Microbial diversity in tropical marine sediments assessed using culture‐dependent and culture‐independent techniques

Demko, Alyssa M.; Patin, Nastassia V.; Jensen, Paul R.

doi:10.1111/1462-2920.15798

Cited by 10 publications

(7 citation statements)

References 137 publications

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“…As expected from other environmental microbiome research, a disparity was observed between the microbial communities identified in the samples using cultivation-independent methods in comparison with the cultured strains [72]. Among the total ASVs detected using a culture-independent approach (metabarcoding), only 2.14% were recovered in the cultured collection (Figure 4).…”

Section: Discussionsupporting

confidence: 55%

Culturable Bacterial Diversity from the Basaltic Subsurface of the Young Volcanic Island of Surtsey, Iceland

et al. 2022

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The oceanic crust is the world’s largest and least explored biosphere on Earth. The basaltic subsurface of Surtsey island in Iceland represents an analog of the warm and newly formed-oceanic crust and offers a great opportunity for discovering novel microorganisms. In this study, we collected borehole fluids, drill cores, and fumarole samples to evaluate the culturable bacterial diversity from the subsurface of the island. Enrichment cultures were performed using different conditions, media and temperatures. A total of 195 bacterial isolates were successfully cultivated, purified, and identified based on MALDI-TOF MS analysis and by 16S rRNA gene sequencing. Six different clades belonging to Firmicutes (40%), Gammaproteobacteria (28.7%), Actinobacteriota (22%), Bacteroidota (4.1%), Alphaproteobacteria (3%), and Deinococcota (2%) were identified. Bacillus (13.3%) was the major genus, followed by Geobacillus (12.33%), Enterobacter (9.23%), Pseudomonas (6.15%), and Halomonas (5.64%). More than 13% of the cultured strains potentially represent novel species based on partial 16S rRNA gene sequences. Phylogenetic analyses revealed that the isolated strains were closely related to species previously detected in soil, seawater, and hydrothermal active sites. The 16S rRNA gene sequences of the strains were aligned against Amplicon Sequence Variants (ASVs) from the previously published 16S rRNA gene amplicon sequence datasets obtained from the same samples. Compared with the culture-independent community composition, only 5 out of 49 phyla were cultivated. However, those five phyla accounted for more than 80% of the ASVs. Only 121 out of a total of 5642 distinct ASVs were culturable (≥98.65% sequence similarity), representing less than 2.15% of the ASVs detected in the amplicon dataset. Here, we support that the subsurface of Surtsey volcano hosts diverse and active microbial communities and that both culture-dependent and -independent methods are essential to improving our insight into such an extreme and complex volcanic environment.

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Section: Discussionsupporting

confidence: 55%

Culturable Bacterial Diversity from the Basaltic Subsurface of the Young Volcanic Island of Surtsey, Iceland

et al. 2022

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“…The characterization of CFU microbiomes by Illumina-based partial 16S rRNA gene amplicon sequencing has recently more often been applied, that is, with CFUs from a raccoon (Junkins and Stevenson 2021 ) and from tropical sediment (Demko et al 2021 ). We used the technology to cover the (whole) diversity on plates and to use the method´s sensitivity to identify bacteria of small colonies that were not visible to the naked eye.…”

Section: Resultsmentioning

confidence: 99%

Particle-associated bacteria in seawater dominate the colony-forming microbiome on ZoBell marine agar

Heins

Harder

2022

FEMS Microbiology Ecology

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Planktonic particle-associated bacteria comprise particle-attached and motile free-living cells. These groups were obtained by settlement in Imhoff cones. Dilution plating on marine agar 2216 (ZoBell marine agar) and microscopic counts indicated a cultivability of 0.7% (0.4–1.2%) of bacteria in coastal seawater collected at Helgoland Roads, North Sea. Particle-associated bacteria presented a minority population in seawater, but had a larger cultivability of 25% (0.9–100%) for populations collected by settlement of particles and 5.7% (0.9–24%) for populations collected by filtration. Partial 16S rRNA gene sequences indicated that 84% of the cultured taxa were either enriched in particle-associated microbiomes or only found in these microbiomes, including Sulfitobacter and other Rhodobacteraceae, Pseudoalteromonas, Psychromonas, Arcobacter and many Flavobacteriaceae. Illumina-based 16S rRNA V3V4 amplicon sequences of plate communities revealed that nearly all operational taxonomic units had a cultivated and described strain in close phylogenetic proximity. This suggested that decades of strain isolation from seawater on ZoBell marine agar had achieved a very good coverage of cultivable genera abundant in nature. The majority belonged to particle-associated bacteria, complementing observations that abundant free-living seawater bacteria often require cultivation conditions closer to their natural habitat like liquid cultivation in oligotrophic medium.

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“…Moreover, the implementation of these techniques allowed the identification of 45% of the total diversity found by culture-independent techniques. In another study, Demko et al analysed the microbial diversity on marine tropical sediments, and the authors were able to culture 3% of the >800 genera detected by culture independent methods using only two culture media and three dilutions of the samples ( Demko et al, 2021 ). Furthermore, 39 genera could only be detected by cultivation.…”

Section: Introductionmentioning

confidence: 99%

Cultivating marine bacteria under laboratory conditions: Overcoming the “unculturable” dogma

Rodrigues

Carvalho²

2022

Front. Bioeng. Biotechnol.

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Underexplored seawater environments may contain biological resources with potential for new biotechnological applications. Metagenomic techniques revolutionized the study of bacterial communities but culture dependent methods will still be important to help the biodiscovery of new products and enzymes from marine bacteria. In this context, we promoted the growth of bacteria from a marine rock pond by culture dependent techniques and compared the results with culture independent methods. The total number of bacteria and diversity were studied in different agar plate media during 6 weeks. Agar plate counting was of the same order of magnitude of direct microscopy counts. The highest efficiency of cultivation was 45% attained in marine agar medium. Molecular analysis revealed 10 different phyla of which only four were isolated by the culture dependent method. On the other hand, four taxonomic orders were detected by cultivation but not by the molecular technique. These include bacteria from the phyla Bacillota and Actinomycetota. Our study shows that it is possible to grow more than the traditionally considered 1% of bacteria from a seawater sample using standard agar plate techniques and laboratorial conditions. The results also demonstrate the importance of culture methods to grow bacteria not detected by molecular approaches for future biotechnological applications.

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Microbial diversity in tropical marine sediments assessed using culture‐dependent and culture‐independent techniques

Cited by 10 publications

References 137 publications

Culturable Bacterial Diversity from the Basaltic Subsurface of the Young Volcanic Island of Surtsey, Iceland

Culturable Bacterial Diversity from the Basaltic Subsurface of the Young Volcanic Island of Surtsey, Iceland

Particle-associated bacteria in seawater dominate the colony-forming microbiome on ZoBell marine agar

Cultivating marine bacteria under laboratory conditions: Overcoming the “unculturable” dogma

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