Abstract
The capacity of microorganisms from water kefir (WK) to control Aspergillus flavus growth during the aerobic phase of ensiled sorghum grains was determined. Sorghum inoculated with A. flavus was treated with filter-sterilized and non-sterilized water kefir, ensiled, and incubated 7 days at 25 °C. A. flavus growth was quantified by qPCR after incubation. Mold growth was inhibited in the presence of water kefir while no inhibition was observed when filter-sterilized water kefir was applied, demonstrating the relevant role of the microorganisms in the kefir water in the biocontrol process. Fungal and bacterial diversity in treated sorghum mini-silos was analyzed by high-throughput sequencing. Firmicutes was the predominant bacterial phyla and Lactobacillus represented the most abundant genus, while Ascomycota was the predominant fungal phyla with Saccharomyces and Pichia as the major genera. Bacterial and yeast counts before and after incubation indicated that the microbial community obtained from WK was able to grow in the sorghum mini-silos in the presence of A. flavus. Results of the present work indicate that the use of a mixed inoculum of microorganisms present in WK may represent an alternative management practice to avoid the growth of A. flavus in ensiled sorghum grains and the concomitant contamination with aflatoxins.
Highlights
Silage plays a significant role as a source of animal feed, especially in areas where the demand for feed is year-round
(MFiicgrouorrega1n)is.mLsi2n0e1a9,r7it,yx FwOaRsPoEbEsRerRvEeVdIEoWver the range between 53 ng to 5.3 × 10−3 ng of DNA per rea3ctoifo1n8, rwehacictihonco, nwsthiitcuhtecdotnhsetidtuytnedamthicerdanygneamoficthreanmgeethoofdt.hBeamseedthoondt.hBisasceadlcuolnattiohnisacnadlctuhleattiroenatamnednthoef tsrraeeamactptmiloeensn,ptwroiofhriscathompcpoelrnefssotrpimtruiiotnergdtoqtPhpCeeRrdf,oytrhnmeamiqnuigcaqnrPtainCfigcRae,titoohfnetlqhimueaimtnoteifftihtchoaedtim.onBetalhismoedditeooxnfptrthehseissmedceaitlnhcoutedlarmteixospnorfeasnsademdtphilnee twtereremaigtsmhtoewfntsaasomf3.sp2almnegpwDleeNisgpAhrt/igow.raEtsoffi3pc.i2eernnfocgyrmDcaiNnlcgAu/qlagPt.eCEdRff,irctohimeenqtchyueacnsaltloicfpuicelaaottifeodtnheflricomumirtvotehfwethaselsom1p0ee2t%ohfo, dtinhedexicpuartreivsnsegewdthaianst 1tthe0re2m%qsP, CionfRdsiaacsamstaipnylgewtwhasaetihgtihhgtehlwqyPaeCsffiR3c.2aiesnnsatg.yDwNaAs h/gig. hElfyfiecifefinccieynct.alculated from the slope of the curve was 102%, indicating that the qPCR assay was highly efficient
Aspergillus flavus biomass was quantified by qPCR in a Rotor-Gene 6000TM (Corbett Life Science, Sydney, Australia) thermocycler according to the protocol described by Shweta et al (2013) [49] with some modifications. qPCR Reactions were performed in duplicate using a total volume of 10 μL for each sample, consisting of 5μL of Rotor-GeneTM SYBR® Green PCR Master Mix (Quiagen, Venlo, Netherlands), 0.5 μL of primer omt-F (5 -GACCAATACGCCCACACAG-3 ) and 0.5 μL primer omt-R (5 -CTTTGGTAGCTGTTTCTCGC-3 ) (10 μM each) (2013) [49], 1 μL of template DNA solution and 3 μL sterile miliQ water
Summary
Silage plays a significant role as a source of animal feed, especially in areas where the demand for feed is year-round. High-moisture grain silage in Uruguay is an important food resource for livestock and dairy animals It represents a higher quality energy source than dry grain, is economically cheaper, and its production is less dependent on weather conditions [2]. Previous work of García y Santos (2012) [7] performed in Uruguay, indicated that Aspergillus flavus was the main contaminant species in sorghum silage. These results are in agreement with Keller et al (2012) [8], who identified Aspergillus flavus as the main spoiler of sorghum silage in the south of Brazil. Consumption of aflatoxin-contaminated feed is associated with reduced animal performance
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