The microbiota of ensiled forages and of bulk tank milk on dairy cattle farms in northern Sweden - a case study.

Factors contributing to variations in the quality and microbiota of ensiled forages and in bulk tank microbiota in milk from cows fed different forages were investigated. Nutritional quality, fermentation parameters and hygiene quality of forage samples and corresponding bulk tank milk samples collected in 3 periods from 18 commercial farms located in northern Sweden were compared. Principal coordinates analysis revealed that the microbiota in forage and bulk milk, analyzed using 16S rRNA gene-based amplicon sequencing, were significantly different. The genera Lactobacillus, Weissella and Leuconostoc dominated in forage samples, whereas Pseudomonas, Staphylococcus and Streptococcus dominated in bulk milk samples. Forage quality and forage-associated microbiota were affected by ensiling method and by use of silage additive. Forages stored in bunker and tower silos (confounded with use of additive) were associated with higher levels of acetic and lactic acid and Lactobacillus. Forage ensiled as bales (confounded with no use of additive) was associated with higher dry matter content, water-soluble carbohydrate content, pH, yeast count and the genera Weissella, Leuconostoc and Enterococcus. For bulk tank milk samples, milking system was identified as the major factor affecting the microbiota and type of forage preservation had little impact. Analysis of common amplicon sequence variants (ASVs) suggested that forage was not the major source of Lactobacillus found in bulk tank milk.

Epiphytic microbiota in Swedish grass-clover herbage and the effect of silage additives on fermentation profiles and bacterial community compositions of the resulting silages.

AIMS: To investigate the epiphytic microbiota in grass-clover herbage harvested at different sites and occasions and to explore the effect of different silage additives on the resulting silage microbiota. METHODS AND RESULTS: Herbage was harvested from grass-clover leys at geographically distributed sites in a long-term field experiment in Sweden, in early and late season of two consecutive years. Different silages were made from the herbage using: (1) no additive, (2) acid-treatment, and (3) inoculation by starter culture. Herbages were analysed for botanical and chemical composition, and the resulting silages for products of fermentation. Bacterial DNA was extracted from herbage and silage samples, followed by sequencing using Illumina 16S rRNA amplicon sequencing. Herbage microbiota showed no clear correlation to site or harvesting time. Silage additives had a major effect on the ensiling process; inoculation resulted in well fermented silages comprising a homogenous microbiota dominated by the genera Lactobacillus and Pediococcus. A minor effect of harvest time was also observed, with generally a more diverse microbiota in second-harvest silages. Untreated silages showed a higher relative abundance (RA) from non-lactic acid bacteria compared to acid-treated silages. In most silages, only a few bacterial amplicon sequence variants contributed to most of the RA. CONCLUSIONS: The epiphytic microbiota in grass-clover herbage were found to be random and not dependent on site. From a microbial point of view, the most predictable and preferable silage outcome was obtained by inoculation with a starter culture. Acid-treatment with formic- and propionic acid surprisingly resulted in a less preferable silage. Silage making without additives cannot be recommended based on our results.

Near-infrared hyperspectral image analysis for monitoring the cheese-ripening process.

Ripening is the most crucial process step in cheese manufacturing and constitutes multiple biochemical alterations that describe the final cheese quality and its perceived sensory attributes. The assessment of the cheese-ripening process is challenging and requires the effective analysis of a multitude of biochemical changes occurring during the process. This study monitored the biochemical and sensory attribute changes of paraffin wax-covered long-ripening hard cheeses (n = 79) during ripening by collecting samples at different stages of ripening. Near-infrared hyperspectral (NIR-HS) imaging, together with free amino acid, chemical composition, and sensory attributes, was studied to monitor the biochemical changes during the ripening process. Orthogonal projection-based multivariate calibration methods were used to characterize ripening-related and orthogonal components as well as the distribution map of chemical components. The results approve the NIR-HS imaging as a rapid tool for monitoring cheese maturity during ripening. Moreover, the pixelwise evaluation of images shows the homogeneity of cheese maturation at different stages of ripening. Among the chemical compositions, fat content and moisture are the most important variables correlating to NIR-HS images during the ripening process.

Milking system and premilking routines have a strong effect on the microbial community in bulk tank milk.

In this study, we investigated the variation in the microbial community present in bulk tank milk samples and the potential effect of different farm management factors. Bulk tank milk samples were collected repeatedly over one year from 42 farms located in northern Sweden. Total and thermoresistant bacteria counts and 16S rRNA gene-based amplicon sequencing were used to characterize microbial community composition. The microbial community was in general heterogeneous both within and between different farms and the community composition in the bulk tank milk was commonly dominated by Pseudomonas, Acinetobacter, Streptococcus, unclassified Peptostreptococcaceae, and Staphylococcus. Principal component analysis including farm factor variables and microbial taxa data revealed that the microbial community in milk was affected by type of milking system. Milk from farms using an automatic (robot) milking system (AMS) and loose housing showed different microbial community composition compared with milk from tiestall farms. A discriminant analysis model revealed that this difference was dependent on several microbial taxa. Among farms using an automatic milking system, there were further differences in the microbial community composition depending on the brand of the milking robot used. On tiestall farms, routines for teat preparation and cleaning of the milking equipment affected the microbial community composition in milk. Total bacteria count (TBC) in milk differed between the farm types, and TBC were higher on AMS than tiestall farms (log 4.05 vs. log 3.79 TBC/mL for AMS and tiestalls, respectively). Among tiestall farms, milk from farms using a chemical agent in connection to teat preparation and a more frequent use of acid to clean the milking equipment had lower TBC in milk, than milk from farms using water for teat preparation and a less frequent use of acid to clean the milking equipment (log 3.68 vs. 4.02 TBC/mL). There were no significant differences in the number of thermoresistant bacteria between farm types. The evaluated factors explained only a small proportion of total variation in the microbiota data, however, despite this, the study highlights the effect of routines associated with teat preparation and cleaning of the milking equipment on raw milk microbiota, irrespective of type of milking system used.

Composition and properties of bovine milk: A study from dairy farms in northern Sweden; Part I. Effect of dairy farming system.

This study was part of a larger project that aimed to understand the causes for increasing variation in cheese ripening in a cheese-producing region in northern Sweden. The influence of different on-farm factors on raw milk composition and properties was investigated and is described in this paper, whereas the monthly variation in the milk quality traits during 1 yr is described in our companion paper. The dairy farming systems on a total of 42 dairy farms were characterized through a questionnaire and farm visits. Milk from farm tanks was sampled monthly over 1 yr and analyzed for quality attributes important for cheese making. On applying principal component analyses to evaluate the variation in on-farm factors, different types of farms were distinguished. Farms with loose housing and automatic milking system (AMS) or milking parlor had a higher number of lactating cows, and predominantly Swedish Holstein (SH) breed. Farms associated with tiestalls had a lower number of lactating cows and breeds other than SH. Applying principal component analyses to study the variation in composition and properties of tank milk samples from farms revealed a tendency for the formation of 2 clusters: milk from farms with AMS or a milking parlor, and milk from farms with tiestall milking. The interaction between the milking system, housing system, and breed probably contributed to this grouping. Other factors that were used in the characterization of the farming systems only showed a minor influence on raw milk quality. Despite the interaction, milk from tiestall farms with various cow breeds had higher concentrations (g/100 g of milk) of fat (4.74) and protein (3.63), and lower lactose concentrations (4.67) than milk from farms with predominantly SH cows and AMS (4.32, 3.47, and 4.74 g/100 g of milk, respectively) or a milking parlor (4.47, 3.54, and 4.79 g/100 g of milk, respectively). Higher somatic cell count (195 × 103/mL) and lower free fatty acid concentration (0.75 mmol/100 g of fat) were observed in milk from farms with AMS than in milk from tiestall systems (150 × 103/mL and 0.83 mmol/100 g of fat, respectively). Type of farm influenced milk gel strength, with milk from farms with predominantly SH cows showing the lowest gel strength (65.0 Pa), but not a longer rennet coagulation time. Effects of dairy farming system (e.g., dominant breed, milking system, housing, and herd size) on milk quality attributes indicate a need for further studies to evaluate the in-depth effects of farm-related factors on milk quality attributes.

Composition and properties of bovine milk: A study from dairy farms in northern Sweden; Part II. Effect of monthly variation.

This study investigated the influence of monthly variation on the composition and properties of raw farm milk collected as part of a full-scale cheese-making trial in a region in northern Sweden. In our companion paper, the contribution of on-farm factors to the variation in milk quality attributes is described. In total, 42 dairy farms were recruited for the study, and farm milk samples were collected monthly over 1 yr and characterized for quality attributes of importance for cheese making. Principal component analysis suggested that milk samples collected during the outdoor period (June-September) were different from milk samples collected during the indoor period. Despite the interaction with the milking system, the results showed that fat and protein concentrations were lower in milk collected during May through August, and lactose concentration was higher in milk collected during April through July than for the other months. Concentrations of free fatty acids were generally low, with the highest value (0.86 mmol/100 g of fat) observed in February and the lowest (0.70 mmol/100 g of fat) observed in June. Plasmin and plasminogen-derived activities varied with sampling month without a clear seasonal pattern. The pH of farm tank milk ranged from 6.60 to 6.82, with the lowest and highest values in September and February, respectively. The highest somatic cell count was observed in August (201 × 103 cells/mL) and the lowest in April (143 × 103 cells/mL). The highest value of gel strength, was recorded in December (88 Pa) and the lowest in July (64 Pa). Rennet coagulation time and gel strength were inversely correlated, with the lowest rennet coagulation time value observed in December. Orthogonal projections to latent structures (OPLS) and discriminant analysis adaptation of OPLS identified casein micelle size and total proteolysis as the milk quality attributes with major responses to sampling month, with smaller casein micelle size and higher total proteolysis associated with the outdoor months. Using discriminant analysis adaptation of OPLS to further investigate causes behind the variation in milk traits revealed that there were factors in addition to feeding on pasture that differed between outdoor and indoor months. Because fresh grass was seldom the primary feed in the region during the outdoor period, grazing was not considered the sole reason for the observed difference between outdoor and indoor periods in raw milk quality attributes.

Associations between the Bacterial Composition of Farm Bulk Milk and the Microbiota in the Resulting Swedish Long-Ripened Cheese.

The maturation of a traditional Swedish long-ripened cheese has shown increasing variation in recent years and the ripening time is now generally longer than in the past. While the cheese is reliant on non-starter lactic acid bacteria for the development of its characteristic flavour, we hypothesised that the observed changes could be due to variations in the microbiota composition and number of bacteria in the raw milk used for production of the cheese. To evaluate associations between microbiota in the raw milk and the resulting cheese, three clusters of commercial farms were created to increase variation in the microbiota of dairy silo milk used for cheese production. Cheese production was performed in three periods over one year. Within each period, milk from the three farm clusters was collected separately and transported to the cheese production facility. Following pasteurisation, the milk was processed into the granular-eyed cheese and matured at a dedicated cheese-ripening facility. For each cheese batch, farm bulk and dairy silo milk samples, a starter culture, early process samples and cheese samples from different stages of maturation (7-20 months) were collected and their microbiota characterised using 16S rRNA amplicon sequencing. The microbiota in the farm bulk milk differed significantly between periods and clusters. Differences in microbiota in dairy silo milk were observed between periods, but not between farm clusters, while the cheese microbiota differed between periods and clusters. The top 13 amplicon sequence variants were dominant in early process samples and the resulting cheese, making up at least 93.3% of the relative abundance (RA). Lactococcus was the dominant genus in the early process samples and, together with Leuconostoc, also dominated in the cheese samples. Contradicting expectations, the RA of the aroma-producing genus Lactobacillus was low in cheese during ripening and there was an unexpected dominance of starter lactic acid bacteria even at the later stages of cheese ripening. To identify factors behind the recent variations in ripening time of this cheese, future studies should address the effects of process variables and the dairy environment.