Characterizing seasonal changes in soil microbial communities on Marion Island
Prudent Mokgokong, Gwynneth Matcher and Rosemary A. Dorrington
1
Department of Biochemistry and Microbiology, Rhodes University, Grahamstown, South Africa
The isolated sub-Antarctic islands have a significant population of unique flora and fauna. The climatic conditions have enabled establishment and survival of unique endemic organisms, however increasingly, changes in environmental conditions due to global climate change, pose a threat to the different habitats. The sub-Antarctic region has been shown to be particularly sensitive to these climate change and therefore provides a good site for studying the response of ecosystems to global warming. An important question is how the rise in temperatures will impact nutrient cycling. Microorganisms (bacteria, archaea, fungi and algae are the foundation of marine and terrestrial food webs, being involved in sequestration of inorganic nutrients as well as the breakdown and re-cycling of organic nutrients in the environment. In polar environments, the microbiota is the major ecosystem drivers responsible for biogeochemical processes such as decomposition of organic matter and nutrient cycling. Their short lifecycle and metabolic diversity enables microbes to respond rapidly to changes in the environment and as such, studies on their diversity and community structure provides valuable information on how ecosystems function and respond to rising temperatures in polar regions.
The aim of this study was to conduct a pilot study to investigate soil microbial diversity in selected biomes on Marion Island and to characterize seasonal changes in community structure
Bacterial Phylogenetic Profiling
Figure 1: Sampling site (Blechnum slope) )
C ONCLUSIONS
• Diversity and structure of bacterial communities change with soil depth
• Microbial communities appear to be stable over time
• Dominance of decomposers indicates that the food web is heterotrophic
METHODOLOGY
Study site
Core samples (1 cm, 5 cm, 10 cm, 20 cm and 30 cm) were collected from the Blechnum Slope outside of the Marion Island Base (S 46°52.596, E 37°51.507) in May 2013 and April 2014.
Sample processing and PCR amplification
DNA was extracted from 30 mg of the soil using the Qiagen’s AllPrep DNA/ RNA mini kit. Variable regions V4 - V5 of the bacterial 16S rRNA gene were PCR amplified using appropriate 454 sequencing primers and the resultant amplicons were pooled for emulsion PCR and subjected to 454 sequencing (GS Junior, Roche, Life Sciences).
Figure 5: Phylogenetic tree of dominant OTUs . Evolutionary relationships of taxa
The evolutionary history was inferred using the Neighbor-Joining method. The bootstrap consensus tree inferred from 100 replicates is taken to represent the evolutionary history of the taxa analyzed The evolutionary distances were computed using the Maximum Composite Likelihood method. Evolutionary analyses were conducted in
MEGA6 .
• High relative abundance of Acidobacteria and Proteobacteria and unclassified bacterial species at all depths
• Similar profiles for 1 cm and 5 cm samples
• Extent of diversity decreases with an increase in depth
0% 20% 40% 60% 80% 100%
30 cm 20 cm 10 cm 5 cm 1 cm
Relative abundance
Soil depth
Acidobacteria Actinobacteria Bacteroidetes
Chloroflexi Nitrospira Planctomycetes
Proteobacteria Unclassified_Bacteria Verrucomicrobia Minor
Figure 2: Analysis of the 16S rRNA diversity from Blechnum slope soil samples at 1 cm to 30 cm. Relative percentages of the total number of 16S rRNA sequence reads for each of the dominant phyla as determined by 454-pyrosequencing
0.0%
0.5%
1.0%
Percentage relative abundance
1 cm 5 cm 10 cm 20 cm 30 cm
(A) MAJOR PHYLA (B) MINOR PHYLA
DNA Sequence analysis
Sequence reads shorter than 200 nt in length, sequences with homopolymeric runs longer than 7 and any reads containing ambiguous nucleotides were removed using Mothur. Ribosomal Database Project Classifier was used for phylogenetic classifcation down to the genus level. Identification of operational taxonomic units (OTUs) at the species level (0.03) using Mothur and the MEGA 6.0 software was used for constructing the phylogenetic tree.
Denaturing Gradient Gel Electrophoresis (DGGE) Profiling
The variable regions V3 - V5 of the bacterial 16S rRNA gene were PCR amplified and the resultant amplicons were analyzed by DGGE using on a 45% - 70% denaturing gradient.
RESULTS
DGGE profiles of bacterial diversity
Figure 6: DGGE analysis of bacterial communities in samples collected from the same site in May 2013 and April 2014. Arrows to the right indicate common bands in the 1 cm and 5 cm samples (A, B) and 10 – 30 cm)
• Similar DGGE profiles for samples from 1 cm and 5 cm
• Profiles of samples collected in 2013 and 2014 show several conserved bands, suggesting that the soil microbial communities are stable over time
30 cm
144 10 cm
193
20 cm 91 27
61
13 48
Figure 3 :Distribution of OTUs at soil depths of (A) 1 cm, - 10 cm and (B) 10 cm – 30 cm
(A) (B)
5 cm 144 1 cm
138
10 cm 123 40
23
36 112
Operational Taxonomic Units (Species Level)
Figure 4: Comparison of the dominant OTUs at different soil depths. (top 20 OTUs frome ach sample)
0 5 10 15 20 25 30
Percentage relative abundance
1 cm 5 cm 10 cm 20cm 30 cm
• Samples from 1– 10 cm contain highest number of conserved OTUs
• Microbial community at 30 cm is quite distinct (dominant OTUs) from the communities found between 1 – 20 cm
• Most abundant is OTU5 (~30% of total reads) & common to all soil samples
Sample Unique OTUs
Total OTUs
1 cm 133 313
5 cm 144 332
10 cm 123 294
20 cm 91 227
30 cm 144 266
• The majority of OTUs belong to the Phyla Acidobacterium and Proteobacterium
• OTU5, which is the most abundant OTU and common to all samples is an Acidobacterium.
2013 2014 2013 2014 2013 2014 2013 2014 2013 2014
1 cm 5 cm 10 cm 20 cm 30 cm
INTRODUCTION
OTU1 OTU13 OTU15
Bacterium_Ellin OTU19
Roseimicrobium_gellanilyticum OTU3
OTU7
Edaphobacter_aggregans Granulicella_tundricola Terriglobus_saanensis OTU16
Acidobacterium_capsulatum OTU21
OTU25 OTU26 OTU9
Ktedonobacter_racemifer Sphaerobacter_thermophilus OTU22
Pelobacter_carbinolicus OTU24
OTU27
Gemmatimonas_aurantiaca OTU2
OTU10
Nitrosococcus_watsonii OTU14
OTU12 OTU4
Candidatus_Solibacter OTU5
OTU6 OTU18 OTU17 OTU20
Candidatus_Koribacter OTU23
Prosthecobacter_fusiformis Brevifollis_gellanilyticus Singulisphaera_acidiphila Rubrobacter_taiwanensis Sulfuritalea_hydrogenivorans Thermolithobacter_ferrireducens Desulfotomaculum_alcoholivorax Steroidobacter_denitrificans OTU11
Azospira_restricta
Thiobacillus_aquaesulis OTU28
Smithella_propionica Opitutus_terrae
Halorhodospira_neutriphila OTU8
Pseudolabrys_taiwanensis Planctomyces_maris
OTU29
Moorella_stamsii
99
97 100
94 98 100
91
90 87 85
84
77 69 69
67
65 60
57 57 50 48 45
57 60 46
100 46
37
31
29 30
23 22 23
18 17
27
97
98 52 37 36 64 96
12 12 17 99 57
18
13
14 14
10 8
