Bacteriocin Production Correlates with Epidemiological Prevalence of Phylotype I Sequevar 18 Ralstonia pseudosolanacearum in Madagascar

Abstract

Bacterial wilt caused by the Ralstonia solanacearum species complex (RSSC) is a major threat to vegetable crops in Madagascar. For more effective disease management, surveys were carried out in the main vegetable production areas of the country, leading to the collection of 401 new RSSC isolates. Phylogenetic assignment of the isolates revealed a high prevalence of phylotype I sequevar 18. This result contrasts sharply with the epidemiological pattern of RSSC in neighboring islands, including Reunion Island, Comoros, Mayotte, Mauritius, Rodrigues, and the Seychelles, where phylotype I sequevar 31 is widespread. Molecular typing characterization of the Malagasy isolates allowed the identification of 96 haplotypes. Some are found in various plots located in different provinces, which suggests that they were probably disseminated via infected plant material. To find out a potential explanation for the observed epidemiological pattern, we examined the capacity of the Malagasy strains to produce bacteriocin. Interestingly, the highly prevalent genetic lineages I-18 produce bacteriocins that are active against all the genetic lineages present in the country. This work sheds light on the potential impact of bacteriocins in the epidemiology of Malagasy RSSC. IMPORTANCE Knowledge of the epidemiology of a plant pathogen is essential to develop effective control strategies. This study focuses on the epidemiological pattern of Ralstonia pseudosolanacearum phylotype I populations responsible for bacterial wilt in Madagascar. We identified, with the newly collected isolates in three provinces, four genetic lineages probably propagated via infected plant material in Madagascar. We revealed that the epidemiological situation in Madagascar contrasts with that of neighboring Indian Ocean islands. Interestingly, our study on the bacteriocin-producing capacity of Malagasy isolates revealed a correlation between the inhibitory activity of the producing strains and the observed epidemiology. These results suggested that the epidemiology of plant pathogens may be impacted by bacteriocin production.

Keywords: Madagascar; Ralstonia pseudosolanacearum; bacteriocins; molecular epidemiology.

FIG 1

Multidimensional scaling (MDS) of the 96 Malagasy phylotype I haplotypes suggests two clusters with a probability of 52.08%. Cluster 1 groups together the majority of the haplotypes (G = 72), while cluster 2 comprises 24 haplotypes.

FIG 2

Minimum spanning tree (MST) of the Malagasy phylotype I shows that, according to the province of collection, the haplotypes are distributed among 11 clonal complexes. The haplotypes were identified by using goeBURST full MST in PHYLOVIZ. Each MLVA type (MT) is displayed as a circle, the size of which is proportional to the number of isolates represented. The different colors indicate the province where samples were collected. The branch thickness depends on the number of locus differences between the neighboring haplotypes. MCC1 to MCC11 represent the Malagasy clonal complexes 1 to 11. A clonal complex is composed of haplotypes that differ only by one VNTR locus. Haplotypes were regrouped into two clusters according to the MDS result. Here, n represents the number of strains and G represents the number of haplotypes.

FIG 3

Minimum spanning tree (MST) of the Malagasy phylotype I shows that numerous haplotypes are shared in different plots suggesting dissemination of these haplotypes via infected plant material. The haplotypes were identified by using goeBURST full MST in PHYLOVIZ. Each MLVA type (MT) is displayed as a circle, the size of which is proportional to the number of isolates represented. The different colors indicate the sampling plot. The branch thickness depends on the number of locus differences between the neighboring haplotypes. MCC1 to MCC11 represent the Malagasy clonal complexes 1 to 11. A clonal complex is composed of haplotypes that differ only by one VNTR locus. Here, n represents the number of strains and G represents the number of haplotypes.

FIG 4

Minimum spanning tree (MST) shows the genetic links between the Malagasy and worldwide phylotype I predominantly from the South-West Indian Ocean and Africa. The haplotypes were identified by using goeBURST full MST in PHYLOVIZ. Each MLVA type (MT) is displayed as a circle, the size of which is proportional to the number of isolates represented. The different colors indicate the sampling country. The reference strain GMI1000 isolated from French Guiana corresponds to the MT034 haplotype. The branch thickness depends on the number of locus differences between the neighboring haplotypes. MCC1 to MCC11 represent the Malagasy clonal complexes 1 to 11. A clonal complex is composed of haplotypes that differ only by one VNTR locus. Here, n represents the number of strains and G represents the number of haplotypes.

FIG 5

Inhibitory activity of 18 bacteriocin-producing Malagasy isolates on 27 target isolates shows that the highly prevalent genetic lineage I-18 produces bacteriocins that are active against all other phylotype I genetic lineages present in Madagascar. The dendrogram was based on the similarity/dissimilarity of the inhibitory activity patterns of the 18 producing isolates. Links between the different patterns are colored according to the sequevar of the isolates (I-18, I-31, I-33, and I-46). The heatmap was based on the target isolates’ sensitivity to the antimicrobial compounds. The target isolates were grouped according to their sequevar.