Identification of a divergent O-acetyltransferase gene oac1b from Shigella flexneri serotype 1b strains

Figures & data

Table 1 Properties of S. flexneri serotype 3a, 3b, 4b and 1b strains analyzed in this study

Serotype	Strain	Regions of isolation	Host	Year of isolation	DNA sequence identity to oac of Sf6	oac genotype
3a	2002061	China/Henan	Human	2002	99	oac
	2002074	China/Henan	Human	2002	99
	2002127	China/Henan	Human	2002	99
	2002133	China/Henan	Human	2002	99
	03HL12	China/Heilongjiang	Human	2003	99
	06GS02	China/Gansu	Human	2006	99
	06GS03	China/Gansu	Human	2006	99
	HB05	China/Hubei	Human	2008	100
	51575	China/Gansu	Human	—	100
3b	2002110	China/Henan	Human	2002	99	oac
	20030363b	China/Henan	Human	2003	99
4b	NCTC8336	England/London	Monkey	1947	100	oac
	NCTC8522	England/Birmingham	Human	1951	100
	NCTC8598	England/London	Monkey	1953	100
	NCTC9726	USA/Atlanta	—	1955	99
	51577	China/Sichuan	Human	—	100
1b	NCTC5	England/London	Human	1920	89	oac1b
	1997005	China/Henan	Human	1997	89
	1997019	China/Henan	Human	1997	89
	1997020	China/Henan	Human	1997	89
	1997021	China/Henan	Human	1997	89
	1997022	China/Henan	Human	1997	89
	2003070	China/Henan	Human	2003	89
	2003071	China/Henan	Human	2003	89
	2003072	China/Henan	Human	2003	89
	2003073	China/Henan	Human	2003	89
	2005020	China/Henan	Human	2005	89
	06HN87	China/Henan	Human	2006	89
	07GS73	China/Gansu	Human	2007	88
	07HN57	China/Henan	Human	2007	89
	09GS62	China/Gansu	Human	2009	89
	09GS70	China/Gansu	Human	2009	88
	09GS119	China/Gansu	Human	2009	88
	51572	China/Shandong	Human	—	89
	M1250	Australia	Human	—	89
	M1349	Unknown^a	Human	—	89

Table 2 Primers used in this study

Primer pairs	Primer sequence (5′-3′)	Target gene or fragments	Amplicon size (bp)
O1	O1U: TCA ATC CAG GGA TAA TTT AGG CG; O1L: ATG CAT AAG AGC AAC TGC TTT GA	oac or oac1b	1002
O2	O2U: TAG AAA CAG AAG CCA CTG GAG CAC C; O2L: GCT GCG TGG AAA AGA ACT CCA CCT T	oac1b, promoter and terminator	1260
O3	O3U: CCG CCA GGA TGG TGA AAA AGA G; O3L: AGA ACG CCA GTC CAC GCA AAG G	yfdC-oac	3034
O4	O4U: ATG CAT AAG AGC AAC TGC TTT GA; O4L: GGT TTA TGG CTG GGT ATT TGA T	oac-tsp	2075
O5	O5U: CCG CCA GGA TGG TGA AAA AGA GC; O5L: GGT TTA TGG CTG GGT ATT TG	yfdC-tsp	4489

Figure 1 Comparison of protein Oac1b and Oac. (A) Plot of variation between Oac1b and Oac. Nucleotide differences were plotted across the horizontal axis with synonymous and non-synonymous mutations plotted as black and red vertical lines. Ticks and numbers across the horizontal axis are base positions. Transmembrance (TM) segments as predicted by Verma et al. are shown in yellow boxes below the gene. The critical residues for Oac function are indicated by green stars. The ratio of non-synonymous and synonymous substitution rate (Ka/Ks) across the gene using a sliding window of 90 bases (30 codons) and overlap of 3 bases (1 codon) is shown above the gene. (B) Pair-wise list the amino acid sequences of protein Oac1b and Oac. Lines represent amino acid residues that are identical, whereas dots represent amino acids that are similar. TM segments are indicated in red color. The three major regions conserved among the inner membrane trans-acylase family proteins are boxed. The three critical residues for Oac function are marked by asterisk.

Figure 2 Genomic structure of the oac1b region. (A) Comparison of chromosomal regions flanking O-acetyltransferase gene oac1b in serotype 1b strains, and oac in serotype 3a, 3b and 4b strains and serotype-converting phage Sf6. Regions sharing >85% sequence identity are indicated by shaded boxes. Genes coding the same function are shown in the same color. Key primers used in this study are marked by arrows. (B) Genomic structure of regions flanking O-acetyltransferase gene oac1b in serotype 1b strain 1997020 and comparison with relevant regions of sequenced S. flexneri strain 301, 2457t and 2002017. The details of ORFs in strain 1997020 are listed in Supplementary Table S1. Genes share high homologies are shown in the same color. (C) Comparison of the genomic structure of oac1b-carrying prophage in serotype 1b strain 1997020 with prophage genomes in Salmonella enterica serovar Paratyphi B strain SPB7 and E. coli O127:H6 strain E2348/69. Genes sharing >40% identity at amino acid level between the strains are marked by color, red, >80% identity; yellow, 60%–80%; blue, 40%–60%.

Figure 3 Schematic diagram of sensitivity of S. flexneri serotypes (1a, X, Y and 4a) to serotype-converting phage Sf6 and serotype 3b to serotype-converting phage SfI. Numbers of strains tested were showed in parentheses.