Nachiket Vaze

Rapid Inactivation of Airborne Bacteria Using Atmospheric Pressure Dielectric Barrier Grating Discharge

Dielectric barrier discharge plasma has been known to inactivate many different microorganisms on surfaces when treatment times are on the order of seconds or minutes in duration. In this paper, a unique plasma air cleaning facility was created which combines a dielectric barrier grating discharge (DBGD) with a filterless laboratory-scale ventilation system and is used to treat concentrated bacterial bioaerosol in a moving air stream at air flow rates of 25 L/s. Results indicate that plasma treatment times on the order of milliseconds corresponding to one pass through the DBGD device can achieve 1.5-log reduction in culturable E. coli immediately after contact with plasma and 5-log reduction totally following in the minutes after the plasma treatment. A numerical characterization study was performed to help predict and understand the mechanism of bacteria inactivation in the DBD plasma from a variety of plasma factors.

The Presence and Impact of Biofilm-Producing Staphylococci in Atopic Dermatitis

IMPORTANCE Atopic dermatitis (AD) is thought to be a double-hit phenomenon with an unknown environmental component and a genetic abnormality likely centered on the filaggrin gene. Biologically, the presence of Staphylococcus aureus in AD was reported more than 2 decades ago, but the relationship to AD has been elusive. OBJECTIVE To explore the bacteria that produce the biofilms in the lesions of AD and the response of the innate immune system to these biofilm occlusions of the sweat ducts by specifically evaluating Toll-like receptor 2. DESIGN, SETTING, AND PARTICIPANTS University hospital dermatologic clinic study involving the environmental component related to the characterization, correlation, and impact of staphylococci and their biofilms in AD. We processed routine skin swabs from lesional and nonlesional skin from 40 patients with AD and performed scrapings and biopsies. We also obtained 20 samples from controls (10 inflamed skin samples and 10 normal skin samples). EXPOSURES Gram staining, bright-field microscopy, hematoxylin and eosin, periodic acid-Schiff, Congo red, and light microscopy. MAIN OUTCOMES AND MEASURES Association of staphylococcal biofilms with AD pathogenesis. RESULTS All AD-affected samples contained multidrug-resistant staphylococci, with S aureus (42.0%) and Staphylococcus epidermidis (20.0%) as the predominant species. All isolates were positive for extracellular polysaccharide and biofilm (85.0% strong biofilm producers and 15.0% moderately to weakly positive). Polymerase chain reaction revealed the biofilm-mediating icaD (93.0%) and aap (12.5%) genes in the isolates (some contained both). We also examined tissues for microbial identification, extracellular biomass formation, biofilm formation, and staphylococcal biofilm in skin tissues. Occlusion of sweat ducts with periodic acid-Schiff-positive and Congo red-positive material was noted on microscopic tissue examination. Toll-like receptor 2 was shown to be activated in AD lesional skin (immediately proximal to the sweat ducts), which likely led to the initiation of proteinase-activated receptor 2-mediated pruritus and MyD88-mediated spongiosis. CONCLUSIONS AND RELEVANCE Biofilm formation by AD-associated staphylococci almost certainly plays a major role in the occlusion of sweat ducts and leads to inflammation and pruritus. We believe the environmental hit in AD relates to staphylococci and their biofilms, which occlude sweat ducts.

Inactivation of Bacteria in Flight by Direct Exposure to Nonthermal Plasma

Plasma treatment is a promising technology for fast and effective sterilization of surfaces, waterflow, and airflow. The treatment of airflow is an important area of healthcare and biodefense that has recently gained the interest of many scientists. In this paper, we describe a dielectric barrier grating discharge (DBGD) which is used to study the inactivation of airborne Escherichia coli inside a closed air circulation system. Earlier published results indicate approximately 5-log reduction (99.999%) in the concentration of the airborne bacteria after single DBGD exposure of 10-s duration. This paper investigates plasma species influencing the inactivation. The two major factors that are studied are the effect of charged and short-lived species (direct exposure to plasma) and the effect of ozone. It is shown that for a 25% reduction in direct exposure, the inactivation falls from 97% to 29% in a single pass through the grating. The influence of ozone was studied by producing ozone remotely with an ozone generator and injecting the same concentration into the system, as that produced by the DBGD plasma. The results show a 10% reduction in the bacterial load after 10-s exposure to ozone; thus, ozone alone may not be one of the major inactivating factors in the plasma.

Plasma for Air and Water Sterilization

This chapter describes the research efforts of Drexel Plasma Institute (DPI) in the area of plasma-based air and water sterilization. Motivation of this research is presented as well as the methods for selection of parameters for the experimental systems. Experimentally obtained results for air sterilization demonstrate that the direct influence of plasma charged particles on airborne bacteria in combination with active chemical substances generated by plasma is the probable reason for high sterilization efficiency of the Dielectric Barrier Grating Discharge (DBGD). Energy input on the level of 13 kJ/m3 is enough to reach a 5-log reduction of viable E. coli bacteria. Experimentally reached D-value (the dosage required for a 90% reduction of the number of viable microorganisms) for E. coli bacteria deactivation in water using spark discharge is very low, about 125 kJ/m3, and UV-radiation is the most plausible sterilization factor in this case. A new semi-numerical model is proposed for initial phase of electrical breakdown in water.

Air and Water Sterilization using Non-Thermal Plasma

The sterilization effect of plasma on air and water were investigated in this study*. For air sterilization, a small scale model of HVAC was designed and Dielectric Barrier Discharge plasma source was used for treatment of air. This PDRF (Pathogen Detection and Remediation Facility) consisted of a circulatory airflow system, a plasma chamber and a sampling system. Air sterilization experiments were performed and the inactivation of Escherichia coli was studied. Conventional water sterilization methods such as chlorination, ozonation, filtration, UV irradiation etc have several drawbacks. Pulsed plasma discharge for the destruction of microorganisms in waste water and potable water is a cost effective technique developed recently. The energy efficiency of different types of plasma discharges in water contaminated with Escherichia coli has been studied. The effect of initial concentration of bacterial solution on the inactivation efficiency has also been studied

Involvement of multiple stressors induced by non-thermal plasma-charged aerosols during inactivation of airborne bacteria

A lab-scale, tunable, single-filament, point-to-point nonthermal dieletric-barrier discharge (DBD) plasma device was built to study the mechanisms of inactivation of aerosolized bacterial pathogens. The system inactivates airborne antibiotic-resistant pathogens efficiently. Nebulization mediated pre-optimized (4 log and 7 log) bacterial loads were challenged to plasma-charged aerosols, and lethal and sublethal doses determined using colony assay, and cell viability assay; and the loss of membrane potential and cellular respiration were determined using cell membrane potential assay and XTT assay. Using the strategies of Escherichia coli wildtype, over-expression mutant, deletion mutants, and peroxide and heat stress scavenging, we analyzed activation of intracellular reactive oxygen species (ROS) and heat shock protein (hsp) chaperons. Superoxide dismutase deletion mutants (ΔsodA, ΔsodB, ΔsodAΔsodB) and catalase mutants ΔkatG and ΔkatEΔkatG did not show significant difference from wildtype strain, and ΔkatE and ΔahpC was found significantly more susceptible to cell death than wildtype. The oxyR regulon was found to mediate plasma-charged aerosol-induced oxidative stress in bacteria. Hsp deficient E. coli (ΔhtpG, ΔgroEL, ΔclpX, ΔgrpE) showed complete inactivation of cells at ambient temperature, and the treatment at cold temperature (4°C) significantly protected hsp deletion mutants and wildtype cells, and indicate a direct involvement of hsp in plasma-charged aerosol mediated E. coli cell death.

Direct exposure to a single filament of DBD plasma leads to the inactivation of airborne bacteria

Airborne transmission is the cause of the spread of many deadly diseases. There are bacteria that can be easily aerosolized and transmitted far distances inside HVAC systems. To control the spread of these contaminants, HEPA filters are used. These filters however, do not kill bacteria and viruses but only trap them. They also cause large pressure drops inside HVAC systems. DBD plasma discharge has been used as a method of inactivating airborne bacteria[1]. The focus now is on determining the major mechanism of this inactivation. The main distinguishing parameter used to evaluate the sterilization of bacteria is whether this effect is due to direct exposure to plasma or the indirect effect of long living species produced by it. The hypothesis here is that the charges produced by DBD are responsible for this inactivation.

Use of fluorescent techniques for understanding the effect of non thermal plasma discharge on the E. coli cell

It has been demonstrated that ambient air non-thermal plasma discharges are very effective in inactivating various microorganisms. Plasma is being considered as a potent sterilization technology. It is important to understand the biological effect of plasma on bacteria. Fluorescent techniques are often employed by cell biologists to detect cellular phenomenon. These phenomena include changes in pH, membrane permeation and certain cellular reactions. The LIVE/DEAD® BacLight™ Bacterial Viability Kit (Molecular Probes, Eugene OR) consists of two stains that are used to detect live and dead bacteria in large bacterial populations. These stains are Propidium Iodide (PI) and SYTO 9. The SYTO 9 (Green) stain permeates the membranes of all bacteria, whereas the PI (Red) stain only permeates the bacteria whose membrane has been irreversibly damaged. Hence the ratio of Green to Red fluorescence is considered the Live/Dead ratio. This kit has been used successfully to detect live bacteria in soil, milk and other environmental samples. It has also been used to determine the effectiveness of bactericidal treatments. E. coli bacteria were grown overnight and suspended in solution. The solution was treated with dielectric barrier discharge (DBD) plasma. The viable counts using plating techniques and colony forming unit (CFU) counting indicate ∼2log reduction in the concentration of live bacteria. For the same treatment dose, the LIVE/DEAD® kit was added to the samples after they were plasma treated and then they were analyzed under a fluorescent microscope. Multiple fields of vision were analyzed and statistical analysis was performed on the samples. The results indicate that the number of bacteria fluorescing red was significantly lower than the expected number from viable counts. This indicates that membrane permeation is not the major cause of plasma induced inactivation of these bacteria. Further experiments are being performed to understand the effect of plasma treatment on the cellular respiration and other potential avenues for inactivation of the bacteria.

Air sterlization of spores with non-thermal plasma

Air quality control is important for facilities such as drug manufacturing plants and hospitals to prevent contaminations. The release of airborne biological weapons into the air circulation system is also a possible route of attack for military bases and large office buildings. Currently the most common method of air quality maintenance is the expensive HEPA filter system installed into the air circulation system. The HEPA filter requires routine maintenance and replacements of filters. But in dealing with deadly pathogens, even if the HEPA filter pores are small enough to filter out the pathogens, the filters do not inactivate them therefore threat is maintained. Also cost of the system and maintenance increases significantly with the level of air quality desired.

Effects of substrate conductivity on dielectric barrier discharge sterilization efficacy

Summary form only given. Non-equilibrium atmospheric pressure plasma is an ideal solution for sterilizing spacecraft before flight and upon return to Earth without thermal or chemical degradation to the surface. Spacecrafts are composed of a variety of conductive and non-conductive components. For this reason, it is important to study sterilization efficacy on varying substrates which are conductive, non-conductive, wet, dry, grounded, and un-grounded. The bacteria chosen for these experiments are Escherichia coli, Bacillus subtilis, Bacillus pumulis, and Deinococcus radiodurans. The results will allow NASA researchers to better design spacecraft for higher sterilization efficacy. Further, it will ultimately aide in preventing forward and reverse contamination for NASA space missions.