Characterization of Ralstonia solanacearum species complex strains causing bacterial wilt of tomato in Louisiana, USA

Figures & data

Table 1. Genotypic and phenotypic diversity of Ralstonia solanacearum species complex strains from Louisiana.

Strain No.	Parish-Field^1	Biovar^2	Phylotype^3	egl PCR^4	Sequevar^5
MLI71-15	Livingston-F1	3	I	+	14
MLI72-15	Livingston-F1	6	II	-	-
MLI73-15	Livingston-F1	6	II	-	-
MLI74-15	Livingston-F2	6	II	-	-
MLI75-15	Livingston-F2	3	I	+	14
MLI76-15	Livingston-F2	6	II	-	-
MLI85-15	Saint Helena-GH1	1	II	+	7
MLI86-15	Saint Helena-GH1	1	II	+	7
MLI87-15	Saint Helena-GH1	1	II	+	7
MLI88-15	East Baton Rouge-F1	6	II	+	UD
WLS-2	Livingston-F3	3	I	-	-

¹ Field number represents a unique field (F) or greenhouse (GH) site within the parish.

² Biovar classification based on published biovar type patterns (He et al., 1983; French et al., 1993; Hayward, 1994; Xue et al., 2011).

³Phylotype characterization based on multiplex PCR according to the protocol described by Fegan & Prior (2005).

⁴ egl was amplified from RSCC strains using the procedure described by Fegan et al. (1998). + indicates the presence of a partial fragment of egl and – indicates the absence of an amplicon. egl sequences for MLI71-15, MLI75-15, MLI85-15, MLI86-15 and MLI87-15 were published in GenBank under the accession numbers MK355504–MK355508 respectively.

⁵Sequevar determined based on analyses of partial egl sequences. Sequences within a sequevar have <1% nucleotide differences among the strains (Poussier et al., 2000; Fegan & Prior, 2005). UD (undetermined) indicates that the strain did not align with a RSCC strain within any published sequevar and – indicates that sequencing was not conducted due to the absence of an egl sequence amplicon.

Table 2. List of Ralstonia solanacearum reference species complex strains used in the phylogenetic analysis.

Strain	GenBank accession no.	Host	Origin	Phylotype^2	Sequevar^3	Reference
A3909	EF371812.1	Heliconia	USA	II	6	Wicker et al. (2007)
ACH732	GQ907150.1	Tomato	Australia	IV	8	Lebeau et al. (2011)
AW1	KC188082.1	Tomato	Alabama	II	7	Hong et al. (2012)
B1	FJ561159.1	Sweet potato	China	I	15	Xu et al. (2009)
B34	GQ907154.1	Banana	Brazil	IIA	24	Lebeau et al. (2011)
Blood disease R233	DQ011542.1	−^1	−^1	IV	UD^4	Fegan and Prior (2005)
CFBP2047 (K60)	AF295262.1	Tomato	USA	II	7	Poussier et al. (2000)
CFBP2957	AF295265.1	Tomato	Martinique	IIA	36	Poussier et al. (2000)
CFBP765	EF371810.1	Tobacco	Japan	I	13	Wicker et al. (2012)
CFBP6783	EF371817.1	Heliconia	Martinique	IIB	4	Wicker et al. (2012)
CFBP734	AF295274.1	Potato	Madagascar	III	19	Poussier et al. (2000)
CIP120	GQ907152.1	Potato	Peru	IIA	38	Prior and Fegan (2005)
CIP365	GQ907151.1	Potato	Philippines	I	45	Prior and Fegan (2005)
CMR134	EF439740.1	Huckleberry	Cameroon	I	13	Mahbou Somo Toukam et al. (2009)
CMR15	EF439743.1	Tomato	Cameroon	III	29	Mahbou Somo Toukam et al. (2009)
CMR34	EF439750.1	Tomato	Cameroon	IIB	1	Mahbou Somo Toukam et al. (2009)
CMR39	EF439726.1	Tomato	Cameroon	IIA	41	Mahbou Somo Toukam et al. (2009)
GMI1000	AF295251.1	Tomato	French Guyana	I	18	Poussier et al. (2000)
ICMP7963	AF295263.1	Potato	Kenya	IIA	7	Poussier et al. (2000)
J25	AF295279.1	Potato	Kenya	III	20	Fegan and Prior (2005)
JT516	AF295258.1	Potato	Reunion	IIB	1	Poussier et al. (2000)
K60 (CFBP2047)	DQ657614.1	Tomato	USA	II	7	Castillo and Greenberg (2007)
MAFF211266	AF295250.1	Tomato	Japan	I	15	Poussier et al. (2000)
MAFF301070	AB508612.1	Tomato	Japan	I	14^5	Liu et al. (2009)
MOLK2	EF371841.1	Banana	Philippines	II	3	Prior and Fegan (2005)
NCPPB332	DQ657649.1	Potato	Zimbabwe	III	19	Prior and Fegan (2005)
PSI07	EF371804.1	Tomato	Indonesia	IV	10	Prior and Fegan (2005)
PSS190	EU407285.1	Tomato	Taiwan	I	15	Lebeau et al. (2011)
PSS219	FJ561167.1	Tomato	Taiwan	I	34	Xu et al. (2009)
R288	GQ907153.1	Mulberry	China	I	12	Fegan and Prior (2005)
RS121	KT355485.1	Pepper	Florida	I	UD	Stulberg and Huang (2015)
SUPP1725	AB508665.1	Bellflower	Japan	I	UD	Liu et al. (2009)
Tb3	FJ561128.1	Tobacco	China	I	17	Xu et al. (2009)
UW21	DQ011546.1	Banana	Honduras	II	6	Fegan and Prior (2005)
UW175	DQ011547.1	Plantain	Columbia	II	4	Fegan and Prior (2005)
Zo4	FJ561156.1	Ginger	Philippines	I	14	Xu et al. (2009)

¹Indicates that no data were available for the host or origin.

²Phylotype characterization based on multiplex PCR according to the protocol described by Fegan & Prior (2005).

³Sequevar determined based on analyses of partial endoglucanase gene sequences. Sequences within a sequevar have <1% nucleotide differences among the strains (Poussier et al., 2000; Fegan & Prior, 2005).

⁴ UD (undetermined); indicates that the strain did not align with a RSCC strain within any published sequevar.

Fig. 1 Pathogenicity of Ralstonia solanacearum species complex (RSCC) strains isolated from tomato in Louisiana on susceptible tomato ‘Roma VF’ and hypersensitive response (HR) on Nicotiana tabacum (tobacco). The dark grey bars indicate the number of days post inoculation that wilt symptoms were first observed and the light grey bars indicate the number of days post inoculation that the plants died from RSCC infection. The + symbol above each bar indicates a positive HR 24 h after infiltration into tobacco and – symbol indicates no HR within 72 h post infiltration.

Fig. 2 Maximum likelihood phylogenetic tree based on the partial (750-bp) endoglucanase gene sequences of five Ralstonia solanacearum species complex from Louisiana (MLI71-15, MLI75-15, MLI85-15, MLI86-15 and MLI87-15 shown in bold text) and 36 reference strains previously deposited in GenBank. Bootstrap support values ≥70 are shown above the branches. The scale bar represents an estimated three nucleotide substitutions per 100 nucleotides. The midpoint of the path between the two most distant taxa on the tree was used to root the phylogeny. Strains within each of the four phylotypes (I, II, II and IV) are shown in brackets. <>

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