Microbe-Mediated Control of Mycotoxigenic Grain Fungi in Stored Rice with Focus on Aflatoxin Biodegradation and Biosynthesis Inhibition

( Mohamed Mannaa ),( Ki Deok Kim ) 한국균학회 2016 Mycobiology Vol.44 No.2

Rice contaminated with fungal species during storage is not only of poor quality and low economic value, but may also have harmful effects on human and animal health. The predominant fungal species isolated from rice grains during storage belong to the genera Aspergillus and Penicillium. Some of these fungal species produce mycotoxins; they are responsible for adverse health effects in humans and animals, particularly Aspergillus flavus, which produces the extremely carcinogenic aflatoxins. Not surprisingly, there have been numerous attempts to devise safety procedure for the control of such harmful fungi and production of mycotoxins, including aflatoxins. This review provides information about fungal and mycotoxin contamination of stored rice grains, and microbe-based (biological) strategies to control grain fungi and mycotoxins. The latter will include information regarding attempts undertaken for mycotoxin (especially aflatoxin) bio-detoxification and microbial interference with the aflatoxin-biosynthetic pathway in the toxin-producing fungi.

Biocontrol Activity of Volatile-Producing Bacillus megaterium and Pseudomonas protegens Against Aspergillus and Penicillium spp. Predominant in Stored Rice Grains: Study II

( Mohamed Mannaa ),( Ki Deok Kim ) 한국균학회 2018 Mycobiology Vol.46 No.1

In our previous studies, Bacillus megaterium KU143, Microbacterium testaceum KU313, and Pseudomonas protegens AS15 have been shown to be antagonistic to Aspergillus flavus in stored rice grains. In this study, the biocontrol activities of these strains were evaluated against Aspergillus candidus, Aspergillus fumigatus, Penicillium fellutanum, and Penicillium islandicum, which are predominant in stored rice grains. In vitro and in vivo antifungal activities of the bacterial strains were evaluated against the fungi on media and rice grains, respectively. The antifungal activities of the volatiles produced by the strains against fungal development and population were also tested using I-plates. In in vitro tests, the strains produced secondary metabolites capable of reducing conidial germination, germ-tube elongation, and mycelial growth of all the tested fungi. In in vivo tests, the strains significantly inhibited the fungal growth in rice grains. Additionally, in I-plate tests, strains KU143 and AS15 produced volatiles that significantly inhibited not only mycelial growth, sporulation, and conidial germination of the fungi on media but also fungal populations on rice grains. GC-MS analysis of the volatiles by strains KU143 and AS15 identified 12 and 17 compounds, respectively. Among these, the antifungal compound, 5-methyl-2-phenyl-1H-indole, was produced by strain KU143 and the antimicrobial compounds, 2-butyl 1-octanal, dimethyl disulfide, 2-isopropyl- 5-methyl-1-heptanol, and 4-trifluoroacetoxyhexadecane, were produced by strain AS15. These results suggest that the tested strains producing extracellular metabolites and/or volatiles may have a broad spectrum of antifungal activities against the grain fungi. In particular, B. megaterium KU143 and P. protegens AS15 may be potential biocontrol agents against Aspergillus and Penicillium spp. during rice grain storage.

Influence of Temperature and Water Activity on Deleterious Fungi and Mycotoxin Production during Grain Storage

( Mohamed Mannaa ),( Ki Deok Kim ) 한국균학회 2017 Mycobiology Vol.45 No.4

Cereal grains are the most important food source for humans. As the global population continues to grow exponentially, the need for the enhanced yield and minimal loss of agricultural crops, mainly cereal grains, is increasing. In general, harvested grains are stored for specific time periods to guarantee their continuous supply throughout the year. During storage, economic losses due to reduction in quality and quantity of grains can become very significant. Grain loss is usually the result of its deterioration due to fungal contamination that can occur from preharvest to postharvest stages. The deleterious fungi can be classified based on predominance at different stages of crop growth and harvest that are affected by environmental factors such as water activity (a_w) and eco-physiological requirements. These fungi include species such as those belonging to the genera Aspergillus and Penicillium that can produce mycotoxins harmful to animals and humans. The grain type and condition, environment, and biological factors can also influence the occurrence and predominance of mycotoxigenic fungi in stored grains. The main environmental factors influencing grain fungi and mycotoxins are temperature and a_w. This review discusses the effects of temperature and a_w on fungal growth and mycotoxin production in stored grains. The focus is on the occurrence and optimum and minimum growth requirements for grain fungi and mycotoxin production. The environmental influence on aflatoxin production and hypothesized mechanisms of its molecular suppression in response to environmental changes are also discussed. In addition, the use of controlled or modified atmosphere as an environmentally safe alternative to harmful agricultural chemicals is discussed and recommended future research issues are highlighted.

Biocontrol Activity of Volatile-Producing Bacillus megaterium and Pseudomonas protegens against Aspergillus flavus and Aflatoxin Production on Stored Rice Grains

( Mohamed Mannaa ),( Ji Yeon Oh ),( Ki Deok Kim ) 한국균학회 2017 Mycobiology Vol.45 No.3

In our previous study, three bacterial strains, Bacillus megaterium KU143, Microbacterium testaceum KU313, and Pseudomonas protegens AS15, were selected as effective biocontrol agents against Aspergillus flavus on stored rice grains. In this study, we evaluated the inhibitory effects of the volatiles produced by the strains on A. flavus growth and aflatoxin production on stored rice grains. The three strains significantly reduced mycelial growth of A. flavus in dual-culture assays compared with the negative control strain, Sphingomonas aquatilis KU408, and an untreated control. Of these tested strains, volatiles produced by B. megaterium KU143 and P. protegens AS15 markedly inhibited mycelial growth, sporulation, and conidial germination of A. flavus on agar medium and suppressed the fungal populations in rice grains. Moreover, volatiles produced by these two strains significantly reduced aflatoxin production in the rice grains by A. flavus. To our knowledge, this is the first report of the suppression of A. flavus aflatoxin production in rice grains using B. megaterium and P. protegens volatiles.

Effect of Temperature and Relative Humidity on Growth of Aspergillus and Penicillium spp. and Biocontrol Activity of Pseudomonas protegens AS15 against Aflatoxigenic Aspergillus flavus in Stored Rice Grains

( Mohamed Mannaa ),( Ki Deok Kim ) 한국균학회 2018 Mycobiology Vol.46 No.3

In this study, we evaluated the effect of different temperatures (10, 20, 30, and 40℃) and relative humidities (RHs; 12, 44, 76, and 98%) on populations of predominant grain fungi (Aspergillus candidus, Aspergillus flavus, Aspergillus fumigatus, Penicillium fellutanum, and Penicillium islandicum) and the biocontrol activity of Pseudomonas protegens AS15 against aflatoxigenic A. flavus KCCM 60330 in stored rice. Populations of all the tested fungi in inoculated rice grains were significantly enhanced by both increased temperature and RH. Multiple linear regression analysis revealed that one unit increase of temperature resulted in greater effects than that of RH on fungal populations. When rice grains were treated with P. protegens AS15 prior to inoculation with A. flavus KCCM 60330, fungal populations and aflatoxin production in the inoculated grains were significantly reduced compared with the grains untreated with strain AS15 regardless of temperature and RH (except 12% RH for fungal population). In addition, bacterial populations in grains were significantly enhanced with increasing temperature and RH, regardless of bacterial treatment. Higher bacterial populations were detected in biocontrol strain-treated grains than in untreated control grains. To our knowledge, this is the first report showing consistent biocontrol activity of P. protegens against A. flavus population and aflatoxin production in stored rice grains under various environmental conditions of temperature and RH.

곡류작물의 수확전후에 발생하는 유해 저장 곰팡이와 균독소에 대한 제어 전략

모하메드 만나(Mohamed Mannaa),김기덕(Ki Deok Kim) 고려대학교 생명자원연구소 2017 생명자원연구 Vol.25 No.1

Abstract Consumption of cereals contaminated with mycotoxigenic fungi is a very serious issue threatening human and animal health. Therefore, the management of the occurrence of such fungi and control of mycotoxin level in cereals are current issues of great significance. These fungi may contaminate the grains and produce mycotoxins in cereal crops during preharvest or at postharvest periods, depending on the prevailing environmental conditions. Thus, efficient mycotoxin control strategies should consider all the factors that contribute to the accumulation of mycotoxins at different stages of the crops and adopt the most suitable control measures in an integrated control strategy. Several control measures have been proposed for the management of grain fungi and mycotoxins in the field during preharvest and at postharvest periods. Such methods include the selection of varieties resistant to the mycotoxigenic fungi, adhering to the proper cultural practices in the field, avoiding kernel damage during the harvesting process, and following the recommended postharvest practices to minimize the presence of mycotoxins in the grains. In the present review, the methods proposed for controlling mycotoxins are discussed, including the use of chemical methods, which are being replaced by other promising environment-friendly methods, such as the use of plant extracts and biocontrol microbial agents. In addition, the various antagonistic microbes as potential biocontrol agents against mycotoxigenic fungi and mycotoxins from previous studies are discussed. Finally, recommendations for future studies are provided, highlighting the importance of the integration of different control practices to minimize mycotoxins in grains and to reduce the use of synthetic chemicals.

Genomics-based Sensitive and Specific Novel Primers for Simultaneous Detection of Burkholderia glumae and Burkholderia gladioli in Rice Seeds

이채영,이현희,Mohamed Mannaa,김남규,박정욱,김주연,서영수 한국식물병리학회 2018 Plant Pathology Journal Vol.34 No.6

Panicle blight and seed rot disease caused mainly by Burkholderia glumae and Burkholderia gladioli is threatening rice cultivation worldwide. The bacteria have been reported as seed-borne pathogens from rice. Accurate detection of both pathogens on the seeds is very important for limiting the disease dissemination. Novel primer pairs targeting specific molecular markers were developed for the robust detection of B. glumae and B. gladioli. The designed primers were specific in detecting the target species with no apparent crossreactions with other related Burkholderia species at the expected product size. Both primer pairs displayed a high degree of sensitivity for detection of B. glumae and B. gladioli separately in monoplex PCR or simultaneously in duplex PCR from both extracted gDNA and directly preheated bacterial cell suspensions. Limit of detection was as low as 0.1 ng of gDNA of both species and 3.86 × 102 cells for B. glumae and 5.85 × 102 cells for B. gladioli. On inoculated rice seeds, the designed primers could separately or simultaneously detect B. glumae and B. gladioli with a detection limit as low as 1.86 × 103 cells per rice seed for B. glumae and 1.04 × 104 cells per rice seed of B. gladioli. The novel primers maybe valuable as a more sensitive, specific, and robust tool for the efficient simultaneous detection of B. glumae and B. gladioli on rice seeds, which is important in combating rice panicle blight and seed rot by early detection and confirmation of the dissemination of pathogen-free rice seeds.

Transcriptomic Insights into Abies koreana Drought Tolerance Conferred by Aureobasidium pullulans AK10

Park Jungwook,Mohamed Mannaa,Gil Han,Jung Hyejung,Jeon Hyo Seong,김진철,Park Ae Ran,서영수 한국식물병리학회 2024 Plant Pathology Journal Vol.40 No.1

The conservation of the endangered Korean fir, Abies koreana, is of critical ecological importance. In our previous study, a yeast-like fungus identified as Aureobasidium pullulans AK10, was isolated and shown to enhance drought tolerance in A. koreana seedlings. In this study, the effectiveness of Au. pullulans AK10 treatment in enhancing drought tolerance in A. koreana was confirmed. Furthermore, using transcriptome analysis, we compared A. koreana seedlings treated with Au. pullulans AK10 to untreated controls under drought conditions to elucidate the molecular responses involved in increased drought tolerance. Our findings revealed a predominance of downregulated genes in the treated seedlings, suggesting a strategic reallocation of resources to enhance stress defense. Further exploration of enriched Kyoto Encyclopedia of Genes and Genomes pathways and protein-protein interaction networks revealed significant alterations in functional systems known to fortify drought tolerance, including the terpenoid backbone biosynthesis, calcium signaling pathway, pyruvate metabolism, brassinosteroid biosynthesis, and, crucially, flavonoid biosynthesis, renowned for enhancing plant drought resistance. These findings deepen our comprehension of how AK10 biostimulation enhances the resilience of A. koreana to drought stress, marking a substantial advancement in the effort to conserve this endangered tree species through environmentally sustainable treatment.

Versatile Roles of Microbes and Small RNAs in Rice and Planthopper Interactions

Abdelaziz Mansour,Mannaa Mohamed Ibrahim Mohamed Ahmed,Omar Hewedy,Mostafa G. Ali,정혜정,서영수 한국식물병리학회 2022 Plant Pathology Journal Vol.38 No.5

Planthopper infestation in rice causes direct and indirect damage through feeding and viral transmission. Host microbes and small RNAs (sRNAs) play essential roles in regulating biological processes, such as metabolism, development, immunity, and stress responses in eukaryotic organisms, including plants and insects. Recently, advanced metagenomic approaches have facilitated investigations on microbial diversity and its function in insects and plants, highlighting the significance of microbiota in sustaining host life and regulating their interactions with the environment. Recent research has also suggested significant roles for sRNA-regulated genes during rice–planthopper interactions. The response and behavior of the rice plant to planthopper feeding are determined by changes in the host transcriptome, which might be regulated by sRNAs. In addition, the roles of microbial symbionts and sRNAs in the host response to viral infection are complex and involve defense-related changes in the host transcriptomic profile. This review reviews the structure and potential functions of microbes and sRNAs in rice and the associated planthopper species. In addition, the involvement of the microbiota and sRNAs in the rice–planthopper–virus interactions during planthopper infestation and viral infection are discussed.

The Roles of Two hfq Genes in the Virulence and Stress Resistance of Burkholderia glumae

Jieun Kim,Mohamed Mannaa,Namgyu Kim,Chaeyeong Lee,Juyun Kim,Jungwook Park,Hyun-Hee Lee,Young-Su Seo 한국식물병리학회 2018 Plant Pathology Journal Vol.34 No.5

The Hfq protein is a global small RNA chaperone that interacts with regulatory bacterial small RNAs (sRNA) and plays a role in the post-transcriptional regulation of gene expression. The roles of Hfq in the virulence and pathogenicity of several infectious bacteria have been reported. This study was conducted to elucidate the functions of two hfq genes in Burkholderia glumae, a causal agent of rice grain rot. Therefore, mutant strains of the rice-pathogenic B. glumae BGR1, targeting each of the two hfq genes, as well as the double defective mutant were constructed and tested for several phenotypic characteristics. Bacterial swarming motility, toxoflavin production, virulence in rice, siderophore production, sensitivity to H2O2, and lipase production assays were conducted to compare the mutant strains with the wildtype B. glumae BGR1 and complementation strains. The hfq1 gene showed more influence on bacterial motility and toxoflavin production than the hfq2 gene. Both genes were involved in the full virulence of B. glumae in rice plants. Other biochemical characteristics such as siderophore production and sensitivity to H2O2 induced oxidative stress were also found to be regulated by the hfq1 gene. However, lipase activity was shown to be unassociated with both tested genes. To the best of our knowledge, this is the first study to elucidate the functions of two hfq genes in B. glumae. Identification of virulence-related factors in B. glumae will facilitate the development of efficient control measures.