Biocontrol of postharvest gray and blue mold decay of apples with Rhodotorula mucilaginosa and possible mechanisms of action

The efficacy of Rhodotorula mucilaginosa against postharvest gray mold, blue mold and natural decay development of apples and the possible mechanisms involved were investigated. The decay incidence and lesion diameter of gray mold and blue mold of apples treated by R. mucilaginosa were significantly reduced compared with the control fruits, and the higher concentration of R. mucilaginosa, the better the efficacy of the biocontrol. R. mucilaginosa also significantly reduced the natural decay development of apples following storage at 20 °C for 35 days or at 4 °C for 45 days followed by 20 °C for 15 days. Germination and survival of spores of Penicillium expansum and Botrytis cinerea were markedly inhibited by R. mucilaginosa in an in vitro test. Rapid colonization of the yeast in apple wounds was observed whether stored at 20 °C or 4 °C. In apples, the activities of peroxidase (POD) and polyphenoloxidase (PPO) were significantly induced and lipid peroxidation (malondialdehyde (MDA) content) was highly inhibited by R. mucilaginosa treatment compared with those of the control fruits. All these results indicated that R. mucilaginosa has great potential for development of commercial formulations to control postharvest pathogens on fruits. Its modes of action were based on competition for space and nutrients with pathogens, inducement of activities of defense-related enzymes such as POD, PPO and inhibition of lipid peroxidation (MDA content) of apples, so as to enhance the resistance and delay the ripening and senescence of apples.     

Methyl jasmonate enhances biocontrol efficacy of Rhodotorula glutinis to postharvest blue mold decay of pears

The effect of Rhodotorula glutinis treatment alone or in combination with methyl jasmonate (MeJA) in controlling blue mold decay, the natural fungal decay of pears and the postharvest quality parameters including fruit firmness, total soluble solids, titratable acidity, and ascorbic acid were investigated. The combination of methyl jasmonate (200 μM) and R. glutinis (1 × 10⁸ CFU/ml) was a more effective approach to reduce the disease incidence and lesion diameter of blue mold decay of pears than the application of MeJA or R. glutinis alone after incubation for 7 d at 20 °C. The natural fungal decay of pears treated with the application of R. glutinis combined with MeJA resulted in reduced average decay incidence of 10.42% or 4.16%, respectively, compared with 27.17% or 20.83% in the control fruits following storage at 20 °C for 15 d or 4 °C for 60 d followed by 20 °C for 15 d. The combined treatment did not impair quality parameters of fruits under both conditions.     

植酸结合胶红酵母对桃果采后根霉病的控制

为了寻找一种能够代替化学杀菌剂控制桃果采后病害的方法,研究了植酸与胶红酵母结合使用对桃果采后根霉病的控制效果。结果表明,0.15%植酸与胶红酵母结合使用能显著降低桃果采后根霉病发病率,防治桃果自然腐烂的发生,并且对果实贮藏品质没有产生不利影响;能诱导桃果多酚氧化酶(PPO)、过氧化物酶(POD)活性的提高,同时可以抑制丙二醛(MDA)含量的增加。     

Combination of Kluyveromyces marxianus and sodium bicarbonate for controlling green mold of citrus fruit

Biocontrol efficacy of an antagonistic yeast Kluyveromyces marxianus was evaluated individually or in combination with sodium bicarbonate (SBC) against green mold of citrus fruit caused by Penicillium digitatum. Their effects on postharvest quality of citrus fruit were also investigated. The results indicated that the antagonistic activity of K. marxianus at 1 × 10⁸ CFU/mL on green mold of citrus fruit was enhanced by 2% SBC treatment. In artificial inoculation trials, disease control after 3 and 6 days, respectively, with the mixture of K. marxianus and 2% SBC (18.33%, 58.33%) was significantly improved over that obtained with K. marxianus (41.67%, 70.00%) or SBC (43.33%, 81.67%) alone. The combination of K. marxianus with SBC was as effective as the imazalil treatment in natural infection trials, which gave about 90% control of green mold. Addition of 2% SBC significantly stimulated the growth of K. marxianus in citrus fruit wounds after 72 h. Moreover, K. marxianus, SBC and their combination did not impair quality parameters including weight loss, fruit firmness, total soluble solids, titratable acidity and ascorbic acid at 4 °C for 30 days followed by 20 °C for 15 days. These results suggested that the use of SBC is a useful approach to improve the efficacy of K. marxianus for the postharvest green mold of citrus fruit.     

苯并噻二唑结合罗伦隐球酵母对桃果采后青霉病的控制

研究了拮抗酵母菌罗伦隐球酵母与植物诱导剂苯并噻二唑(BTH)结合使用控制桃果青霉病的效果。结果显示,罗伦隐球酵母与100μg/mL的BTH结合使用时,不仅其生防效力显著提高,还可以显著提高桃果的抗性相关酶活性,包括过氧化物酶(POD)和过氧化氢酶(CAT),并且抑制了丙二醛(MDA)含量的增加。因此,BTH与罗伦隐球酵母结合使用,集成了生物防治酵母和植物生长调节剂的生物双重活性,可能成为防治桃果采后病害的一个新途径。      

Salicylic acid enhances biocontrol efficacy of Rhodotorula glutinis against postharvest Rhizopus rot of strawberries and the possible mechanisms involved

The potential of using Rhodotorula glutinis alone or in combination with salicylic acid (SA) for the control of postharvest Rhizopus rot of strawberries, and their effects on enzyme activities of fruits were investigated. The combination of R. glutinis (1 × 10⁸ CFU ml⁻¹) with SA (100 μg ml⁻¹) resulted in a significant reduction in the disease incidence and lesion diameter of Rhizopus rot on the strawberry fruits at 20 °C and 4 °C, and more so than with SA or yeast alone. SA at the concentration of 100–1000 μg ml⁻¹ significantly inhibited spore germination of Rhizopus stolonifer. About 100 μg ml⁻¹ of SA did not inhibit the growth of the antagonistic yeast, and could significantly increase the population growth of R. glutinis in strawberry wounds at 20 °C. SA, combined with R. glutinis, increased the activity of strawberry host defence enzymes (POD) and cell wall lytic enzymes (β-1,3-glucanase).     

热空气复合茉莉酸甲酯处理对杨梅果实采后腐烂和品质的影响

目的 分析热空气复合茉莉酸甲酯(methyl jasmonate, MeJA)处理对杨梅果实采后腐烂和品质的影响。方法 将“乌种”杨梅在48 °C下进行10 μmol/L MeJA的熏蒸处理, 随后在20 °C下贮藏3 d或者1 °C下贮藏5、10 d、15 d后再20 °C贮藏1 d以模拟货架条件, 贮藏期间或结束后测定果实发病率以及硬度、可滴定酸(titratable acid, TA)、可溶性固形物(Total Soluble Solid, TSS)、维生素C和总酚等品质指标。同时, 采用芽管离体培养法测定该复合处理对Verticicladiella abietina(杨梅轮帚霉)孢子萌发和菌丝生长的影响。结果 10 μmol/L MeJA处理对V. abietina孢子萌发无明显抑制作用, 但可显著抑制病原菌菌丝的生长; 48 °C热空气处理则可显著抑制病原菌孢子萌发和菌丝生长。单一热空气或茉莉酸甲酯处理均可抑制杨梅果实采后贮藏期间绿霉病发病率, 但两者复合处理较单一处理更为显著的抑制了杨梅果实病害发生, 且经复合处理的杨梅果实其硬度、TA、TSS、维生素C和总酚含量均显著高于单一处理果实。结论 热空气复合茉莉酸甲酯处理可通过抑制V. abietina的生长从而控制杨梅果实采后腐烂并同时延缓果实品质下降, 从而为该复合处理的实际应用提供理论依据。     

A combination of marine yeast and food additive enhances preventive effects on postharvest decay of jujubes (Zizyphus jujuba)

We investigated the effects of marine yeast Rhodosporidium paludigenum in combination with a food additive, carboxymethylcellulose sodium (CMC-Na), on prevention of postharvest decay and food quality of Chinese winter jujubes. R. paludigenum (1 × 10⁸ cells/ml) combined with CMC-Na (0.3%) significantly increased the inhibition of black rot on jujubes at 25 °C when compared with R. paludigenum-alone treatment (5.8% vs. 20%, p < 0.05). The combination also reduced natural rot from 86% (control) to 56%. The combination caused transient changes in enzyme activities or contents of some oxidation reactive markers such as peroxidase (POD), superoxide dismutase (SOD), and malondialdehyde (MDA) of jujubes. The combination had no significant effect on the food qualities such as colour (chroma and hue angle), total soluble solid (TSS) and titratable acidity (TA) of the fruit. While enhancing these effects, CMC-Na did not affect the survival of R. paludigenum in nutrient yeast dextrose agar (NYDA) culture. Thus, we conclude that the combination of R. paludigenum and CMC-Na is a promising formulation to control postharvest decay of Chinese winter jujubes.     

Cloning, characterization, expression and antifungal activity of an alkaline serine protease of Aureobasidium pullulans PL5 involved in the biological control of postharvest pathogens

An alkaline protease gene was amplified from genomic DNA and cDNA of the antagonistic yeast-like fungus Aureobasidium pullulans PL5, a biocontrol agent effective against Monilinia laxa on stone fruit and Botrytis cinerea and Penicillium expansum on pome fruits. An open reading frame of 1248 bp encoding a 415-amino acid (aa) protein with a calculated molecular weight (M_r) of 42.9 kDa and an isoelectric point (pI) of 4.5 was characterized. The cDNAALP5 gene had an 18-amino acid signal peptide, one N-gylcosylation, one histidine active site, and one serine active site. The ALP5 gene with a M_r of 1351 bp contained two introns. One intron was of 54 bp, while the other was of 50 bp. Protein BLAST and phylogenetic tree analysis of the deduced amino sequences from the cDNAALP5 gene showed that the encoded protein had 100% homology to a protease enzyme (ALP2) of a sea strain of A. pullulans, suggesting that the protein ALP5 was an alkaline serine protease. Expression of ALP5 in Escherichia coli BL21 (DE3), followed by identification with Western-blotting, purification with Ni-NTA and analysis of enzymatic activity, yielded an homogeneous recombinant ALP5 which hydrolysed the substrate casein and inhibited the mycelial growth of the pathogens. At its optimal pH of 10.0 and reaction temperature of 50 °C, the recombinant protease exhibited the highest activity towards the substrate casein, though the highest stability was at lower temperatures and pH between 7.0 and 9.0. This study provided the direct evidence that extracellular proteases secreted by the antagonist A. pullulans PL5 played a role in the biocontrol activities against some postharvest pathogens of apple and peach.     

膜醭毕赤酵母对杨梅果实采后绿霉病的抑制与抗病性的诱导

为研究膜醭毕赤酵母(Pichia membranaefaciens)对杨梅果实绿霉病的抑制及其对抗病性的诱导作用,将杨梅果实先接种15μL浓度为1×109cfu/mL的膜醭毕赤酵母菌悬液,风干1h后再接种15μL的1×105cfu/mL的桔青霉(Penicillium citrinum)孢子悬浮液,然后转入(1±1)℃下贮藏8d,每隔2d测定果实腐烂率、病斑直径、抗病相关酶几丁质酶、β-1,3-葡聚糖酶、苯丙氨酸解氨酶(PAL)、过氧化物酶(POD)、多酚氧化酶(PPO)活性以及总酚含量;同时分析了离体条件下P.membranaefaciens对P.citrinum孢子萌发和芽管伸长的影响。结果显示,P.membranaefaciens可显著抑制果实贮藏期间绿霉病的发生,并可诱导几丁质酶、β-1,3-葡聚糖酶、PAL、POD和PPO活性和总酚含量的升高;同时P.membranaefaciens在离体条件下也明显抑制病原菌的生长。因此,P.membranaefaciens抑制杨梅果实采后腐烂与直接抑制病原菌生长以及间接诱导抗病性密切相关。     

Effect of chitin on the antagonistic activity of Rhodotorula glutinis against Botrytis cinerea in strawberries and the possible mechanisms involved

The influence of chitin on the efficacy of Rhodotorula glutinis in controlling postharvest grey mould of strawberries and the possible mechanisms involved were investigated. The results showed that the antagonistic activity of R. glutinis harvested either from the culture media of NYDB amended with chitin at 0.5% or from the culture media in which chitin was the sole carbon source (NYCB) was improved greatly compared with that without chitin. The application of R. glutinis cultivated in the culture media of the chitin-supplement (0.5%) induced higher β-1,3-glucanase activity and reduced more MDA content of strawberries compared with that R. glutinis cultivated in the NYDB. Moreover, the control efficacy of the cell-free filtrate of the chitin-supplement culture media (0.5%) and NYCB were higher than that of cell-free culture filtrates of NYDB in 2 days incubation, with the associated high level of chitinase activity.     

Combination of salicylic acid and ultrasound to control postharvest blue mold caused by Penicillium expansum in peach fruit

The effect of ultrasound (40 kHz, 10 min) and salicylic acid (SA, 0.05 mM) either separately, or combined on blue mold caused by Penicillium expansum in peach fruit was investigated. The results showed that the application of SA alone could reduce blue mold, while the use of ultrasound had no effect. Our results also revealed that SA combined with ultrasound treatment was more effective in inhibiting fungal decay during storage than the SA treatment alone. The combined treatment increased the activities of defense enzymes such as chitinase, β-1,3-glucanase, phenylalanine ammonia-lyase, polyphenol oxidase and peroxidase, which were associated with higher disease resistance induced by the combined treatment. Furthermore, the combined treatment did not impair the quality parameters of peach fruit after 6 days of storage at 20 °C. These results suggested that the combination of ultrasound and SA treatment may be a useful technique to reduce blue mold in peach fruit.

Industrial relevance

This paper investigates the effect of ultrasound combined with SA on decay incidence of peach fruit. The results presented demonstrate that the effect of the combined treatment on the disease resistance and fruit quality should be considered by processors prior to its adoption as a preservation technique.     

Control of Listeria monocytogenes growth in a ready-to-eat poultry product using a bacteriophage

A bacteriophage (phage) that infected strains of the species Listeria monocytogenes as well as Listeria ivanovii and Listeria welshimeri, but not Listeria grayi or Listeria innocua, was isolated from sheep faeces. The phage had a contractile tail and an icosohedral head indicating that it was a myovirus, and was morphologically similar to phage A511. At 30 °C, phages added at 5.2 × 10⁷ PFU ml⁻¹ prevented the growth in broth of L. monocytogenes present at approximately twice this concentration for 7 h, but re-growth occurred such that the concentration after 24 h incubation was similar in both control and phage-treated cultures. At the same temperature, but on the surface of vacuum-packed ready-to-eat chicken breast roll, there was an immediate 2.5 log₁₀ CFU cm⁻² reduction in pathogen concentration following addition of phages and then re-growth. However, at a temperature reflecting that at which a chilled food might be held (5°C), this re-growth was prevented over 21 days incubation. The data suggest a dose-dependent rapid reduction in pathogen concentration followed by no continued phage-mediated effect. These results, alongside other published data, indicate that a high concentration of phages per unit area is required to ensure significant inactivation of target pathogens on food surfaces.     

Effect of water activity on stress tolerance and biocontrol activity in antagonistic yeast Rhodosporidium paludigenum

The aims of this study were to optimize the salt-adaptation conditions of the marine antagonistic yeast Rhodosporidium paludigenum and investigate the biocontrol activity of salt-induced cell suspensions of R. paludigenum on postharvest pathogens in fruits. Low water activity (a_w = 0.98, 0.97, 0.96, and 0.95) inhibited the growth of R. paludigenum in nutrient yeast dextrose broth, but the yeast grew better in the medium modified with NaCl solute than other nonionic solutes. R. paludigenum grown in 6.6% NaCl-modified medium had higher viabilities (92.1%) at low water activity (a_w = 0.95) than control (81.1%) after 48 h incubation. The salt-adapted R. paludigenum also showed better viability than the un-adapted cells after being frozen, which may be related to the accumulation of intracellular trehalose. Moreover, the best biocontrol inhibition in pears and Chinese winter jujubes was obtained when R. paludigenum was grown in NaCl-modified medium. Therefore, this study implies that improving physiological inducement methods may be a promising strategy for accelerating commercialization of biocontrol agents.     

Biocontrol of fungal decay of citrus fruit by Pichia pastoris recombinant strains expressing cecropin A

Cecropin A gene was cloned into the expression vector pPIC9k and was successfully expressed in methylotrophic yeast, Pichia pastoris GS115. The yeast had effective antimicrobial activity on Geotrichum citri-aurantii spores by the thiazolyl blue (MTT) assay. There was no large growth difference between nontransformed strain GS115 and recombinant strain GS115/CEC in citrus fruits wounds. Yeast transformants could significantly inhibit growth of germinated G. citri-aurantii spores and inhibited decay development caused by G. citri-aurantii in citrus fruits compared to the yeast strain GS115/pPIC. This study demonstrates the potential of expression of an antifungal peptide in yeast for enhancing suppression of postharvest diseases and represents a new approach for the biological control of postharvest diseases.     

Red mold dioscorea: A potentially safe traditional function food for the treatment of hyperlipidemia

A study was undertaken to evaluate whether the interaction between Monascus-fermented products and lovastatin contributes to increased risk of rhabdomyolysis. Rhabdomyolysis is a potentially dangerous side effect of statin drugs. In this study with hyperlipidemic hamsters fed lovastatin only, lovastatin with 1-fold red mold dioscorea (RMD), and lovastatin, the functional components of red mold fermented products, HMG-CoA reductase inhibitors, did not exacerbate pre-existing diseases, and actually helped in improving existing disease conditions, respectively, as compared with the control. Administration of RMD, alone or in combination with lovastatin did not cause significant rhabdomyolysis as assessed by measuring the levels of creatinine phosphokinase. Further, we did not find any study that clearly implicates the involvement of RMD, which have long been considered a food product, in liver and kidney toxicity. RMD alone or in combination with lovastatin, does not increase the risk of rhabdomyolysis, even when administered at a high dosage (including HMG-CoA reductase inhibitors >75 mg/day/adult).     

Phenolic content and antioxidative capacity of green and white tea extracts depending on extraction conditions and the solvent used

The efficiencies of different solvents in the extraction of phenolics from bagged and loose leaves of white and green tea, after different extraction times, as well as the antioxidative capacity of the obtained extracts, were investigated. The developed HPLC method has the potential to separate and determinate 17 phenolics widely distributed in plants, but in investigated tea extracts only four catechins and traces of three flavonols and one flavone were separated and detected based on comparison with authentic standards. The extraction efficiency of phenolics depended strongly on the time of extraction and the solvents used. The extraction of catechins from green tea was significantly affected by the form (bagged or loose) of the tea, whereas this effect was shown not to be statistically significant for white tea. Green tea was a richer source of phenolics than was white tea. The extraction of phenolics from white tea by water could be accelerated by the addition of lemon juice. Aqueous ethanol (40%) was most effective in the prolonged extraction of catechins. The antioxidative capacity of the investigated tea extracts correlated with their phenolic content.     

Effects of curing sodium nitrite additive and natural meat fat on growth control of Listeria monocytogenes by the bacteriocin-producing Lactobacillus curvatus strain CWBI-B28

In realistic model meat systems, the separate and combined effects of fat content and sodium nitrite on the antilisterial activity of the bacteriocin of Lactobacillus curvatus CWBI-B28 were studied. In laboratory fermentations where Listeria monocytogenes was co-cultured at 4 °C with bacteriocin-producing CWBI-B28 in lean pork meat (fat content: 13%) without added nitrite, a strong antilisterial effect was observed after one week. The effect was maintained for an additional week, after which a slight and very gradual rebound was observed. Both added nitrite (20 ppm) and a high-fat content (43%) were found to antagonise this antilisterial effect, the Listeria cfu count reached after six weeks being 200 times as high in high-fat meat with added nitrite than in lean meat without nitrite. This antagonism could not be attributed to slower growth of the bacteriocin-producing strain, since CWBI-B28 grew optimally in fat-rich meat with 20 ppm sodium nitrite. Bacteriocin activity was also measured in the samples. The observed activity levels are discussed in relation to the degree of antilisterial protection conferred.