In Vitro Selection of Turmeric Somaclone Resistant to Fusarium oxysporum f.sp. Zingiberi

Fusarium wilt disease tolerant clones of Curcuma longa L. cv. Suroma were isolated using continuous in vitro selection technique against pure culture filtrate of Fusarium oxysporum f.sp.zingiberi. In vitro shoot buds were challenged with pure culture filtrate of F. oxysporoum to isolate viable plants within 30 days of culture, which were further subjected to pure culture filtrate treatment. After four cycles of treatment, 15 cell lines, tolerant to culture filtrate were isolated through continuous in vitro selection and subcultured on Murashige and Skoog basal medium with Benzylaminopurine (3 mg/l) along with the control plants. Both tolerant and control plants were screened for disease tolerance by adopting in vitro sick plot technique. The data obtained from this experiment revealed a ratio of 188:68 tolerant: susceptible in vitro clones retrieved from tolerant plants. However, plants from T1a₁ and control were susceptible under in vitro sick plot technique. The Fusarium wilt disease tolerant clones were hardened and established in soil with 90 % survival frequency.     

Evaluating the Impact of Trichoderma brevicompactum 31636 on Root Rot of Atractylodes macrocephala and the Fungal Community in the Rhizosphere Soil

BackgroundTrichoderma brevicompactum can control of plant diseases, but there are few studies about the control of root rot by T. brevicompactum 31636 and its biocontrol mechanism.ObjectiveTo explore the impact of T. brevicompactum 31636 on root rot and the structure of fungal communities of rhizosphere soil.MethodsThe effect of T. brevicompactum 31636 on controlling root rot was assessed, the growth of plants was observed by pot experiments, and the fungal structure was detected by high-throughput sequencing analysis.ResultsCompared with the CK (healthy plants) group, the plant height, fresh weight, and dry weight increased by 55.3%, 48.55% and 57.8%, respectively. While the concentration of atractylenolide I, II, III, and atractylone increased by 19.35%, 16.96%, 89.42% and 89.32%, respectively. The results of pot experiments indicated that T. brevicompactum 31636 could significantly improve the survival rate of Atractylodes macrocephala. The results of principal coordinates analysis (PCoA) showed that the sample of CK and Fo+Th (adding Fusdrium oxysporum before T. brevicompactum 31636) group on one side, and samples of Th and Fo group on the other. Furthermore, among the top 20 dominant fungi detected, the fungal genus with 1% increase of relative abundance included Trichoderma (1%), Sordariomycetes (1.5%), Mortierella (1.3%), Penicillium (1%), meanwhile the fungal genus with 1% decrease of relative abundance included Fusarium (30%) and Pleosporales (1%).ConclusionIn summary, T. brevicompactum 31636 can control the root rot of A. macrocephala by regulating of the soil fungal community and inhibiting pathogen.     

Effects of an additive (hydroxyapatite–bentonite–biochar) on Cd and Pb stabilization and microbial community composition in contaminated vegetable soil

A two-year pot experiment was conducted with a pimiento–celery cabbage (Capsicum annuum L.–Brassica pekinensis) rotation in acidic soil contaminated with Cd and Pb, which was amended with 0.0, 1.0, 2.5, 5.0 and 10.0% (w/w) premixtures of hydroxyapatite, bentonite and biochar combinations (HTB, in a ratio of 1 : 2 : 2). The results showed that the application of HTB at 2.5–10.0% significantly increased soil pH and organic carbon by an average of 10.38–17.60% and 35.60–55.34% during the two years, respectively. Compared to the control treatment, 1.0–10.0% HTB decreased the available Cd and Pb concentrations by 40.92–77.53% and 41.60–82.79% on average, respectively. In addition, the diversity and richness of the soil bacterial community improved after the two-year application of HTB. The relative abundances of Acidobacteria, Bacteroidetes and Chloroflexi increased under the HTB treatments, while those of Proteobacteria and Actinobacteria decreased. Redundancy analysis (RDA) and regression analysis indicated that soil pH and Cd and Pb availability were important factors shaping the soil bacterial community. The Cd and Pb concentrations in the edible parts of pimiento and celery cabbage decreased as the HTB application rate increased and met the Food Quality Standard in each season when the HTB application rate was 5.0% or higher. Higher rates of HTB (5.0% and 10.0%) not only ensured the quality of vegetables, but also significantly promoted pimiento and celery cabbage growth. Overall, these results indicated that the application of HTB, especially at a rate of 5.0%, could be an effective way to immobilize Cd and Pb, improve soil quality and ensure vegetables produced in acidic contaminated soil are safe for human consumption.

A two-year pot experiment was conducted with an amendment of hydroxyapatite, bentonite and biochar combinations (HTB) to study the influence of HTB on soil physicochemical properties, soil Cd and Pb availability and microbial community composition.     

Combined use of the biocontrol bacterium Pseudomonas fluorescens strain LRB3W1 with reduced fungicide application for the control of tomato Fusarium wilt

Pseudomonas fluorescens strain LRB3W1 inhibited the mycelial growth of Fusarium oxysporum f. sp. lycopersici and suppressed the Fusarium wilt of tomato. The chemical fungicide, benomyl, did not suppress the disease incidence at low concentrations. However, the disease incidence was decreased by the combined application of benomyl at low concentrations with strain LRB3W1. Combined application of benomyl with the bacterium was more effective than treatment with the bacterium alone. The survival of strain LRB3W1 was not influenced by the presence of benomyl. This combined use of the biocontrol bacterium, strain LRB3W1, and a fungicide, benomyl, should be an attractive approach for suppressing tomato wilt.     

EFFECT OF AMMONIUM NITRATE AND LIME ON GERMINATION AND EARLY GROWTH OF PINUS CARIBAEA

Depressive effects of nitrogen (N) salts on germination and survival of Caribbean pine (Pinus caribaea Mor.) were investigated in the glasshouse by following changes in mineral soil N after the application of lime and fertilizers. The effect of partial soil sterilization was also studied.

Immobilization and subsequent nitrification of ammonium nitrate was greatly stimulated by liming. Germination and survival declined as the soil nitrate level was raised, apparently because of increased pre—and post-emergence damping-off by Fusarium spp.

Seedling yield at 20 weeks was reduced by high soil nitrate levels, but this effect was reversed at 31 weeks except for the highest level of nitrate.

The percentage of total N in seedling tissues was closely correlated with the concentration of soil nitrate.

The effects of partial soil sterilization on plant behaviour were slight compared to those of ammonium nitrate and lime.     

Fusarium species causing invasive fungal pneumonia in an immunocompetent patient: a case report

Fusarium is a large genus of filamentous fungi that are rarely associated with disease in humans. In the clinical setting, Fusarium species are often difficult to distinguish from other fungal organisms, particularly Aspergillus species. Invasive fungal pneumonia caused by Fusarium species has rarely been reported, especially in immunocompetent patients. In this study, we reported a case of invasive Fusarium pneumonia in a previously healthy 68-year-old woman. The disease was initially misdiagnosed as invasive Aspergillus pneumonia because of the similarity in radiologic and histopathologic findings between these conditions. After Fusarium was identified via microbiological analysis, the antifungal agent was changed, and the patient recovered fully.     

Various antibacterial mechanisms of biosynthesized copper oxide nanoparticles against soilborne Ralstonia solanacearum

The substantial antimicrobial efficacy of nanoparticles against phytopathogens has been extensively investigated for advanced agricultural applications. However, few reports have focused on soilborne pathogenic bacteria. The aim of this study was to obtain sustainably synthesized copper oxide nanoparticles (CuONPs) using papaya leaf extracts and investigate the bactericidal activity of these CuONPs against Ralstonia solanacearum, the cause of bacterial wilt, under laboratory and greenhouse conditions. The results showed that CuONPs possessed strong antibacterial activity and that all R. solanacearum were killed after exposure to 250 μg mL⁻¹ CuONPs. CuONPs could interact with bacterial cells to prevent biofilm formation, reduce swarming motility and disturb ATP production. Ultrastructural observations by transmission electron microscopy (TEM) revealed that after interactions with CuONPs, bacterial cells suffered significantly from nanomechanical damage to the cytomembrane, accompanied by the absorption of multiple nanoparticles. In addition, molecular studies identified the downregulation mechanism of a series of genes involving pathogenesis and motility. The control efficiency of CuONPs in tobacco bacterial wilt disease management under greenhouse conditions was verified by root irrigation application, demonstrating that as-prepared CuONPs significantly reduced the disease occurrence and disease index. Our studies focused on developing biosynthesized nanoparticles as a biocompatible alternative for soilborne disease management.

Green synthesized CuONPs disturb cell metabolism, biofilm formation, physical motility and gene expression in Ralstonia solanacearum, thereby effectively controlling bacterial wilt.     

Effects of Dextranases on Attachment of Streptococcus mutans to Hydroxyapatite

A Fusarium dextranase and a Penicillium dextranase were compared for their relative ability to quantitatively reduce the adsorption of ³H-labeled Steptococcus mutans cells onto hydroxyapatite. Fusarium dextranase-treated hydroxyapatite disks caused a statistically significant decrease in the hydroxyapatite adsorption of both the OMZ 176 and NCTC 10449 strains of S. mutans relative to untreated control disks. The extent of initial bacterial adsorption was not promoted by sucrose-dependent glucan synthesis. Since the Fusarium dextranase has a much greater affinity for hydroxyapatite than the Penicillium dextranase, it could represent an enzyme with improved decay-preventive therapeutic properties. This was concluded because the Fusarium dextranase may interfere with both the initial attachment and later glucan-dependent accumulation of dental plaque microorganisms.     

Co-inoculation of an antibiotic-producing bacterium and a lytic enzyme-producing bacterium for the biocontrol of tomato wilt caused by Fusarium oxysporum f. sp. lycopersici

The antifungal compound 2,4-diacetylphloroglucinol-producing bacterium, Pseudomonas fluorescens strain LRB3W1, inhibits the growth of Fusarium oxysporum f. sp. lycopersici, and controls Fusarium wilt of tomato caused by F. oxysporum f. sp. lycopersici. On the other hand, Serratia marcescens strain B2, which produces cell wall-degrading enzyme chitinases, did not inhibit fungal growth and the suppressive effect of strain B2 against tomato Fusarium wilt was less than that of strain LRB3W1. Combined inoculation of strain LRB3W1 with strain B2 was more effective than treatment with strain LRB3W1 alone. When 2,4-diacetylphloroglucinol and the chitinolytic enzymes were applied in combination, a synergistic inhibitory effect against the pathogen was observed. It was possible that bacteria which produce cell wall-degrading enzymes enhanced the biocontrol effect of the antibiotic-producing bacterium against tomato Fusarium wilt.     

Antifungal effect of ophthalmic preservatives phenylmercuric nitrate and benzalkonium chloride on ocular pathogenic filamentous fungi

In the present study, the antifungal effects of phenylmercuric nitrate and benzalkonium chloride versus those of natamycin and ketoconazole were assessed against 216 filamentous fungi isolates from cases of fungal keratitis. They included 112 Fusarium isolates, 94 Aspergillus isolates, and 10 Alternaria alternata isolates. The strains were tested by broth dilution antifungal susceptibility testing of filamentous fungi approved by the Clinical and Laboratory Standards Institute M38-A document. The results showed that the MIC₅₀ values of phenylmercuric nitrate were 0.0156, 0.0156, and 0.0313 μg/mL for Fusarium spp., Aspergillus spp., and A. alternata, respectively. The MIC₉₀ values of phenylmercuric nitrate were 0.0313, 0.0313, and 0.0313 μg/mL for Fusarium spp., Aspergillus spp., and A. alternata, respectively. The MIC₅₀ values of benzalkonium chloride were 16, 32, and 8 μg/mL for Fusarium spp., Aspergillus spp., and A. alternata, respectively. The MIC₉₀ values of benzalkonium chloride were 32, 32, and 16 μg/mL for Fusarium spp., Aspergillus spp., and A. alternata, respectively. The study indicates that phenylmercuric nitrate has considerable antifungal activity and its effect is significantly superior to those of benzalkonium chloride, natamycin, and ketoconazole against ocular pathogenic filamentous fungi in vitro, deserving further investigation for treating fungal keratitis as a main drug.     

Occurrence of Zearalenone, Zearalenol and Deoxynivalenol in Some Market Samples of Dried Vegetables in J&K State

In Jammu and Kashmir state, there is a trend of sun-drying the surplus vegetables and marketing them for consumption in off-seasons. Unfortunately, warm and humid weather conditions prevailing during dehydration, followed by faulty storage practices may allow proliferation of such microbes, which could be degradative and toxigenic. In view of this, an investigation was conducted on market samples of J&K state to determine the association of toxigenic fusarial species with the surface of commonly consumed sun-dried vegetables viz., brinjal, tomato, cauliflower, bottle gourd, bitter gourd and turnip. All the samples of dried vegetables excepting that of bottle gourd were found to be contaminated with two species of Fusarium viz., F. solani and F. subglutinans. HPLC analysis conducted for estimating natural incidence of fusarial toxins revealed the presence of zearalenone (ZEN), zearalenol (ZOL) and deoxynivalenol (DON). Contamination of ZEN was detected in market samples of dried brinjal, tomato, cauliflower, turnip and bitter gourd with highest concentration (4.75?ppm) in one of the tomato samples. However, while investigating the marketed dried vegetables for ZOL contamination, only bitter gourd samples were found to be positive with contamination varying from 1.79 to 2.16?ppm. Similarly, contamination of DON was detected from market samples of dried brinjal, turnip and cauliflower. Its concentration was detected to be maximum in a cauliflower sample (2.12?ppm), whereas least contamination (0.09?ppm) was detected from a turnip sample. Some market samples of dried brinjal, bitter gourd, turnip and cauliflower even showed co-occurrence of fusarial toxins.     

Development of a Translatable Ultrasound Molecular Imaging Agent for Inflammation

This study details the development, characterization and non-clinical efficacy of an ultrasound molecular imaging agent intended for molecular imaging of P-selectin in humans. A targeting ligand based on a recently discovered human selectin ligand was manufactured as fusion protein, and activity for human and mouse P- and E-selectin was evaluated by functional immunoassay. The targeting ligand was covalently conjugated to a lipophilic anchor inserted into a phospholipid microbubble shell. Three lots of the targeted microbubble drug product, TS-07-009, were produced, and assays for size distribution, zeta potential and morphology were established. The suitability of TS-07-009 as a molecular imaging agent was evaluated in vitro in a flow-based adhesion assay and in vivo using a canine model of transient myocardial ischemia. Selectivity for P-selectin over E-selectin was observed in both the human and murine systems. Contrast agent adhesion increased with P-selectin concentration in a dynamic adhesion assay. Significant contrast enhancement was observed on ultrasound imaging with TS-07-009 in post-ischemic canine myocardium at 30 or 90 min of re-perfusion. Negligible enhancement was observed in resting (no prior ischemia) hearts or with a control microbubble 90 min after ischemia. The microbubble contrast agent described here exhibits physiochemical properties and in vivo behavior suitable for development as a clinical imaging agent.     

Identification and sensitivity of two rare fungal species isolated from two patients with Fusarium keratomycosis

We report two cases of Fusarium keratomycosis in which molecular analysis was used to identify two rare causative Fusarium species. A 65-year-old woman was diagnosed with Fusarium keratomycosis caused by F. equiseti, confirmed by sequencing the internal transcribed spacer (ITS) region and morphological characteristics. She was initially treated with topical corticosteroid but the keratitis did not improve. Even after aggressive antifungal treatment, there was a severe reduction of vision. In the end, the eye perforated. Our second case was a 75-year-old man who had diabetes mellitus. He developed keratitis after his cornea was injured by corn stalks. The ITS region identified the causative organism as F. verticillioides. Unlike the first case, he had good visual recovery after early antifungal therapy. Our findings demonstrate that both ITS and morphological characteristics can be used to identify the exact Fusarium species causing Fusarium keratomycosis. This allowed us to determine the sensitivity of these species to antifungal drugs that can be used to treat patients with these species of Fusarium.     

Patho-Biochemical Investigations of Blight and Wilt Diseases of Tomato (Lycopersicon esculentum Mill)

Tomato (Lycopersicon esculentum Mill) is an important vegetable, which is an integral component of the regular daily diet of human beings world over, consumed both in raw and cooked form. The alternaria blight and fusarium wilt diseases of tomato, earlier reported from several countries, were observed occurring in moderate to severe form in all farmer’s fields surveyed extensively in south Rajasthan. In the present study, attempts have been made to partially purify the toxic metabolites produced by the blight and wilt inducing pathogens and the effects of the toxins thus obtained were examined using different bio-assay methods. Results obtained in the present study clearly show that the toxins affected the germination of seeds, the root and shoot length of seedlings and caused significant leakage of electrolytes.     

Inhibition of RNA Synthesis as a Therapeutic Strategy against Aspergillus and Fusarium: Demonstration of In Vitro Synergy between Rifabutin and Amphotericin B

We investigated the in vitro antifungal activity of amphotericin B, alone and in combination with rifabutin, an inhibitor of bacterial RNA polymerase, against 26 clinical isolates of Aspergillus and 25 clinical isolates of Fusarium. Synergy or additivism between these drugs was demonstrated against all isolates tested. Amphotericin B MICs were reduced upon combination with rifabutin from a mean of 0.65 μg/ml to a mean of 0.16 μg/ml against Aspergillus, and from a mean of 0.97 μg/ml to a mean of 0.39 μg/ml against Fusarium (P < 0.000001 for both). Similarly, the MICs of rifabutin were reduced upon combination with amphotericin B from a mean of >32 μg/ml to a mean of 1.1 μg/ml against both fungi (P < 0.000001 for both). These positive interactions were corroborated by a colony count study with two Fusarium isolates, for which treatment with the combination of subinhibitory concentrations of amphotericin B (at concentrations 2- and 4-fold less than the MIC) and rifabutin (at concentrations ranging from 4- to 64-fold less than the MIC) resulted in 3.2-log reductions in colony counts compared to those after treatment with either drug alone. Inhibition of RNA synthesis was shown to be the mechanism of antifungal activity. These results suggest that inhibition of fungal RNA synthesis might be a potential target for antifungal therapy.     

Evaluation of Some <Emphasis Type="Italic">Trichoderma harzianum</Emphasis> Isolates for the Management of Soilborne Diseases of Brinjal and Okra

Present study investigated the efficacy of 6 Trichoderma harzianum isolates against damping off and wilt diseases of brinjal and okra caused by Fusarium oxysporium (FO) and Pythium aphanidermatum (PA), respectively in order to get more effective isolates for disease management. Isolated pathogens from brinjal were named as Fo^b and Pa^b and from okra Fo^o and Pa^o for F. oxysporum and P. aphanidermatum, respectively. In vitro analysis revealed that T. harzianum isolates inhibited the radial growth of both pathogens and isolate Th-Sks was most effective with 78.44 and 74.16% growth inhibition of Fo^b and Pa^b, and 78.19 and 78.11% of Fo^o and Pa^o, respectively. Production of volatile and non-volatile metabolites by isolate Th-Sks was more aggressive in mycelial growth inhibition of all test pathogens than that of other isolates. Isolate Th-Sks had highest suppression efficacy i.e. 95.92 and 98.93% for Fo^b and Pa^b, 96.43 and 93.33% for Fo^o and Pa^o under glasshouse conditions, 96.18 and 95.73% for Fo^b and Pa^b, and 94.66 and 96.18% for Fo^o and Pa^o in the field conditions, respectively. Additionally, seed treatment of brinjal and okra with isolate Th-Sks gave a significant stimulatory effect on plant height (brinjal 52.9 cm; okra 110.5 cm) and increased fruit yield (brinjal 328.3 g; okra 294.6 g) in comparison to other isolates and control sets during field trials. Thus, it is concluded that isolate Th-Sks can be used as biocontrol agent for management of wilt and damping off diseases of okra and brinjal after multilocation trials.     

Fertilizer Response of Maritime Pine on a Lateritic Soil

Superphosphate and urea produced large volume increases in 13-year-old maritime pine, Pinus pinaster (Aiton) growing on a lateritic podzolic soil. Growth curves over two years show the detailed effects of the fertilizers on seasonal growth. Volume increments were increased three-fold by the application of 800 lb superphosphate and 400 lb urea per acre, at a cost in fertilizers of 12 cents per cubic foot. Further volume increments can be expected from the original fertilizer application.     

Evaluation of Cultivated and Wild <Emphasis Type="Italic">Allium</Emphasis> Accessions for Resistance to <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">cepae</Emphasis>

Fusarium basal rot (FBR) caused by Fusarium oxysporum f. sp. cepae (FOC) is a highly destructive soil borne disease incurring heavy damage in pre and post harvest onion and garlic crops worldwide. Only a few onion lines exhibit partial resistance against the pathogen and there is a need for identification of more effective resistance sources for use in breeding programmes. Selected sets of wild onion and garlic accession and seven related Allium species were screened for resistance to Fusarium basal rot using three FOC isolates. FOC infection revealed significant variation among the evaluated Allium species (at P = 0.001). A. sativum accession ‘CBT-As153’ showed high level of resistance to each isolate while A. cepa accession ‘CBT-Ac77’ exhibited intermediate resistance. Among related Allium species, A. fistulosum, A. roylei and A. schoenoprasum were highly resistant, A. tuberosum had mixed response while A. griffithianum was susceptible. Further, the root density of Allium species negatively correlated with disease incidence for different FOC isolates. Thus, the present study suggests that besides related Allium species, A. sativum ‘CBT-As153’ can be used as a potential donor of FBR resistance for genetic improvement of onion and garlic in India.     

A novel nanoparticle loaded with methyl caffeate and caffeic acid phenethyl ester against Ralstonia solanacearum—a plant pathogenic bacteria

Developing a novel agent and understanding the interaction model between multipolymer nanoparticles and bacteria could be worthwhile to induce the protection of crops with the prevalence of frequent hazards because of the use of pesticides and chemical resistance. Unlike metal nanoparticles, multipolymer nanoparticles have bacteriostatic properties against Ralstonia solanacearum that can trigger bacterial wilt by infecting the plant. Therefore, a novel poly(lactic-co-glycolic acid) nanoparticle containing caffeic acid phenethyl ester (CAPE) and methyl caffeate (MC) was prepared with the sustained-release property (for 10 d at pH 6.5); here, 50% of the cumulative release rate was achieved. It was observed that the cytomembrane of R. solanacearum was jeopardized by the nanoparticle by the creation of large holes on the bacterial surface. The nanoparticle has an approximate EC₅₀ value of 0.285 mg mL⁻¹ with active pharmaceutical ingredients (APIs), while the drug dosage could be reduced by 2/3. Furthermore, to reveal the possible mechanism of interaction between the multipolymer nanoparticles and bacteria, a formidable inhibition effect was observed; the pathogenicity-related genes, namely, phcA, phcB, pehC, egl, pilT, and polA, of R. solanacearum were downregulated by 1/2, 1/42, 1/13, 1/6, 1/2, and 1/8, respectively, showing significant effects on the major virulence-related genes. Hence, a novel nanoparticle with excellent antibacterial and sustained-release properties has been prepared, possessing the potential to replace chemical pesticides and serve as a new control strategy for mulberry blight disease.

Developing a novel agent and understanding an interaction model between multipolymer nanoparticles and bacteria could be worthwhile to induce the protection of crops with the prevalence of frequent hazards because of the use of chemical pesticides.     

Identification and rapid detection of bacterial wilt in plantation Eucalyptus in China

Bacterial wilt caused by Ralstonia solanacearum is one of the most significant diseases of plantation Eucalyptus in southern China. In this study, 11 isolates were obtained from symptomatic individual trees from eucalypt plantations located in the key eucalypt-growing regions in China: Guangdong, Guangxi and Hainan. Cultures of these isolates were subject to morphological examination followed by DNA sequence comparisons to provide positive identification; all were confirmed to be R. solanacearum. Identification of the biovar types of all isolates obtained was then carried out using an improved biovar testing technique established in the course of this study. Ten of the isolates were shown to be biovar 3 of R. solanacearum, whilst the one other isolate appeared similar to one reported from a previous Chinese study on crop plants and designated as subtype 1 of biovar 4. This study represents the first detailed scientific investigation on the causal agent of Eucalyptus bacterial wilt in southern China.