Comparative genome analysis provides deep insights into Aeromonas hydrophila taxonomy and virulence-related factors

Abstract

Background: Aeromonas hydrophila is a potential zoonotic pathogen and primary fish pathogen. With overlapping characteristics, multiple isolates are often mislabelled and misclassified. Moreover, the potential pathogenic factors among the publicly available genomes in A. hydrophila strains of different origins have not yet been investigated.

Results: To identify the valid strains of A. hydrophila and their pathogenic factors, we performed a pan-genomic study. It revealed that there were 13 mislabelled strains and 49 valid strains that were further verified by Average nucleotide identity (ANI), digital DNA-DNA hybridization (dDDH) and in silico multiple locus strain typing (MLST). Multiple numbers of phages were detected among the strains and among them Aeromonas phi 018 was frequently present. The diversity in type III secretion system (T3SS) and conservation of type II and type VI secretion systems (T2SS and T6SS, respectively) among all the strains are important to study for designing future strategies. The most prevalent antibiotic resistances were found to be beta-lactamase, polymyxin and colistin resistances. The comparative analyses of sequence type (ST) 251 and other ST groups revealed that there were higher numbers of virulence factors in ST-251 than in other STs group.

Conclusion: Publicly available genomes have 13 mislabelled organisms, and there are only 49 valid A. hydrophila strains. This valid pan-genome identifies multiple prophages that can be further utilized. Different A. hydrophila strains harbour multiple virulence factors and antibiotic resistance genes. Identification of such factors is important for designing future treatment regimes.

Keywords: Aeromonas; Antibiotic resistance patterns; Mislabelled strains; Pan-genome; Prophages; Virulence factors.

Fig. 1

Core genome-based phylogeny: a The core genome-based phylogeny was verified by in silico MLST, ANI and dDDH. The red dotted line represents the cut-off line that separates the 13 mislabelled strains based on ANI and dDDH results. The remaining 49 strains are considered to be valid strains. b Core genome-based un-rooted phylogenetic tree. All the mislabelled strains are shown to be distinct from other valid strains. The remaining 49 strains are closely related to each other

Fig. 2

Pan-genome tree of 62 strains based on the presence or absence of genes with the mislabelled 13 strains represented as a separate group

Fig. 3

Heatmap chart generated from distances calculated based on the presence or absence of genes in all valid 49 strains. All ST-251 strains are grouped into one clade, while the UBA0705 strain from Australia showed little relation to other strains

Fig. 4

Pan-genome versus core genome plot to indicate the openness and closeness of the valid 49 A. hydrophila strains. The pan-genome increased continuously due to the addition of gene families, while the core genome stabilized

Fig. 5

COG-based functional classification of core, accessary and unique genes. The distribution of functional genes among these core genes represents the abilities of all the strains in this pan-genome, whereas the remaining dispensable genes provides of increasing trend of genes

Fig. 6

KEGG-based functional annotation of all the protein datasets of the core, accessary and unique genes. Different classes of genes are found among the strains

Fig. 7

Predictions of the core genome and pan-genome of the ST-based groups. a Prediction of the core and pan-genome of the A. hydrophila strains with ST-251 and (b) other STs. ST-251 represents a closed pan-genome, while the other STs represent an open pan-genome

Fig. 8

Comparison of functionally annotated proteins among the ST-251 and other ST groups. a Comparison of A. hydrophila strains with ST-251 and other STs based on COG functional categories. Unique and accessary classes are different, while there is little difference between the core genes. b Distribution of KEGG superfamilies among the various gene fractions of ST-251 and the other STs

Fig. 9

Number of prophages with their completeness profiles in A. hydrophila strains. Only the ATCC-7966 strain did not have any prophages, while the 2JBN101 strain had 7 prophages

Fig. 10

Type of prophages present across the A. hydrophila strains. Multiple prophages were detected in different strains. A maximum of two phages belonging to one prophage type were detected within one strain

Fig. 11

Antibiotic resistance genes and their distribution across the A. hydrophila strains. The red colour represents the presence of that particular gene from that antibiotic class in the strain, while the light yellow colour represents the absence that gene

Fig. 12

Distribution of virulence factors across all 49 genomes and among the various ST groups. The red colour represents the presence of the gene, and the light yellow colour represents the absence of the gene