Hypervirulent R20291 Clostridioides difficile spores show disinfection resilience to sodium hypochlorite despite structural changes

Malyshev, Dmitry; Jones, Imogen Anne; McKracken, Matthew; Öberg, Rasmus; Harper, Glenn; Joshi, Lovleen Tina; Andersson, Magnus

doi:10.1186/s12866-023-02787-z

Cited by 19 publications

(33 citation statements)

References 64 publications

(73 reference statements)

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“…In this study, SEM was used to determine whether CRA use physically damaged the outer spore layers. The results clearly demonstrated no visible effect on spore morphology after biocide exposure, which correlates with results from our previous study [20]. C.…”

Section: Discussionsupporting

confidence: 90%

“…Growth conditions, C. difficile ribotypes and spore production C. difficile strains from two clinically relevant ribotypes (RTs) were used in this study: the hypervirulent epidemic Stoke-Mandeville R20291 strain (RT 027), clinical isolate DS1813 (RT 027) and the type strain CD630 (RT012). These strains have been widely tested for susceptibility to disinfection [20,29,30]. Strains used in this study and their PCR RTs are described in Table 1 and were obtained from the Anaerobic Reference Unit, University Hospital Wales, Cardiff, UK.…”

Section: Methodsmentioning

confidence: 99%

“…Spores from strains R20291, DS1813 and CD630 (Table 1) at a concentration of 1×10 8 spores ml -1 were independently exposed to 1000, 5000 and 10 000 p.p.m. NaOCl in liquid form for 10 min (recommended contact time) and biocide activity was neutralized with an equal volume of 5 g l -1 sodium thiosulphate for 10 min contact time to remove any residual chlorine biocide activity [20,28,29]. Spores were enumerated on supplemented BHI agar after anaerobic incubation at 37 °C for 48 h and growth of a single C. difficile colony on the agar correlated to evidence of single spore germination [31,32].…”

Section: Exposure Of Spores To Sodium Hypochlorite Disinfectantmentioning

confidence: 99%

“…Current UK guidelines recommend use of CRAs at 1000 p.p.m. for ~10 min contact time to disinfect surfaces soiled with C. difficile; CRAs are a low-cost, efficient disinfection method used in healthcare facilities to disinfect surfaces and include sodium hypochlorite bleach (NaOCl) and sodium dichloroisocyanurate (NaDCC) [19][20][21]. However, recent studies have shown emerging sporicidal resistance to 1000 p.p.m.…”

Section: Introductionmentioning

confidence: 99%

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Clostridioides difficile spores tolerate disinfection with sodium hypochlorite disinfectant and remain viable within surgical scrubs and gown fabrics

Ahmed,

Joshi

2023

Microbiology

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Clostridioides difficile is the most common cause of antibiotic-associated diarrhoea globally. Its spores have been implicated in the prevalence of C. difficile infection due to their resistance and transmission ability between surfaces. Currently, disinfectants such as chlorine-releasing agents (CRAs) and hydrogen peroxide are used to decontaminate and reduce the incidence of infections in clinical environments. Our previous research demonstrated the ability of C. difficile spores to survive exposure to recommended concentrations of sodium dichloroisocyanurate in liquid form and within personal protective fabrics such as surgical gowns; however, the present study examined the spore response to clinical in-use concentrations of sodium hypochlorite. Spores were exposed to a 10 min contact time of 1000, 5000 and 10 000 p.p.m. sodium hypochlorite, and spore recovery was determined. To understand whether biocide-exposed spores transmitted across clinical surfaces in vitro, biocide-exposed spores were spiked onto surgical scrubs and patient gowns and recovery was determined by a plate transfer assay. Scanning electron microscopy was used to establish if there were any morphological changes to the outer spore coat. The results revealed that viable biocide-exposed C. difficile spores can be recovered from surgical scrubs and patient gowns, with no observable changes to spore morphology, highlighting the potential of these fabrics as vectors of spore transmission. This study demonstrates that alternative strategies should be urgently sought to disinfect C. difficile spores to break the chain of transmission in clinical environments.

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Section: Discussionsupporting

confidence: 90%

Section: Methodsmentioning

confidence: 99%

Section: Exposure Of Spores To Sodium Hypochlorite Disinfectantmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Clostridioides difficile spores tolerate disinfection with sodium hypochlorite disinfectant and remain viable within surgical scrubs and gown fabrics

Ahmed,

Joshi

2023

Microbiology

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“…The sporicidal assay was performed as previously described [96]. Spore suspensions were exposed for 10 min to 0.1% HClO or to water.…”

Section: Spore Production Germination and Sporicidal Assaysmentioning

confidence: 99%

The multiplicity of Thioredoxin systems meets the specific needs of Clostridia

Anjou,

Lotoux,

Zhukova

et al. 2023

Preprint

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Oxidative stress is a highly common stress for cells, which targets proteins with oxidation of cysteine residues. The thioredoxin (Trx) system, which is a ubiquitous system for thiol- and protein-repair, is composed of a thioredoxin (TrxA) and a thioredoxin-reductase (TrxB). TrxAs reduce disulfide bonds of oxidized proteins and are then usually recycled by a single pleiotropic NAD(P)H-dependent TrxB (NTR). However, some Clostridia have also ferredoxin-dependent TrxBs. In this work, we first analyzed the composition of Trx systems across Bacteria. Most of bacteria have only one NTR, but organisms in some Phyla including Firmicutes have several TrxBs. In Firmicutes, this multiplicity of TrxBs is observed only in Clostridia. We thus used Clostridioides difficile as a model to investigate the biological relevance of TrxB multiplicity by studying the physiological roles of the Trx systems in this gut pathogen. Three TrxAs and three TrxBs are present in the 630Δerm strain. We showed that two systems were involved in response to infection-related stresses, allowing survival of vegetative cells to exposure to oxygen, inflammation-related molecules and bile salts. A supplementary TrxB copy present in some C. difficile strains also contributes to this stress-response arsenal. One of the conserved stress-response Trx system was also found to be present in the spore via a dual transcriptional control by different sigma factors. This system contributes to spore survival to hypochlorite and ensure proper germination in the presence of oxygen. Finally, we found that the third Trx system was contributing to sporulation. This involvement was likely linked to the recycling of the glycine-reductase, a Stickland pathway enzyme that allows consumption of glycine, a spore co-germinant. Altogether, our results showed that the multiplicity of Trx systems produced under the control of different regulatory signals and networks and the diversity of TrxBs meet specific needs of Clostridia, i.e., adaptation to strong stress exposure, sporulation and Stickland pathways. More broadly, this multiplicity responds to cell compartmentation and differentiation, which can be transposed to other multiple-TrxBs organisms such as Cyanobacteria or eukaryotes.

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Single‐cell analysis reveals microbial spore responses to sodium hypochlorite

Yang,

Yuan,

et al. 2024

Journal of Biophotonics

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Pollution from toxic spores has caused us a lot of problems because spores are extremely resistant and can survive most disinfectants. Therefore, the detection of spore response to disinfectant is of great significance for the development of effective decontamination strategies. In this work, we investigated the effect of 0.5% sodium hypochlorite on the molecular and morphological properties of single spores of Bacillus subtilis using single‐cell techniques. Laser tweezers Raman spectroscopy showed that sodium hypochlorite resulted in Ca2+‐dipicolinic acid release and nucleic acid denaturation. Atomic force microscopy showed that the surface of treated spores changed from rough to smooth, protein shells were degraded at 10 min, and the permeability barrier was destroyed at 15 min. The spore volume decreased gradually over time. Live‐cell imaging showed that the germination and growth rates decreased with increasing treatment time. These results provide new insight into the response of spores to sodium hypochlorite.

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Hypervirulent R20291 Clostridioides difficile spores show disinfection resilience to sodium hypochlorite despite structural changes

Cited by 19 publications

References 64 publications

Clostridioides difficile spores tolerate disinfection with sodium hypochlorite disinfectant and remain viable within surgical scrubs and gown fabrics

Clostridioides difficile spores tolerate disinfection with sodium hypochlorite disinfectant and remain viable within surgical scrubs and gown fabrics

The multiplicity of Thioredoxin systems meets the specific needs of Clostridia

Single‐cell analysis reveals microbial spore responses to sodium hypochlorite

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