Molecular analyses reveal stability of bacterial communities in bulk soil of a Japanese paddy field: Estimation by denaturing gradient gel electrophoresis of 16S rRNA genes amplified from DNA accompanied with RNA

Figures & data

Figure 1  Denaturing gradient gel electrophoresis pattern of bacterial communities in the plow layer soil based on DNA analysis. Bands with arrows were excised and subjected to sequencing (see Table 1).

Figure 2  Principal component analysis of the denaturing gradient gel electrophoresis patterns of the bacterial communities in the plow layer soil based on DNA analysis. (a) Plots of first and second principal components and (b) plots of first and third principal components. Contribution rates of first, second and third principal components were 29.7%, 24.9% and 18.9%, respectively.

Figure 3  Denaturing gradient gel electrophoresis pattern of the bacterial communities in the plow layer soil based on DNA and RNA analyses. Bands with arrows were excised and subjected to sequencing (see Table 1). D, DNA; R, RNA.

Table 1 Close relatives of excised denaturing gradient gel electrophoresis bands (see Figs 1,3) that were commonly present or characterized the bacterial communities in the plow layer soil at the respective periods based on DNA and RNA analyses

	DGGE band	DNA/RNA	Close relatives			Similarity (%)	Alignment
			Microorganisms	Phylogenetic affiliations	Accession number
Common	f	DNA	Uncultured soil bacterium	Bacteria	AY242815	99	137/138
			Uncultured Verrucomicrobia bacterium	Verrucomicrobia	AY214795	98	136/138
	i	DNA	Uncultured bacterium	Bacteria	AY221053	94	142/151
			Uncultured Chlorobi bacterium	Chlorobi	AJ519407	87	132/151
	k	DNA	Uncultured bacterium	Bacteria	AF407193	90	99/109
			Uncultured Chloroflexales bacterium	Chloroflexi	AJ577123	89	98/109
	n	DNA	Uncultured bacterium	Bacteria	AF523972	100	159/159
			Uncultured Nitrospira sp.	Nitrospira	AF351231	97	155/159
	n	RNA	Uncultured bacterium	Bacteria	AF498727	98	155/158
			Uncultured Nitrospira sp.	Nitrospira	AF351231	95	151/158
	p	DNA	Uncultured bacterium	Bacteria	AY711541	97	132/136
			Uncultured Chloroflexi bacterium	Chloroflexi	AY211664	93	127/136
	2l	DNA	Kouleothrix aurantiaca	Chloroflexi	AB079640	96	130/135
	2k	DNA	Uncultured soil bacterium	Bacteria	AY289395	97	145/148
			Uncultured Acidobacteria bacterium	Acidobacteria	AF431515	97	144/148
	2f	DNA	Uncultured bacterium	Bacteria	AF432717	99	134/135
			Uncultured Chloroflexi bacterium	Chloroflexi	AY922047	97	131/135
	2f	RNA	Methylomonas sp. GYJ3	Gammaproteobacteria	DQ149125	100	135/135
Before midseason drainage	j	DNA	Uncultured Acidobacteria bacterium	Acidobacteria	AY281355	100	157/157
	s	DNA	Uncultured Alphaproteobacterium	Alphaproteobacteria	AF431210	100	146/146
	x	DNA	Uncultured bacterium	Bacteria	AB127643	97	133/136
			Dehalococcoides sp. BHI80-15	Chloroflexi	AJ431246	94	129/136
Flooded periods	t	DNA	Uncultured bacterium	Bacteria	AY222300	96	131/136
			Uncultured bacterium GC55	candidate division OP10	AJ271048	93	127/136
	u	DNA	Uncultured cyanobacterium	Cyanobacteria	AY437428	98	134/136
Midseason drainage	2j	DNA	Uncultured bacterium	Chloroflexi	AJ009453	96	129/134
	2t	DNA	Uncultured actinobacterium	Actinobacteria	AJ519398	88	133/150
	2t	RNA	Uncultured actinobacterium	Actinobacteria	AJ519398	88	133/150
After midseason drainage	g	DNA	uncultured Verrucomicrobia bacterium	Verrucomicrobia	AY694581	98	157/159
	y	DNA	Uncultured Bradyrhizobium sp.	Alphaproteobacteria	AB207831	100	135/135
			Afipia sp. SP3302452	Alphaproteobacteria	AY599913	100	135/135
			Nitrobacter sp.	Alphaproteobacteria	L35511	100	135/135
			Photorhizobium sp. ‘MKAa 3’	Alphaproteobacteria	L23409	100	135/135
	y	RNA	Uncultured Bradyrhizobium sp.	Alphaproteobacteria	DQ267119	99	134/135
Upland period	2h	DNA	Arthrobacter sp.	Actinobacteria	AB167248	99	139/140
	2i	DNA	Knoellia sinensis	Actinobacteria	AY167851	100	140/140
	2o	DNA	Streptomyces sp. 316D09	Actinobacteria	AB123946	100	140/140
	2o	RNA	Streptomyces sp. 316D10	Actinobacteria	AB123946	99	139/140
	2q	DNA	Terrabacter sp. Ellin102	Actinobacteria	AF408944	100	140/140
			Kitasatospora sp. PU22869	Actinobacteria	AJ427923	100	140/140
			Streptomyces sp. 331G08	Actinobacteria	AB124208	100	140/140
	2q	RNA	Terrabacter sp. Ellin102	Actinobacteria	AF408944	100	140/140
	2r	DNA	Uncultured bacterium	Actinobacteria	AJ277697	99	135/136
RNA specific	R-1	RNA	Phalaenopsis aphrodite subsp.	Tracheophyta (chloroplast)	AY916449	100	137/137
			Calycanthus fertilis	Tracheophyta (chloroplast)	DQ187922	100	137/137
	R-2	RNA	Uncultured Anaeromyxobacter sp.	Deltaproteobacteria	AJ504433	100	128/128
	R-3	RNA	Anaeromyxobacter dehalogenans	Deltaproteobacteria	AF382398	96	156/161
Microorganisms of the top BLAST hits for the respective band are shown. In the case that the top hit had no taxonomic description at the pylum level, the highest hit which could be affiliated to any bacterial phyla is also shown.

Table 2 Comparison of the distribution of characteristic bacteria at the phylum and class levels among bulk soil (this study) and other habitats in the plow layer

Phylum	Class	Bulk soil	Rice straw	Rice straw compost	Plant residues	Rice roots	Percolating water
Actinobacteria	Actinobacteria	26 (6)	3 (1)	11 (2)	–	–	18 (4)
Firmicutes	Clostridia	–	–	11 (2)	19 (5)	13 (6)	–
	Mollicutes	–	–	–	–	–	–
	Bacilli	–	–	–	–	–	–
	Unclassified	–	–	–	4 (1)	–	9 (2)
Proteobacteria	Alphaproteobacteria	7 (2)	14 (5)	–	19 (5)	9 (4)	9 (2)
	Betaproteobacteria	–	6 (2)	–	7 (2)	30 (14)	–
	Gammaproteobacteria	–	14 (5)	11 (2)	7 (2)	2 (1)	14 (3)
	Deltaproteobacteria	4 (1)	14 (5)	22 (4)	11 (3)	24 (11)	36 (8)
Acidobacteria	Acidobacteria	7 (2)	11 (4)	–	7 (2)	4 (2)	–
Cyanobacteria	Cyanobacteria	4 (1)	–	–	–	–	–
Verrucomicrobia	Verrucomicrobiae	7 (2)	6 (2)	11 (2)	4 (1)	2 (1)	5 (1)
Bacteroidetes	Bacteroidetes	–	6 (3)	–	4 (1)	–	5 (1)
	Flavobacteria	–	3 (1)	6 (1)	–	7 (3)	–
	Sphingobacteria	–	9 (3)	–	–	2 (1)	5 (1)
Spirochaetes	Spirochaetes	–	9 (3)	11 (2)	15 (4)	7 (3)	–
Chlorobi	Chlorobia	4 (1)	3 (1)	17 (3)	–	–	–
Chloroflexi	Chloroflexi	26 (6)	–	–	–	–	–
Nitrospira	Nitrospira	4 (1)	–	–	4 (1)	–	–
Candidate division OP10		4 (1)	–	–	–	–	–
Total		100 (23)	100 (35)	100 (18)	100 (27)	100 (46)	100 (22)
Figures show proportion (percentage) of the respective group of the total. Values in parentheses denote number of denaturing gradient gel electrophoresis bands. Data for rice straw, rice straw compost, plant residues, rice roots and percolating water are from Sugano et al. (2005), Tanahashi et al. (2005), Matsuyama et al. (2007), Ikenaga et al. (2003) and Murase et al. (2005), respectively.

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