The physicochemical properties
The physicochemical properties of the soil samples varied with the deterioration degree of the earthen wall (Table 1). Moisture was higher in the moderately and severely deteriorated samples S3 and S4 than in the not or slightly deteriorated samples S1 and S2 (p>0.05), and pH ranged from 7.32 in S4 to 7.48 in S1 (p>0.05). EC value and the concentrations of Mg2+, Ca2+ and SO42- were higher in the more deteriorated samples (P<0.05), whereas the concentrations of NH4+ and K+ were lower (p>0.05). In the SEM-EDS spot analysis, the relative proportions of C, S, O and Mg elements were higher in samples S3 and S4 than in samples S1 and S2, and those of Al, Si and K were lower (p>0.05) (Additional File 1: Table S1).
Table 1 Physicochemical properties of soil with different degree of deterioration from Jinsha earthen site.
Sample
|
Moisture
(%)
|
pH
|
EC
|
Contents of soluble salts (mmol kg-1)
|
(μs cm-1)
|
Na+
|
NH4+
|
K+
|
Mg2+
|
Ca2+
|
Cl-
|
NO3-
|
SO42-
|
S1
|
4.56±
0.11b
|
7.48±
0.04a
|
6.74±
0.05d
|
12.53±3.66b
|
3.80±
0.02a
|
4.94±
0.04a
|
193.933±
0.99d
|
4.45±
0.08d
|
21.12±0.19c
|
10.44±0.46b
|
111.39±
0.91d
|
S2
|
4.69±
0.04b
|
7.41±
0.03ab
|
10.45±
0.40c
|
18.59±0.02a
|
3.62±
0.02b
|
4.70±
0.02b
|
668.29±
1.44c
|
11.25±
0.02c
|
51.43±0.56a
|
33.46±0.42a
|
205.04±
0.92c
|
S3
|
4.84±
0.10a
|
7.35±
0.09ab
|
11.44±
0.67b
|
10.41±0.05c
|
2.45±
0.04c
|
4.56±
0.02c
|
682.74±
2.19b
|
43.15±
0.33b
|
22.80±0.79b
|
7.25±
0.16c
|
326.42±
2.24b
|
S4
|
4.98±
0.04a
|
7.32±
0.08b
|
13.92±
0.63a
|
10.40±0.16c
|
0.0014±
0.0003d
|
4.55±
0.04c
|
1519.70±
1.56a
|
143.07±
0.94a
|
13.67±0.30d
|
9.87±
0.08b
|
946.48±
2.12a
|
The results are average ± standard deviation (n=3). Different superscript letters in a column indicate statistical significant difference (p < 0.05) in the least significant difference test. S1, no obvious deterioration; S2, mild deterioration; S3, moderate deterioration; S4, severe deterioration.
The diversity of microbial communities
The bacterial and fungal communities in the soil samples were analyzed using amplicon sequencing targeting the 16S rRNA gene and ITS, respectively. The 816336 16S rRNA gene amplicons were divided into 2555 bacterial operational taxonomic units (OTUs) at ≥ 97% similarity. The Good’s coverage was above 99.7% and 99.9% for 16S rRNA gene and ITS amplicons, respectively, and all the rarefaction curves reached an asymptote, showing that the amplicons represented well the sampled populations (Figure S1). The average number of bacterial OTUs per sample group ranged from 553 to 718 (Table 2). The 957322 ITS amplicons were divided into 899 fungal OTUs. The average number of fungal OTUs per sample group ranged from 249 to 355 (Table 2). For bacteria, richness was higher in S4 than in the other sample groups, and diversity was higher in S3 and S4 than in S1 and S2 (p < 0.05) (Table 2). For fungi, richness was higher in S1 than in S4 (p < 0.05) (Table 2), and diversities were on the same level in all the sample groups.
Altogether 330 bacterial and 200 fungal OTUs were detected in all the four sample groups (Figure 1). The highest number of unique bacterial OTUs was detected in S4 and the lowest in S2. The highest number of unique fungal OTUs was detected in S4 (Figure 1). Both the bacterial and fungal communities in the not deteriorated and severely deteriorated sample groups S1 and S4, respectively, were clearly separated in the principal component analysis (PCA) (Figure 2).
Table 2 Richness and diversity of bacterial and fungal communities in soil with different degree of deterioration.
|
Bacteria
|
|
|
Fungi
|
|
|
Sample
|
OTUs
|
Chao1
|
Shannon
|
OTUs
|
Chao1
|
Shannon
|
S1
|
570±76b
|
641±70b
|
3.456±0.25b
|
355±26a
|
392±15a
|
5.86±0.05a
|
S2
|
553±163b
|
598±158b
|
3.76±0.974b
|
288±29ab
|
344±61ab
|
4.603±0.49a
|
S3
|
656±128b
|
672±128b
|
6.771±0.046a
|
325±18ab
|
361±11ab
|
5.67±0.088a
|
S4
|
718±375a
|
741±362a
|
6.649±1.142a
|
249±80b
|
264±83b
|
4.255±1.863a
|
The results are average ± standard deviation (n=3). Different superscript letters in a column indicate statistical significant difference (p < 0.05) in the least significant difference test. S1, no obvious deterioration; S2, mild deterioration; S3, moderate deterioration; S4, severe deterioration.
Distribution of microbial community in sample groups
The bacterial OTUs were assigned into 36 phyla and 617 genera. Actinobacteria, Bacteroidetes, Proteobacteria and Firmicutes were the most abundant phyla (Figure 3a). The relative abundances of Actinobacteria were highest in the sample groups S1 and S2, and those of Proteobacteria and Firmicutes in S3 and S4 (p < 0.05) (Additional File 1: Table S2).
At the genus level, the relative abundances of Rubrobacter were highest in all sample groups except S4 where that of Bacteroides was highest (Figure 3b). Compared with the S1 and S2, the relative abundances of Shewanella were higher in S3 and S4 (p>0.05). The results showed that the bacterial community compositions in sample groups with different degree of deterioration were significantly different (Additional File 1: Table S3).
The fungal communities were assigned into 5 phyla and 205 genera. In all sample groups, Ascomycota was the most abundant phylum and Basidiomycota the second most abundant with relative abundances ranging from 95.9% to 98.8% and 1.1% to 2.7%, respectively (Figure 4a, Additional File 1: Table S4). At the genus level, the relative abundances of Toxicocladosporium and Alternaria were higher in sample groups S1 and S3 than in S2 and S4 (p < 0.05), that of Fusarium was highest in S2 (p < 0.05) and that of Candida in S4 (Figure 4b, Additional File 1: Table S5).
Liner discriminant analysis (LDA) coupled with effect size (LEfSE) was used to identify differentially abundant taxa. Forty-six bacterial taxa were differentially abundant among the four sample groups. Three taxa were significantly more abundant in S1 than in the other three sample groups, six taxa in S2, sixteen taxa in S3, and 21 taxa in S4 (Additional File 1: Figure S2a). Thirty fungal taxa were differentially abundant. Fifteen taxa were significantly more abundant in S1 than in the other three sample groups, ten taxa in S3, and five taxa in S4 (Additional File 1: Figure S2b). The LDA scores of bacterial phylum Bacteroidetes and genus Bacteroides, and that of fungal class Saccharomycetes that includes Candida sp. were approximately five in the S4 samples, suggesting that these organisms were specific for the most deteriorated samples.
The correlation between the microbial community and environmental factors
The relationship between community compositions and environmental factors was analyzed using redundancy analysis (RDA). For bacteria, the RDA axes 1 and 2 accounted for 24.8% and 14.8%, respectively, of the total variation (Figure 5a); for fungi, 30.0% and 15.0%, respectively (Figure 5b). For both bacteria and fungi, the differences in community composition were associated with differences in EC, moisture, pH, and the concentrations of NH4+, K+, Mg2+, Ca2+ and SO42-.