Population structure-guided profiling of antibiotic resistance patterns in clinical Listeria monocytogenes isolates from Germany identifies pbpB3 alleles associated with low levels of cephalosporin resistance

Figures & data

Figure 1. Identification of phylogenetic groups with reduced ceftriaxone resistance. (A) Phylogeny of isolates shown as Neighborhood-Joining tree based on the 1701 locus cgMLST scheme for the model population used for the antibiotic susceptibility testing. Starting from the centre, the rings represent the serogroups and the antibiotics tested (CRO, DAP, CIP, RAM, TET, LNZ, VAN, GEN, MEP, SXT, AMP, PEN, ERY, TGC). The colour code for the antibiotics represent resistant (red), intermediate susceptible (yellow) and susceptible (blue) strains for the individual antibiotics. The two outer rings show MIC values determined for CRO from 4 mg/L (green) to >128 mg/L (red), as well as the positions of the isolates belonging to the STs further investigated. Data was visualized using iTOL v4 [41]. (B) Ceftriaxone resistance levels among 544 selected L. monocytogenes isolates according to MLST STs. Only STs for which MICs of ≥4 isolates were available were considered in this analysis.

Table 1. Antibiotic resistance profiles of the L. monocytogenes model population.

	MIC (mg/L)
	0.03	0.06	0.125	0.25	0.5	1	2	4	8	16	32	64	128	>128
AMP		17	132	265	128	2	0	0	0
PEN		0	135	250	157	2	0	0	0
CRO^1						0	0	1	0	16	32	61	141	293
CIP^2				0	22	451	69	2	0
DAP^2					6	45	302	183	8	0
ERY				544	0	0	0	0	0
GEN^2					488	55	1	0	0	0
LNZ^1					2	27	253	262	0	0
MEP		46	481	17	0	0	0	0	0
RAM^1		528	16	0	0	0	0
TET^1				0	25	361	157	1	0
TGC^2	9	212	321	2	0	0	0	0
SXT	541	3	0	0	0	0	0	0
VAN^1						341	203	0	0	0

544 clinical L. monocytogenes strains were tested against 14 antibiotics. Numbers of isolates with a certain MIC are given. Underlined values indicate no observable growth at the lowest tested concentration. Vertical lines indicate resistance breakpoints as defined by EUCAST for Listeria monocytogenes, Streptococcus pneumoniae¹ or Staphylococcus aureus². Intermediate resistance is marked by a grey background. All values below the grey area are considered fully susceptible, all values right of the vertical bar are considered fully resistant.

Figure 2. Identification of pbpB3 alleles associated with reduced ceftriaxone resistance. (A) Ceftriaxone resistance levels among 544 selected L. monocytogenes isolates according to their pbpB3 allele in the Ruppitsch cgMLST scheme. Only those pbpB3 alleles for which MICs of ≥4 isolates were available were considered in this analysis. (B) Scheme illustrating PBP B3 domains and position of the amino acid exchanges found in the pbpB3 alleles no. 4 (Ala172Val) and 20 (Thr53Ser), which are associated with reduced ceftriaxone resistance. Abbreviations: TM – transmembrane helix, NTD – N-terminal domain; AD – allosteric domain.

Table 2. Effect of pbpB3 on ceftriaxone resistance.

Strain	genotype	ST	pbpB3 allele	ceftriaxone MIC [mg/L]
EGD-e	wt	35	1	64 (64,64,64)
LMJR41	ΔpbpB3	35	-	2 (2,2,2)
LMMF1	ΔpbpB3 + pbpB3-1	35	1	32 (32,32,32)
LMMF2	ΔpbpB3 + pbpB3-4	35	4	16 (16,16,16)
LMMF5	ΔpbpB3 + pbpB3-49	35	49	32 (32,32,32)
LMMF3	ΔpbpB3 + pbpB3-13	35	13	64 (64,64,64)
LMMF6	ΔpbpB3 + pbpB3-20	35	20	16 (16,16,16)
LMMF7	ΔpbpB3 + pbpB3-56	35	56	32 (32,32,32)
18-00792	wt	4	4	16 (16,16,16)
17-04405	wt	217	49	128 (128,128,128)
18-04540	wt	6	13	>128 (>128, >128, >128)
18-00287	wt	403	20	16 (16, 16, 16)
18-02573	wt	296	56	>128(>128, >128, >128)

MIC of ceftriaxone for L. monocytogenes ΔpbpB3 strains complemented with pbpB3 alleles from clinical L. monocytogenes strains with different levels of ceftriaxone resistance. All measurements were performed in triplicates and average values are shown with the individual values given in parentheses.

Letunic I, Bork P. Interactive tree of life (iTOL) v4: recent updates and new developments. Nucleic Acids Res. 2019;47(W1):W256–W259.

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