Bovine Milk Microbiota: Comparison among Three Different DNA Extraction Protocols To Identify a Better Approach for Bacterial Analysis

Cremonesi, Paola; Romanò, Alicia; Sala, Lorenza; Luini, M.; Castiglioni, Bianca

doi:10.1128/spectrum.00374-21

Cited by 8 publications

(7 citation statements)

References 32 publications

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“…The profile of the decontaminated milk microbiome data in this study showed that Staphylococcus and Acinetobacter were the predominant taxa, which is in line with previous studies [53][54][55]. However, Bacteroidetes, Corynebacterium, Pseudomonas and Staphylococcus might be overestimated in previous bovine milk microbiome studies [4,17,24,25,53,56], as they were identified as contaminants in our study.…”

Section: Characterization Of the De-contaminated Milk Microbiomesupporting

confidence: 91%

The impact of kit, environment and sampling contamination on the observed microbiome of bovine milk

Dean,

Deng,

Wehri

et al. 2023

Preprint

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Contaminants can easily outnumber bacteria that originate within the milk itself, milk microbiome research currently suffers from a critical knowledge gap; namely, does non-mastitis bovine milk contain a native microbiome? In this study, we sampled external and internal mammary epithelium, stripped and cisternal milk, used numerous negative controls to identify potential sources of microbial contamination. Two algorithms were used to mathematically remove this contamination and to track potential movement of microbes among our samples. Our results suggest that majority (i.e., >75%) of the sequence data generated from bovine milk and mammary epithelium samples represents contaminating DNA. The contaminants in milk samples were primarily sourced from the DNA extraction kits and the internal and external skin of the teat, while the teat canal and apex samples were mainly contaminated during the sampling process. After decontamination, the milk microbiome displayed a more dispersed, less diverse and compositionally distinct bacterial profile compared with the teat skin samples. Similar microbial compositions were observed between cisternal and stripped milk samples, as well as between teat apex and canal samples.StaphylococcusandAcinetobacterwere the predominant genera detected in the sequences of milk samples, and bacterial culture showed growth ofStaphylococcusandCorynebacteriumspp. in 50% (7/14) of stripped milk samples and growth ofStaphylococcusspp. in 7% (1/14) of cisternal milk samples. Our study suggests that microbiome data generated from milk samples obtained from clinically healthy bovine udders may be heavily biased by contaminants that enter the sample during the sample collection and processing workflows.ImportanceObtaining a non-contaminated sample of bovine milk is challenging due to the nature of the sampling environment and the route by which milk is typically extracted from the mammary gland. Furthermore, the very low bacterial biomass of bovine milk exacerbates the impacts of contaminant sequences in downstream analyses, which can lead to severe biases. Our finding showed that bovine milk contains very low bacterial biomass, and each contamination event (including sampling procedure and DNA extraction process) introduces bacteria and/or DNA fragments that easily outnumber the native bacterial cells. This finding has important implications for our ability to draw robust conclusions from milk microbiome data, especially if the data have not been subjected to rigorous decontamination procedures. Based on these findings, we strongly urge researchers to include numerous negative controls into their sampling and sample processing workflows; and to utilize several complementary methods for identifying potential contaminants within the resulting sequence data. These measures will improve the accuracy, reliability, reproducibility, and interpretability of milk microbiome data and research.

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Section: Characterization Of the De-contaminated Milk Microbiomesupporting

confidence: 91%

The impact of kit, environment and sampling contamination on the observed microbiome of bovine milk

Dean,

Deng,

Wehri

et al. 2023

Preprint

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“…Another issue that makes the bacteria in milk important for healthcare is the potential to carry antimicrobial resistance genes, because antibiotic consumption in animal husbandry can be unconscious in most developing countries [ 47 , 48 ]. Antibiotic resistance is not only a major crisis for non-healthy animals (with mastitis), but also a potential problem for healthy animals.…”

Section: Discussionmentioning

confidence: 99%

The Characterization and Beta-Lactam Resistance of Staphylococcal Community Recovered from Raw Bovine Milk

et al. 2023

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Staphylococci is an opportunistic bacterial population that is permanent in the normal flora of milk and poses a serious threat to animal and human health with some virulence factors and antibiotic-resistance genes. This study was aimed at identifying staphylococcal species isolated from raw milk and to determine hemolysis, biofilm, coagulase activities, and beta-lactam resistance. The raw milk samples were collected from the Düzce (Türkiye) region, and the study data represent a first for this region. The characterization of the bacteria was performed with MALDI-TOF MS and 16S rRNA sequence analysis. The presence of coa, icaB, blaZ, and mecA was investigated with PCR. A nitrocefin chromogenic assay was used for beta-lactamase screening. In this context, 84 staphylococci were isolated from 10 different species, and the dominant species was determined as S. aureus (32.14%). Although 32.14% of all staphylococci were positive for beta hemolysis, the icaB gene was found in 57.14%, coa in 46.42%, mecA in 15.47%, and blaZ in 8.33%. As a result, Staphylococcus spp. strains that were isolated from raw milk in this study contained some virulence factors at a high level, but also contained a relatively low level of beta-lactam resistance genes. However, considering the animal–environment–human interaction, it is considered that the current situation must be monitored constantly in terms of resistance concerns. It must not be forgotten that the development of resistance is in constant change among bacteria.

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“…The relationship between cows and microorganisms has traditionally been related to intramammary infections [64]. However, recent metagenomic studies indicate that mammary secretions from clinically healthy quarters may harbor genomic markers for detecting several bacterial groups, the majority of which were not associated with mastitis but were defined as "commensal mammary microbiota" [64,98,99]. Whether disruption of udder microbiota diversity (dysbiosis) plays a role in determining susceptibility to mastitis is still unknown, and little is known about the contributions of various biotic and abiotic factors on the composition of udder microbiota diversity [64,100,101].…”

Section: Relationship Between Genomic Genotypic and Phenotypic Udder ...mentioning

confidence: 99%

Genomic and Phenotypic Udder Evaluation for Dairy Cattle Selection: A Review

Gutiérrez-Reinoso

Aponte

García-Herreros

2023

Animals

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The traditional point of view regarding dairy cattle selection has been challenged by recent genomic studies indicating that livestock productivity prediction can be redefined based on the evaluation of genomic and phenotypic data. Several studies that included different genomic-derived traits only indicated that interactions among them or even with conventional phenotypic evaluation criteria require further elucidation. Unfortunately, certain genomic and phenotypic-derived traits have been shown to be secondary factors influencing dairy production. Thus, these factors, as well as evaluation criteria, need to be defined. Owing to the variety of genomic and phenotypic udder-derived traits which may affect the modern dairy cow functionality and conformation, a definition of currently important traits in the broad sense is indicated. This is essential for cattle productivity and dairy sustainability. The main objective of the present review is to elucidate the possible relationships among genomic and phenotypic udder evaluation characteristics to define the most relevant traits related to selection for function and conformation in dairy cattle. This review aims to examine the potential impact of various udder-related evaluation criteria on dairy cattle productivity and explore how to mitigate the adverse effects of compromised udder conformation and functionality. Specifically, we will consider the implications for udder health, welfare, longevity, and production-derived traits. Subsequently, we will address several concerns covering the application of genomic and phenotypic evaluation criteria with emphasis on udder-related traits in dairy cattle selection as well as its evolution from origins to the present and future prospects.

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Bovine Milk Microbiota: Comparison among Three Different DNA Extraction Protocols To Identify a Better Approach for Bacterial Analysis

Cited by 8 publications

References 32 publications

The impact of kit, environment and sampling contamination on the observed microbiome of bovine milk

The impact of kit, environment and sampling contamination on the observed microbiome of bovine milk

The Characterization and Beta-Lactam Resistance of Staphylococcal Community Recovered from Raw Bovine Milk

Genomic and Phenotypic Udder Evaluation for Dairy Cattle Selection: A Review

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