Growth characterization and predictive behavior of Eurotium species in a feedstuff matrix / Caracterización del crecimiento y comportamiento predictivo de especies de Eurotium en matriz de alimento balanceado

ABSTRACT Animal feeds are characterized by low water activity values. Nevertheless, fungal contamination withEurotiumspecies are quite common, causing nutritional depletion, spoilage and economic losses. The aim of this work was to assessEurotiumamstelodami,E. chevalieri,E. repensandE. rubrumgrowth in a feed matrix at different conditions of water activity (0.71-0.97) and temperature (5, 15, 25, 30 and 37°C). It was found thatEurotiumspecies are able to grow in a wide range of water activity and temperature in a short period of time (7 days) and faster than in synthetic media. Rosso and probabilistic models were applied in order to determine the limiting and optimum growth conditions as well as growth probability at certain combinations of environmental factors. Both models provided an accurate fit to the cardinal parameters and good performance for growth/no growth cases. This is the first report assessing the growth parameters ofEurotiumspecies directly in animal feed. Data obtained in the present study is useful to predict and avoidEurotiumspecies growth in animal feed.

RESUMEN Los alimentos balanceados se caracterizan por tener bajos valores de actividad de agua. Sin embargo, la contaminación por hongos con especies deEurotiumes bastante común y causa agotamiento nutricional, deterioro y pérdidas económicas. El objetivo de este trabajo fue evaluar el crecimiento deEurotiumamstelodami,E. chevalieri,E. repensyE. rubrumen una matriz de alimento balanceado a diferentes condiciones de actividad de agua (0,71-0,97) y temperatura (5, 15, 25, 30 y 37°C). Se determinó que las especies deEurotiumpueden crecer en un amplio rango de actividades de agua y temperatura en un corto período de tiempo (7 días), y a mayor velocidad que en medio sintético. Se utilizaron los modelos de Rosso y probabilísticos para determinar las condiciones de crecimiento limitantes y óptimas, así como la probabilidad de crecimiento en ciertas combinaciones de factores ambientales. Ambos modelos proporcionaron un ajuste preciso a los parámetros cardinales y una buenaperformancepara los casos de crecimiento/sin crecimiento. Este es el primer trabajo que evalúa los parámetros de crecimiento de las especies deEurotiumdirectamente en alimento balanceado. Los datos obtenidos en el presente estudio son útiles para predecir y evitar el crecimiento de especies deEurotiumen este tipo de alimentos.

Factores extrínsecos e intrínsecos asociados a poblaciones fúngicas micotoxigénicas de granos de maíz (Zea mays L) almacenados en silos bolsa en Argentina / Extrinsic and intrinsic factors associated with mycotoxigenic fungi populations of maize grains (Zea mays L.) stored in silobags in Argentina

Con el objeto de caracterizar las poblaciones fúngicas, en particular las especies potencialmente micotoxigénicas, que pueden contaminar los granos de maíz almacenados en silos bolsa con un contenido de humedad superior al recomendado como seguro, se evaluaron 270 muestras extraídas al inicio, a los 90 días y al final de un período de almacenamiento de 5 meses. En dichas muestras se cuantificó e identificó la biota fúngica y se determinó la contaminación con fumonisinas y aflatoxinas. Asimismo, se evaluó el efecto de factores extrínsecos (ambiente), intrínsecos (granos) y tecnológicos (ubicación de los granos en el perfil del silo bolsa) sobre las poblaciones totales y micotoxigénicas. El pH de los granos y el nivel de O2 se redujeron significativamente a los 5 meses, mientras que la concentración de CO2 se incrementó en igual período. Los recuentos totales de la micobiota fueron significativamente mayores en los granos ubicados en el estrato superior del silo bolsa. Se identificaron especies micotoxigénicas de Fusarium, Aspergillus, Penicillium y Eurotium. La frecuencia de aislamiento de Fusarium verticillioides se redujo al final del almacenamiento y Aspergillus flavus solo se aisló en el inicio del almacenamiento. Los recuentos de Penicillium spp. y Eurotium spp. se incrementaron al final del almacenamiento. El 100 % de las muestras presentaron contaminación con fumonisinas, con niveles máximos de 5,707 mg/kg, mientras que las aflatoxinas contaminaron el 40 % de las muestras con niveles máximos de 0,0008 mg/kg. Las condiciones ambientales y de sustrato generadas durante el almacenamiento produjeron cambios en la composición de las poblaciones fúngicas y limitaron el desarrollo de hongos micotoxigénicos y la producción de micotoxinas.

In order to determine the behavior of mycotoxin-producing fungal populations linked with silobags stored corn grains with a moisture content greater at the recommended as safe, 270 samples taken in three times (beginning, 90 days, final) over a five month period of storage were evaluated. The fungal biota was quantified and identified and the contamination with fumonisin and aflatoxin was determined. Extrinsic factors (environment), intrinsic factors (grains) and technological factors (location of the grains in the profile of silobag) were taken into account to evaluate the presence and quantity of total and mycotoxigenic fungal populations. The pH of grains and O2 levels were significantly reduced after five months, while CO2 concentration increased in the same period. The total counts of mycobiota were significantly higher in grains located in the top layer of silobag. Mycotoxigenic species of Fusarium, Aspergillus, Penicillium and Eurotium were identified. The frequency of isolation of Fusarium verticillioides decreased at the end of storage and Aspergillus flavus was isolated only at the beginning of storage. The counts of the Penicillium spp. and Eurotium spp. were increased at the end of storage. Fumonisin contamination was found in all the samples (100 %) with maximum levels of 5.707 mg/kg whereas aflatoxin contaminated only 40 % with maximum levels of 0.0008 mg/kg. The environmental and substrate conditions generated during the storage limited the development of mycotoxigenic fungi and mycotoxin production.

The Mycobiota of Air Inside and Outside the Meju Fermentation Room and the Origin of Meju Fungi

The fungi on Meju are known to play an important role as degrader of macromolecule of soybeans. In order to elucidate the origin of fungi on traditional Meju, mycobiota of the air both inside and outside traditional Meju fermentation rooms was examined. From 11 samples of air collected from inside and outside of 7 Meju fermentation rooms, 37 genera and 90 species of fungi were identified. In outside air of the fermentation room, Cladosporium sp. and Cladosporium cladosporioides were the dominant species, followed by Cladosporium tenuissimum, Eurotium sp., Phoma sp., Sistotrema brinkmannii, Alternaria sp., Aspergillus fumigatus, Schizophyllum commune, and Penicillium glabrum. In inside air of the fermentation room, Cladosporium sp., Aspergillus oryzae, Penicillium chrysogenum, Asp. nidulans, Aspergillus sp., Cla. cladosporioides, Eurotium sp., Penicillium sp., Cla. tenuissimum, Asp. niger, Eur. herbariorum, Asp. sydowii, and Eur. repens were collected with high frequency. The concentrations of the genera Aspergillus, Eurotium, and Penicillium were significantly higher in inside air than outside air. From this result and those of previous reports, the origin of fungi present on Meju was inferred. Of the dominant fungal species present on Meju, Lichtheimia ramosa, Mucor circinelloides, Mucor racemosus, and Scopulariopsis brevicaulis are thought to be originated from outside air, because these species are not or are rarely isolated from rice straw and soybean; however, they were detected outside air of fermentation room and are species commonly found in indoor environments. However, Asp. oryzae, Pen. polonicum, Eur. repens, Pen. solitum, and Eur. chevalieri, which are frequently found on Meju, are common in rice straw and could be transferred from rice straw to Meju. The fungi grow and produce abundant spores during Meju fermentation, and after the spores accumulate in the air of fermentation room, they could influence mycobiota of Meju fermentation in the following year. This could explain why concentrations of the genera Aspergillus, Eurotium, and Penicillium are much higher inside than outside of the fermentation rooms.

Mycoflora of Soybeans Used for Meju Fermentation

Diverse fungi are present in Korean traditional meju and they are known to play an important role in fermented soybean products. To determine the origin of the fungi in meju, we examined the mycoflora of soybeans from 10 traditional meju factories. The samples were untreated or treated with sodium hypochlorite, and placed on malt extract agar (MEA), dichloran 18% glycerol agar (DG18), and dichloran rose bengal chloramphenicol agar (DRBC) medium. A total of 794 fungal strains were isolated and they were identified as 41 genera and 86 species. From sodium hypochlorite untreated soybeans, the genera, Cladosporium (55%), Eurotium (51%), Fusarium (33%), Penicillium (22%), and Aspergillus (exclusion of Eurotium) (20%), were mainly isolated, and Eurotium herbariorum (22%), Eurotium repens (18%), Cladosporium tenuissimum (18%), F. fujikuroi (18%), Aspergillus oryzae/flavus (7%), and Penicillium steckii (6%) were the predominant species. In case of sodium hypochlorite-treated soybeans, Eurotium (31%) and Cladosporium (5%) were frequently isolated, but Aspergillus (excluding Eurotium), Penicillium and Fusarium which were frequently isolated from untreated soybeans, were rarely isolated. Eurotium herbariorum (21%), Eurotium repens (8%), and Cladosporium tenuissimum (3%) were the predominant species. Of the 41 genera and 86 species isolated from soybeans, 13 genera and 33 species were also found in meju. These results suggest that the fungi on soybeans may influence the mycoflora of meju.

Regulation of eurotium repens reproduction and secondary metabolite production

Eurotium repens [Anamorph Aspergillus repens] was isolated 1 from spoiled fruit. It reproduced sexually at different sucrose concentrations up to 50% [w/v]; water activity, 0.79. It reproduced asexually at high sucrose concentration 60% [w/v] water activity, 0.75. The concentrations of all detected amino acids were higher in the teleomorph than the anamorph except that of glycine, while alpha aminoadipic acid and alanine were detected in teleomorph only. The extracellular secondary metabolites produced by the teleomorph and anamorph stages were variable and different except epoxysuccinic acid and 2-pyruvylamino benzamide which were produced by the two stages. Glycine, arginine and calcium chloride unlike glutamic acid, aspartic acid and alanine, plays an important role in the induction of teleomorph stage formation at high sucrose concentration 60% [w/v]

Identification and Characterization of Eurotium rubrum Isolated from Meju in Korea

We isolated and identified a strain of Eurotium rubrum from Meju that has not been reported in Korea. This fungus is yellowish brown; reverse dark brown on CYA and PDA while yellow on 2% MEA at 25degrees C. Cleistothecia are first bright yellow and gradually turned brown. Mycerial growth on CYA attained a diameter of 30 mm at 20degrees C, 37 mm at 25degrees C and 32 mm at 30degrees C after 15 days. The isolate grew slower on 2% MEA (< 20 mm 15 days at 25degrees C) compared to CYA and PDA (< 40 mm 15 days at 25degrees C). Cleistothecia are superficial, yellow to light brown, globose to subglobose, 40~75 microm in diameter. Asci are 8-spored and globose to subglobose 8~11 microm. Ascospores are disciform, 4.0~5.0 microm in length and 4.2~4.5 microm in width. Conidia are ovate or bacillar, finely roughened to densely spinulose, 4.6~6.0 microm in length and 3.0~4.3 microm in width. Compared to known Eurotium rubrum, the Korean isolate showed 99% sequence similarity in ITS rDNA (554 bp) and calmodulin (750 bp) gene and 100% in beta-tubulin (1016 bp) gene. The E. rubrum isolate also had weak beta-glucosidase and protease activities.

Detection of Fungus and Bacteria in Otitis Media with Effusion of Children Using Polymerase Chain Reaction and Its Correlation of Clinical Factors / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: Several hypotheses have been proposed in the etiopathogenesis of otitis media with effusion (OME). The bacterial or fungal infection may also play a major role in the pathogenesis of OME. To investigate the relationship between bacteria and fungi as pathogens in OME and to analyze the incidence, the authors evaluated the pathogens of OME using polymerase chain reaction (PCR) technique, which is known to be more sensitive and specific than conventional bacterial and fungal culture. SUBJECTS AND METHOD: Twenty-three children (thirty ears), who were diagnosed with OME and had underwent ventilation tube insertion, were evaluated in the department of ORL-HNS, University Hospital, from May 2006 to March 2007. The middle ear effusion, obtained during the procedure, was evaluated for the identification of bacteria and fungi by PCR. RESULTS: Among 30 ears, viable pathogenic bacteria were detected in 20 ears (66.6%) and fungi in 13 ears (43.3%). The bacterial pathogens included Haemophilus influenzae (13 ears), followed by Streptococcus pneumonize (6 ears). The fungi detected from PCR were Candida albicans (4 ears-30.8%), Aspergillus niger (2 ears-15.4%) and Paecilomyces lilacinus (2 ears-15.4%). Additional pathogens include Basidiomycete yeast, Saccharamycete sp., Eurotium rubrum, Dothioraceae sp. and Stemphylium solani. Detection of fungal DNA was more common in patients with cleft palate and in recurrent cases with statistical significance. CONCLUSION: The use of PCR of middle ear effusion is effective for the detection of pathogens in patients with OME. While bacteria were thought to be the causative pathogen, this study suggests the etiological role of fungi in the pathogenesis of OME. However, the relationship between fungi and OME requires further studies.