Effects Of Sodium Hypochlorite Research Articles

Exploration of optimal disinfection model based on groundwater risk assessment in disinfection process

Under the influence of different types of disinfectants and disinfection environments, the removal level of pathogens and the formation potential of disinfection by-products (DBPs) will have a dual impact on the groundwater environment. The key points for sustainable groundwater safety management are how to balance the positive and negative relationship and formulate a scientific disinfection model in combination with risk assessment. In this study, the effects of sodium hypochlorite (NaClO) and peracetic acid (PAA) concentrations on pathogenic E. coli and DBPs were investigated using static-batch and dynamic-column experiments, as well as the optimal disinfection model for groundwater risk assessment was explored using quantitative microbial risk assessment and disability-adjusted life years (DALYs) models. Compared to static disinfection, deposition and adsorption were the dominant factors causing E. coli migration at lower NaClO levels of 0–0.25 mg/L under dynamic state, while disinfection was its migration factor at higher NaClO levels of 0.5–6.5 mg/L. In contrast, E. coli removed by PAA was the result of the combined action of deposition, adsorption, and disinfection. The disinfection effects of NaClO and PAA on E. coli differed under dynamic and static conditions. At the same NaClO level, the health risk associated with E. coli in groundwater was higher, whereas, under the same PAA conditions, the health risk was lower. Under dynamic conditions, the optimal disinfectant dosage required for NaClO and PAA to reach the same acceptable risk level was 2 and 0.85 times (irrigation) or 0.92 times (drinking) of static disinfection, respectively. The results may help prevent the misuse of disinfectants and provide theoretical support for managing twin health risks posed by pathogens and DBPs in water treatment.

Effects of sodium hypochlorite and ethylenediaminetetraacetic acid on proliferation, osteogenic/odontogenic differentiation, and mechanosensitive gene expression of human dental pulp stem cells

Sodium Hypochlorite (NaOCl) and Ethylene Diamine Tetraacetic Acid (EDTA) can change the biochemical and biophysical properties of dentin. However, the response of human dental pulp stem cells (hDPSCs) to NaOCl and EDTA-treated dentin remains unknown. This study was conducted to investigate the effect of NaOCl and EDTA on cell proliferation, osteogenic/odontogenic differentiation, and the response to mechanosensitive gene expression in hDPSCs. Dentin slices were treated with 5.25% NaOCl, 17% EDTA, and saline (0.9% NaCl) separately. The cell viability and osteogenic/odontogenic differentiation of hDPSCs were analyzed using scanning electron microscopy, cell counting assay, alkaline phosphatase (ALP) staining, and quantitative polymerase chain reaction (qPCR). Besides, the hardness was measured by a Vickers microhardness tester. The expression of mechanosensitive genes was detected by the qPCR assay. All the irrigant-treated dentin allowed cell attachment. The EDTA-treated dentin significantly boosted the ALP and osteogenic/odontogenic differentiation, followed by NaCl and NaOCl groups. Remarkably, these trends were similar to the expression of mechanosensitive genes but were different from the trends of hardness values. The effect of irrigant-treated dentin on regulating hDPSCs differentiation might correlate with mechanosensitive signals. Whereas, the hardness changes between groups might not produce significant roles in regulating osteogenic/odontogenic differentiation of stem cells on dentin surfaces.

Effects of sodium hypochlorite on the potential infectivity of human norovirus GII .4 using propidium monoazide with RT‐qPCR and quality assessments in Manila clams ( Ruditapes philippinarum )

This study investigated the effect of NaOCl on the inactivation of human norovirus (HNoV) GII.4 infectivity in Manila clams treated with NaOCl solution at 100-1,000 mg/L chlorine for 10 min using propidium monoazide (PMA)/RT-qPCR. HNoV GII.4 detected using non-PMA/PMA were significantly (p < .05) reduced to 0.15/0.37, 0.33/0.58, 0.59/ 0.85, 0.69/0.99, and 1.05/1.54 log10copy number/μL upon a 10 minute treatment with 100, 300, 500, 700, and 1,000 mg/L of NaOCl, respectively. The D-value of HNoV GII.4 infectivity in non-PMA and PMA-treated samples was calculated as 1,000 and 714.3 mg/L of NaOCl, respectively, using the first-order kinetics model. The pH value increased significantly as chlorine concentration increased. Changes in color and appearance in quantitative analysis evaluation did not appear after treatment with 100-1,000 mg/L chlorine although all sensorial parameters were above 4 points in all NaOCl treatments. The highest score of overall acceptability was observed in 100-500 mg/L chlorine treated Manila clams.

Effects of sodium hypochlorite on corrosion of the rotary nickel-titanium endodontic instruments - SEM analysis

Introduction/Objective. of this study is to use scanning electron microscopy (SEM) analysis to examine surfaces of new and same sets of Ni-Ti instruments after canal preparations, to check their suspectability to corrosion. Methods. In this study, we used five different endodontic Ni-Ti instruments: K3, Mtwo, ProTaper Universal, HyFlex and BioRaCe. Instruments were analyzed before and after preparation of canals of different curvature, using SEM (150?2000 ?). Results. Corrosion of the working part was observed in 5.5% of new Ni-Ti instruments of the K3 system (apical and middle segment), in 5.5% of Mtwo instruments (apical third), and in 11.1% of ProTaper Universal systems (apical and middle third). Corrosion was not observed on the new instruments of the HyFlex and BioRaCe kits. After instrumentation, disinfection, and sterilization, corrosion was observed in all sets of K3 and ProTaper Universal systems and in all HyFlex instruments of the first group. Corrosion was observed in the HyFlex system in the second group in 16.7% of instruments (apical and middle third) and in the third 83.3% in the apical and 66.7% in the middle segment. In the Mtwo set, corrosion was observed in 16.7% of instruments in the first (apical and middle third), in the second group in 33.3% of instruments in the apical part and 50% in the middle third, while in the third group, corrosion was observed in 16.7% of instruments in the middle third of instruments. Conclusion. Rotary Ni-Ti instruments K3 and ProTaper Universal are susceptible to corrosion in a very high percentage. Ni-Ti systems with post-heat treatment of the working part (HyFlex) are somewhat more resistant to corrosion, while in Ni-Ti systems with electropolished surface (BioRaCe), corrosion is not observed.

Treatment of ammonia‑nitrogen wastewater by the ultrasonic strengthened break point chlorination method

Ammonia nitrogen wastewater has the potential to cause significant harm to the environment and human health and cannot be directly discharged, making it a problem in the wastewater industry. A novel ultrasonic strengthened break point chlorination method (USBM) was used to treat ammonia nitrogen wastewater produced by hot galvanized enterprises, and sodium hypochlorite was selected as the oxidizer. The effects of sodium hypochlorite (NaClO) dosage, initial pH, temperature, process time and ultrasonic power on the decomposition of ammonia nitrogen were studied. The experimental results show that under the optimal conditions of 4%, with a temperature of 308 K, an initial pH of 7.8, a process time of 15 min, and a ultrasonic power of 150 W, the ammonia nitrogen removal efficiency could reach more than 80.31% by USBM. Compared with the alone break point chlorination method (ABM), the ammonia nitrogen removal efficiency can increase by 29.86%. A kinetic difference analysis of ABM and USBM was conducted and the result indicated that both processes conform to the pseudo-second-order kinetic model, and the apparent rate constants of ABM and USBM were 0.00035 and 0.00119, respectively. To explore the removal mechanism of ammonia nitrogen, gas chromatography was used to analyze the gas produced in the process. The results show that the produced gas is nitrogen, which is a nontoxic and harmless gas for the environment. The USBM technique has the advantages of a simple process, a high ammonia nitrogen removal rate, low energy loss and is environmentally friendly.

Real-time co-site optical microscopy study on the morphological changes of the dentine’s surface after citric acid and sodium hypochlorite: a single-tooth model

BackgroundTo this day, the effects of sodium hypochlorite and chelating agents on the smear layer and on the dentine’s surface, remain not fully examined. The study is aimed to analyze the dentine's surface treated with 40% citric acid and 5.25% sodium hypochlorite according to two irrigation protocols.Materials and methodsThe study employed a computer-controlled Nikon Eclipse LV100 optical microscope. Ten roots split longitudinally with canals prepared mechanically using the MTwo system to a size of 40/04 were observed. The root halves were divided into two study groups, one half in each of the groups. According to two irrigation protocols, the dentine's surface was irrigated with 40% citric acid and 5.25% sodium hypochlorite, separated with water. Dentine surface was observed in a fixed place and photographed in 500 × magnification after each irrigation stage. The obtained images were then analyzed using computer software (NIS-Elements AR, GIMP-2.6, ImageJ 1.45s).ResultsVarious speed of removal of the smear layer and varied morphological changes of the dentine's surface were observed in both examined groups. Double irrigation of the dentine with citric acid for 30 s resulted in complete removal of the smear layer, and double irrigation for 60 s resulted in increased mean diameter of the dentine tubules (degradation of the dentine's surface).ConclusionsDouble alternating irrigation with citric acid and sodium hypochlorite for 30 s yielded satisfactory results, while double irrigation for 60 s resulted in increased mean diameter of the dentine tubules. A real time assessment of the dentine’s surface changes after both tested solutions can improve understanding of the phenomena that occur during the irrigation and as a result it might help to improve clinical outcomes in root canal treatment.

The Effects of Sodium Hypochlorite on Organic Matters: Influences of Concentration, Renewal Frequency and Contact Area.

Sodium hypochlorite (NaOCl) reacts mainly with proteins and its effectiveness depends on the substances chemical reactivity. It has been reported that volume, concentration, renewal, time, temperature and contact area affect the diffusion of NaOCl in the root canal. However, the relationship between some of these factors is not clear. The purpose of this study was to test the effect of volume, contact area, concentration and renewal frequency of 2.5% and 9.8% NaOCl solutions on their organic matter dissolving-capacity. Pieces of gelatine (18% w/v) with standardized weight, form and structure were either fully or partially exposed to a 2.5% or 9.8% NaOCl solution. In three successive studies, biological dissolution-capacity of NaOCl was tested under different conditions. In experiment 1 the effect of volume/time, in experiment 2 the time/concentration/renewal frequency and in experiment 3 the contact area/renewal frequency/concentration/time of 2.5% or 9.8% NaOCl solutions on dissolving-capacity of organic matter were studied. The weight loss of gelatine pieces over time was registered. The non-parametric tests of Mann-Whitney and Kruskal-Wallis at the 5% threshold were used for statistical analysis. The differences between the two concentrations of NaOCl solution (2.5% and 9.8%) are statistically significant in the effects of different volumes on total dissolution time (P<0.05). Differences in weight loss according to the concentration of the NaOCl solution used (2.5% or 9.8%) were significant after 2 min of contact time (P<0.05). Differences in weight loss between the model and the tube are significant (P<0.05) when the solution is repeated every 30 sec and every 1 min after 2 min of contact. This in vitro study showed that using a more concentrated NaOCl solution would certainly improve the endodontic disinfection, but the biological risk in case of apical extrusion should be considered.

Effects of sodium hypochlorite as dentin deproteinizing agent and aging media on bond strength of two conventional adhesives.

This study evaluated the effect of 10% sodium hypochlorite (NaOCl) as deproteinizing agent and storage media on bond strength (BS) of two etch-and-rinse adhesive systems to dentin. Twenty-eight sound extracted human third molars were divided in four groups (n = 7), according to dentin treatment (conventional etching or etching followed by 10% NaOCl application) and adhesive systems (GB-Gluma 2Bond and OS-One-Step). After dentin treatments and adhesive application, a composite block was built-up on dentin surface and teeth were serially sectioned to obtain bonded sticks specimens. The sticks were submitted to three aging conditions: (24H) 24 hr in water (immediate), (SH) 3 hr of NaOCl accelerated-aging or (1Y) 1 year of water storage. Afterward, submitted to microtensile bond strength test (μTBS), failure modes and adhesive interfaces analyzes. Data were analyzed by two-way analysis of variance (ANOVA) and Tukey's test (α = .05). Dentin deproteinization before bonding significantly reduced μTBS for GB-treated group (p < .05), regardless the aging conditions. Water storage for 1 year (1Y) and NaOCl accelerated-aging (SH) decreased μTBS for both adhesives. Yet, the groups stored in NaOCl (SH) exhibited the lowest BS results (p < .05). Bond strength of deproteinized dentin was dependent on the adhesive system composition and NaOCl accelerated-aging promoted decreased bond strength and further degradation than water storage for 1 year.

Effect of Sodium Hypochlorite and Benzalkonium Chloride on the Structural Parameters of the Biofilms Formed by Ten Salmonella enterica Serotypes.

The influence of the strain on the ability of Salmonella enterica to form biofilms on polystyrene was investigated by confocal laser scanning microscopy. The effects of sodium hypochlorite with 10% active chlorine (SHY; 25,000, 50,000, or 100,000 ppm), and benzalkonium chloride (BZK; 1000, 5000, or 10,000 ppm) on twenty-four-hour-old biofilms was also determined. The biofilms of ten Salmonella enterica isolates from poultry (S. Agona, S. Anatum, S. Enteritidis, S. Hadar, S. Infantis, S. Kentucky, S. Thompson, S. Typhimurium, monophasic variant of S. Typhimurium 1,4,(5),12:i:-, and S. Virchow) were studied. Biofilms produced by S. Anatum, S. Hadar, S. Kentucky, and S. Typhimurium showed a trend to have the largest biovolume and the greatest surface coverage and thickness. The smallest biofilms (P < 0.01) in the observation field (14.2 × 103 µm2) were produced by S. Enteritidis and S. 1,4,(5),12:i:- (average 12.9 × 103 ± 9.3 × 103 µm3) compared to the rest of the serotypes (44.4 × 103 ± 24.7 × 103 µm3). Biovolume and surface coverage decreased after exposure for ten minutes to SHY at 50,000 or 100,000 ppm and to BZK at 5000 or 10,000 ppm. However, the lowest concentrations of disinfectants increased biovolume and surface coverage in biofilms of several strains (markedly so in the case of BZK). The results from this study suggest that the use of biocides at low concentrations could represent a public health risk. Further research studies under practical field conditions should be appropriate to confirm these findings.

Effects of sodium hypochlorite on some physiological and cytogenetical parameters in Allium cepa L. exposed to salt stress

The effects of sodium hypochlorite (NaClO) on the seed germination, seedling growth (radicle length, radicle number and fresh weight), mitotic activity and chromosomal aberrations of Allium cepa L. germinated under salt stress were studied. Salt stress considerably inhibited the seed germination and seedling growth of A. cepa. Furthermore, it markedly reduced the mitotic index in root tip meristems of the seeds and increased the number of chromosomal aberrations. Whereas, the detrimental effects of salt on the seed germination, seedling growth, mitotic activitiy and chromosomal aberrations were dramatically alleviated in varying degrees by NaClO application.

The effects of sodium hypochlorite and ethylenediaminetetraacetic acid on the microhardness of Mineral Trioxide Aggregate and TotalFill Bioceramic Putty.

This study aimed to evaluate the effects of 5% NaOCl and 17% EDTA on the Vickers microhardness of Mineral Trioxide Aggregate, MTA Angelus (MTAA), and TotalFill Bioceramic Putty (BCP) after 24h and 8days. Twenty-four samples of MTAA and BCP were tested for baseline microhardness at 24h. They were divided into four subgroups (5% NaOCl or 17% EDTA, 24h, 5% NaOCl or 17% EDTA at 8days) and microhardness was evaluated at different time-points. Results were recorded and analysed statistically via one-way anova Kruskal-Wallis test and post hoc uncorrected Dunn's multiple comparison test. MTAA had a higher baseline microhardness than BCP. NaOCl reduced the microhardness of MTAA but increased that of BCP. EDTA reduced the microhardness for both materials at 24h. At 8days, EDTA reduced the microhardness of MTAA, while BCP exhibited an increase in microhardness. Enabling these materials to set for a week prior to exposure to irrigants improves their microhardness.

차아염소산나트륨 (NaOCl, Sodium hypochlorite) 에 노출된 굴 (Crassostrea gigas Thunberg, 1793) 의 면역 반응 조사

차아염소산나트륨 (NaOCl, Sodium hypochlorite) 에 노출된 굴 (Crassostrea gigas Thunberg, 1793) 의 면역 반응 조사

Primum non nocere - The effects of sodium hypochlorite on dentin as used in endodontics.

The antimicrobial and tissue-dissolution capacities of NaOCl render it a well-accepted agent for root canal debridement. These highly desirable properties, however, appear to be intertwined with the untoward effect of collagen matrix degradation within mineralized dentin. Because of its small size, the hypochlorite anion is capable of infiltrating mineralized collagen and destroying the collagen fibrils, producing a mineral-rich, collagen sparse ghost mineral matrix with reduced flexural strength. Findings from the present work challenge the biosafety of NaOCl when it is used in high concentrations and for lengthy time periods during root canal treatment, and laid the background work for future biomaterials design in debridement of the canal space.

Effects of different concentrations and exposure time of sodium hypochlorite on the structural, compositional and mechanical properties of human dentin.

This study evaluated the effects of sodium hypochlorite (NaOCl) with different concentrations and exposure time on the structural, compositional and mechanical properties of human dentin in vitro. Sixty dentin slabs were obtained from freshly extracted premolars, randomly distributed into four groups (n=15), and treated with 1%, 5%, 10% NaOCl and distilled water (control group), respectively, for a total of 60 min. Attenuated total reflection infrared (ATR-IR) spectroscopy, Raman spectroscopy and X-ray diffraction (XRD) were carried out before, 10 min and 60 min after the treatment. Scanning electron microscopy (SEM) and flexural strength test were conducted as well. The results showed that dentins experienced morphological alterations in the NaOCl groups, but not in the control group. Two-way repeated-measures analysis of variance revealed that the carbonate:mineral ratio (C:M), Raman relative intensity (RRI), a-axis, c-axis length and full width at half maximum (FWHM) with the increase of time and concentration in the NaOCl groups were not significantly different from those in the control group (P>0.05). Nevertheless, the mineral:matrix ratio (M:M) increased and the flexural strength declined with the increase of concentration and the extension of time in the NaOCl groups (P<0.05). Additionally, it was found that the M:M and the flexural strength remained unchanged after 1% NaOCl treatment (P>0.05), and the morphology changes were unnoticeable within 10 min in 1% NaOCl group. These results indicated that NaOCl has no significant effects on the inorganic mineral of human dentin; but it undermines and eliminates the organic content concentration- and time-dependently, which in turn influences the flexural strength and toughness of dentins. In addition, an irrigation of 1% NaOCl within 10 min can minimize the effects of NaOCl on the structural and mechanical properties of dentin during root canal treatment.

Lifting the biofilm lid on the antibacterial and antibiofilm effects of sodium hypochlorite against Staphylococcus aureus in atopic dermatitis.

Lifting the biofilm lid on the antibacterial and antibiofilm effects of sodium hypochlorite against Staphylococcus aureus in atopic dermatitis.

Effects of disinfectants against norovirus virus-like particles predict norovirus inactivation.

Human noroviruses (NoVs) are a major cause of epidemic and sporadic acute gastroenteritis worldwide. Public and personal hygiene is one of the most important countermeasures for preventing spread of NoV infection. However, no a practicable cell culture system for NoV had been developed, initial tests of the virucidal effectiveness of anti-NoV disinfectants and sanitizers have been performed using surrogate viruses. In this study, NoV virus-like particles (VLPs) were used as a new surrogate for NoVs and a method for evaluating NoV inactivation using them developed. This method is based on morphological changes in VLPs after treatment with sodium hypochlorite. VLP specimens were found to become deformed and degraded in a concentration-dependent manner. Based on these results, the effects of sodium hypochlorite on VLPs were classified into four phases according to morphological changes and number of particles. Using the criteria thus established, the efficacy of ethanol, carbonates and alkali solutions against VLPs was evaluated. Deformation and aggregation of VLPs were observed after treatment with these disinfectants under specific conditions. To determine the degradation mechanism(s), VLPs were examined by SDS-PAGE and immunoblotting after treatment with sodium hypochlorite and ethanol. The band corresponding to the major capsid protein, VP1, was not detected after treatment with sodium hypochlorite at concentrations greater than 500 ppm, but remained after treatment with ethanol. These results suggest that VLPs have excellent potential as a surrogate marker for NoVs and can be used in initial virucidal effectiveness tests to determine the mechanism(s) of chemical agents on NoVs.

Effects of sodium hypochlorite on structural/surface characteristics, filtration performance and fouling behaviors of PVDF membranes

Sodium hypochlorite (NaClO) is a common chemical cleaning agent in water treatment due to its effective removal of irreversible pollutants that cannot be removed by physical cleaning. In this research, PVDF ultrafiltration (UF) membranes were exposed to NaClO for 120–960h, and then the structural/surface characteristics of the control and exposed membranes were compared. Results showed that different exposure doses lead to variations in PVDF membrane structural/surface characteristics according to different mechanisms and further influence the permeability and fouling behaviors of PVDF membranes during ultrafiltration (UF) processes. Reversible fouling was the most common fouling behavior of membranes and exposure to NaClO resulted in higher degree of irreversible fouling. The irreversible flux decline of the membranes could not only caused by physically irremovable fouling but also irreversible compaction of the membranes due to the formation of macro-voids in the membrane skin layer. The changes in membrane mechanical properties caused a sharp increase in the inherent membrane resistance (Rm) which was generally considered as a constant. ‘Dynamic resistance mechanism’ could be proposed based on the impact of different NaClO exposure doses on the total membrane resistance (Rt ), which is the sum of the changes in Rm and the variation of fouling resistance (Rf) resulted from the changes in surface characteristics of the membranes. According to this concept, the filtration operating conditions and cleaning processes could be optimized in water treatment.

The effects of sodium hypochlorite against selected drinking water-isolated bacteria in planktonic and sessile states

Chlorine is the most commonly used agent for general disinfection, particularly for microbial growth control in drinking water distribution systems. The goals of this study were to understand the effects of chlorine, as sodium hypochlorite (NaOCl), on bacterial membrane physicochemical properties (surface charge, surface tension and hydrophobicity) and on motility of two emerging pathogens isolated from drinking water, Acinetobacter calcoaceticus and Stenotrophomonas maltophilia. The effects of NaOCl on the control of single and dual-species monolayer adhered bacteria (2h incubation) and biofilms (24h incubation) was also assessed. NaOCl caused significant changes on the surface hydrophobicity and motility of A. calcoaceticus, but not of S. maltophilia. Planktonic and sessile S. maltophilia were significantly more resistant to NaOCl than A. calcoaceticus. Monolayer adhered co-cultures of A. calcoaceticus-S. maltophilia were more resilient than the single species. Oppositely, dual species biofilms were more susceptible to NaOCl than their single species counterparts. In general, biofilm removal and killing demonstrated to be distinct phenomena: total bacterial viability reduction was achieved even if NaOCl at the higher concentrations had a reduced removal efficacy, allowing biofilm reseed. In conclusion, understanding the antimicrobial susceptibility of microorganisms to NaOCl can contribute to the design of effective biofilm control strategies targeting key microorganisms, such as S. maltophilia, and guarantying safe and high-quality drinking water. Moreover, the results reinforce that biofilms should be regarded as chronic contaminants of drinking water distribution systems and accurate methods are needed to quantify their presence as well as strategies complementary/alternative to NaOCl are required to effectively control the microbiological quality of drinking water.

Sodium Hypochlorite Dental Emergency-A Review

Root canal irrigants play an important role in the success of root canal treatment. As the major objective in root canal treatment is to disinfect the entire root canal system, it requires that the pulpal contents be eliminated as sources of infection. This goal may be accomplished using mechanical instrumentation and chemical irrigation, in conjunction with medication of the root canal between treatment sessions. However there remains a risk of extrusion of these irrigants beyond or into the surrounding tissues and cause severe complications. Sodium hypochlorite is widely used in dental practice during root canal treatment. Although generally regarded as being highly effective as an irrigant, potentially severe complications occur when it comes in contact with soft tissues. This article discusses the use, inadvertent effects of sodium hypochlorite and considers the appropriate management for a practitioner when faced with potential adverse incident with this agent.

In vitro seed germination and seedling growth of Calanthe discolor Lindl

Abstract We investigated the effects of sodium hypochlorite (NaOCl) and culture medium on embryo swelling and germination of Calanthe discolor Lindl., and established a method for determining the swelling and protocorm formation of C. discolor seeds via in vitro examination of immature seeds. Treatment of immature seeds with NaOCl greatly enhanced the extent of embryo swelling and protocorm formation of immature zygote embryos compared to seeds without NaOCl treatment. The effects of the culture media were also evaluated with regard to embryo swelling and protocorm formation of in vitro cultured seeds with and without NaOCl treatment. Additionally, the effects of white fluorescent light and red and blue LED lights on seedling growth in in vitro culture were examined. The most suitable condition for seedling growth after 12 weeks of culture was the red LED light with POM medium. These results show effective asymbiotic germination and growth of C. discolor.