Genome Plasticity among RelatedLactococcus Strains: Identification of Genetic Events Associated with Macrorestriction Polymorphisms

Pascal Le Bourgeois; Marie-Line Daveran-Mingot; Paul Ritzenthaler

doi:10.1128/JB.182.9.2481-2491.2000

DOI: 10.1128/JB.182.9.2481-2491.2000

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Fig. 1.
Biological and genomic relationship of strains derived from L. lactis subsp. cremoris strain NCDO712. Parentheses indicate strains that were not included in this study. I, Inversion of half of the chromosome in regions 1 and 3; II, excision of the uncharacterized prophage in region 2; III, excision of the complete conjugative sex factor in region 5; IV, excision of the φT712 prophage in region 4; V, chromosomal deletion of a fragment containing the opp-pepO operon in region 3; VI, partial deletion of the conjugative sex factor in region 5. EMS, ethyl methanesulfonate; NTG, nitrosoguanidine; Str, streptomycin.
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Fig. 2.
Macrorestriction patterns of L. lactischromosomes. Lanes: 1, NCDO763; 2, MMS36; 3, NCDO712; 4, NCDO505; 5, NCDO2031; 6, C2; 7, LM2301; 8, LM0230; 9, MG1363; λ, lambda DNA concatemers. (A) ApaI digestion. Electrophoresis conditions were 7.5 s for 13.5 h. (B) SmaI digestion. In the upper panel, electrophoresis conditions for separation of the largerSmaI fragments were 15 s for 13.5 h. In the lower panels, electrophoresis conditions were 8 s for 13.5 h. (C) Dendogram of similarity values.
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Fig. 3.
Locations of the five polymorphic regions of the MG1363 chromosome. Parentheses indicate the sizes (in kilobases) of the restriction fragments. The genome map of strain MG1363 is drawn according to Le Bourgeois et al. (35). Abbreviations; Sm,SmaI; Ap, ApaI; No, NotI; Ce, I-CeuI.
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Fig. 4.
Chromosome map of the different lactococcal strains deduced from the physical map of the reference strain MG1363. Abbreviations; Sm, SmaI; Ap, ApaI; No,NotI; Ce, I-CeuI.

Tables

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Table 1.

Sizes of polymorphic SmaI and ApaI restriction fragments^a

Fragment size^b(kb)	Presence of fragment in strain^c:
Fragment size^b(kb)	MG1363^d	NCDO712	NCDO505	NCDO763	MMS36	NCDO2031	C2	LM0230	LM2301
SmaI fragments
610	+ (Sm1)	+	+	−	−	−	−	−	−
580	−	−	−	+	+	−	−	−	−
550	−	−	−	−	−	+	+	+	+
325	+ (Sm2)	+	+	−	−	+	+	+	+
290	−	−	−	+	+	−	−	−	−
220	−	+	+	−	−	+	+	−	−
180	+ (Sm5A)	−	−	+	+	−	−	+	+
160	−	−	−	+	+	−	−	−	−
130	+ (Sm6)	+	+	−	−	+	+	+	−
105	+ (Sm8)	+	+	+	+	+	+	−	+
58	−	−	−	−	−	−	−	+	+
55^*	−	+	+	−	−	+	+	−	−
45	−	−	−	−	−	−	+	−	−
42	+ (Sm11A)	+	+	+	+	+	−	+	+
36^*	−	+	+	−	−	−	−	−	−
30^*	−	+	+	−	−	−	−	−	−
24^*	−	−	−	+	+	−	−	−	−
ApaI fragments
165	−	+	+	−	−	+	+	−	−
160	+ (Ap3)	−	−	+	+	−	−	+	+
145	+ (Ap4)	+	+	−	−	−	−	−	−
120	−	−	−	+	+	−	−	−	+
100	−	−	−	−	−	+	+	+	+
75	+ (Ap10C)	+	+	+	+	+	+	+	−
69	+ (Ap11)	+	+	−	−	+	+	−	−
67	−	−	−	−	−	−	−	+	−
65	+ (Ap12B)	+	+	−	−	+	+	+	+
55^*	−	+	+	+	+	+	+	−	−
49	+ (Ap14A)	+	+	+	+	+	+	−	−
29	−	−	−	−	−	−	−	+	+
19	−	−	−	+	+	−	−	−	−
16	+ (Ap26)	+	+	+	+	−	−	−	−
13	+ (Ap27B)	+	+	+	+	+	+	−	−
12	+ (Ap28)	+	+	+	+	+	+	−	−
11^*	−	+	+	−	−	+	+	−	−

↵a A detailed table containing the number and size of the ApaI and SmaI restriction fragments of each strain is available upon request to the authors.
↵b Asterisks indicate fragments assigned to plasmid DNA (see text).
↵c The mean genome size for a given strain was calculated assuming an error of the order of 5%: MG1363, 2.5 ± 0.1 Mb; NCDO712 and NCDO505, 2.6 ± 0.1 Mb; NCDO763 and MMS36, 2.6 ± 0.1 Mb; NCDO2031, 2.6 ± 0.1 Mb; C2, 2.6 ± 0.1 Mb; LM0230, 2.4 ± 0.1 M; LM2301, 2.4 ± 0.1 Mb.
↵d Restriction fragments of strain MG1363 were labeled according to reference 35.

Table 2.

Sizes of hybridizing restriction fragments

Probe used	Enzyme tested	Fragment^a(kb)	Presence of fragment in strain^b:
Probe used	Enzyme tested	Fragment^a(kb)	MG1363	NCDO712	NCDO763	NCDO2031	LM0230	LM2301
ISS1	SmaI	130 [Sm6]	+	+	+ (160)	+	+	+ (105)
		55	−	+	−	+	−	−
	ApaI	75 [Ap10C]	+	+	+	+	+	+ (120)
		55	−	+	+	+	−	−
φT712	SmaI	220^* [Sm4]	+	+	+	+	+	+
		180^* [Sm5]	+	−	+	−	+	+
		105 [Sm8]	+	+	+	+	−	−
		22 [Sm14]	+	+	+	+	+	+
	ApaI	165^*	−	+	−	+	−	−
		160^* [Ap3]	+	−	+	−	+	+
		49 [Ap14]	+	+	+	+	−	−
		13 [Ap27]	+	+	+	+	−	−
		12 [Ap28]	+	+	+	+	−	−
		33 [Ap19]	+	+	+	+	+	+

Sm8	SmaI	105 [Sm8]	+	+	+	+	+ (58)	+ (58)
	ApaI	205 [Ap1]	+	+	+	+	+	+
		49 [Ap14A]	+	+	+	+	+ (29)	+ (29)
		13 [Ap27B]	+	+	+	+	−	−
		12 [Ap28]	+	+	+	+	−	−
		8 [Ap29]	+	+	+	+	+	+
Ap3	SmaI	180 [Sm5A]	+	+ (220)	+	+ (220)	+	+
	ApaI	160 [Ap3]	+	+ (165)	+	+ (165)	+	+
No2	NotI	230 [No2]	+	+	+ (200)	+ (175)	+ (175)	+ (175)
	SmaI	610 [Sm1]	+	+	+ (580)	+ (550)	+ (550)	+ (550)
	ApaI	205 [Ap1]	+	+	+	+	+	+
		145 [Ap4]	+	+	+ (120)	+ (100)	+ (100)	+ (100)
		16 [Ap26]	+	+	+	−	−	−
5′-cluA	SmaI	610 [Sm1]	+	+	+ (580)	−	−	−
	ApaI	50^*	+	+	+ (25)	−	−	−
		16 [Ap26]	+	+	+	−	−	−

5′-oppD	SmaI	130 [Sm6]	+	+	+ (160)	+	+	−
		55	−	+	−	+	+	−
	ApaI	75 [Ap10C]	+	+	+	+	+	−
		69^* [Ap11]	+	+	+ (19)	+	+ (67)	−
		55	−	+	−	+	+ (65)	−
oppFB^c	SmaI	130 [Sm6]	+	+	+ (160)	+	+	−
		55	−	+	−	+	+ (58)	−
	ApaI	75 [Ap10C]	+	+	+	+	+	−
		55	−	+	−	+	+ (65)	−

↵a Fragment names are given in brackets. Asterisks indicate a weak hybridization signal. Sizes different from the size given in this column are shown in parentheses.
↵b NCDO712 refers to NCDO712 and NCDO505 genomes; NCDO763 refers to NCDO763 and MMS36 genomes; and NCDO2031 refers to NCDO2031 and C2 genomes.
↵c Probes oppCA and pepOgave the same hybridization pattern as oppFB.