Microbial Associations of Four Species of Algal Symbiont-bearing Foraminifers from the Florida Reef Tract, Usa

2019 ◽  
Vol 49 (2) ◽  
pp. 178-190
Author(s):  
Makenna M. Martin ◽  
Christina A. Kellogg ◽  
Pamela Hallock
Keyword(s):  
Dna Extraction ◽  
Variable Region ◽  
Florida Keys ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Microbiome Composition ◽  
Algal Symbiont ◽  
Reef Tract ◽  
Bacterial Dna Extraction

Abstract While microbiome research is a rapidly expanding field of study, relatively little is known of the microbiomes associated with Foraminifera. This preliminary study investigated microbes associated with four species of Foraminifera, representing two taxonomic orders, which host three kinds of algal endosymbionts. A major objective was to explore potential influences on the microbiome composition, including phylogenetic relatedness among the host species, similarities in algal symbionts hosted, and environmental conditions from which the specimens were collected. Samples examined from two locations along the middle Florida Keys reef tract included 45 foraminiferal specimens and four environmental samples. Bacterial DNA extraction from individual specimens was followed by amplification and amplicon sequencing of the V4 variable region of the 16S rRNA gene; results were obtained from 21 specimens. The Order Miliolida, Family Soritidae, was represented by 5–8 specimens of each of three species: Archaias angulatus and Cyclorbiculina compressa, which both host chlorophyte symbionts, and Sorites orbiculus, which hosts dinoflagellate symbionts. Three Ar. angulatus specimens from which the microbiome was successfully sequenced shared 177 OTUs. Six C. compressa specimens successfully sequenced shared 58 OTUs, of which 31 were also shared by the three specimens of Ar. angulatus. Four successfully sequenced S. orbiculus specimens shared 717 unique OTUs. The 13 soritid specimens shared 26 OTUs, 23 of which represented Proteobacteria, predominantly of the bacterial family Rhodobacteraceae. The fourth foraminiferal species, Amphistegina gibbosa (Order Rotaliida) hosts diatom endosymbionts. Bacterial DNA extraction was attempted on 16 Am. gibbosa, including both normal-appearing and partly-bleached specimens. Only six OTUs, four of which represented Proteobacteria, were found in all eight specimens successfully sequenced. The partly bleached specimens shared nearly twice as many unique microbial OTUs (32) as the normal-appearing specimens (19). All Am. gibbosa specimens shared only four microbial OTUs with the soritid species, three of which may have been contaminants, indicating minimal commonality between the microbiomes of Am. gibbosa and the soritid taxa.

scholarly journals Amplicon-sequencing of raw milk microbiota: impact of DNA extraction and library-PCR

2021 ◽  
Author(s):  
Annemarie Siebert ◽  
Katharina Hofmann ◽  
Lena Staib ◽  
Etienne V. Doll ◽  
Siegfried Scherer ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Dna Extraction ◽  
Raw Milk ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Microbiome Analysis ◽  
Eukaryotic Dna ◽  
Selective Lysis ◽  
The Impact

Abstract The highly complex raw milk matrix challenges the sample preparation for amplicon-sequencing due to low bacterial counts and high amounts of eukaryotic DNA originating from the cow. In this study, we optimized the extraction of bacterial DNA from raw milk for microbiome analysis and evaluated the impact of cycle numbers in the library-PCR. The selective lysis of eukaryotic cells by proteinase K and digestion of released DNA before bacterial lysis resulted in a high reduction of mostly eukaryotic DNA and increased the proportion of bacterial DNA. Comparative microbiome analysis showed that a combined enzymatic and mechanical lysis procedure using the DNeasy® PowerFood® Microbial Kit with a modified protocol was best suitable to achieve high DNA quantities after library-PCR and broad coverage of detected bacterial biodiversity. Increasing cycle numbers during library-PCR systematically altered results for species and beta-diversity with a tendency to overrepresentation or underrepresentation of particular taxa. To limit PCR bias, high cycle numbers should thus be avoided. An optimized DNA extraction yielding sufficient bacterial DNA and enabling higher PCR efficiency is fundamental for successful library preparation. We suggest that a protocol using ethylenediaminetetraacetic acid (EDTA) to resolve casein micelles, selective lysis of somatic cells, extraction of bacterial DNA with a combination of mechanical and enzymatic lysis, and restriction of PCR cycles for analysis of raw milk microbiomes is optimal even for samples with low bacterial numbers. Key points • Sample preparation for high-throughput 16S rRNA gene sequencing of raw milk microbiota. • Reduction of eukaryotic DNA by enzymatic digestion. • Shift of detected microbiome caused by high cycle numbers in library-PCR.

scholarly journals The microbial community of the gut differs between piglets fed sow milk, milk replacer or bovine colostrum

2017 ◽  
Vol 117 (7) ◽  
pp. 964-978 ◽  
Author(s):  
Ann-Sofie R. Poulsen ◽  
Nadieh de Jonge ◽  
Sugiharto Sugiharto ◽  
Jeppe L. Nielsen ◽  
Charlotte Lauridsen ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Amplicon Sequencing ◽  
Bovine Colostrum ◽  
Milk Replacer ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Faecal Samples ◽  
Milk Replacers ◽  
The 16S Rrna Gene ◽  
Gut Microbiota Composition

AbstractThe aim of this study was to characterise the gut microbiota composition of piglets fed bovine colostrum (BC), milk replacer (MR) or sow milk (SM) in the post-weaning period. Piglets (n36), 23-d old, were randomly allocated to the three diets. Faecal samples were collected at 23, 25, 27 and 30 d of age. Digesta from the stomach, ileum, caecum and mid-colon was collected at 30 d of age. Bacterial DNA from all samples was subjected to amplicon sequencing of the 16S rRNA gene. Bacterial enumerations by culture and SCFA analysis were conducted as well. BC-piglets had the highest abundance ofLactococcusin the stomach (P<0·0001) and ileal (P<0·0001) digesta, whereas SM-piglets had the highest abundance ofLactobacillusin the stomach digesta (P<0·0001). MR-piglets had a high abundance of Enterobacteriaceae in the ileal digesta (P<0·0001) and a higher number of haemolytic bacteria in ileal (P=0·0002) and mid-colon (P=0·001) digesta than SM-piglets. BC-piglets showed the highest colonic concentration of iso-butyric and iso-valeric acid (P=0·02). Sequencing and culture showed that MR-piglets were colonised by a higher number of Enterobacteriaceae, whereas the gut microbiota of BC-piglets was characterised by a change in lactic acid bacteria genera when compared with SM-piglets. We conclude that especially the ileal microbiota of BC-piglets had a closer resemblance to that of SM-piglets in regard to the abundance of potential enteric pathogens than did MR-piglets. The results indicate that BC may be a useful substitute for regular milk replacers, and as a feeding supplement in the immediate post-weaning period.

scholarly journals Choice of bacterial DNA extraction method from fecal material influences community structure as evaluated by metagenomic analysis

Microbiome ◽  
2014 ◽  
Vol 2 (1) ◽  
pp. 19 ◽  
Author(s):  
Agata Wesolowska-Andersen ◽  
Martin Bahl ◽  
Vera Carvalho ◽  
Karsten Kristiansen ◽  
Thomas Sicheritz-Pontén ◽  
...  
Keyword(s):  
Community Structure ◽  
Dna Extraction ◽  
Extraction Method ◽  
Metagenomic Analysis ◽  
Fecal Material ◽  
Bacterial Dna ◽  
Dna Extraction Method ◽  
Bacterial Dna Extraction

scholarly journals Necessity of purification during bacterial DNA extraction with environmental soils

2017 ◽  
Vol 32 ◽  
pp. e2017013 ◽  
Author(s):  
Hyun Jeong Lim ◽  
Jung-Hyun Choi ◽  
Ahjeong Son
Keyword(s):  
Dna Extraction ◽  
Bacterial Dna ◽  
Bacterial Dna Extraction

scholarly journals A New, Rapid, Efficient and Non-Toxic Method for Bacterial DNA Extraction

2019 ◽  
Vol 116 (3) ◽  
pp. 360a
Author(s):  
Semire Uzun Gocmen ◽  
Ahmet Aslan ◽  
Muhyittin Temiz
Keyword(s):  
Dna Extraction ◽  
Bacterial Dna ◽  
Bacterial Dna Extraction

scholarly journals The Influence of DNA Extraction and Lipid Removal on Human Milk Bacterial Profiles

2020 ◽  
Vol 3 (2) ◽  
pp. 39 ◽  
Author(s):  
Anna Ojo-Okunola ◽  
Shantelle Claassen-Weitz ◽  
Kilaza S. Mwaikono ◽  
Sugnet Gardner-Lubbe ◽  
Heather J. Zar ◽  
...  
Keyword(s):  
Human Milk ◽  
Dna Extraction ◽  
Bacterial Population ◽  
Illumina Miseq ◽  
Molecular Techniques ◽  
Rrna Gene ◽  
Bacterial Dna ◽  
Dna Recovery ◽  
Culture Independent ◽  
Zymo Research

Culture-independent molecular techniques have advanced the characterization of environmental and human samples including the human milk (HM) bacteriome. However, extraction of high-quality genomic DNA that is representative of the bacterial population in samples is crucial. Lipids removal from HM prior to DNA extraction is common practice, but this may influence the bacterial population detected. The objective of this study was to compare four commercial DNA extraction kits and lipid removal in relation to HM bacterial profiles. Four commercial DNA extraction kits, QIAamp® DNA Microbiome Kit, ZR Fungal/Bacterial DNA MiniPrep™, QIAsymphony DSP DNA Kit and ZymoBIOMICS™ DNA Miniprep Kit, were assessed using milk collected from ten healthy lactating women. The kits were evaluated based on their ability to extract high quantities of pure DNA from HM and how well they extracted DNA from bacterial communities present in a commercial mock microbial community standard spiked into HM. Finally, the kits were evaluated by assessing their extraction repeatability. Bacterial profiles were assessed using Illumina MiSeq sequencing targeting the V4 region of the 16S rRNA gene. The ZR Fungal/Bacterial DNA MiniPrep™ and ZymoBIOMICS™ DNA Miniprep (Zymo Research Corp., Irvine, CA, USA) kits extracted the highest DNA yields with the best purity. DNA extracted using ZR Fungal/Bacterial DNA MiniPrep™ best represented the bacteria in the mock community spiked into HM. In un-spiked HM samples, DNA extracted using the QIAsymphony DSP DNA kit showed statistically significant differences in taxa prevalence from DNA extracted using ZR Fungal/Bacterial DNA MiniPrep™ and ZymoBIOMICS™ DNA Miniprep kits. The only difference between skim and whole milk is observed in bacterial profiles with differing relative abundances of Enhydrobacter and Acinetobacter. DNA extraction, but not lipids removal, substantially influences bacterial profiles detected in HM samples, emphasizing the need for careful selection of a DNA extraction kit to improve DNA recovery from a range of bacterial taxa.

Development of a bacterial DNA extraction modular chip using a magnetic particle and portable vibration motor

2020 ◽  
Vol 12 (9) ◽  
pp. 1197-1202
Author(s):  
Sun Young Lim ◽  
Tae Jae Lee ◽  
Seol Yi Shin ◽  
Nam Ho Bae ◽  
Seok Jae Lee ◽  
...  
Keyword(s):  
Power System ◽  
Electric Power ◽  
Dna Extraction ◽  
Magnetic Particles ◽  
Magnetic Particle ◽  
Electric Power System ◽  
Bacterial Dna ◽  
System P ◽  
3D Printed ◽  
Bacterial Dna Extraction

The bacterial DNA was simply purified by magnetic particles with a portable vibration motor. To effectively extract DNA in the field, the 3D-printed device was employed with low electric power system.

Sign in / Sign up

Export Citation Format

Share Document