The volatile organic compounds of milk contaminated with psychrotrophic bacteria were studied by HS‐SPME and GC/MS. Pseudomonas fluorescens PS14, Pseudomonas fragi PS55, Pseudomonas mosselii PS39, Pseudomonas rhodesiae PS62 and Serratia marcescens S92 were inoculated in sterilised milk (2.5% fat) stored at either 5 °C or 10 °C. A total of 47 volatile organic compounds (VOCs) belonging to seven chemical groups were identified in the spoiled milk. Volatile organic compound patterns peculiar to the inoculate bacterial strains were highlighted. 3‐Methylbutan‐1‐ol, 2 methylpropan‐1‐ol, 3‐hydroxybutan‐2‐one, butan‐2,3‐dione, butanoic and hexanoic acids were revealed as potential chemical spoilage indexes of milk spoilage due to the activity of the five psychrotrophic strains studied.

Clostridia in the milk can lead to late blowing, a cheese defect. Clostridia are ubiquitous, deriving from both the farm environment and the feed ingested by the cows, and are transferred into the milk through faecal contamination. Our aim was to investigate the effect of different in-parlour practices on the content of anaerobic spore-forming bacteria in milk, and to monitor the variation in spore content in the feed and environment. The experiment, conducted in an experimental dairy during autumn, was repeated in exactly the same way for two consecutive years. The experimental design applied three different milking routines in three consecutive 7-d periods: forestripping alone (F); forestripping and post-dipping (F+Post); pre-dipping, wiping, forestripping and post-dipping (Pre+F+Post). Teat skin swabs and samples of feed, faeces, bedding materials and milk were collected for microbiological analyses. The dietary forage of the lactating cows included maize silage, which, in both years, was found to have the highest level of clostridial spore contamination. Pre-dipping with a detergent/emollient solution, and drying with a disposable paper towel, proved much more efficient in reducing spore contamination than forestripping alone, both on the teats (1·30 vs. 2·20 log 10 MPN/swab; P < 0·001) and in the milk (1·82 vs. 2·47 log 10 MPN/L, P < 0·02), while post-dipping had little influence on spore count. The standard plate count in milk was significantly lower with Pre+F+Post treatment than with F (3·80 vs. 4·51 log 10 CFU/mL, P < 0·01). The teat preparation procedure did not influence the lactic acid bacterial levels in the milk, which is very positive in that decreased lactic acid bacterial content can lessen raw milk cheese quality.

The ability of 15 psychrotrophic strains to release fatty acids C‐8:0 to C‐20:1 from cream by GC/MS and the substrate specificity of their lipases against selected triacylglycerols (TAG) were studied. The free fatty acids (FFAs) detected in the highest quantities were, in general, those of long (C‐16; C‐18) and intermediate (C‐12; C‐14) even‐numbered carbon chain and unsaturated C‐18:1 and C‐18:2. The ability of lipolytic strains to release fatty acids from milk fat varies according to the bacterial genus, among species and even with the strain of a given species. TAG breakdown by psychrotrophs may occur in a greater or lesser extent, but type and amount of released FFAs are not easily predictable. Pseudomonas rhodesiae showed the highest hydrolytic resistance towards all tested fatty acid triglycerides.

An autochthonous starter culture was used in the production of Protected Designation of Origin (PDO) Silter raw milk cheese, and compared to a control cheese in order to study its influence on microbial population dynamics during ripening. Curd and cheese at different ripening periods (0, 30, 60 and 200 days) from two independent dairies were analysed combining culture-based microbiological analysis, Length Heterogeneity-PCR and 16S microbiome profiling.The autochthonous starter determined higher lactic acid bacteria (LAB) levels for the first 30 days of ripening, without interfering with secondary microbiota that determines the tipicality of this cheese. Only a few genera and species persisted in the cheese despite the microbial richness of the curd. In addition, the high levels of different LAB reduced harmful microorganisms. The various analytic methods used resulted in discrepancies in the proportions of Enterococcus, Lactococcus and Lactobacillus spp., but the pivotal role of Streptococcus and Lactococcus genera was evident. The dominant species included those selected to formulate the starter (St thermophilus, Lc. lactis, Lit mesenteroides) as well as Lb. paracasei.The addition of autochthonous starter proved to he effective in controlling the first phases of the cheese-making, without compromising cheese typicality. (C) 2017 Elsevier Ltd. All rights reserved.

Sustainable exploitation of agro-industrial by-products has attracted great interest in cereal bran valorization. In this research, a polyphasic approach has been carried out to characterize maize bran at microbiological and chemical level during a sourdough like fermentation process, in order to enhance its technological and nutritional properties. Autochthonous microbiota was isolated at different refreshment steps and subjected to identification and molecular characterization. Fermentation was characterized by a rapid increase in lactic acid bacteria and yeasts, with a co-dominance, at the initial stage, of Weissella spp., Pediococcus spp. and Wickerhamomyces anomalus. At the end of the fermentation, a natural selection was produced, with the prevalence of Lactobacillus plantarum, Lactobacillus brevis and Kazachstania unispora. This is the first time that a specific association between LAB and yeasts is reported, during the maize bran fermentation process. Enzymatic activities related to this microbial consortium promoted a "destructuration" of the fiber fraction, an increase in soluble dietary fiber and a reduction of phytic acid content. Our data also evidenced a noticeable increment in ferulic acid. The results obtained indicate that fermentation processes represent an efficient biotechnological approach to increase nutritional and functional potential of maize bran. Moreover, the characterization of microbiota involved in natural fermentation process will allow the selection of specific biotypes, with appropriate metabolic and enzymatic activities, to conduct "tailored" fermentation processes and improve brans or whole-meal flours from both nutritional and technological points of view.

Bacterial resistance to antibiotics is a major global health problem and resistance of Pseudomonadaceae and Enterobacteriaceae is a serious concern. We investigated the prevalence of drug-resistance in a total of 80 psychrotrophic strains from bulk milk belonging to Pseudomonas genus (n. 63) and Enterobacteriaceae group (n. 17). All the strains were tested against 16 antibiotics. Pseudomonas were further investigated for their sensitivity against 12 additional antibiotics. Pseudomonas showed a high susceptibility toward fluoroquinolones, aminoglycosides, and piperacillin and, to a lesser extent, to imipenem, ceftazidime, cefepime. Thirty-five out of 63 Pseudomonas strains were susceptible to meropenem, while among antibiotics for which recommended breakpoints are not yet available, 55% of Pseudomonas strains had no inhibition halo in presence of nitrofurantoin, highlighting a resistance toward this drug. The results obtained in this study indicate a high efficiency of fluoroquinolones, chloramphenicol (94%), and kanamycin (76%) for Enterobacteriaceae while a high prevalence of resistant strains was found to ampicillin (13/17). Serratia marcescens is highly susceptible to fluoroquinolones, chloramphenicol, and kanamycin. Moreover, mupirocin seems to be the new antibiotic with the less efficacy for Enterobacteriaceae, with 41% of strains without halo, pointing out an important resistance. Further knowledge on resistance to known and new antibiotics among Pseudomonas species and Enterobacteriaceae of milk origin was acquired.

The aim of the study is to develop a scoring system for dairy farms in order to give specific information about the product and production process of milk. The scoring system, based on a multi-dimensional approach, was developed on the basis of data collected in 29 Italian dairy farms and included different aspects. For the evaluation of animal welfare, a selection of indicators set up in the European Project® assessment protocol for cattle 2009 was used. Environmental sustainability of milk production was assessed by a cradle-to-farm-gate Life Cycle Assessment. Laboratory analyses were carried out on bulk tank milk to evaluate microbiological, nutritional and nutraceutical quality. Nineteen variables were selected and retained to define six quality aspects: animal welfare, environmental and economic sustainability of farms, microbiological, nutritional and nutraceutical quality of milk. Each farm was visited twice; each visit received, for each variable, a score between 1 and 3 based on the frequency distribution of that variable in the farm sample. The relation among farm characteristics and quality aspects showed the importance to maximise dairy efficiency to improve environmental and economic sustainability of the farms and the inclusion of hay in dairy cows’ ration to enhance the nutraceutical and nutritional quality of milk. The proposed multi-dimensional scoring system is a practical tool: for the farmer, to support decisions for improving the quality of the product and the productive process; for the dairy company, as a value-added opportunity; for the consumer, who receives detailed information about nutritional characteristics and production system of dairy products.

Eighty psychrotrophic bacterial strains, isolated from different northwest Italian bulk tank milks destined for Grana Padano cheese production, were identified by 16S rRNA gene amplification and partial sequence analysis of the rpoB gene. Pseudomonas spp. were the most commonly occurring contaminants, P. fluorescens being the predominant isolated species, along with Enterobacteriaceae, primarily Serratia marcescens. RAPD-PCR was used to study genetic variability and distinguish closely related strains; a high degree of genetic heterogeneity among the strains was highlighted. All the strains were characterized for their ability to produce proteases, lipases and lecithinases at different temperatures (7, 22, and 30 °C). Forty-one of the psychrotrophic strains were positive for all the enzymatic activities. The highest number of positive strains for all the incubation temperatures was found for lipolytic activity (59), followed by proteolytic (31) and lecithinase (28) activities, and the enzymatic traits varied among the Pseudomonas and Enterobacteriaceae strains. The proteolytic psychrotrophic strains were screened for the presence of the aprX gene, coding for a heat-resistant metalloprotease in Pseudomonas spp. The aprX gene was detected in 19 of 63 Pseudomonas strains, and was widespread in the P. fluorescens strains (14/19). PRATICAL APPLICATION: The study provides new data on the enzymatic activity of Gram-negative psychrotrophic bacteria, useful in developing strategies to control the proteo-lipolytic spoilage of raw and processed milk that causes gelation, off-flavors, and loss of sensory quality and shelf life.

BACKGROUND Anaerobic spore-forming bacteria (ASFB) in milk derive from the farm environment, and the use of silages and management practices are the main responsible of milk ASFB contamination. The aim of this study was to evaluate the relationships between feeding, milking routine and cow hygiene and milk and Grana Padano cheese (produced with and without lysozyme) ASFB contamination. RESULTS The study involved 23 dairy farms. ASFB in corn silage were on average 2.34 ± 0.87 log10 MPN g(-1). For grass, Italian ryegrass and alfalfa, ASFB (log10 MPN g(-1)) were numerically higher for silages (3.22) than hays (2.85). The use of corn silages of high quality (high lactic and acetic acids concentrations) decreased the milk ASFB contamination, whilst the use of herbage silages did not affect it. The presence (>40%) of cows with dirty udders increased the ASFB contamination of milk, while forestripping had a positive effect (-9% ASFB). Ripened Grana Padano had an ASFB count below the analytical limit; Clostridium tyrobutyricum DNA was found only in wheels produced without lysozyme, which also showed late blowing. CONCLUSION The factors increasing milk spore contamination were corn silage quality, cow udder hygiene and inadequate milking routine. Late blowing was present only in cheeses without lysozyme.