The occurrence of Paenibacillus and Clostridium spores in silage is of great concern for dairy producers because their spores can contaminate milk and damage processed milk and semi-hard cheeses. Spoiled silage is considered to be the main contamination source of the total mixed ration (TMR), feces of dairy cows, and consequently bulk tank milk via the contamination of cow teats by dirt during milking. The presence of an anaerobic and facultative anaerobic sporeformer population in different matrices (soil, corn silage, other feeds, TMR, feces, and milk) and its transmission pathway has been studied on 49 dairy farms by coupling plate count data with 16S-DNA identification. The different matrices have shown a high variability in the anaerobic and facultative anaerobic spore count, with the highest values being found in the aerobically deteriorated areas of corn silages. Clostridium tyrobutyricum, Paenibacillus macerans, and Paenibacillus thermophilus were detected in all the matrices. The TMR spore count was influenced by the amount of spoiled corn silage in the TMR and by the care taken when cleaning the spoiled silage before feed-out. Most of the farms that prevent the presence of visible moldy silage in the silo and carefully clean to remove molded spots were able to maintain their TMR spore counts below 4.0 log spores/g. When a level of 4.5 log spores/g of TMR was exceeded, the feces presented a greater contamination than 3.0 log spores/g. Moreover, the higher the number of spores in the feces was, the higher the number of spores in the milk. Most of the farms that presented a feces contamination greater than 5.0 log spores/g had a higher milk spore contamination than 1,000 spores/L. Careful animal cleaning and good milking practices have been found to be essential to maintain low levels of contamination in bulk tank milk, but it has emerged that only by coupling these practices with a correct silage management and cleaning during TMR preparation can the contamination of milk by spores be kept at a low level. It has been found that aerobically deteriorated silage has a great capacity to contaminate TMR and consequently to increase the risk of milk spore contamination, even when routine milking practices are adopted correctly.

The environment temperature and its effect on the temperature of silage is very important for the fermentation and subsequent quality of a silage. Obligate heterofermentative lactic acid bacteria (LAB) inocula, because of their ability to inhibit yeasts, have been developed to prevent the aerobic deterioration of silages. The temperature during silage conservation may also play an important role in the fermentation profile of silages. This study has evaluated the effect of temperature, during the conservation of whole crop corn silage, untreated or treated with different LAB inocula, on the fermentation profile and on the aerobic stability of the silage. Corn was harvested at 42% dry matter and either not treated (control) or treated with Lactobacillus buchneri NCIMB 40788 (LB) at 300,000 cfu/g fresh matter (FM); Lactobacillus hilgardii CNCM I-4785 at 150,000 cfu/g FM (LH150); L. hilgardii CNCM I-4785 at 300,000 cfu/g FM (LH300); or LB+LH at 150,000 cfu/g FM each. In an attempt to experimentally simulate temperature fluctuations in the mass or at the periphery of a silage bunker, corn was conserved in laboratory silos at a constant temperature (20 ± 1°C; MASS) or at lower and variable outdoor temperatures (PERIPH; ranging from 0.5 to 19°C), and the silos were opened after 15, 30, and 100 d of conservation. Lactic acid, acetic acid, and ethanol contents increased in all the silages over the conservation period. The lactic acid content was higher (+10%) in the silages kept at a constant temperature than those conserved at the lower and variable outdoor temperatures. The acetic acid was higher in the treated silages than in the control ones conserved at a constant temperature for 100 d. Moreover, 1,2-propanediol was only detected in the treated silages after at least 30 d at a constant temperature, whereas only traces were detected in the LB+LH treatment for the other temperature conditions. The yeast count decreased during conservation at a slower rate in PERIPH than in MASS and on average reached 2.96 and 4.71 log cfu/g for MASS and PERIPH, respectively, after 100 d of conservation. The highest aerobic stability values were observed for LH300 (191 h) in the MASS silage after 100 d of conservation, whereas the highest aerobic stability was observed in LB+LH (150 h) in the PERIPH silages. After 7 d of air exposure, a pH higher than 4.5 and a higher yeast than 8.0 log cfu/g were detected in all the silages opened after 15 and 30 d of conservation. A pH value close to that of silo opening was detected in the LB, LH150, and LH300 silages conserved under MASS conditions after 100 d, whereas LB+LH was the most effective under PERIPH conditions. The temperature and its fluctuation during conservation of silage in laboratory silos influenced the fermentation, which in turn had an effect on the quality of silage and on the extent of the effect of LAB inocula.

The growth of Aspergillus flavus and the production of aflatoxins (AF) during the aerobic deterioration of corn silage represent a problem for animal and human health. This experiment was conducted to evaluate whether the presence of A. flavus and AF production originate from the field or additional AF are produced during the fermentation phase or during aerobic deterioration of corn silage. The trial was carried out in northern Italy on corn at a dry matter (DM) level of 34%. The fresh herbage was either not treated (C) or treated with a Lactobacillus buchneri (LB) NCIMB 40788 [(at 3 × 105 cfu/g of fresh matter (FM)], Lactobacillus hilgardii (LH) CNCM I-4785 (at 3 × 105 cfu/g of FM), or their combination (LB+LH; at 1.5 × 105 cfu/g of FM of each strain) ensiled in 20-L silos and opened after 250 d of ensiling. After silo opening, the aerobic stability was evaluated and samples were taken after 7 and 14 d of air exposure. The pre-ensiled material, the silages at silo opening, and the aerobically exposed silages were analyzed for DM content, fermentative profiles, microbial count, nutritive characteristics, DM losses, and AFB1, AFB2, AFG1, and AFG2 contents. Furthermore, a subsample of colonies with macromorphological features of Aspergillus section Flavi was selected for AF gene pattern characterization and in vitro AF production. The presence of A. flavus was below the detection limit (<1.00 log10 cfu/g) in the fresh forage before ensiling, whereas it was found in 1 out of 16 silage samples at silo opening at a level of 1.24 log10 cfu/g. The AF were found in both the fresh forage and at opening in all the samples, with a predominance of AFB2 (mean value of 1.71 μg/kg of DM). The inoculation of lactic acid bacteria determined a reduction in the lactic-to-acetic ratio compared with the control. A larger amount of acetic acid resulted in a lower yeast count and higher aerobic stability in the treated silages than in the control ones. At the beginning of aerobic deterioration, the yeasts increased to over 5 log10 cfu/g, whereas the molds were close to the value observed at silo opening. When the inhibiting conditions were depleted (pH and temperature higher than 5 and 35°C, respectively), both the total molds and A. flavus reached higher values than 8.00 and 4.00 log10 cfu/g, respectively, thus determining the ex novo production of AFB1 during aerobic deterioration, regardless of treatments. The analysis of gene pattern showed that 64% of the selected colonies of A. flavus showed the presence of all 4 AF gene patterns, and 43% of the selected colonies were able to produce AF in vitro. During air exposure, after 1,000°C·h have been cumulated, starch content decreased (below 10% DM) and concentration of neutral detergent fiber, acid detergent fiber, hemicelluloses, crude protein, and ash increased. The inoculation with LB and LB+LH increased the aerobic stability of the silages and delayed the onset of aerobic microbial degradation, which in turn indirectly reduced the risk of A. flavus outgrowth and AFB1 production after silage opening.

Heterofermentative Lentilactobacillus hilgardii isolated from sugarcane silage, has recently been proposed as a silage inoculant to increase aerobic stability. Various conditions can influence the activity of LAB and their ability to alter silage quality (e.g., DM content and length of conservation). The aim of this study has been to evaluate the effect of L. hilgardii on the fermentation quality and aerobic stability of whole crop corn silage with different DM contents (from 26 to 45%), conserved for various conservation lengths (13–272 days). The silages were analyzed for their DM content, pH, fermentative profile, microbial count, and aerobic stability. L. hilgardii showed a positive effect on improving the aerobic stability of silages, due its ability to produce acetic acid, and reduced the yeast count. The acetic acid content increased as the conservation period increased and decreased as the DM content increased. The yeast count was reduced during conservation in a DM dependent manner and the inoculation with LH determined a reduction in the count of 0.48 log cfu/g. The aerobic stability increased as the conservation period increased, and the treatment with LH on average increased the aerobic stability by 19 h. The results of this experiment suggest that higher aerobic stability could be achieved in corn silages by ensiling at medium or low DM contents, or by increasing the length of conservation if a higher DM content at ensiling is needed. The inoculation with LH helps to improve the aerobic stability of corn silages by reducing the yeast count.

Computer Vision is an approach of Artificial Intelligence (AI) that conceptually enables "computers and systems to derive useful information from digital images" giving access to higher-level information and "take actions or make recommendations based on that information". Comprehensive two-dimensional chromatography gives access to highly detailed, accurate, yet unstructured information on the sample's chemical composition, and makes it possible to exploit the AI concepts at the data processing level (e.g., by Computer Vision) to rationalize raw data explorations. The goal is the understanding of the biological phenomena interrelated to a specific/diagnostic chemical signature. This study introduces a novel workflow for Computer Vision based on pattern recognition algorithms (i.e., combined untargeted and targeted UT fingerprinting) which includes the generation of composite Class Images for representative samples' classes, their effective re-alignment and registration against a comprehensive feature template followed by Augmented Visualization by comparative visual analysis. As an illustrative application, a sample set originated from a Research Project on artisanal butter (from raw sweet cream to ripened butter) is explored, capturing the evolution of volatile components along the production chain and the impact of different microbial cultures on the finished product volatilome. The workflow has significant advantages compared to the classical one-step pairwise comparison process given the ability to realign and pairwise compare both targeted and untargeted chromatographic features belonging to Class Images resembling chemical patterns from many different samples with intrinsic biological variability.