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Indoor mold due to water damage causes serious human respiratory disorders, and the remediation to homes, schools, and businesses is a major expense. Prevention of mold infestation of building materials would reduce health problems and building remediation costs. Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit yeasts and a limited number of filamentous fungi. The purpose of this research was to determine the possible inhibitory activity of nonsteroidal anti-inflammatory drugs (NSAIDs) on germination, fungal growth, and reproduction of Chaetomium globosum and other important filamentous fungi that occur in water-damaged buildings. Several NSAIDs were found to inhibit C. globosum germination, growth, and reproduction. The most effective NSAIDs inhibiting C. globosum were ibuprofen, diflunisal, and diclofenac. Fusarium oxysporum, Fusarium solani, Aspergillus niger, and Stachybotrys atra were also tested on the various media with similar results obtained. However, F. oxysporum and A. niger exhibited a higher level of resistance to aspirin and NaSAL when compared to the C. globosum isolates. The inhibition exhibited by NSAIDs was variable depending on growth media and stage of fungal development. These compounds have a great potential of inhibiting fungal growth on building materials such as gypsum board. Formulations of sprays or building materials with NSAID-like chemical treatments may hold promise in reducing mold in homes and buildings.

Chaetomium globosum Kunze:Fr is a dermatophytic, dematiaceous fungus that is ubiquitous in soils, grows readily on cellulolytic materials, and is commonly found on water-damaged building materials. Chlorate affects nitrogen metabolism in fungi and is used to study compatibility among anamorphic fungi by inducing nit mutants. The effect of chlorate toxicity on C. globosum was investigated by amending a modified malt extract agar (MEA), oat agar, and carboxymethyl cellulose agar (CMC) with various levels of potassium chlorate (KClO(3)). C. globosum perithecia production was almost completely inhibited (90-100 %) at low levels of KClO(3) (0.1 mM) in amended MEA. Inhibition of perithecia production was also observed on oat agar and CMC at 1 and 10 mM, respectively. However, hyphal growth in MEA was only inhibited 20 % by 0.1-100 mM KClO(3) concentrations. Hyphal growth was never completely inhibited at the highest levels tested (200 mM). Higher levels of KClO(3) were needed on gypsum board to inhibit perithecia synthesis. In additional experiments, KClO(3) did not inhibit C. globosum, Fusarium oxysporum, Aspergillus niger, Penicillum expansum, and airborne fungal spore germination. The various fungal spores were not inhibited by KClO(3) at 1-100 mM levels. These results suggest that C. globosum perithecia synthesis is more sensitive to chlorate toxicity than are hyphal growth and spore germination. This research provides basic information that furthers our understanding about perithecia formation and may help in developing control methods for fungal growth on building materials.

Greenhouse experiments were conducted to determine disease progression of phytophthora root rot on nonwounded and wounded pepper plants (Capsicum annuum) and to determine whether susceptibility to Phytophthora capsici decreases with wound aging. Two isolates of P. capsici were used in this study, one less aggressive than the other. Trimming the roots immediately prior to inoculation with either isolate increased susceptibility significantly (P ≤ 0·05) compared with plant roots that were not trimmed. Both isolates caused a higher level of disease severity on disturbed/trimmed than on disturbed/nontrimmed roots. Disease also occurred earlier with the more aggressive isolate on both wounded and nonwounded roots. Disease severity was three to four times more severe on plants treated with the aggressive isolate (NM6011) than on those inoculated with the less aggressive isolate (NM6040), regardless of root treatment. In separate experiments, pepper roots were wounded and allowed to age for up to 5 days before inoculation. Resistance to P. capsici increased as the wounds aged, resulting in significantly (P ≤ 0·001) lower disease severity on plants with 3- and 5-day-old wounds than on those inoculated at the time of wounding and the nondisturbed/nontrimmed controls. Wounding of the roots followed by immediate inoculation with zoospores resulted in significantly higher levels of attachment than when roots were inoculated with zoospores 48 h after wounding. The 48-h postwounding inoculation treatment showed the same amount of zoospore attachment as nonwounded roots. Increase in plant resistance correlated (P ≤ 0·01) with an increase in total peroxidase activity. Isoelectric focusing-polyacrylamide gel electrophoresis (IEF-PAGE) indicated increased band intensity of three acidic and one basic isozyme as wounds aged. These data suggest that wound repair plays a role in decreasing infection and resultant disease symptoms of pepper to P. capsici.

Fusarium solani is one of the more important fungal pathogens involved in pre- and post-harvest decay of muskmelon fruit. Production of polygalacturonase (PG), by F. solani was studied in vitro and in vivo . The fungus produced at least 14 PG isozymes with pIs of 4.5 to 9.5 in shake culture using pectin as the sole carbon source. When glucose and pectin were used in combination as the carbon source, total PG activity decreased substantially as compared to pectin alone, suggesting that glucose may suppress PG production in vitro . Only one PG isozyme, designated as PG1, was detected in extracts from infected fruit tissue. PG1 from decayed fruit was purified to homogeneity by protein extraction, ultrafiltration, gel filtration chromatography, and cation exchange chromatography. The molecular weight of PG1 was estimated at 38 kDa based on SDS-PAGE with a pI of 9.5 according to IEF-PAGE. PG1 exhibited only endo-PG activity based on viscosity reduction and thin layer chromatography analysis of products released by enzymatic action. The optimum pH for PG1 activity was 6. The K m and V max of PG1 using polygalacturonic acid as the substrate were 1.34 mg ml -1 and 0.30 unit μg protein -1 , respectively. PG1 effectively macerated fruit tissue which suggests that it may play an important role in decay of muskmelon fruit caused by F. solani .

Phomopsis cucurbitae is a latent infecting pathogen that infects unripe muskmelon fruit, but causes decay after harvest. This fungus causes severe losses during muskmelon fruit storage and marketing in the U.S., Japan, and some Central American countries. Previous studies showed that the fungus produced the cell wall-degrading enzyme polygalacturonase (PG) in both culture and muskmelon fruit tissue. The role of P. cucurbitae PG in the fruit decay process and its relation to latent infection is not well-understood. A prominent PG isozyme produced by the fungus in decayed fruit was purified to homogeneity by a sequence of extraction, ultrafiltration, preparative isoelectric focusing, anion exchange, and gel filtration chromatography. This isozyme exhibited endo-activity, a molecular weight of 54 kDa according to SDS-PAGE, and a pI of 4.2 based on IEF-PAGE. Isozyme activity was optimal at 40–45°C and pH 5.0. It had a Km of 44.7 g/ml and a Vmax of 0.313. The purified isozyme also effectively macerated mature muskmelon fruit tissues. This isozyme was the most prominent of the PG isozymes produced by P. cucurbitae in decaying fruit, and may play an important role in postharvest decay.

Activity of β-galactosidase (EC 3.2.1.23) and isozymes of the New Mexican type pepper were assayed in ripening fruit and characterised in order to further understand the possible role this enzyme plays in fruit ripening. Total β-galactosidase activity increased in New Mexican type peppers (NM chile) as they ripened. To a lesser extent, xylosidase and β-glucosidase increased from the mature green stage to the turning stage of ripening and then decreased in the red ripe stage. Cation exchange chromatography indicated two prominent β-galactosidase peaks depending on stage of ripeness. In mature NM green chile, one large anionic, and in NM red chile, one large cationic peak was found. Isoelectric focusing-polyacrylamide gel electrophoresis (IEF-PAGE) separated one isozyme from mature green NM chile (pI 4.6) and an additional β-galactosidase isozyme as NM chile turned from green to red (pI 8.0). The cationic β-galactosidase isozyme from red NM chile extract had an apparent native-molecular weight of 51 kDa. The optimum pH for β-galactosidase activity in both red and green NM chile was between 3 and 4.5. Stable enzyme activity was observed from 25 to 50°C. β-Galactosidase from red NM chile had a K m of 0.87 mM and V max of 12.3 nKat, while the values for green NM chile β-galactosidase were 2.5 mM and 8.3 nKat. The heavy metal ions Ag 2+ and Hg 2+ inhibited β-galactosidase activity by 100% and 90%, respectively.

Peroxidase activity is used by the food processing industry as an indi- cator of adequacy of the blanching process and to predict off-flavour develop- ment in frozen food. Research reported herein showed that total peroxidase activity did not parallel flavour changes in frozen unblanched supersweet (sh2) or sugary enhanced (sullse) sweet corn genotypes. Frozen corn-on-the-cob of 'Florida Staysweet' (sh2), 'Merit' (standard sweet-sul) and 'Bodacious' (sullse), blanched and unblanched, was subjected to sensory evaluation and peroxidase analysis following frozen storage up to 12 months. Trained taste panellists rated the unblanched sh2 and sullse corn as acceptable up to 8 months of frozen storage. Kernels were cut from cobs after 0 and 12 months of storage. Proteins were extracted from acetone powders of kernels and separated by isoelectric focusing and native PAGE. Banding patterns differed according to genotype and storage duration. These results suggested molecular differences in peroxidase iso- zymes among the sweet corn genotypes which could be involved in off-flavour development. All genotypes contained a peroxidase isozyme having a molecular mass of 80 kDa and PI of 4-5. The sullse and sh2 genotypes produced an addi- tional peroxidase band of 13.8 kDa. An additional peroxidase isozyme (PI 5.4) appeared in extracts from the sul genotype after 12 months storage. Although changes in total peroxidase activity may not predict flavour changes in all geno- types, the presence or absence of certain peroxidase isozymes may be useful in predicting off-flavour development in sul frozen corn.

New Mexican Chile peppers (Capsicum annuum L. `New Mexico 6-4') were harvested at weekly intervals beginning 20 days after flowering (DAF), and were evaluated for ethylene (C2 H4) production, respiration rates, chlorophyll content, degradative enzyme activity (cellulase, polygalacturonase, ß-galactosidase), and fruit firmness. Morphological and physiological changes were most apparent in peppers harvested 54 to 69 DAF. ß-galactosidase activity increased rapidly beginning 54 DAF and reached a peak by 89 DAF. Fruit firmness was highest (36 newtons) at 54 DAF and had decreased significantly by 69 DAF. Carbon dioxide production and chlorophyll content were highest in young pods harvested 20 DAF and decreased steadily thereafter. A climacteric increase of CO, was absent. There were two peaks in C2 H4 production: one associated with rapid fruit growth and the other with color change (61 to 69 DAF). Fruit harvested on the same day but at different developmental stages (green to red) were similar to those observed in fruit harvested over the season for the physiological characteristics tested. Separation of pepper fruit soluble proteins on SDS-PAGE demonstrated increased intensity in protein bands at 27, 35, and 40 kDa and decreased intensity of 51 kDa band as the fruit matured. Several biochemical processes appeared to be enhanced in Chile pepper fruit from 47 to 69 DAF.

Field experiments were conducted to test the effects of alternate-row irrigation, every-row irrigation, and chemical treatments with alternate-row irrigation on yield of chile (chilli) peppers ( Capsicum annuum ) and disease incidence of chile pepper root rot caused by Phytophthora capsici . Field trials were conducted in 1989 and 1990 near Las Cruces, New Mexico. In 1989, chemical treatments were Telone C-17, metalaxyl, copper sulphate and metam-sodium; in 1990, chemical treatments were copper sulphate and metam-sodium. Disease incidence in plots receiving alternate-row irrigation without fungicides was significantly less than irrigation of every row without fungicides in 1989 and 1990. Disease incidence was also significantly less in the metam-sodium and metalaxyl plots than the every-row irrigated plots in 1989, but not significantly less than control plots irrigated with alternate rows without chemicals. Yield of red chile was significantly higher in plots treated wth metam-sodium and given alternate-row irrigation than all other treatments in 1989. Yield of green chile pepper from plots given alternate-row irrigation was significantly higher than plots given every-row irrigation in 1990. Metam-sodium with alternate-row irrigation was as good as, or better than, alternate-row irrigation alone in reducing disease incidence of P. capsici and in increasing yield.

An isolate (87) of the yeast Pichia guilliermondii, protects apples from postharvest fruit rotting fungi Botrytis cinerea and Penicillium expansum. In order to examine the yeast-pathogen interaction, B. cinerea was grown on agar plates overlayed with cellophane. Effective and non-effective yeast isolates were applied near the young hyphal growth. Samples were taken 24 h later from the area where the fungi and yeast had intersected. Light microscopy revealed a general attachment of the effective biocontrol agent P. guilliermondii (isolate 87) and a non-effective isolate (117) of Debaryomyces hansenii. Low temperature scanning electron microscopy (LTSEM) indicated that both species of yeast attached to the fungal hyphae, but the 87 isolate attached fastidiously. Twenty-four hours after applying the 87 isolate to B. cinerea, pitting and collapse of the hyphae were observed. These observations were confirmed using transmission electron microscopy. These features were not observed with the ineffective isolate of D. hansenii. Further experiments indicated that attachment of P. guilliermondii to hyphae of B. cinerea could be blocked by agents that alter protein integrity (salts, proteases, etc.) and certain sugars. Isolates of both species produced β-(1–3) glucanase when cultured in various carbon sources and on cell walls of fruit rotting pathogens. Culture supernatants from P. guilliermondii, however, yielded two- to five-fold more β-(1–3) glucanase activity compared with D. hansenii. Data indicate that tenacious attachment, along with secretion of cell wall degrading enzymes, may play a role in the biocontrol activity of this yeast antagonist.

Changes in cellulase, polygalacturonase and total protein were measured during the growth and ripening of three apple cultivars (Malus sylvestris Mill ‘Stark Lodi’, ‘Paulared’ and ‘Golden Delicious’) with the view of establishing the role of these cell wall-degrading enzymes in fruit development. In agreement with earlier work, no endo-polygalacturonase activity was evident at any stage of apple development. Cellulase was present in young expanding fruit and decreased in activity as the fruit reached full size and ripened. Separation of cellulase on MonoQ columns indicated the presence of three isoforms, two of which had molecular weights of 67 kDa. Electrophoresis of proteins from developing fruit indicated that proteins with molecular weights of 24, 26 and 67 kDa increased during ripening. Unlike other climacteric fruit, ripening in apples was not associated with an increase in extractable cellulase or polygalacturonase. The sodium dodecylsulfate-polyacrylamide gel electrophoresis studies suggest that a 67 kDa protein increased during ripening. The decrease in enzyme activity may be due to the inability of available extraction techniques to release this enzyme from apple cell walls.

Xylem sap from apple (Malus domestica Borkh), peach (Prunus persica Batsch), and pear (Pyrus communis L.) twigs was collected by means of pressure extrusion. This sap contained a number of acidic peroxidases and other proteins. Two other sources of xylem sap used in this study were stem exudates and guttation fluid. Similar peroxidases were also found in stem exudates and guttation fluids of strawberry (Fragaria x ananassa Duch.), tomato (Lycopersicum esculentum L.), and cucumber (Cucumis sativus L.). Isoelectric focusing activity gels showed that two peroxidases (isoelectric point [pl] 9 and pl 4.6) were present in initial stem exudates collected in the first 30 minutes after excision. Subsequent samples of stem exudate collected contained only the pl 4.6 isozyme. The pl 4.6 peroxidase isozyme was also found in root tissue and guttation fluid. These observations suggest that roots produce and secrete the pl 4.6 peroxidase into xylem sap. Cucumber seedlings were treated with 100 microliters per liter ethylene for 16 hours and the exudate from decapitated hypocotyl stumps was collected over a 3 hour period. Ethylene increased the peroxidase activity of stem exudates and inhibited the amount of exudate released. These observations suggest that xylem sap peroxidase may play a role in plugging damaged vascular tissue.

Developing peach ( Prunus persica L. Batsch `Redskin9) fruit were used to characterize the role of peroxidases in lignification. During development, the endocarp of these drupes becomes lignified while the mesocarp remains parenchymatous. Acidic peroxidase from lignifying endocarp were similar to those of the fleshy mesocarp. The endocarp had a larger amount and number of basic peroxidases than the mesocarp. Cultured peach leaf cells are thought to be lignified because their walls give a positive reaction with phloroglucinol-HCI. These cells also secreted a basic peroxidase. Peroxidases were difficult to extract from endocarp tissue as they lignified. This was also demonstrated by tissue printing on nitrocellulose. Flesh, but not endocarp peroxidase was evident in tissue prints. This suggests that tissue printing may fail to reveal the presence of enzymes which are firmly attached to the cell.

Production of polygalacturonase (PG), a cell wall-degrading enzyme, by Phomopsis cucurbitae (latent infection fungus) was studied in relation to different carbon sources and various stages of cantaloupe fruit development. P. cucurbitae produced multiple PG isozymes both in vitro and in vivo. The fungus produced the highest PG activity and the greatest number of isozymes on pectin compared with those produced on glucose, galactose, and sucrose. Eight P. cucurbitae PG isozymes (pIs 3.7, 4.2, 6.6, 7.0, 7.3, 7.5, 7.8, and 8.6) were detected in extract from inoculated mature fruit (40 days after anthesis) by isoelectric focusing. Isozyme bands with pIs of 4.2, 7.3, and 7.8 were the most prominent. A similar set of PG isozymes was produced by P. cucurbitae in autoclaved mature fruit tissue (mesocarp). When tissue discs taken from 20-, 30-, 40-, and 50-day postanthesis fruit were inoculated with P. cucurbitae, PG activity and the number of PG isozymes extracted from the macerated fruit tissue discs increased with the degree of fruit maturity and ripening. Increases in PG activity and PG isozymes were also correlated with reactivation of latent infections and the beginning of tissue maceration. An anionic PG isozyme (pI 4.2) was only visualized on decayed 50-day-old fruit exocarp, as well as 40- and 50-day-old fruit mesocarp. The experimental results support the hypotheses that P. cucurbitae PG isozymes play an important role in fruit decay once latent infection becomes active following harvest.

Cucumber (Cucumis sativus L.) seed require soil temperature to be around 20C for efficient germination. This hinders early planting in cool soils. This study was conducted to determine how germinating seed of cucumber cultivars Earlipik 14 and Arkansas Little Leaf at 13.9, 15.6, and 20C in the dark affected protein formation. Seed were removed from moist chambers at 0, 12, 24, 48, 72, 84, 96, 120, and 168 h. Germination, defined as the radicle being at least 5 mm long, was determined at each time. Germinated and ungerminated seed was prepared for polyacrylamide gel electrophoresis. At 20C, 90% to 100% of seed had germinated by 48 h. At 15.6C, 20% to 50% of seed germinated by 168 h, and at 13.9C, ≈2% of seed had germinated by 168 h. For seed incubated at 20C, concentrations of proteins at 70.1 kDa decreased, while those at 37.4, 43.4, and 50 kDa increased after 24 h, which corresponded to formation and elongation of the majority of radicles. These changes were expressed later for seed germinated at 15.6 and 13.9C. Identification of the proteins is being attempted. The importance of these proteins in germination and early development will be discussed.

Increased peroxidase activity is used to predict development of off-flavor in frozen sweet corn. However, peroxidase activity was not indicative of flavor changes in frozen supersweet (sh2) or sugar enhanced (sul/se) sweet corn genotypes. These results suggested an inactivation or absence of certain peroxidase isozymes. Frozen `Florida Staysweet' (sh2), `Merit' (sul), and `Bodacious' (sul/se) kernels were cut from cobs after 0 and 12 months of storage. Proteins extracted from acetone powders were separated by isoelectric focusing (IEF) and Native-PAGE. Banding patterns differed according to cultivar and storage duration. All cultivars contained a peroxidase isozyme having a molecular weight of 99 kD and pI of 4.5. The sul/se and su2 cultivars expressed an additional peroxidase band of 17.9 kD. An additional peroxidase isozyme (pI 5.0) appeared after 12 months of storage in the sul cultivar. This isozyme did not appear in sul/se or sh2 and is a possible marker for predicting off-flavor in corn. This isozyme may also catalyze off-flavor reactions in sul corn genotypes. Although changes in total peroxidase activity may not predict flavor loss in all genotypes, certain peroxidase isozymes may be useful in predicting and catalyzing off-flavor reactions in sul corn cultivars.

Experiments were conducted to determine the relationship between pepper fruit susceptibility to Phytophthora fruit rot and fruit maturity. Pepper fruits were harvested weekly throughout the 1991 growing season and were dipped in a suspension of Phytophthora capsici zoospores (5×10 3 zoosporesl/ml). The susceptibility of nonwounded fruit to infection decreased with increased ripening. Lesion elongation in nonwounded fruit decreased from 14.1 mml/day in green fruit to 10.7 mm/day in nondehydrated red fruit. Nonwounded fruit (immature green, mature green, and red) inoculated with 5×10 3 zoospores in a 100-μl drop also exhibited less infection as the fruit ripened [...]

New Mexican chile peppers were harvested at weekly intervals beginning 105 days after planting (DAP), and evaluated for ethylene (C2H4) production, respiration rates, chlorophyll content, beta-galactosidase activity, polygalacturonase (PG) activity, and fruit firmness. Physiological changes were most apparent in peppers harvested 139-154 DAP. Beta-galactosidase activity increased rapidly beginning 147 DAP, and reached a peak of 24.5 mmol·gfw-1 when peppers were harvested 160 DAP. Polygalacturonase was not detectable at any stage of maturation. Fruit firmness was greatest (35.8 N) at 139 DAP and decreased significantly at 160 DAP. Carbon dioxide production and chlorophyll content were highest in young pods harvested 105 DAP, and decreased steadily thereafter. Ethylene production peaked (0.185-0.202 nl·gfw-1·h-1) in peppers harvested between 146-154 DAP.

Ethylene enhanced the levels of peroxidases in the roots, stems, leaves, and cotyledons of 2-week-old cucumber Cucumis sativus cv Poinsett 76 seedlings. Antibodies to the isoelectric point (pl) 9 and pl 4 isoenzymes were used in a radial immuno-diffusion assay to demonstrate that ethylene induced similar peroxidases in other cultivars of C. sativus, other species of Cucumis and other genera of Cucurbitaceae. Examination of ethylene-induced peroxidases, using isoelectric focusing gels, demonstrated the presence of a series of other peroxidases, mostly slightly acidic, whose isoelectric focusing pH was approximately 6. These pl 6 peroxidases were partially purified on a cation exchange column. Ouchterlony double diffusion gels indicated that these proteins cross-reacted with antibodies to both the pl 9 and pl 4 peroxidase. The data presented here suggest that the induction of peroxidase isoenzymes during ethylene-induced senescence is a common response in this family of plants. In addition, antibody and isoelectric focusing studies indicate that both acidic and basic peroxidase are highly conserved in members of this family.