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![Phylogeny of Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) and Salmonella 1,4,[5],12:i:-isolates from the United Kingdom and Italy, 2005-2010. Maximum-likelihood tree of 212 Salmonella Typhimurium and monophasic isolates was constructed by using 12,793 single-nucleotide polymorphisms (SNPs) outside of prophage elements, insertion sequence elements and sequence repeats identified by reference to the whole-genome sequence of Salmonella Typhimurium strain SL1344. The tree is rooted with Salmonella Enteritidis whole-genome sequence as an outgroup (note shown). The lineage containing the Salmonella 1,4,[5],12:i:-current UK epidemic group is conflated for simplicity (filled triangle). The designation of the isolates (left column) and phage type are shown (right column). *Monophasic isolates outside of the main epidemic clade. †Monophasic clade closely related to the monophasic clone CVM23701 from North America (9). DT, definitive (phage) type; ND, not determined. Scale bar indicates the approximate number of SNPs determined by genetic distance and the number of SNPs used to construct the tree. An expanded version of this figure is available online (http://wwwnc.cdc. gov/EID/article/22/4/15-0531-F1.htm).](profile/Robert-Kingsley-4/publication/295834129/figure/fig1/AS:613879923089417@1523371787594/Phylogeny-of-Salmonella-enterica-serovar-Typhimurium-Salmonella-Typhimurium-and.png)
Phylogeny of Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) and Salmonella 1,4,[5],12:i:-isolates from the United Kingdom and Italy, 2005-2010. Maximum-likelihood tree of 212 Salmonella Typhimurium and monophasic isolates was constructed by using 12,793 single-nucleotide polymorphisms (SNPs) outside of prophage elements, insertion sequence elements and sequence repeats identified by reference to the whole-genome sequence of Salmonella Typhimurium strain SL1344. The tree is rooted with Salmonella Enteritidis whole-genome sequence as an outgroup (note shown). The lineage containing the Salmonella 1,4,[5],12:i:-current UK epidemic group is conflated for simplicity (filled triangle). The designation of the isolates (left column) and phage type are shown (right column). *Monophasic isolates outside of the main epidemic clade. †Monophasic clade closely related to the monophasic clone CVM23701 from North America (9). DT, definitive (phage) type; ND, not determined. Scale bar indicates the approximate number of SNPs determined by genetic distance and the number of SNPs used to construct the tree. An expanded version of this figure is available online (http://wwwnc.cdc. gov/EID/article/22/4/15-0531-F1.htm).
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Microevolution associated with emergence and expansion of new epidemic clones of bacterial pathogens holds the key to epidemiologic success. To determine microevolution associated with monophasic Salmonella Typhimurium during an epidemic, we performed comparative whole-genome sequencing and phylogenomic analysis of isolates from the United Kingdom...
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... determined that contemporary Salmonella 4, [5],12:i:- strains in the United Kingdom are part of a single clonally expanding clade. We constructed a maximum-likelihood phylogeny of all 97 monophasic and 142 Salmonella Ty- phimurium strains (online Technical Appendix 1 Table) by using 12,793 variable sites in the genome, with reference to the whole-genome sequence of reference strain SL1344, excluding single-nucleotide polymorphisms (SNPs) in prophage, insertion sequence elements, and repetitive se- quences ( Figure 1). Most (77 of 97) monophasic strains were from a single distinct clade that seemed to be part of the current monophasic Salmonella Typhimurium epidem- ic because they were the most abundant and most recently isolated strains. However, older monophasic isolates were also found in at least 3 other clades within the Salmonella Typhimurium tree ( Figure 1, indicated with *). A clade con- taining 8 isolates including 2 DT191a ( Figure 1, indicated with †) was closely related to a Salmonella 1,4, [5],12:i:-iso- late from the North American epidemic strain CVM23701 (9). Only 6 SNPs distinguished this isolate from strain H07 474 0455. In addition, a clade containing 6 Salmo- nella Typhimurium var. Copenhagen (4,12:i:1,2) strains (e.g., H070160417) and a clade containing 4 isolates (e.g., H103720606) contained monophasic ...
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... determined that contemporary Salmonella 4, [5],12:i:- strains in the United Kingdom are part of a single clonally expanding clade. We constructed a maximum-likelihood phylogeny of all 97 monophasic and 142 Salmonella Ty- phimurium strains (online Technical Appendix 1 Table) by using 12,793 variable sites in the genome, with reference to the whole-genome sequence of reference strain SL1344, excluding single-nucleotide polymorphisms (SNPs) in prophage, insertion sequence elements, and repetitive se- quences ( Figure 1). Most (77 of 97) monophasic strains were from a single distinct clade that seemed to be part of the current monophasic Salmonella Typhimurium epidem- ic because they were the most abundant and most recently isolated strains. However, older monophasic isolates were also found in at least 3 other clades within the Salmonella Typhimurium tree ( Figure 1, indicated with *). A clade con- taining 8 isolates including 2 DT191a ( Figure 1, indicated with †) was closely related to a Salmonella 1,4, [5],12:i:-iso- late from the North American epidemic strain CVM23701 (9). Only 6 SNPs distinguished this isolate from strain H07 474 0455. In addition, a clade containing 6 Salmo- nella Typhimurium var. Copenhagen (4,12:i:1,2) strains (e.g., H070160417) and a clade containing 4 isolates (e.g., H103720606) contained monophasic ...
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... determined that contemporary Salmonella 4, [5],12:i:- strains in the United Kingdom are part of a single clonally expanding clade. We constructed a maximum-likelihood phylogeny of all 97 monophasic and 142 Salmonella Ty- phimurium strains (online Technical Appendix 1 Table) by using 12,793 variable sites in the genome, with reference to the whole-genome sequence of reference strain SL1344, excluding single-nucleotide polymorphisms (SNPs) in prophage, insertion sequence elements, and repetitive se- quences ( Figure 1). Most (77 of 97) monophasic strains were from a single distinct clade that seemed to be part of the current monophasic Salmonella Typhimurium epidem- ic because they were the most abundant and most recently isolated strains. However, older monophasic isolates were also found in at least 3 other clades within the Salmonella Typhimurium tree ( Figure 1, indicated with *). A clade con- taining 8 isolates including 2 DT191a ( Figure 1, indicated with †) was closely related to a Salmonella 1,4, [5],12:i:-iso- late from the North American epidemic strain CVM23701 (9). Only 6 SNPs distinguished this isolate from strain H07 474 0455. In addition, a clade containing 6 Salmo- nella Typhimurium var. Copenhagen (4,12:i:1,2) strains (e.g., H070160417) and a clade containing 4 isolates (e.g., H103720606) contained monophasic ...
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... maximum-likelihood phylogenetic tree, reconstructed by using variable sites within the whole-genome sequence with reference to the draft genome sequence of a represen- tative strain from within the epidemic (strain SO4698-09), indicated a clonally expanding clade with a maximum root- to-tip distance of ≈70 SNPs. This finding indicated that all strains in the tree shared a common ancestor in the recent past ( Figure 2). All isolates from this monophasic clade were of sequence type 34. The phage type of monopha- sic epidemic isolates varied according to phylogeny. Most isolates were DT193 (38 of 62 typed) or DT120 (9) and various other phage types including DT7 (3), DT191a (1), DT21 (1), DT21var (1), U311 (3), U302 (2), and RDNC (3). However, although virtually all isolates in subclades A and B were DT193, the phage type was highly variable in subclade C. Biphasic DT193 strains (e.g., 4061-1997; Figure 1) isolated before 2005 were not direct ancestors of the current monophasic Salmonella Typhimurium epidem- ic because they were present on a distinct lineage. Indeed, DT193 isolates were present on 4 distinct lineages within the phylogenetic tree, highlighting the polyphyletic nature of this phage type (Figure 1). Isolates from UK animals in subclade C were relatively scarce; 1 of 21 isolates in this subclade was from a UK animal. Instead, isolates from this subclade came predominantly from humans in the United Kingdom and humans and animals in Italy. In contrast, iso- lates from subclade A were mostly (18 of 32) of livestock origin; only 5 were of human origin. Clade B contained an approximately equal number of human and livestock iso- lates. Furthermore, although isolates from UK pigs were (Salmonella Typhimurium) and Salmonella 1,4, [5],12:i:- isolates from the United Kingdom andItaly, 2005-2010. Maximum-likelihood tree of 212 Salmonella Typhimurium and monophasic isolates was constructed by using 12,793 single-nucleotide polymorphisms (SNPs) outside of prophage elements, insertion sequence elements and sequence repeats identified by reference to the whole-genome sequence of Salmonella Typhimurium strain SL1344. The tree is rooted with Salmonella Enteritidis whole-genome sequence as an outgroup (note shown). The lineage containing the Salmonella 1,4, [5],12:i:- current UK epidemic group is conflated for simplicity (filled triangle). The designation of the isolates (left column) and phage type are shown (right column). *Monophasic isolates outside of the main epidemic clade. †Monophasic clade closely related to the monophasic clone CVM23701 from North America (9). DT, definitive (phage) type; ND, not determined. Scale bar indicates the approximate number of SNPs determined by genetic distance and the number of SNPs used to construct the tree. An expanded version of this figure is available online (http://wwwnc.cdc. ...
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... maximum-likelihood phylogenetic tree, reconstructed by using variable sites within the whole-genome sequence with reference to the draft genome sequence of a represen- tative strain from within the epidemic (strain SO4698-09), indicated a clonally expanding clade with a maximum root- to-tip distance of ≈70 SNPs. This finding indicated that all strains in the tree shared a common ancestor in the recent past ( Figure 2). All isolates from this monophasic clade were of sequence type 34. The phage type of monopha- sic epidemic isolates varied according to phylogeny. Most isolates were DT193 (38 of 62 typed) or DT120 (9) and various other phage types including DT7 (3), DT191a (1), DT21 (1), DT21var (1), U311 (3), U302 (2), and RDNC (3). However, although virtually all isolates in subclades A and B were DT193, the phage type was highly variable in subclade C. Biphasic DT193 strains (e.g., 4061-1997; Figure 1) isolated before 2005 were not direct ancestors of the current monophasic Salmonella Typhimurium epidem- ic because they were present on a distinct lineage. Indeed, DT193 isolates were present on 4 distinct lineages within the phylogenetic tree, highlighting the polyphyletic nature of this phage type (Figure 1). Isolates from UK animals in subclade C were relatively scarce; 1 of 21 isolates in this subclade was from a UK animal. Instead, isolates from this subclade came predominantly from humans in the United Kingdom and humans and animals in Italy. In contrast, iso- lates from subclade A were mostly (18 of 32) of livestock origin; only 5 were of human origin. Clade B contained an approximately equal number of human and livestock iso- lates. Furthermore, although isolates from UK pigs were (Salmonella Typhimurium) and Salmonella 1,4, [5],12:i:- isolates from the United Kingdom andItaly, 2005-2010. Maximum-likelihood tree of 212 Salmonella Typhimurium and monophasic isolates was constructed by using 12,793 single-nucleotide polymorphisms (SNPs) outside of prophage elements, insertion sequence elements and sequence repeats identified by reference to the whole-genome sequence of Salmonella Typhimurium strain SL1344. The tree is rooted with Salmonella Enteritidis whole-genome sequence as an outgroup (note shown). The lineage containing the Salmonella 1,4, [5],12:i:- current UK epidemic group is conflated for simplicity (filled triangle). The designation of the isolates (left column) and phage type are shown (right column). *Monophasic isolates outside of the main epidemic clade. †Monophasic clade closely related to the monophasic clone CVM23701 from North America (9). DT, definitive (phage) type; ND, not determined. Scale bar indicates the approximate number of SNPs determined by genetic distance and the number of SNPs used to construct the tree. An expanded version of this figure is available online (http://wwwnc.cdc. ...
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... monophasic phenotype results from the absence of phase-2 flagellin monomer FljB. The presence of the fljBA genes and the neighboring genome sequence of Salmonella Typhimurium and monophasic variants, determined by mapping raw sequence read data to the fljB locus region of the SL1344 whole-genome sequence (online Techni- cal Appendix 2 Figure 5, panel A), indicated that the UK epidemic strains are monophasic because of multiple in- dependent deletion events that occurred during clonal ex- pansion. Four Salmonella Typhimurium isolates (2 DT7 isolates [SO5416-06 and H09164 0090], 1 DT135 isolate [SO6221-07], and 1 DT177 isolate [H08390 0191]) that were closely related and shared a common ancestor with the monophasic epidemic strains (Figure 1) fljBA locus, indicating that the MRCA with these strains and the epidemic strains was biphasic. In contrast, 67 of 77 monophasic Salmonella Typhimurium strains from the epidemic clade lacked at least part of the fljBA locus, result- ing from deletions ranging in size and with a distribution that was consistent with the phylogenetic relationship of the strains (online Technical Appendix 2 Figure 5, panel A). The 8 epidemic strains that did not have a deletion in the fljB locus were deeply rooted in the tree, consistent with multiple deletion events (1-36 kb) occurring since clonal expansion of the clade. Most deletions shared a common junction in the intergenic region of fljB and iroB. Because it was not possible to assemble short read sequence data across the fljB locus deletion region, to investigate the na- ture of the deletion, we generated long read sequence data for a representative isolate SO4698-09 by using the PacBio sequencing platform (Pacific Biosciences, Menlo Park, CA, USA). A single contig assembly of these data revealed a 15,726-bp deletion of the genome relative to SL1344 and a 27,473-bp insertion of a novel sequence (online Techni- cal Appendix 2 Figure 5, panel B). The inserted sequence was similar to sequences of several genes from transposon Tn21, mercury resistance genes (merTABCDE and merR), and antimicrobial drug resistance genes, consistent with the resistance profile of this strain (strA, strB, sul2, tet [B], and bla TEM-1 ). The composite transposon insertion was not pres- ent in closely related isolates (e.g., SO5416-06) (Figure 1) that were outside of the monophasic clade, suggesting that it was acquired by the MRCA of the monophasic clade and not before clonal expansion. The deletions in the fljB locus of monophasic strains from outside the main clade from the United Kingdom were distinct from that in the UK mono- phasic clade but identical to those described for strains from epidemics in North America (e.g., CVM23701) (9) and Spain (e.g., 1115/25) (10) (online Technical Appendix 2 Figure 5, panel ...
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... monophasic phenotype results from the absence of phase-2 flagellin monomer FljB. The presence of the fljBA genes and the neighboring genome sequence of Salmonella Typhimurium and monophasic variants, determined by mapping raw sequence read data to the fljB locus region of the SL1344 whole-genome sequence (online Techni- cal Appendix 2 Figure 5, panel A), indicated that the UK epidemic strains are monophasic because of multiple in- dependent deletion events that occurred during clonal ex- pansion. Four Salmonella Typhimurium isolates (2 DT7 isolates [SO5416-06 and H09164 0090], 1 DT135 isolate [SO6221-07], and 1 DT177 isolate [H08390 0191]) that were closely related and shared a common ancestor with the monophasic epidemic strains (Figure 1) fljBA locus, indicating that the MRCA with these strains and the epidemic strains was biphasic. In contrast, 67 of 77 monophasic Salmonella Typhimurium strains from the epidemic clade lacked at least part of the fljBA locus, result- ing from deletions ranging in size and with a distribution that was consistent with the phylogenetic relationship of the strains (online Technical Appendix 2 Figure 5, panel A). The 8 epidemic strains that did not have a deletion in the fljB locus were deeply rooted in the tree, consistent with multiple deletion events (1-36 kb) occurring since clonal expansion of the clade. Most deletions shared a common junction in the intergenic region of fljB and iroB. Because it was not possible to assemble short read sequence data across the fljB locus deletion region, to investigate the na- ture of the deletion, we generated long read sequence data for a representative isolate SO4698-09 by using the PacBio sequencing platform (Pacific Biosciences, Menlo Park, CA, USA). A single contig assembly of these data revealed a 15,726-bp deletion of the genome relative to SL1344 and a 27,473-bp insertion of a novel sequence (online Techni- cal Appendix 2 Figure 5, panel B). The inserted sequence was similar to sequences of several genes from transposon Tn21, mercury resistance genes (merTABCDE and merR), and antimicrobial drug resistance genes, consistent with the resistance profile of this strain (strA, strB, sul2, tet [B], and bla TEM-1 ). The composite transposon insertion was not pres- ent in closely related isolates (e.g., SO5416-06) (Figure 1) that were outside of the monophasic clade, suggesting that it was acquired by the MRCA of the monophasic clade and not before clonal expansion. The deletions in the fljB locus of monophasic strains from outside the main clade from the United Kingdom were distinct from that in the UK mono- phasic clade but identical to those described for strains from epidemics in North America (e.g., CVM23701) (9) and Spain (e.g., 1115/25) (10) (online Technical Appendix 2 Figure 5, panel ...
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... in all 3 subclades, isolates from UK cattle were present only in subclade A, consistent with epidemiologic reports that the epidemic originated in pig herds and later spread to cattle herds (17). Despite analysis inclusion of only 6 isolates from birds, these were distributed through- out the tree, suggesting multiple transmission events into these animal populations. The distribution of isolates from humans and livestock (pigs, cattle, and sheep) within sub- clades of the phylogenetic tree of UK monophasic isolates was also strikingly uneven. Most (64 of 77) isolates were ASSuT tetraresistant, and the corresponding resistance genes were detected in de novo assembled sequences (on- line Technical Appendix 2 Figure 1), suggesting that the most recent common ancestor (MRCA) of the clade had this complement of resistance genes. However, during clonal expansion, 7 strains had lost their resistance genes entirely and another 7 had an altered complement of resis- tance ...
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... sopE virulence gene was acquired on a novel pro- phage, mTmV (monophasic Salmonella Typhimurium V), by multiple independent events during clonal expan- sion of the epidemic clade. The thrW locus of contem- porary monophasic Salmonella Typhimurium isolates has been reported to harbor either a prophage, a novel genetic island, or neither (11). In strain SO4698-09, the thrW locus contains the novel genetic island described previ- ously but also an additional prophage element encoding the sopE gene that together total 55 kb. Determination of the accessory genome by using the Roary pan genome pipeline (22) indicated that 23 of 77 monophasic isolates from the epidemic clade contained the sopE gene ( Figure 2). SopE is a guanine exchange factor involved in subver- sion of the host enterocyte cytoskeleton, a key component of the infection process (11,23,24). The sopE gene was present in 6 distinct clusters of the monophasic clade, and ancestral state reconstruction indicated that multiple in- dependent acquisitions followed by clonal expansion of the sopE -positive variant was the most likely explana- tion for their distribution (online Technical Appendix 2 Figure 3, panel B). The sopE gene of strain SO4698-09 is present on a 55-kb region, designated mTmV phage, which was absent from strain SL1344 and shared the greatest similarity with the Shigella flexneri V prophage (online Technical Appendix 2 Figure 6) (25). The mTmV phage from SO4698-09 was not related to the FELS-2 prophage of Salmonella Typhimurium strain SL1344, which also encodes the sopE gene, except in a 2,443-bp region that encoded the sopE gene and flanking sequence. Examination of partial assemblies of other monophasic strains encoding sopE revealed that the gene was associ- ated with the same prophage and inserted between the ge- nome region corresponding to the thrW locus. These data indicated that a novel sopE phage entered the genome on at least 6 occasions during the clonal expansion of the epidemic clade. Because the sopE gene was present in phylogenetic clusters toward the terminal branches of the monophasic clade tree and subsequently exhibited clon- al expansion, we addressed the question of whether the proportion of strains that encoded the sopE gene in our strain collection each year changed during 2005-2010. The frequency distribution for each year was determined from collated data from 59 strains for which date of isola- tion and sequence data were available and an additional 41 randomly selected monophasic strains from the United Kingdom for which the presence of the sopE gene was determined by PCR (Figure 3; online Technical Appen- dix 2 Table). Increased frequency, ranging from none in 2005 and 2006 to 40% in 2010, suggested that acquisition of this gene may have conferred a competitive ...
Citations
... This approach revealed that the S. Typhimurium isolates belong to a single clonal complex, and the most abundant sequence type (ST) is the ST19 sequence type 3 . Nonetheless, recently, there has been an increase in the reports of emerging sequence types dominating geographical areas, such as the ST34 sequence type, which was initially associated with gastrointestinal infection in Europe and is currently found to be globally distributed as a pandemic clone 4,5 . Other sequence types of interest are ST313 and ST213. ...
... Part of the relevance of the NA-ST213 sequence type is their potential ability to cause invasive infections in humans lacking wellcharacterized virulence determinants, including the virulence plasmid (pSTV) 10 and their virulence-related genes ( Fig. 1 and Supplementary Fig. 3). Similar observations have been made for the epidemic clones of monophasic S. Typhimurium ST34 mainly linked to gastrointestinal infections in humans and animals 4,45,46 . Our findings suggest that the lack of virulence determinants does not reduce the NA-ST213 strains capabilities to cause disease (Fig. 4a-e). ...
Salmonella enterica serovar Typhimurium ST213 is an emergent multidrug-resistant sequence type associated with the food chain, and gastrointestinal and invasive infections in North America. Here, we applied genomic and phenotypic analyses to illustrate the diversity and evolution of sequence type ST213. The population structure and evolutionary history of ST213 strains, particularly the North American isolates (NA-ST213) distinguish them from other S . Typhimurium sequence types, including European ST213 strains. NA-ST213 isolates were distributed in four co-circulating lineages with distinct multidrug resistance profiles and unique phage and CRISPR spacers patterns that could have shaped their local microevolution. Compared to the SL1344 reference strain, NA-ST213 demonstrated reduced adherence and internalization in cultured eukaryotic cell lines but exhibited more efficient replication and intracellular survival. This study underscores the relevance of studying an emergent S . Typhimurium sequence type and the events leading to its diversification beyond the well-characterized reference strains and worldwide predominant sequence types. However, it must also serve as a cautionary tale of the potential health risk the NA-ST213 may represent; particularly when there is a close relationship with pandemic sequence types such as the monophasic ST34.
... We also observed that even very closely related Salmonella strains exhibited diversity in sensitivity to SPLA phages. For example, S. Typhimurium strains S04698-09 and A53 were both part of the monophasic S. Typhimurium ST34 epidemic clade that emerged in the last three decades (32,33), yet these strains exhibited distinct differences in susceptibility to at least five SPLA phages. Furthermore, a S. Typhimurium strain of ST4/74 that was genetically modified to remove prophage elements from its genome exhibited a moderate decrease in sensitivity to several SPLA phages. ...
The ability of virulent bacteriophages to lyse bacteria influences bacterial evolution, fitness, and population structure. Knowledge of both host susceptibility and resistance factors is crucial for the successful application of bacteriophages as biological control agents in clinical therapy, food processing, and agriculture. In this study, we isolated 12 bacteriophages termed SPLA phage which infect the foodborne pathogen Salmonella enterica . To determine phage host range, a diverse collection of Enterobacteriaceae and Salmonella enterica was used and genes involved in infection by six SPLA phages were identified using Salmonella Typhimurium strain ST4/74. Candidate host receptors included lipopolysaccharide (LPS), cellulose, and BtuB. Lipopolysaccharide was identified as a susceptibility factor for phage SPLA1a and mutations in LPS biosynthesis genes spontaneously emerged during culture with S . Typhimurium. Conversely, LPS was a resistance factor for phage SPLA5b which suggested that emergence of LPS mutations in culture with SPLA1a represented collateral sensitivity to SPLA5b. We show that bacteria-phage co-culture with SPLA1a and SPLA5b was more successful in limiting the emergence of phage resistance compared to single phage co-culture. Identification of host susceptibility and resistance genes and understanding infection dynamics are critical steps in the rationale design of phage cocktails against specific bacterial pathogens.
IMPORTANCE
As antibiotic resistance continues to emerge in bacterial pathogens, bacterial viruses (phage) represent a potential alternative or adjunct to antibiotics. One challenge for their implementation is the predisposition of bacteria to rapidly acquire resistance to phages. We describe a functional genomics approach to identify mechanisms of susceptibility and resistance for newly isolated phages that infect and lyse Salmonella enterica and use this information to identify phage combinations that exploit collateral sensitivity, thus increasing efficacy. Collateral sensitivity is a phenomenon where resistance to one class of antibiotics increases sensitivity to a second class of antibiotics. We report a functional genomics approach to rationally design a phage combination with a collateral sensitivity dynamic which resulted in increased efficacy. Considering such evolutionary trade-offs has the potential to manipulate the outcome of phage therapy in favor of resolving infection without selecting for escape mutants and is applicable to other virus-host interactions.
... A number of outbreaks caused by Salmonella 4, [5],12:i:-have been reported in many countries, and most of these cases have been linked to the consumption of contaminated pork and other sources, such as cattle and poultry (Arnott et al. 2018, Bone et al. 2010, Elnekave et al. 2018, Mossong et al. 2007, Tavechio et al. 2004, Trüpschuch et al. 2010, Yang et al. 2015. Sequence type 34, which is genotyed by multilocus sequence typing based on the nucleotide sequences of seven housekeeping genes, namely, aroC, dnaN,hemD,hisD,purE,sucA,and thrE,Salmonella 4,[5],12:i:-, called European clone, is one of the beststudied clones due to its worldwide dissemination (Elnekave et al. 2018, Mather et al. 2018, Petrovska et al. 2016, Yang et al. 2015. This clone has been characterized JARQ 58 (1) 2024 14 by the possession of a composite transposon and integrative and conjugative elements (ICEs) that are responsible for antimicrobial resistance and heavy metal tolerance, respectively (Petrovska et al. 2016). ...
... Sequence type 34, which is genotyed by multilocus sequence typing based on the nucleotide sequences of seven housekeeping genes, namely, aroC, dnaN,hemD,hisD,purE,sucA,and thrE,Salmonella 4,[5],12:i:-, called European clone, is one of the beststudied clones due to its worldwide dissemination (Elnekave et al. 2018, Mather et al. 2018, Petrovska et al. 2016, Yang et al. 2015. This clone has been characterized JARQ 58 (1) 2024 14 by the possession of a composite transposon and integrative and conjugative elements (ICEs) that are responsible for antimicrobial resistance and heavy metal tolerance, respectively (Petrovska et al. 2016). ...
... Thus, clade 9 isolates typically showed resistance to ampicillin, streptomycin, sulfonamides, and tetracycline. In addition, all clade 9 isolates had an integrative and conjugative element, ICEmST, which is a specific genomic island in the European clone (Petrovska et al. 2016). The ST34-specific ICE was first identified in Salmonella 4,[5],12:i:-and was designated as Salmonella genomic island 3 (SGI-3) in 2016 (Petrovska et al. 2016). ...
Salmonella enterica subsp. enterica serovar Typhimurium and its monophasic variant (Salmonella 4,[5],12:i:-) are two of the most frequently isolated serovars in food animals worldwide. Especially, human and swine infections by Salmonella 4,[5],12:i:- have increased considerably in Western countries since the mid-1990s. Although bovine and swine salmonellosis by Salmonella 4,[5],12:i:- has been increasing in Japan, the genetic background of the serovar has been unclear. In our study, whole genome sequence (WGS)-based phylogenetic analysis was performed to reveal the characteristics of Salmonella Typhimurium and Salmonella 4,[5],12:i:- isolated in Japan. Nine distinctive clades were identified and clonal replacements were observed several times in the last four decades. Among the nine clades, clade 9, which is a multidrug-resistant and predominant clone after 2010, has similar characteristics to the European clone that has been a major threat to public and animal health worldwide in recent years. Furthermore, the two distinctive clades in clade 9 were identified by phylogenetic analysis. Each clade has diverged through microevolution mediated by the acquisition or deletion of mobile genetic elements such as plasmids, prophages, composite transposons, and integrative and conjugative elements. The present review focuses on WGS-based phylogenetic studies of Salmonella Typhimurium and Salmonella 4,[5],12:i:- isolated from food animals in Japan.
... We contend that this method is pertinent to virulence loci found with SPI as the regions are subject to horizontal gene transfer [46,47]. Common methods to differentiate serovars typically rely on the alignment of core genes or single nucleotide polymorphisms (SNP) identified against reference assemblies [7,25,48,49]. These methods must rely on post hoc analysis to determine if two evolutionary similar strains have acquired virulence factor genes which may correspond to differences in case presentation as witnessed in S. Infantis and S. Typhimurium. ...
The serovars of Salmonella enterica display dramatic differences in pathogenesis and host preferences. We developed a process (patent pending) for grouping Salmonella isolates and serovars by their public health risk. We collated a curated set of 12,337 S. enterica isolate genomes from human, beef, and bovine sources in the US. After annotating a virulence gene catalog for each isolate, we used unsupervised random forest methods to estimate the proximity (similarity) between isolates based upon the genomic presentation of putative viru-lence traits We then grouped isolates (virulence clusters) using hierarchical clustering (Ward's method), used non-parametric bootstrapping to assess cluster stability, and externally validated the clusters against epidemiological virulence measures from FoodNet, the National Outbreak Reporting System (NORS), and US federal sampling of beef products. We identified five stable virulence clusters of S. enterica serovars. Cluster 1 (higher viru-lence) serovars yielded an annual incidence rate of domestically acquired sporadic cases roughly one and a half times higher than the other four clusters combined (Clusters 2-5, lower virulence). Compared to other clusters, cluster 1 also had a higher proportion of infections leading to hospitalization and was implicated in more foodborne and beef-associated outbreaks, despite being isolated at a similar frequency from beef products as other clusters. We also identified subpopulations within 11 serovars. Remarkably, we found S. Infantis and S. Typhimurium subpopulations that significantly differed in genome length and clinical case presentation. Further, we found that the presence of the pESI plasmid accounted for the genome length differences between the S. Infantis subpopulations. Our results show that S. enterica strains associated with highest incidence of human infections share a common virulence repertoire. This work could be updated regularly and used in combination with foodborne surveillance information to prioritize serovars of public health concern.
... We contend that this method is pertinent to virulence loci found with SPI as the regions are subject to horizontal gene transfer [46,47]. Common methods to differentiate serovars typically rely on the alignment of core genes or single nucleotide polymorphisms (SNP) identified against reference assemblies [7,25,48,49]. These methods must rely on post hoc analysis to determine if two evolutionary similar strains have acquired virulence factor genes which may correspond to differences in case presentation as witnessed in S. Infantis and S. Typhimurium. ...
... The epidemic of human infections with DT104 appeared to peak in the late 1990s in England, Wales, and Ireland, although it was somewhat later in other countries, including Scotland [6]. Although now overtaken by the monophasic S. Typhimurium ST34, which emerged in pig populations and spread globally [7], likely due to enhanced resistance to heavy metals over other variants [8,9], DT104 continues to circulate in animal populations, causing sporadic outbreaks in both animals and humans. A zoonotic outbreak in 2016, first identified in sheep, cattle, and horses in Anglesey, North Wales [10], presented the basis for this study. ...
SNP addresses are a pathogen typing method based on whole-genome sequences (WGSs), assigning groups at seven different levels of genetic similarity. Public health surveillance uses it for several gastro-intestinal infections; this work trialled its use in veterinary surveillance for salmonella outbreak detection. Comparisons were made between temporal and spatio-temporal cluster detection models that either defined cases by their SNP address or by phage type, using historical data sets. Clusters of SNP incidents were effectively detected by both methods, but spatio-temporal models consistently detected these clusters earlier than the corresponding temporal models. Unlike phage type, SNP addresses appeared spatially and temporally limited, which facilitated the differentiation of novel, stable, or expanding clusters in spatio-temporal models. Furthermore, these models flagged spatio-temporal clusters containing only two to three cases at first detection, compared with a median of seven cases in phage-type models. The large number of SNP addresses will require automated methods to implement these detection models routinely. Further work is required to explore how temporal changes and different host species may impact the sensitivity and specificity of cluster detection. In conclusion, given validation with more sequencing data, SNP addresses are likely to be a valuable addition to early warning systems in veterinary surveillance.
... Despite the fact that all groups displayed diarrhea, no fever was detected in the group challenged with S. 1,4, [5],12:i:-, and only piglets challenged with S. 1,4, [5],12:i:-shed Salmonella continuously throughout the trial, with higher excretion levels than the group challenged with S. Typhimurium [66]. These data allow for hypothesizing a competitive and selective advantage of S. 1,4, [5],12:i:-over S. Typhimurium, which may be aided by microevolution affecting antigenicity, pathogenicity, and transmission [67]. Because of its high transmission ability, this serovar has become a global public and animal health hazard [56]. ...
Salmonella is one of the most spread foodborne pathogens worldwide, and Salmonella
infections in humans still represent a global health burden. The main source of Salmonella infections
in humans is represented by contaminated animal-derived foodstuffs, with pork products being
one of the most important players. Salmonella infection in swine is critical not only because it is one
of the main causes of economic losses in the pork industry, but also because pigs can be infected
by several Salmonella serovars, potentially contaminating the pig meat production chain and thus
posing a significant threat to public health globally. As of now, in Europe and in the United States,
swine-related Salmonella serovars, e.g., Salmonella Typhimurium and its monophasic variant Salmonella
enterica subsp. enterica 1,4,[5],12:i:-, are also frequently associated with human salmonellosis cases.
Moreover, multiple outbreaks have been reported in the last few decades which were triggered
by the consumption of Salmonella-contaminated pig meat. Throughout the years, changes and
evolution across the pork industry may have acted as triggers for new issues and obstacles hindering
Salmonella control along the food chain. Gathered evidence reinforces the importance of coordinating
control measures and harmonizing monitoring programs for the efficient control of Salmonella in
swine. This is necessary in order to manage outbreaks of clinical disease in pigs and also to protect
pork consumers by controlling Salmonella subclinical carriage and shedding. This review provides
an update on Salmonella infection in pigs, with insights on Salmonella ecology, focusing mainly
on Salmonella Choleraesuis, S. Typhimurium, and S. 1,4,[5],12:i:-, and their correlation to human
salmonellosis cases. An update on surveillance methods for epidemiological purposes of Salmonella
infection in pigs and humans, in a “One Health” approach, will also be reported.
... The evolution and epidemiology of ST34 have been investigated extensively in high-income settings, including the UK, USA, Japan and Australia [9][10][11][12] . These showed that ST34 has likely diverged from the ancestral ST19 in the mid 1990s 10,13 , and its expansion is characterised by several genetic features: (a) frequent deletion(s) of fljB and the IncFII S. Typhimurium virulence plasmid, (b) acquisition of the genomic island SGI-4 enhancing resistance to copper 14 and (c) chromosomal integration of AMR genes conferring the ASSuT resistance pattern (to ampicillin, streptomycin, sulphonamides and tetracycline) 10 . ...
... genomes. The four major clones carried comparative amount of acquired AMR genes (median: [8][9][10][11][12], significantly lower than that of the VBSI clone (p < 0.05, ANOVA-Tukey test; Fig. 2c and Supplementary Data 2). Animal-derived ST34 carried as many AMR genes as those isolated from human, both when accounting for all Vietnamese isolates (p = 0.815, Wilcoxon signed rank test) or only those in five major clones (p = 0.304). ...
Salmonella enterica serotype 1,4,[5],12:i:- (Typhimurium monophasic variant) of sequence type (ST) 34 has emerged as the predominant pandemic genotype in recent decades. Despite increasing reports of resistance to antimicrobials in Southeast Asia, Salmonella ST34 population structure and evolution remained understudied in the region. Here we performed detailed genomic investigations on 454 ST34 genomes collected from Vietnam and diverse geographical sources to elucidate the pathogen’s epidemiology, evolution and antimicrobial resistance. We showed that ST34 has been introduced into Vietnam in at least nine occasions since 2000, forming five co-circulating major clones responsible for paediatric diarrhoea and bloodstream infection. Most expansion events were associated with acquisitions of large multidrug resistance plasmids of IncHI2 or IncA/C2. Particularly, the self-conjugative IncA/C2 pST34VN2 (co-transferring blaCTX-M-55, mcr-3.1, and qnrS1) underlies local expansion and intercontinental spread in two separate ST34 clones. At the global scale, Southeast Asia was identified as a potential hub for the emergence and dissemination of multidrug resistant Salmonella ST34, and mutation analysis suggests of selection in antimicrobial responses and key virulence factors.
... Therefore, our study makes a substantial contribution to the understanding of S. Typhimurium diversity with the identification of three new lineages associated with avian hosts (i.e., passerine lineage 1, larid lineage, and water bird lineage). Previous work on S. Typhimurium population structure focused on specific lineages formed by isolates from humans and domestic animals, and the geographic locations of these isolates were restricted to certain countries or regions (14,49,50,(52)(53)(54)66). As a result, the genetic diversity of this bacterial pathogen was underestimated due to a lack of representative isolates from wild animals, and the phylogenetic relationship of individual lineages remains largely unexplored on a global scale. ...
Within-host evolution of bacterial pathogens can lead to host-associated variants of the same species or serovar. Identification and characterization of closely related variants from diverse host species are crucial to public health and host-pathogen adaptation research. However, the work remained largely underexplored at a strain level until the advent of whole-genome sequencing (WGS). Here, we performed WGS-based subtyping and analyses of Salmonella enterica serovar Typhimurium (n = 787) from different wild birds across 18 countries over a 75-year period. We revealed seven avian host-associated S. Typhimurium variants/lineages. These lineages emerged globally over short timescales and presented genetic features distinct from S. Typhimurium lineages circulating among humans and domestic animals. We further showed that, in terms of virulence, host adaptation of these variants was driven by genome degradation. Our results provide a snapshot of the population structure and genetic diversity of S. Typhimurium within avian hosts. We also demonstrate the value of WGS-based subtyping and analyses in unravelling closely related variants at the strain level.
... In the preceding two decades, three other significant S. Typhimurium and its monophasic variant clones have been spreading: the European, Spanish and US clones (Nale et al., 2021). The S. 1,4, [5],12:i:-ST34 European clone encodes a chromosomal element that confers resistance to four antimicrobials (Petrovska et al., 2016). The Spanish S. 1,4,12:i:-ST19 clone harbors a plasmid that provides resistance to seven antimicrobials (Antunes et al., 2011;Duggett et al., 2016), while the US ST19 clone is rarely resistant to antimicrobials (Cadel-Six et al., 2021). ...
... A European lineage of S. 1,4,[5],12:i:-ST34, harboring SGI-4, mercury, and ASSuT resistance genes, has been reported previously (Petrovska et al., 2016;Cadel-Six et al., 2021). The genetic relatedness between BL708 and two previously reported isolates (S01861-18 and 12CEB4916SAL) was studied. ...
... The ST34 monophasic MDR variant of S. 1,4,[5],12:i:-is connected to human and animal infections (Hengkrawit and Tangjade, 2022) and has already been reported in the UK (Cadel-Six et al., 2021) and Thailand (Patchanee et al., 2020). Concerningly, not only is its prevalence increasing but an mcrcarrying variant conferring resistance to the last resort antibiotic colistin has been reported (Petrovska et al., 2016;Elnekave et al., 2018;Biswas et al., 2019;Patchanee et al., 2020;Ingle et al., 2021;Hengkrawit and Tangjade, 2022). ...
Salmonella is a food-borne pathogen often linked to poultry sources, causing gastrointestinal infections in humans, with the numbers of multidrug resistant (MDR) isolates increasing globally. To gain insight into the genomic diversity of common serovars and their potential contribution to disease, we characterized antimicrobial resistance genes, and virulence factors encoded in 88 UK and 55 Thai isolates from poultry; the presence of virulence genes was detected through an extensive virulence determinants database compiled in this study. Long-read sequencing of three MDR isolates, each from a different serovar, was used to explore the links between virulence and resistance. To augment current control methods, we determined the sensitivity of isolates to 22 previously characterized Salmonella bacteriophages. Of the 17 serovars included, Salmonella Typhimurium and its monophasic variants were the most common, followed by S. Enteritidis, S. Mbandaka, and S. Virchow. Phylogenetic analysis of Typhumurium and monophasic variants showed poultry isolates were generally distinct from pigs. Resistance to sulfamethoxazole and ciprofloxacin was highest in isolates from the UK and Thailand, respectively, with 14–15% of all isolates being MDR. We noted that >90% of MDR isolates were likely to carry virulence genes as diverse as the srjF , lpfD , fhuA , and stc operons. Long-read sequencing revealed the presence of global epidemic MDR clones in our dataset, indicating they are possibly widespread in poultry. The clones included MDR ST198 S . Kentucky, harboring a Salmonella Genomic Island-1 (SGI)-K, European ST34 S . 1,4,[5],12:i:-, harboring SGI-4 and mercury-resistance genes, and a S . 1,4,12:i:- isolate from the Spanish clone harboring an MDR-plasmid. Testing of all isolates against a panel of bacteriophages showed variable sensitivity to phages, with STW-77 found to be the most effective. STW-77 lysed 37.76% of the isolates, including serovars important for human clinical infections: S . Enteritidis (80.95%), S . Typhimurium (66.67%), S . 1,4,[5],12:i:- (83.3%), and S . 1,4,12: i:- (71.43%). Therefore, our study revealed that combining genomics and phage sensitivity assays is promising for accurately identifying and providing biocontrols for Salmonella to prevent its dissemination in poultry flocks and through the food chain to cause infections in humans.
