Comparison of commercially-available preservatives for maintaining the integrity of bacterial DNA in human milk

Lackey, Kimberly A.; Williams, Janet E.; Price, William J.; Carrothers, Janae M; Brooker, Sarah L; Shafii, Bahman; McGuire, Mark A.; McGuire, Michelle K.

doi:10.1016/j.mimet.2017.08.002

Cited by 19 publications

(25 citation statements)

References 32 publications

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“…PCR was conducted using 96‐well plates (USA Scientific, Ocala, FL). The reaction mix was comprised of 12.5 μL Q5® Hot Start High‐Fidelity 2X Master Mix (New England Biolabs®, Inc., Ipswich, MA), 0.25 μL each of forward and reverse primers (27F and 534R, respectively; Carrothers et al, ; Frank et al, ; Lackey et al, ), 2 μL template DNA, and 8 μL nuclease‐free, sterile water to bring the first reaction volume to 23 μL. For each plate, 2 μL of nuclease‐free water was used as a negative control; 2 μL of Escherichia coli DNA (221 ng/mL) was used as a positive control.…”

Section: Methodsmentioning

confidence: 99%

Household composition and the infant fecal microbiome: The INSPIRE study

Lane

McGuire

et al. 2019

American J Phys Anthropol

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Objectives: Establishment and development of the infant gastrointestinal microbiome (GIM) varies cross-culturally and is thought to be influenced by factors such as gestational age, birth mode, diet, and antibiotic exposure. However, there is little data as to how the composition of infants' households may play a role, particularly from a cross-cultural perspective. Here, we examined relationships between infant fecal microbiome (IFM) diversity/composition and infants' household size, number of siblings, and number of other household members. Materials and methods:We analyzed 377 fecal samples from healthy, breastfeeding infants across 11 sites in eight different countries (Ethiopia, The Gambia, Ghana, Kenya, Peru, Spain, Sweden, and the United States). Fecal microbial community structure was determined by amplifying, sequencing, and classifying (to the genus level) the V1-V3 region of the bacterial 16S rRNA gene. Surveys administered to infants' mothers identified household members and composition.Results: Our results indicated that household composition (represented by the number of cohabitating siblings and other household members) did not have a measurable impact on the bacterial diversity, evenness, or richness of the IFM. However, we observed that variation in household composition categories did correspond to differential relative abundances of specific taxa, namely: Lactobacillus, Clostridium, Enterobacter, and Klebsiella.Discussion: This study, to our knowledge, is the largest cross-cultural study to date examining the association between household composition and the IFM. Our results indicate that the social environment of infants (represented here by the proxy of household composition) may influence the bacterial composition of the infant GIM, although the mechanism is unknown. A higher number and diversity of cohabitants and potential caregivers may facilitate social transmission of beneficial bacteria to the infant gastrointestinal tract, by way of shared environment or through direct

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Section: Methodsmentioning

confidence: 99%

Household composition and the infant fecal microbiome: The INSPIRE study

Lane

McGuire

et al. 2019

American J Phys Anthropol

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“…Participants hand expressed milk (∼15 mL) into sterile containers, and a representative aliquot of milk (∼2 mL) was chemically preserved with Milk Preservation Solution (Norgen Biotek Corporation, Thorold, ON) in a 1:1 ratio. Previous work has shown that this is an appropriate method to preserve the bacterial community structure of milk samples for 2–4 weeks (Lackey et al, ). Preserved milk samples remained in ambient temperature (regional mean temperature = 25°C [Bahuchet, ]; ranging from approximately 16–35°C) for up to 4 weeks, after which they were shipped on dry ice to Washington State University, and then stored at −20°C upon arrival.…”

Section: Methodsmentioning

confidence: 99%

“…Briefly, DNA from 0.5 mL of preserved milk was extracted using the Milk DNA Preservation and Isolation Kit (Norgen Biotek Corporation), including an additional 2 h lysozyme lysis (20 mg/mL) targeting Gram‐positive genera as described previously for preserved samples (Lackey et al, ). We also included a negative control of nuclease‐free water with each set of samples that were extracted.…”

Section: Methodsmentioning

confidence: 99%

“…Barcoded PCR amplicons generated from each sample were pooled to create a composite with equal mass of each sample (50 ng DNA) and were multiplexed for sequencing at the University of Idaho Institute for Bioinformatics and Evolutionary Studies Genomics Core facility using an Illumina MiSeq v3 protocol for 600 cycles and paired end 300‐bp reads. Sequences were demultiplexed and assigned to individual samples, trimmed, merged, and classified using default parameters in the custom python application dbcAmplicons (https://github.com/msettles/dbcAmplicons/blob/master/docs/manual/DBC_ampliconsUserManual.pdf) as described and used previously by our group and others (Carrothers et al, ; Dai, Gliniewicz, Settles, Coats, & McDonald, ; Lackey et al, ; Liang et al, ; Ma et al, ; Williams et al, ). Briefly, during preprocessing barcodes were allowed ≤1 mismatch (hamming distance) and primers ≤4 mismatches (Levenshtein distance) if the final 4 bases of the primer perfectly matched the target sequence.…”

Section: Methodsmentioning

confidence: 99%

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Social networks, cooperative breeding, and the human milk microbiome

Meehan

Lackey

Hagen

et al. 2018

American J Hum Biol

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“…Although it has been shown that storage temperatures such as freezing at −80 • C had no significant impact on the microbial composition of faecal samples when compared to fresh samples, further investigations are needed to determine how storage temperature influences the microbiome of milk [286]. The addition of different commercially available preservation solutions to preserve the microbiota in milk was investigated by Lackey et al, with some solutions maintaining the bacterial communities and diversity in milk [287]. Moreover, different DNA extraction methods are shown to substantially influence the milk microbiome, showing the importance of how different methodologies introduce variability in the milk microbiome, which is apparent across many studies [204].…”

Section: Future Of Breast Milk Researchmentioning

confidence: 99%

Breast Milk, a Source of Beneficial Microbes and Associated Benefits for Infant Health

et al. 2020

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Human breast milk is considered the optimum feeding regime for newborn infants due to its ability to provide complete nutrition and many bioactive health factors. Breast feeding is associated with improved infant health and immune development, less incidences of gastrointestinal disease and lower mortality rates than formula fed infants. As well as providing fundamental nutrients to the growing infant, breast milk is a source of commensal bacteria which further enhance infant health by preventing pathogen adhesion and promoting gut colonisation of beneficial microbes. While breast milk was initially considered a sterile fluid and microbes isolated were considered contaminants, it is now widely accepted that breast milk is home to its own unique microbiome. The origins of bacteria in breast milk have been subject to much debate, however, the possibility of an entero-mammary pathway allowing for transfer of microbes from maternal gut to the mammary gland is one potential pathway. Human milk derived strains can be regarded as potential probiotics; therefore, many studies have focused on isolating strains from milk for subsequent use in infant health and nutrition markets. This review aims to discuss mammary gland development in preparation for lactation as well as explore the microbial composition and origins of the human milk microbiota with a focus on probiotic development.

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Comparison of commercially-available preservatives for maintaining the integrity of bacterial DNA in human milk

Cited by 19 publications

References 32 publications

Household composition and the infant fecal microbiome: The INSPIRE study

Household composition and the infant fecal microbiome: The INSPIRE study

Social networks, cooperative breeding, and the human milk microbiome

Breast Milk, a Source of Beneficial Microbes and Associated Benefits for Infant Health

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