Limits...
The Shiga toxin 2 production level in enterohemorrhagic Escherichia coli O157:H7 is correlated with the subtypes of toxin-encoding phage.

Ogura Y, Mondal SI, Islam MR, Mako T, Arisawa K, Katsura K, Ooka T, Gotoh Y, Murase K, Ohnishi M, Hayashi T - Sci Rep (2015)

Bottom Line: We further show that one of the two subclades in the clade 8, a proposed hyper-virulent lineage of O157, carries the Stx2 phage subtype that confers the highest Stx2 production to the host strain.The presence of this subclade may explain the proposed high virulence potential of clade 8.These results provide novel insights into the variation in virulence among O157 strains and highlight the role of phage variation in determining the production level of the virulence factors that phages encode.

View Article: PubMed Central - PubMed

Affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.

ABSTRACT
Enterohemorrhagic E. coli (EHEC) causes diarrhea and hemorrhagic colitis with life-threatening complications, such as hemolytic uremic syndrome. Their major virulence factor is Shiga toxin (Stx), which is encoded by bacteriophages. Of the two types of Stx, the production of Stx2, particularly that of Stx2a (a subtype of Stx2), is a major risk factor for severe EHEC infections, but the Stx2 production level is highly variable between strains. Here, we define four major and two minor subtypes of Stx2a-encoding phages according to their replication proteins. The subtypes are correlated with Stx2a titers produced by the host O157 strains, suggesting a critical role of the phage subtype in determining the Stx2a production level. We further show that one of the two subclades in the clade 8, a proposed hyper-virulent lineage of O157, carries the Stx2 phage subtype that confers the highest Stx2 production to the host strain. The presence of this subclade may explain the proposed high virulence potential of clade 8. These results provide novel insights into the variation in virulence among O157 strains and highlight the role of phage variation in determining the production level of the virulence factors that phages encode.

No MeSH data available.


Related in: MedlinePlus

Highly variable Stx2 production levels among O157 strains.The Stx2 production levels of the 65 O157 strains are shown. The supernatants of the polymixin B-treated cultures were prepared from the 65 O157 strains without or with the MMC treatment (panels (A,B), respectively). The Stx2 concentration in each supernatant was measured using the VTEC-Reversed Passive Latex Agglutination (VTEC-RPLA) assay kit, and the resulting RPLA titers are shown. The Stx2 prophages of the strains indicated by the dots were sequenced in this study. The strains were categorized according to their stx2 subtypes and Stx2 phage integration sites.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4645166&req=5

f1: Highly variable Stx2 production levels among O157 strains.The Stx2 production levels of the 65 O157 strains are shown. The supernatants of the polymixin B-treated cultures were prepared from the 65 O157 strains without or with the MMC treatment (panels (A,B), respectively). The Stx2 concentration in each supernatant was measured using the VTEC-Reversed Passive Latex Agglutination (VTEC-RPLA) assay kit, and the resulting RPLA titers are shown. The Stx2 prophages of the strains indicated by the dots were sequenced in this study. The strains were categorized according to their stx2 subtypes and Stx2 phage integration sites.

Mentions: From the 123 strains, we selected 65 that represented various combinations of Stx2 subtypes, Stx2 phage integration sites and phylogenetic clades, and we measured their Stx2 production levels in the presence and absence of MMC by means of a reversed passive latex agglutination (RPLA) assay. Even in the absence of MMC, Stx2 production was detected in all the strains. A remarkable strain-to-strain variation in toxin titer (which ranged from 2 to 1,600 in RPLA titer) was also observed (Fig. 1 and Supplementary Table S1). Consistent with previous reports171819, MMC treatment did not strongly enhance Stx2 production in the strains carrying stx2c alone. In contrast, Stx2 production was remarkably enhanced by MMC in almost all the strains carrying stx2a, and the observed Stx2 production levels were highly variable among the strains. The levels of MMC-induced Stx2 production by these strains were significantly correlated with those in the absence of MMC (Supplementary Fig. S1), but there was no significant relation between Stx2 production and the clade of the strain or the integration site of the Stx2a phage. Recently, Neupane et al. showed that clade 8 strains overexpressed stx2 in both RNA and protein levels compared to clade 1–3 strains11. In this study, however, only four clade 8 strains were analyzed and one of them produced a very low level of Stx2. Thus, although the authors concluded that the increased stx2 expression is characteristics of clade8 strains, it is not exclusive to this clade and not all strains within clade 8 overexpress stx2. The Stx2 production patterns of stx2a/2c double-positive strains were similar to those of stx2a single-positive strains. This is consistent with a previous study by Kawano et al.18, and suggests that the Stx2 production levels of the stx2a/2c double-positive strains are largely representative of the levels of Stx2a production by each strain.


The Shiga toxin 2 production level in enterohemorrhagic Escherichia coli O157:H7 is correlated with the subtypes of toxin-encoding phage.

Ogura Y, Mondal SI, Islam MR, Mako T, Arisawa K, Katsura K, Ooka T, Gotoh Y, Murase K, Ohnishi M, Hayashi T - Sci Rep (2015)

Highly variable Stx2 production levels among O157 strains.The Stx2 production levels of the 65 O157 strains are shown. The supernatants of the polymixin B-treated cultures were prepared from the 65 O157 strains without or with the MMC treatment (panels (A,B), respectively). The Stx2 concentration in each supernatant was measured using the VTEC-Reversed Passive Latex Agglutination (VTEC-RPLA) assay kit, and the resulting RPLA titers are shown. The Stx2 prophages of the strains indicated by the dots were sequenced in this study. The strains were categorized according to their stx2 subtypes and Stx2 phage integration sites.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4645166&req=5

f1: Highly variable Stx2 production levels among O157 strains.The Stx2 production levels of the 65 O157 strains are shown. The supernatants of the polymixin B-treated cultures were prepared from the 65 O157 strains without or with the MMC treatment (panels (A,B), respectively). The Stx2 concentration in each supernatant was measured using the VTEC-Reversed Passive Latex Agglutination (VTEC-RPLA) assay kit, and the resulting RPLA titers are shown. The Stx2 prophages of the strains indicated by the dots were sequenced in this study. The strains were categorized according to their stx2 subtypes and Stx2 phage integration sites.
Mentions: From the 123 strains, we selected 65 that represented various combinations of Stx2 subtypes, Stx2 phage integration sites and phylogenetic clades, and we measured their Stx2 production levels in the presence and absence of MMC by means of a reversed passive latex agglutination (RPLA) assay. Even in the absence of MMC, Stx2 production was detected in all the strains. A remarkable strain-to-strain variation in toxin titer (which ranged from 2 to 1,600 in RPLA titer) was also observed (Fig. 1 and Supplementary Table S1). Consistent with previous reports171819, MMC treatment did not strongly enhance Stx2 production in the strains carrying stx2c alone. In contrast, Stx2 production was remarkably enhanced by MMC in almost all the strains carrying stx2a, and the observed Stx2 production levels were highly variable among the strains. The levels of MMC-induced Stx2 production by these strains were significantly correlated with those in the absence of MMC (Supplementary Fig. S1), but there was no significant relation between Stx2 production and the clade of the strain or the integration site of the Stx2a phage. Recently, Neupane et al. showed that clade 8 strains overexpressed stx2 in both RNA and protein levels compared to clade 1–3 strains11. In this study, however, only four clade 8 strains were analyzed and one of them produced a very low level of Stx2. Thus, although the authors concluded that the increased stx2 expression is characteristics of clade8 strains, it is not exclusive to this clade and not all strains within clade 8 overexpress stx2. The Stx2 production patterns of stx2a/2c double-positive strains were similar to those of stx2a single-positive strains. This is consistent with a previous study by Kawano et al.18, and suggests that the Stx2 production levels of the stx2a/2c double-positive strains are largely representative of the levels of Stx2a production by each strain.

Bottom Line: We further show that one of the two subclades in the clade 8, a proposed hyper-virulent lineage of O157, carries the Stx2 phage subtype that confers the highest Stx2 production to the host strain.The presence of this subclade may explain the proposed high virulence potential of clade 8.These results provide novel insights into the variation in virulence among O157 strains and highlight the role of phage variation in determining the production level of the virulence factors that phages encode.

View Article: PubMed Central - PubMed

Affiliation: Department of Bacteriology, Faculty of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.

ABSTRACT
Enterohemorrhagic E. coli (EHEC) causes diarrhea and hemorrhagic colitis with life-threatening complications, such as hemolytic uremic syndrome. Their major virulence factor is Shiga toxin (Stx), which is encoded by bacteriophages. Of the two types of Stx, the production of Stx2, particularly that of Stx2a (a subtype of Stx2), is a major risk factor for severe EHEC infections, but the Stx2 production level is highly variable between strains. Here, we define four major and two minor subtypes of Stx2a-encoding phages according to their replication proteins. The subtypes are correlated with Stx2a titers produced by the host O157 strains, suggesting a critical role of the phage subtype in determining the Stx2a production level. We further show that one of the two subclades in the clade 8, a proposed hyper-virulent lineage of O157, carries the Stx2 phage subtype that confers the highest Stx2 production to the host strain. The presence of this subclade may explain the proposed high virulence potential of clade 8. These results provide novel insights into the variation in virulence among O157 strains and highlight the role of phage variation in determining the production level of the virulence factors that phages encode.

No MeSH data available.


Related in: MedlinePlus