Genomic epidemiology of a national outbreak of post-surgical Mycobacterium abscessus wound infections in Brazil

Abstract

An epidemic of post-surgical wound infections, caused by a non-tuberculous mycobacterium, has been on-going in Brazil. It has been unclear whether one or multiple lineages are responsible and whether their wide geographical distribution across Brazil is due to spread from a single point source or is the result of human-mediated transmission. 188 isolates, collected from nine Brazilian states, were whole genome sequenced and analysed using phylogenetic and comparative genomic approaches. The isolates from Brazil formed a single clade, which was estimated to have emerged in 2003. We observed temporal and geographic structure within the lineage that enabled us to infer the movement of sub-lineages across Brazil. The genome size of the Brazilian lineage was reduced relative to most strains in the three subspecies of Mycobacterium abscessus and contained a novel plasmid, pMAB02, in addition to the previously described pMAB01 plasmid. One lineage, which emerged just prior to the initial outbreak, is responsible for the epidemic of post-surgical wound infections in Brazil. Phylogenetic analysis indicates that multiple transmission events led to its spread. The presence of a novel plasmid and the reduced genome size suggest that the lineage has undergone adaptation to the surgical niche.

Keywords: Genomic epidemiology; Mycobacterium abscessus; nosocomial infections; outbreak; transmission.

Fig. 1.

Midpoint rooted maximum-likelihood phylogeny of the global population of the Mycobacterium abscessus species complex. The scale bar represents the number of nucleotide substitutions per site. This shows that the Brazilian isolates form a single clade (dark blue) closely related to a recently described dominant circulating clone (DCC) (light blue).

Fig. 2.

Maximum-likelihood tree, rooted to an outgroup, of the clades containing the Brazilian lineage and dominant circulating clone (DCC). The scale bar represents the number of nucleotide substitutions per site. The first two metadata columns show the state and time (year) when the isolates were collected and reveals evidence of isolates clustering by state and year of collection. Metadata columns 3 and 4 indicate the presence (red) or absence (blue) of the two plasmids associated with the Brazilian lineage, pMAB01 and the newly described pMAB02. This shows that pMAB02 is found throughout the Brazilian lineage, but pMAB01 is not present in all the Brazilian isolates. *GO-06 was re-sequenced in this study, BRA_GO-06, and found to contain pMAB02, however, the current publicly available version did not contain pMAB02.

Fig. 3.

Maximum Clade Credibility tree of the Brazilian lineage inferred from the 27 003 trees produced by beast after removing burnin, showing the estimated date of the emergence of the Brazilian lineage to be around 2003. The colours show the state from which the isolates were collected.

Fig. 4.

Minimum Spanning Trees (MSTs) showing the transmission route suggested by the genetic data. The colours represent the state from which the isolates were collected and the node sizes are representative of the number of isolates with that genotype. Open circles represent isolates with a known location, but no recorded date. From this analysis, it can be seen that transmission events from Pará introduced the Brazilian lineage to Goiás, Mato Grosso and either Rio de Janeiro or Espírito Santo in 2006, Rio Grande do Sul and Paraná in 2007 and Amazonas in 2010. The outbreak in São Paulo in 2008 was seeded either from Goiás or Pará. A transmission event from Rio de Janeiro to Paraná in 2007 is shown to be responsible for the majority of cases in this region.

Fig. 5.

Boxplots comparing the genome sizes of 352 M . a. abscessus genomes, 30 M . a. bolletii genomes, 130 M . a. massiliense genomes and 188 Brazilian lineage genomes. The borders of the boxplot represent the 25^th and 75^th percentile. The bold line represents the median genome size. The whiskers mark the 5^th and 95^th percentile. This shows that the genome size of the Brazilian lineage (light blue) is significantly smaller than the genome sizes of the three subspecies.

Fig. 6.

A nucleotide blast comparison between a representative of the Brazilian lineage, a representative of the dominant circulating clone (DCC) cluster 1 and cluster 2 and a sporadic M. a. massiliense isolate. This shows the deletions that have occurred in the Brazilian lineage that could be indicative of its adaption to a novel environment.

Fig. 7.

(a) Plasmid map showing the 121 predicted CDSs encoded by pMAB02. The type VII secretion system (T7SS) is shown in red, genes associated with the stable inheritance of pMAB02 are shown in blue, whilst genes that are associated with the mobility of pMAB02 are shown in green. (b) A comparison of the gene order of the T7SS found on pMAB02 with the esxP1 T7SS found on Mycobacterium marinum plasmid pRAW and the T7SS that have been described on other M. abscessus plasmids plsm2 (M. a. bolletii 50 594) and pMBOL1 (M. a. bolletii 5625) and the ESX-5 T7SS found on the chromosome of Mycobacterium tuberculosis H37Rv. *The assembly of pMBOL is not available and therefore the gene order shown here was based upon that reported by Dumas et al. [33].