Inactivation Of Fungal Spores Research Articles

Selection of inactivation medium for fungal spores in clinical wastes by supercritical carbon dioxide.

The present study aimed to select the best medium for inactivation of Aspergillus fumigatus, Aspergillus spp. in section Nigri, A. niger, A. terreus var. terreus, A. tubingensis, Penicillium waksmanii, P. simplicissimum, and Aspergillus sp. strain no. 145 spores in clinical wastes by using supercritical carbon dioxide (SC-CO2). There were three types of solutions used including normal saline, seawater, distilled water, and physiological saline with 1% of methanol; each solution was tested at 5, 10, and 20mL of the water contents. The experiments were conducted at the optimum operating parameters of supercritical carbon dioxide (30MPa, 75°C, 90min). The results showed that the inactivation rate was more effective in distilled water with the presence of 1% methanol (6 log reductions). Meanwhile, the seawater decreases inactivation rate more than normal saline (4.5 vs. 5.1 log reduction). On the other hand, the experiments performed with different volumes of distilled water (5, 10, and 20mL) indicated that A. niger spores were completely inactivated with 10mL of distilled water. The inactivation rate of fungal spores decreased from 6 to 4.5 log as the amount of distilled water increased from 10 to 20mL. The analysis for the spore morphology of A. fumigatus and Aspergillus spp. in section Nigri using scanning electron microscopy (SEM) has revealed the role of temperature and pressure in the SC-CO2 in the destruction of the cell walls of the spores. It can be concluded that the distilled water represent the best medium for inactivation of fungal spores in the clinical solid wastes by SC-CO2.

Inactivation of three genera of dominant fungal spores in groundwater using chlorine dioxide: Effectiveness, influencing factors, and mechanisms

Fungi in aquatic environments received more attention recently; therefore, the characteristics of inactivation of fungal spores by widely used disinfectants are quite important. Nonetheless, the inactivation efficacy of fungal spores by chlorine dioxide is poorly known. In this study, the effectiveness of chlorine dioxide at inactivation of three dominant genera of fungal spores isolated from drinking groundwater and the effects of pH, temperature, chlorine dioxide concentration, and humic acid were evaluated. The inactivation mechanisms were explored by analyzing the leakage of intracellular substances, the increase in extracellular adenosine triphosphate (ATP), deoxyribonucleic acid (DNA), and proteins as well as the changes in spore morphology. The kinetics of inactivation by chlorine dioxide fitted the Chick-Watson model, and different fungal species showed different resistance to chlorine dioxide inactivation, which was in the following order: Cladosporium sp.>Trichoderma sp. >Penicillium sp., which are much more resistant than Escherichia coli. Regarding the three genera of fungal spores used in this study, chlorine dioxide was more effective at inactivation of fungal spores than chlorine. The effect of disinfectant concentration and temperature was positive, and the impact of pH levels (6.0 and 7.0) was insignificant, whereas the influence of water matrices on the inactivation efficiency was negative. The increased concentration of characteristic extracellular substances and changes of spore morphology were observed after inactivation with chlorine dioxide and were due to cell wall and cell membrane damage in fungal spores, causing the leakage of intracellular substances and death of a fungal spore.

Inactivation of four genera of dominant fungal spores in groundwater using UV and UV/PMS: Efficiency and mechanisms

Outbreaks of fungi in groundwater sources of drinking water can produce taste and odor problems, and cause ill health in immunocompromised individuals. The groundwater treatment process is simple; generally, only disinfection is used in China. Efficient disinfection methods to control fungi are required. In the present study, we investigated the inactivation efficiency of 4 dominant genera of fungal spores (a Trichoderma sp., Acremonium sp., Penicillium sp., and Cladosporium sp.) using ultraviolet irradiation (UV) and UV-based advanced oxidation processes, and explored the mechanisms of inactivation by monitoring the leakage of intracellular contents and changes in spore morphology. The inactivation of fungal spores is consistent with first-order Chick–Watson kinetics. The resistance of each of the fungi to UV (Cladosporium sp.>Penicillium sp.>Acremonium sp.>Trichoderma sp.) was greater than that of E. coli, due to their larger sizes and more complex structures. UV/peroxymonosulfate (UV/PMS) treatment had a markedly better inactivation rate constant than UV alone. Furthermore, UV/PMS exhibited similar effects on fungal inactivation in phosphate buffer and in groundwater, whereas UV inactivation efficiency markedly decreased in groundwater. UV/PMS treatment resulted in cell wall and cell membrane damage, the leakage of intracellular contents, and the shriveling of fungal spores, due to the reactive radicals produced by the treatment. Thus, UV/PMS is a promising method to control fungi in drinking water sources.

A novel photosensitization treatment for the inactivation of fungal spores and cells mediated by curcumin

The global concerns regarding the emergence of fungicide-resistant strains and the impact of the excessive use of fungicidal practises on our health, food, and environment have increased, leading to a demand for alternative clean green technologies as treatments. Photosensitization is a treatment that utilises a photosensitiser, light and oxygen to cause cell damage to microorganisms. The effect of photosensitization mediated by curcumin on Aspergillus niger, Aspergillus flavus, Penicillium griseofulvum, Penicillium chrysogenum, Fusarium oxysporum, Candida albicans and Zygosaccharomyces bailii was investigated using three methods. The viability of spores/cells suspended in aqueous buffer using different concentrations of curcumin solution (100–1000μM) and light dose (0, 24, 48, 72 and 96J/cm2) were determined. Spraying curcumin solution on inoculated surfaces of agar plates followed by irradiation and soaking spores/cells in curcumin solution prior to irradiation was also investigated. In aqueous mixtures, photosensitised spores/cells of F. oxysporum and C. albicans were inhibited at all light doses and curcumin concentrations, while inactivation of A. niger, A. flavus P. griseofulvum, P. chrysogenum and Z. bailii were highly significant (P<0.001) reduced by 99%, 88.9%, 78%, 99.7% and 99.2% respectively. On the surface of agar plates, spores/cells exposed to a light dose of 360J/cm2 sprayed with curcumin at 800μM showed complete inhibition for A. niger, F. oxysporum, C. albicans and Z. bailii, while A. flavus P. griseofulvum, and P. chrysogenum reduced by 75%, 80.4% and 88.5% respectively. Soaking spores/cells with curcumin solution prior to irradiation did not have a significant effect on the percentage reduction. These observations suggest that a novel photosensitization mediated curcumin treatment is effective against fungal spores/cells and the variation of percentage reduction was dependent on curcumin concentration, light dosage and fungal species.

Inactivation Efficiency and Mechanism of Three Dominant Fungal Spores in Drinking Groundwater by Chlorine

The outbreak of fungi in drinking groundwater sources can produce odor, cause toxicity and form lots of visible flocs, which seriously affect the drinking water quality. In this study, the inactivation efficiency of three dominant fungal spores Trichoderma, Penicillium, Cladosporium by chlorine was conducted, and the inactivation mechanism was explored by monitoring the hydrophobicity, the leakage of intracellular substances, the increase of extracellular adenosine triphosphate (ATP), deoxyribonucleic acid (DNA) and proteins, and the change of spores' morphology. The results showed that the inactivation of fungal spores was consistent with first-order kinetics and satisfied the Chick model. The resistance to chlorine was in sequence of Trichoderma > Penicillium > Cladosporium, the larger size the fungal spores and the more hydrophilicity the fungal spores, the higher the inactivation efficiency. Chlorination resulted in the remarkable leakage of intracellular compounds, the increase of extracellular characteristic compounds (ATP, DNA and protein), the damage of the cell surface, and the fungal spores recessed and wrinkled. In summary, chlorine firstly reacted with spores' surface and reduced their cultivability, and then resulted in the damage to the permeability barrier of the spores and the release of intracellular characteristic compounds, and finally the viability of spores was damaged.

Supercritical Fluid CO2 Technique for Destruction of Pathogenic Fungal Spores in Solid Clinical Wastes

The present study deals with the inactivation of fungal spores in clinical waste using the advanced supercritical carbon‐dioxide (SC‐CO2) treatment technique. The process of inactivation was investigated under different conditions of pressure, temperature, and processing time. The efficiency of SC‐CO2 was determined in terms of log reduction and inactivation rate of fungal spores. Almost 100% inactivation of initial fungal spores was obtained. Predicted and experimental log reduction of fungal spores was found to be 5.93 and 6.00, respectively, under optimal conditions (35 MPa, 75°C in 90 min). SC‐CO2 treated Aspergillus niger and Penicillium simplicissimum spores were examined using scanning electron microscopy which showed spore damage in the form of rupture, tear, and shape distortion. These damages confirmed complete spore inactivation when no renewed fungal growth occurred after the treated samples were cultured onto fresh media. The inactivation of fungal spores using SC‐CO2 and autoclave inhibits the production of lipase, cellulase, amylase, and protease enzymes. Thus, without addition of any chemicals, the SC‐CO2 sterilization method can be potentially used for the inactivation of microorganisms in clinical waste as well as in food and pharmaceutical products.

Identifying the role of reactive oxygen species (ROSs) in Fusarium solani spores inactivation.

The inactivation mechanism of photocatalytic disinfectants on bacteria is well known. In contrast, the potential inactivation of fungal spores by visible-light induced photocatalysis has been recognized, but the inactivation mechanism is poorly understood. We hypothesize that photocatalytically generated reactive oxygen species (ROSs) are directly involved in this mechanism. To test this hypothesis, we identified the roles of ROSs in the inactivation of Fusarium solani spores. As the photocatalysts, we doped TiO2 with 3 typical dopants, forming Ag/TiO2, N/TiO2 and Er3+:YAlO3/TiO2. The Ag/TiO2 photocatalysis was dominated by H2O2, with the longest lifetime among the investigated ROSs. Ag/TiO2 photocatalysis yielded almost 100 % inactivation efficiency and preserved the cell-wall shape of the spores, thus minimizing the biomolecule leakage. Er3+:YAlO3/TiO2 was dominated by h+ ROSs, yielding an inactivation efficiency of 91 %; however, the severe leakage released large numbers of molecular bio-products. Severe damage to the cell walls by the h+ species was confirmed in micrograph observations. Subsequent to cell wall breakage, the Er3+:YAlO3/TiO2 nanoparticles entered the spore cells and directly oxidized the intracellular material. The N/TiO2 photocatalysis, with •O2− dominated ROSs, delivered intermediate performance. In conclusion, photocatalysts that generate H2O2-dominated ROSs are most preferred for spore inactivation.

Differential Inactivation of Fungal Spores in Water and on Seeds by Ozone and Arc Discharge Plasma.

Seed sterilization is essential for preventing seed borne fungal diseases. Sterilization tools based on physical technologies have recently received much attention. However, available information is very limited in terms of efficiency, safety, and mode of action. In this study, we have examined antifungal activity of ozone and arc discharge plasma, potential tools for seed sterilization. In our results, ozone and arc discharge plasma have shown differential antifungal effects, depending on the environment associated with fungal spores (freely submerged in water or infected seeds). Ozone inactivates Fusarium fujikuroi (fungus causing rice bakanae disease) spores submerged in water more efficiently than arc discharge plasma. However, fungal spores associated with or infecting rice seeds are more effectively deactivated by arc discharge plasma. ROS generated in water by ozone may function as a powerful fungicidal factor. On the other hand, shockwave generated from arc discharge plasma may have greatly contributed to antifungal effects on fungus associated with rice seeds. In support of this notion, addition of ultrasonic wave in ozone generating water has greatly increased the efficiency of seed disinfection.

Effects of the physical parameters of a microwave plasma jet on the inactivation of fungal spores

In spite of their importance in human life, filamentous fungi have not been actively explored in the application of plasma to them. A plasma jet source at the atmospheric pressure was excited by 2.45GHz microwaves and operated at low energy regime with an average power of 0.8W–1.6W. This microwave plasma was applied to examine fungal inactivation and find physical conditions of plasma (electrical power, pulse widths, and fed gases) at which the highest inhibition effects on fungal growth was achieved. Spore germination and hyphal growth of the fungus were dramatically decreased when oxygen was used in the plasma discharge, and this might be due to the elevation in the level of Oxygen (O) radical. The level of O radical in the plasma generated from Ar and oxygen was also enhanced by the increased power and pulse width. Hyphal growth of the fungus was more inhibited when greater power or longer pulse was applied. It appears that plasma effects were varied among different fungal species. Different levels of inhibition on spore germination and growth of three filamentous fungi, Neurospora crassa, Fusarium graminearum, and Fusarium oxysporum was observed.

Inactivation of Aspergillus niger spores from indoor air by photocatalytic filters

The effects of UV-A and UV-C radiation on fungal spores were investigated before and after their germination in photocatalytic and non-photocatalytic air filters commonly used in heating, ventilating, and air conditioning (HVAC) systems.Immediately after the coating of spores on filters, exposure to both types of UV radiation induced the appearance of an inactivation threshold for long durations of exposure probably resulting from the presence of Aspergillus niger spores inside the activated charcoal layer. The use of a thin photocatalytic filter without activated charcoal demonstrated a better disinfection efficiency with total inactivation of the spores, due to an optimal contact between spores and TiO2 coating.The effects of UV radiation were then assessed on spore germination for both types of filters. The inactivation of spores in illuminated photocatalytic filters resulted in an irreversible inhibition of the fungal germination under UV-A or UV-C radiation. In contrast, fungal spores were able to germinate in non-photocatalytic filters despite previous exposure to both types of UV radiation. The monitoring of ergosterol amounts, the major sterol of fungal membranes, corroborated these results.Finally, UV-A or UV-C radiation exposure of filters after spore germination had a lesser disinfection efficiency than experiments whereby spores had just been applied onto the filters, due to the absence of contact between the biological pollutants and the photocatalyst coating.Our results thus demonstrated the interest to use photocatalytic filters ensuring optimal contact between pollutants and TiO2 coating to lead to a total inactivation of fungal spores in filters of HVAC systems.

Inactivation of Alternaria brassicicola spores by supercritical carbon dioxide with ethanol entrainer

Supercritical carbon dioxide (SC–CO 2) was used to inactivate fungal spores of Alternaria brassicicola. The inactivation conditions were optimized using response surface methodology (RSM). When the SC–CO 2–entrainer (ethanol) system was applied to fungal spores, the treatment time required for the complete inactivation of fungal spores was substantially reduced.

Inactivation of fungal spores in apple juice by high pressure homogenization.

High pressure homogenization (HPH) at 300 MPa in apple juice provides more than 5-log kill of ascospores of Saccharomyces cerevisiae, conidiospores of filamentous fungi, and sporulated black yeasts. HPH and heat treatment were more effective against vegetative cells than against the spores of yeasts used in this study. Ascospores of Talaromyces macrosporus and Neosartorya spinosa were resistant to HPH at 300 MPa. HPH of ascospores of T. macrosporus may result in activation.

Ultraviolet germicidal irradiation inactivation of airborne fungal spores and bacteria in upper-room air and HVAC in-duct configurations

The efficacy of ultraviolet germicidal irradiation (UVGI) for inactivating airborne fungal spores and bacterial vegetative cells was evaluated under three configurations — intrinsic, upper-room air, and in-duct. Correspondingly, experiments were performed in (1) a pilot-scale chamber (0.8 m3), fitted with four corner UV lamps that irradiated the entire chamber (average UV fluence rate 10.6 ± 0.8 µJ s–1cm–2); (2) a full-scale room (87 m3), fitted with a UVGI system that irradiated the top 30 cm of the room (5 fixtures, 216 W total lamp power, average upper-zone UV fluence rate 26 ± 1 µJ s–1cm–2); and (3) the supply air duct of a heating ventilation and air-conditioning (HVAC) system. Fungal spores of Aspergillus versicolor and vegetative cells of bacterium Mycobacterium parafortuitum were aerosolized continuously such that their numbers and physiologic state were comparable both with and without the UVGI lamps operating. The Z value (UVGI inactivation rate normalized to UVGI fluence rate) was estimated to be 1.2 ± 0.4 x 10–4 cm2 µJ–1 for aerosolized A. versicolor. Upper-room air UVGI inactivated culturable airborne fungal spores with a first-order rate constant of 0.4 ± 0.2 h–1. Ultraviolet lamps enclosed in ventilation system ductwork inactivated fungal spores and vegetative bacterial cells at single-pass efficiencies of 75% and 87%, respectively, at an air stream velocity of 2.2 m s–1. There was no detected inactivation of fungal spores and vegetative bacterial cells at an air stream velocity of 5.1 m s–1. Key words: bioaerosols, UVGI, air disinfection.

Photodynamic Treatment of the Dermatophyte Trichophyton rubrum and its Microconidia with Porphyrin Photosensitizers¶

ABSTRACTThe application of photosensitizers for the treatment of fungal infections is a new and promising development within the field of photodynamic treatment (PDT). Dermatophytes, fungi that can cause infections of the skin, hair and nails, are able to feed on keratin. Superficial mycoses are probably the most prevalent of infectious diseases in all parts of the world. One of the most important restrictions of the current therapeutic options is the return of the infection and the duration of the treatment. This is especially true in the case of infections of the nail (tinea unguium) caused by Trichophyton rubrum, an anthropophilic dermatophyte with a worldwide distribution. Recently, we demonstrated that 5,10,15‐tris(4‐methylpyridinium)‐20‐phenyl‐[21H,23H]‐porphine trichloride (Sylsens B) and deuteroporphyrin monomethylester were excellent photosensitizers toward T. rubrum when using broadband white light. This study demonstrates the photodynamic activity of these photosensitizers with red light toward both a suspension culture of T. rubrum and its isolated microconidia. The higher penetration depth of red light is important for the PDT of nail infections. In addition, we tested the photodynamic activity of a newly synthesized porphyrin, quinolino‐[4,5,6,7‐efg]‐7‐demethyl‐8‐deethylmesoporphyrin dimethylester, displaying a distinct peak in the red part of the spectrum. However, its photodynamic activity with red light toward a suspension culture of T. rubrum appeared to be only fungistatic. Sylsens B was the best photosensitizer toward both T. rubrum and its microconidia. A complete inactivation of the fungal spores and destruction of the fungal hyphae was found. In studies into the photostability, Sylsens B appeared to be photostable under the conditions used for fungal PDT. A promising result of this study is the demonstration of the complete degradation of the fungal hyphae in the time after the PDT and the inactivation of fungal spores, both with red light. These results offer the ingredients for a future treatment of fungal infections, including those of the nail.

Photodynamic treatment of the dermatophyte Trichophyton rubrum and its microconidia with porphyrin photosensitizers.

The application of photosensitizers for the treatment of fungal infections is a new and promising development within the field of photodynamic treatment (PDT). Dermatophytes, fungi that can cause infections of the skin, hair and nails, are able to feed on keratin. Superficial mycoses are probably the most prevalent of infectious diseases in all parts of the world. One of the most important restrictions of the current therapeutic options is the return of the infection and the duration of the treatment. This is especially true in the case of infections of the nail (tinea unguium) caused by Trichophyton rubrum, an anthropophilic dermatophyte with a worldwide distribution. Recently, we demonstrated that 5,10,15-tris(4-methylpyridinium)-20-phenyl-[21H,23H]-porphine trichloride (Sylsens B) and deuteroporphyrin monomethylester were excellent photosensitizers toward T. rubrum when using broadband white light. This study demonstrates the photodynamic activity of these photosensitizers with red light toward both a suspension culture of T. rubrum and its isolated microconidia. The higher penetration depth of red light is important for the PDT of nail infections. In addition, we tested the photodynamic activity of a newly synthesized porphyrin, quinolino-[4,5,6,7-efg]-7-demethyl-8-deethylmesoporphyrin dimethylester, displaying a distinct peak in the red part of the spectrum. However, its photodynamic activity with red light toward a suspension culture of T. rubrum appeared to be only fungistatic. Sylsens B was the best photosensitizer toward both T. rubrum and its microconidia. A complete inactivation of the fungal spores and destruction of the fungal hyphae was found. In studies into the photostability, Sylsens B appeared to be photostable under the conditions used for fungal PDT. A promising result of this study is the demonstration of the complete degradation of the fungal hyphae in the time after the PDT and the inactivation of fungal spores, both with red light. These results offer the ingredients for a future treatment of fungal infections, including those of the nail.

AN INTEGRATED DYNAMIC MODEL FOR FUNGAL SPORE INACTIVATION BY SELECTIVE INFRARED HEATING

The effect of selective infrared radiation (IR) heating for the inactivation of fungal spores based on a dynamictemperature profile was explored by an integrated model that combined the thermal death kinetics with heat transfer duringIR heating. Selective IR heating was found to differentially contribute to a higher degree of lethality compared to normal IRheating. Differential heating of fungal spores was validated by demonstrating that dry fungal spores were heated by up to 6.Chigher than pure corn meal after 300 s of heating. An R2-value greater than 0.98 was observed between the model-predictedand measured survival ratios. The denaturation of protein components in the selective IR range also contributed to anadditional increase in the degree of lethality of fungal spores, compared to model prediction.

Pressure inactivation of fungal spores in aqueous model solutions and in real food systems

Conidiospores from Penicillium expansum and ascospores from Eurotium repens were exposed to high hydrostatic pressure in isotonic salt solution, apple and broccoli juice. Kinetic measurements were done at 4,25 and 40 or 45°C. The shape of the inactivation curves was strongly dependent on the temperature. Asco- and conidiospores were found to behave in a contrary way. The fastest reduction for conidiospores was found at 4°C, for ascospores best inactivation was achieved at 45°C. High pressure inactivation of spores in apple or broccoli juice was nearly the same as in isotonic salt solution.

Chlorine inactivation of fungal spores on cereal grains

Although 0.4% chlorine for 2 min has been recommended for surface disinfection of food samples before direct plating for fungal enumeration, this procedure may not be adequate for highly contaminated products. The effectiveness of a range of chlorine solutions was investigated using barley samples artificially contaminated with four different concentrations of Aspergillus flavus, A. niger, A. ochraceus, Eurotium repens, Penicillium brevicompactum P. chrysogenum and Cladosporium cladosporioides. At initial contamination levels greater than 10 4/g, 0.4% chlorine did not inactivate sufficient spores to produce less than 20% contamination. Of the test fungi, ascospores of E. repens were the most resistant to chlorine inactivation, whereas the conidia of C. cladosporioides were the most sensitive. Rinsing the samples with 70% ethanol improved the effectiveness of the recommended surface disinfection procedure. However, some ethanol appears to permeate into the grains and may inactivate sensitive internal fungi, although a minimal effect only was observed on wheat infected with Alternaria.