisters equipped with active carbon were tested for the filtration efficiencies against Serratia marcescens aerosol. The filtration efficiency of 1-1, 1-2, 1-3 canisters against Serratia marcescens, Bacillus subtilis var niger and phage f(2) aerosol was 100.000%. The filtration efficiency of 1-4 canister filtration efficiency against Bacillus subtilis var niger spores aerosol was 99.997% and efficiency of the other two aerosol was 100.000%. The filtration efficiency of the two canisters of 543 and 544 to those attached with active carbon against Serratia marcescens aerosol was 100.000%. The testing method might be used to evaluate the protective performance of the canister against microbiological aerosol. The effect of the canisters (including those equipped with active carbon) against microbiological aerosol should be reliable.

Flow Cytometry Sorting Protocol of Bacillus Spore Using Ultraviolet Laser and Autofluorescence as Main Sorting Criterion

The ultraviolet (UV) Fluorescent Aerodynamic Particle Sizer (FLAPS), a flow cytometer-like apparatus was developed by the Canadian Department of National Defence for real-time detection of autofluorescence of biological aerosol particles such as bacterial spores. The direct relation between autofluorescence intensity and viability has recently been reported and viable spore are more autofluorescent in UV (Laflamme, Frontiers in Bioscience). The goal of this manuscript is to describe a flow cytometry sorting protocol based on UV-induced autofluorescence. An EPICS ELITE ESP flow cytometer equipped with a UV laser and cell sorter was used to mimic the optical properties of FLAPS and to study the two extremes of a spore population according to its autofluorescence (lower level of autofluorescence (LLA) and higher level of autofluorescence (HLA) spores). Bacillus subtilis var niger was used as a surrogate for Bacillus anthracis spores and sorted using autofluorescence intensity as the main criterion. The protocol developed in our laboratory to sort Bacillus spores according to their autofluorescence properties is described. Purity of each sorted population was greater than 95%. Using autofluorescence as the main criterion, we demonstrate that it is possible to separate two distinct spore populations.

In situ decontamination of medical wastes using oxidative agents: a 16-month study in a polyvalent intensive care unit

Over a 16-month period from September 1997 to December 1998, a prospective study was made of an on-site treatment of medical wastes in a 10-bed intensive care unit. First, the wastes were ground and then, a high concentration of ozone in air was repeatedly injected into the ground wastes. The study analysed the practical application of the system and its microbiological efficiency. Inactivation experiments were made with reference strains of Staphylococcus aureus,Enterococcus hirae , Pseudomonas aeruginosa, Escherichia coli, Mycobacterium smegmatis,Bacillus subtilis var niger, Bacillus stearothermophilus, Candida albicans and Aspergillus niger. Two thousand eight hundred treatment cycles, i.e. 84000 grindings and 140000 ozone injections gave a treatment capacity of 50kg of waste per day with a good staff acceptability. All kinds of medical devices used in an intensive care unit were treated. In untreated ground wastes, the median bacterial load was 105.86 (range 102.35–108.05) cfu/g. After ozone treatment, bacteria and fungi were reduced by a factor of 105. Aero-contamination of the ward was unchanged. Computer control allowed all events to be tracked. On-site medical waste treatment appears to be an efficient alternative to the usual centralized collection and treatment.

The effect of drinking water treatment on the spatial heterogeneity of micro-organisms: implications for assessment of treatment efficiency and health risk

The effect of drinking water treatment (ferric coagulation, floc blanket clarification, rapid sand filtration) on the spatial heterogeneity of five species of micro-organism was studied at pilot scale. It was found that the spatial heterogeneity of vegetative bacteria (namely total coliform and heterotrophic (22°C; 3d) bacteria) was little affected by treatment. Indeed, counts of total coliform bacteria within 500l volumes of treated water were Poisson distributed (i.e. showed minimum variation). In contrast, treatment appeared to increase the spatial heterogeneity (or clustering) of both aerobic spores indigenous to the raw water and Bacillus subtilis var niger spores added to the raw water. Furthermore, B. subtilis var niger spores added to the raw water were detected in the treated water 25h after termination of spiking to the raw water. The effect on C. parvum oocysts added to the raw water could not be determined because few oocysts broke through treatment into the treated water. Indeed oocyst removals of 5–6 logs were apparent. “Species-specific” differences in the removal ratios were also demonstrated. It is concluded that audits for treatment processes based on single 100ml “spot” samples for spores will tend to over-estimate the net spore removal and hence underestimate the public health risk. Spatial heterogeneity of counts in treated water contributes to explaining why no “ideal” surrogate has been identified for treatment plant performance.

Measured infrared spectral extinction for aerosolized Bacillus subtilis var niger endospores from 3 to 13 µm

We measured spectral extinction in situ for aerosolized Bacillus subtilis var. niger endospores using Fourier-transform infrared spectroscopy from 3.0 to 13.0 mum. Corresponding aerosol size distributions were measured with a commercially available elastic light-scattering probe and verified by direct particle capture and subsequent counting by video microscopy. Aerosol mass density was monitored simultaneously with conventional dosimetry and was used to mass normalize the measured spectral extinction. Mie theory calculations based on measured distributions and available complex indices of refraction agreed well. We also present resultant Mie calculations for the absorption, total scattering, and backscatter. For comparison, measured spectral extinction for three common environmental aerosols is also presented, i.e., for water fog, diesel soot, and Arizona road dust.

Survival of Bacteria on Respirator Filters

The survival and potential growth of two different kinds of bacteria were investigated on NIOSH-certified polypropylene respirator filters. Bacillus subtilis var niger (B. subtilis) spores represented environmentally resistant bacteria, whereas Pseudomonas fluorescens (P. fluorescens) vegetative cells represented stress-sensitive bacteria. The bacteria were aerosolized and loaded on the respirator filters under three nutritional conditions: water, saliva, and tryptic soy broth (TSB). The loaded filters were incubated for 0-13 days and analyzed for culturable and total bacteria count. The analysis was optimized through the evaluation of three methods for eluting bacteria from respirator filter samples: low frequency shaking, vortexing, and ultrasonic vibrating. Vortexing was found to be the most effective with the highest total and culturable count and was therefore used for the analysis of bacterial survival and growth. Neither of the test bacteria was able to grow on the respirator filters even under optimal nutrition and incubation conditions. It therefore appears that reuse of a polypropylene respirator poses minimal risk of bacterial growth, provided the respirator has been carefully handled and stored. The data on respirator survival show that sensitive P. fluorescens cells lost their viability in less than three days, whereas resistant B. subtilis spores remained viable on the filter for over thirteen days of testing. It appears that resistant bacteria may pose risk of infection to the people near the respirator wearer if the bacteria are reaerosolized from the respirator. However, reaerosolization of a very small fraction of collected bacteria can occur only under extreme conditions of violent sneezing or coughing.

Collection of Airborne Microorganisms by Electrostatic Precipitation

The applicability of electrostatic precipitation as a method for bioaerosol collection was investigated by using a modified Electrostatic Aerosol Sampler (EAS) (Model 3100, TSI Inc., St. Paul, MN). The physical and biological efficiencies of this method were determined. The tests were performed using three bacterial species which were collected onto agar, into water, and onto filters. The physical collection efficiency was higher than 80% when using a sampling flow rate of 1 L/min. When the Bacillus subtilis var niger (BG) spores were collected on agar, about 50-60% of the collected culturable organisms formed colonies. The bioefficiency exceeded 90% when the BG spores were collected on a filter, but was only 15-22% when collected into water. The Mycobacterium bovis BCG bacteria recovered at the 0-8% level on all three collection media. The least number of colonies were formed when Pseudomonas fluorescens bacteria were collected on any of the collection media. The data show that the process of electrostatic collection is very complex for sensitive airborne bacteria and thus several effects should be considered when assessing its bioefficiency. In separate tests conducted without aerosol flow through the sampler, bacteria placed onto the collection media did not show any significant reduction in bacterial recovery while exposed to a strong electric field. It was found that evaporation from the collection media, such as agar or water, increases the humidity inside the EAS and may affect the size distribution of the particles being collected, resulting in decreased physical and biological efficiencies of the electrostatic precipitation method. For hardy microorganisms such as BG spores, the bioefficiency for electrostatic collection is high, thus encouraging further explorations of the electrostatic method for sampling bioaerosols.

Detection of Bioaerosols Using Multiwavelength UV Fluorescence Spectroscopy

This article describes the development of an aerosol generation apparatus to investigate the fluorescence spectra of bioaerosols. The experimental system was set up in a Biosafety Level II Laboratory. The system included an aerosol generator, chamber, aerosol monitoring instrumentation, and laser-induced-fluorescence detection system. The aerosol generators, chamber, and monitors were housed in an enclosure with the exhaust vented through a double HEPA filtration system. A Hospitak nebulizer using aqueous suspensions generated aerosols of bacteria. Aerosols of pollens were generated using a small-scale dry powder generator. The aerosol chamber, with four windows for optical access, was designed with the aid of a computational fluid dynamics code to optimize the generation of aerosol beams with a well-defined geometry for reproducible fluorescence measurements. Aerosol concentrations and aerodynamic diameters in the chamber were determined using a filter, an impinger, a modified Andersen impactor, and an Aerodynamic Particle Sizer. Geometric size and particle shapes were determined using microscopy and imaging analysis. The well-characterized aerosol stream allowed reproducible fluorescence measurements to be made with the aerosol generation methods developed in this work. Fluorescence spectra of four bacteria, Escherichia coli, Staphylococcus aureus , Bacillus subtilis var niger , and Bacillus thuringiensis , were found to be very similar. Calibrations of the fluorescence instrumentation allowed cross sections of live cells, killed cells, and spores to be measured with an uncertainty of about 2 to 5.

Evaluation of the antimicrobial activity of a new super-oxidized water, Sterilox®, for the disinfection of endoscopes

The antimicrobial activity of a new super-oxidized water, Sterilox®, has been tested against Mycobacterium tuberculosis, Mycobacterium avium-intracellulare, Mycobacterium chelonae, Escherichia coli (including type 0157), Enterococcus faecalis, Pseudomonas aeruginosa, Bacillus subtilis var niger spores, methicillin-resistant Staphylococcus aureus, Candida albicans, poliovirus type 2 and human immunodeficiency virus HIV-1. Under clean conditions, freshly generated Sterilox was found to be highly active against all these micro-organisms giving a 5 log 10 (99.999%) or greater reduction in two minutes or less.

Bacteriological evaluation of a down-draught necropsy table ventilation system.

To evaluate the microbiological efficacy of a down-draught necropsy table ventilation system (which surrounds the cadaver with a "curtain" of air under continuous extraction) during post mortem procedures. Air sampling was carried out both in the presence and absence of staff and cadaver and during a full post mortem procedure, with functioning and non-functioning table air extraction. The penetration of the air "curtain" was also examined during the use of an oscillating bone saw by means of a tracer organism, Bacillus subtilis var niger, painted on to the skull. There was little difference between bacterial counts obtained in the presence of staff only, staff plus cadaver, or during a post mortem examination. With all counts obtained, however, there was a two to three-fold reduction when the ventilation was in operation compared with when the extract duct was occluded. Using the tracer organism, a two to three log reduction in counts was shown when the "curtain" was in operation during the use of the oscillating bone saw. These results suggest that the system provides potential protection for post mortem room staff against airborne infections.

The biosynthesis of 1-deoxynojirimycin in Bacillus subtilis var niger

Bacillus subtilis var niger ATCC 9372 when grown on a glucose containing soyabean medium produces the amino-sugar 1-deoxynojirimycin (DNJ). The C2 epimer 1-deoxy-mannojirimycin (DMJ) present in Streptomyces subrutilus ATCC 27467 is absent. 13C and 2H labelling studies confirm that glucose is the precursor of DNJ. Randomisation of the 13C label from 1-[ 13C]-D-glucose and the incorporation of 1-[ 13C]-D,L-glyceraldehyde into DNJ also suggest that a minor part of the biosynthetic route involves cleavage of the glucose skeleton. Mannojirimycin (MJ) and nojirimycin (NJ) are postulated intermediates in the biosynthesis as a result of both enzyme assay and isotope labelling studies.

Evaluation of ethylene oxide sterilization of tissue implants

the ability of ethylene oxide (ETO) gas to penetrate tissue matrices was evaluated using Bacillus subtilis var niger (NCTC 10073) spore strips sandwiched between sheets of various tissues exposed to ETO for varying periods. The decimal reduction times (min) obtained using commercially prepared spores were: 7·97 for naked spores, 7·34 between amnion, and 6·39 between amnio-chorion, indicating complete penetration of the matrix by the gas. Similar conclusions were reached using skin and dura mater. The possible protective effect of serum was evaluated by drying spores suspended in serum in different ways, followed by exposure to the gas. Decimal reduction times obtained were: 8·67 (dried at 37 °C); 8·09 (dried at 55 °C) and 8·97 (freeze-dried) confirming slight protection. However, in all cases sterilization was achieved following 100 min ETO exposure, giving a 2·5 times safety factor for the commercial ethylene oxide sterilizer.

Evaluation of air filters with submicron viral aerosols and bacterial aerosols.

Abstract Velocity, aerosol particle size, aerosol charge, and exposure to high humidity were found to affect the performance of air filters. Filter papers and the DOP scan-tested filter units (HEPA filters) fabricated from these papers were evaluated with submicron T1 bacteriophage aerosols having a number median diameter (NMD) of 0.12 micron and with Bacillus subtilis var niger spore aerosols with a NMD of 1 micron. Penetration of submicron phage aerosols through filter papers increased markedly with an increase in velocity. Neutralizing phage aerosol with bipolar air ions resulted in 2- to 5-fold increases in penetration. Penetration of bacterial spore aerosols through the filter papers was essentially zero. Phage aerosol penetration through HEPA filters averaged 0.00095%. Spore aerosol penetration was 0.00005%. Exposure to high humidity (>95%) resulted in roughly a 3-fold increase in HEPA penetration. HEPA filters can be expected to provide excellent protection against submicron viral aerosols as well ...

Foam separation of bacteria with a cationic surfactant

AbstractAn experimental investigation is presented of the foam separation of six species of bacteria: Escherichia coli, Serratia marcescens, Proteus vulgaris, Pseudomonas fluorescens, Bacillus cereus, and Bacillus subtilis var niger. A cationic surfactant, ethylhexadeeyldimethylammonium bromide is used and results are evaluated in terms of total cell count, using a membrane filtration technique. From similar neutral distilled water suspensions of the pure cultures (approximately 107 cells/ml.) and using the same operating conditions, ratios of cell concentrations in the residual suspensions to those in the initial suspensions range from 0.0013 for Bacillus subtilis var niger to 0.25 for Serratia marcescens. The presence of bacteria, compared to pure surfactant solutions, produces lower collapsed foam volumes; the foam volumes have a strong influence on the separations achieved with the various species, with enrichment ratios ranging from 27 to 3088 and residual ratios ranging from 0.001 to 0.247.