Retrospective Investigation of the Whole Genome of the Hypovirulent Listeria monocytogenes Strain of ST201, CC69, Lineage III, Isolated from a Piglet with Fatal Neurolisteriosis

Abstract

Listeria monocytogenes (Lm), the causative agent for both human and animal listeriosis, is considered to be a rare but potentially fatal foodborne pathogen. While Lm strains associated with current cases of human listeriosis are now being intensely investigated, our knowledge of this microorganism which has caused listerial infection in the past is still extremely limited. The objective of this study was a retrospective whole-genome sequence analysis of the Lm collection strain, 4/52-1953, isolated in the middle of the 20th century from a piglet with listerial neuroinfection. The multi-locus sequence typing (MLST) analysis based on seven housekeeping genes (abcZ, bglA, cat, dapE, dat, ldh, and lhkA) showed that the Lm strain 4/52-1953 was assigned to the sequence type 201 (ST201), clonal complex 69 (CC69), and phylogenetic lineage III. The strain 4/52-1953, similarly to other ST201 strains, probably originated from the ST9, CC69 via ST157. At least eight different STs, ST69, ST72, ST130, ST136, ST148, ST469, ST769, and ST202, were identified as the descendants of the first generation and a single one, ST2290, was proved to be the descendant of the second generation. Among them there were strains either associated with some sporadic cases of human and animal listerial infection in the course of more than 60 years worldwide or isolated from food samples, fish and dairy products, or migratory birds. Phylogenetic analysis based on whole genomes of all the Lm strains available in the NCBI GenBank (n = 256) demonstrated that the strain 4/52-1953 belonged to minor Cluster I, represented by lineage III only, while two other major Clusters, II and III, were formed by lineages I and II. In the genome of the strain 4/52-1953, 41 virulence-associated genes, including the Listeria pathogenicity island 1 (LIPI-1), and LIPI-2 represented by two internalin genes, the inlA and inlB genes, and five genes related to antibiotic resistance, were found. These findings can help to make the emergence of both hyper- and hypovirulent variants, including those bearing antibiotic resistance genes, more visible and aid the aims of molecular epidemiology as well.

Complete genome of the Listeria monocytogenes strain AUF, used as a live listeriosis veterinary vaccine

“…The Lm strain AUF was stored in lyophilized form. The Lm strain AUF was routinely cultivated on Tryptone Soy Yeast Extract Agar (TSYE Agar) (Merck, EU) overnight prior to the experiments as described 45 , 46 .…”

“…The isolated DNA from the Lm strain AUF strain was used in two parallel sequencing platforms, Illumina HiSeq 2500 (Illumina Inc., USA) and Nanopore MinION (Oxford Nanopore, UK). For this purpose, the preparation of DNA libraries was conducted using either Nextera XT DNA Library Preparation Kit (Illumina Inc., USA) or Nanopore Kit SQK-LSK109 (Oxford Nanopore, UK), respectively, as we described recently 45 , 46 . In the first case, the DNA was sequenced commercially as 250-bp single-Read (Illumina Inc., USA) at Genoanalytica LLC ( https://www.genoanalytica.ru/ , Moscow, Russia).…”

“…The Lm strain AUF was also distinguished from the Lm 4/52-1953, had also been isolated on the territory of the former USSR, although a decade earlier 46 . In fact, recently phylogenetic analysis based on whole genomes of Lm strains (n = 257) available in the NCBI GenBank ( https://www.ncbi.nlm.nih.gov/ ) showed that these two Lm strains belonged to different Clusters, the Lm strain AUF to the Cluster II, represented by genetic lineages I and II, whereas the Lm strain 4/52-1953 - to the Cluster I, formed by genetic lineage III only 46 . Interestingly, the Lm strain AUF was phylogenetically closer to the Lm strain 10403S derived from the clinical sample of a human with listeriosis 46 .…”

“…In fact, recently phylogenetic analysis based on whole genomes of Lm strains (n = 257) available in the NCBI GenBank ( https://www.ncbi.nlm.nih.gov/ ) showed that these two Lm strains belonged to different Clusters, the Lm strain AUF to the Cluster II, represented by genetic lineages I and II, whereas the Lm strain 4/52-1953 - to the Cluster I, formed by genetic lineage III only 46 . Interestingly, the Lm strain AUF was phylogenetically closer to the Lm strain 10403S derived from the clinical sample of a human with listeriosis 46 . Both strains, the Lm strain AUF and the Lm strain 10403S, formed a single Cluster II 46 .…”

Polarization- and Chaos-Game-Based Fingerprinting of Molecular Targets of Listeria Monocytogenes Vaccine and Fully Virulent Strains

“…# NZ_CP048401.1. This is the oldest strain isolated from a piglet in the current territory of Russian Federation in 1953 in the USSR [41]. 8.…”