scholarly journals Phylogenomic analysis of Clostridioides difficile ribotype 106 strains reveals novel genetic islands and emergent phenotypes

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Bryan Angelo P. Roxas ◽  
Jennifer Lising Roxas ◽  
Rachel Claus-Walker ◽  
Anusha Harishankar ◽  
Asad Mansoor ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Diarrheal Disease ◽  
The United States ◽  
Genomic Islands ◽  
Rapid Expansion ◽  
Phylogenomic Analysis ◽  
Community Settings ◽  
Hamster Model ◽  
Clostridioides Difficile ◽  
Healthcare Associated

AbstractClostridioides difficile infection (CDI) is a major healthcare-associated diarrheal disease. Consistent with trends across the United States, C. difficile RT106 was the second-most prevalent molecular type in our surveillance in Arizona from 2015 to 2018. A representative RT106 strain displayed robust virulence and 100% lethality in the hamster model of acute CDI. We identified a unique 46 KB genomic island (GI1) in all RT106 strains sequenced to date, including those in public databases. GI1 was not found in its entirety in any other C. difficile clade, or indeed, in any other microbial genome; however, smaller segments were detected in Enterococcus faecium strains. Molecular clock analyses suggested that GI1 was horizontally acquired and sequentially assembled over time. GI1 encodes homologs of VanZ and a SrtB-anchored collagen-binding adhesin, and correspondingly, all tested RT106 strains had increased teicoplanin resistance, and a majority displayed collagen-dependent biofilm formation. Two additional genomic islands (GI2 and GI3) were also present in a subset of RT106 strains. All three islands are predicted to encode mobile genetic elements as well as virulence factors. Emergent phenotypes associated with these genetic islands may have contributed to the relatively rapid expansion of RT106 in US healthcare and community settings.

scholarly journals Phylogenomic analysis of Clostridiodes difficile ribotype 106 strains reveals novel genetic islands and emergent phenotypes

2020 ◽  
Author(s):  
Bryan Angelo P. Roxas ◽  
Jennifer Lising Roxas ◽  
Rachel Claus-Walker ◽  
Anusha Harishankar ◽  
Asad Mansoor ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Genomic Island ◽  
Genomic Islands ◽  
Phylogenomic Analysis ◽  
Molecular Type ◽  
Phenotypic Properties ◽  
Hamster Model ◽  
Clostridioides Difficile ◽  
Genes Encoding ◽  
The Usa

ABSTRACTBackgroundClostridioides difficile RT106 has emerged as a dominant molecular type in the USA in recent years, but the underlying factors contributing to its predominance remain undefined. As part of our ongoing C. difficile infection (CDI) surveillance in Arizona, we monitored RT106 frequency and characterized the genomic and phenotypic properties of the recovered isolates.ResultsFrom 2015-2018, RT106 was the second-most prevalent molecular type isolated from CDI patients in our surveillance. A representative RT106 strain displayed robust virulence and 100% lethality in the hamster model of acute CDI. We identified a unique 46 KB genomic island (GI1) in all RT106 strains, including those in public databases. GI1 was not found in its entirety in any other C. difficile clade, or indeed in any other microbial genome; however, smaller segments were detected in select Enterococcus faecium strains. Molecular clock analyses suggest that GI1 was horizontally acquired and sequentially assembled over time. Consistent with the presence of genes encoding homologs of VanZ and a SrtB-anchored collagen-binding adhesin in GI1, all tested RT106 strains had increased teicoplanin resistance and a majority displayed collagen-dependent biofilm formation. Two additional genomic islands (GI2 and GI3) were also present in a subset of RT106 strains. All three islands have features of mobile genetic elements and encode several putative virulence factors.ConclusionsConsistent with the known genetic plasticity of C. difficile, strains belonging to the RT106 clade harbor unique genetic islands. Correspondingly, emergent phenotypic properties may contribute to the relatively rapid shifts of strain distribution in patient populations.

scholarly journals Spo0A Suppresses sin Locus Expression in Clostridioides difficile

mSphere ◽  
2020 ◽  
Vol 5 (6) ◽  
Author(s):  
Babita Adhikari Dhungel ◽  
Revathi Govind
Keyword(s):  
Diarrheal Disease ◽  
Toxin Production ◽  
The United States ◽  
Upstream Region ◽  
Pcr Analysis ◽  
Bacterial Cells ◽  
Master Regulator ◽  
Content Type ◽  
Clostridioides Difficile

ABSTRACT Clostridioides difficile is the leading cause of nosocomial infection and is the causative agent of antibiotic-associated diarrhea. The severity of the disease is directly associated with toxin production, and spores are responsible for the transmission and persistence of the organism. Previously, we characterized sin locus regulators SinR and SinR′ (we renamed it SinI), where SinR is the regulator of toxin production and sporulation. The SinI regulator acts as its antagonist. In Bacillus subtilis, Spo0A, the master regulator of sporulation, controls SinR by regulating the expression of its antagonist, sinI. However, the role of Spo0A in the expression of sinR and sinI in C. difficile had not yet been reported. In this study, we tested spo0A mutants in three different C. difficile strains, R20291, UK1, and JIR8094, to understand the role of Spo0A in sin locus expression. Western blot analysis revealed that spo0A mutants had increased SinR levels. Quantitative reverse transcription-PCR (qRT-PCR) analysis of its expression further supported these data. By carrying out genetic and biochemical assays, we show that Spo0A can bind to the upstream region of this locus to regulates its expression. This study provides vital information that Spo0A regulates the sin locus, which controls critical pathogenic traits such as sporulation, toxin production, and motility in C. difficile. IMPORTANCE Clostridioides difficile is the leading cause of antibiotic-associated diarrheal disease in the United States. During infection, C. difficile spores germinate, and the vegetative bacterial cells produce toxins that damage host tissue. In C. difficile, the sin locus is known to regulate both sporulation and toxin production. In this study, we show that Spo0A, the master regulator of sporulation, controls sin locus expression. Results from our study suggest that Spo0A directly regulates the expression of this locus by binding to its upstream DNA region. This observation adds new detail to the gene regulatory network that connects sporulation and toxin production in this pathogen.

scholarly journals Defining the black box: a narrative review of factors associated with adverse outcomes from severe Clostridioides difficile infection

2021 ◽  
Vol 14 ◽  
pp. 175628482110481
Author(s):  
Adam Ressler ◽  
Joyce Wang ◽  
Krishna Rao
Keyword(s):  
The United States ◽  
Adverse Outcomes ◽  
Chronic Illnesses ◽  
Narrative Review ◽  
Healthcare Associated Infection ◽  
Clostridioides Difficile ◽  
Clostridioides Difficile Infection ◽  
Patients At Risk ◽  
Hospital Deaths ◽  
Healthcare Associated

In the United States, Clostridioides difficile infection (CDI) is the leading cause of healthcare-associated infection, affecting nearly half a million people and resulting in more than 20,000 in-hospital deaths every year. It is therefore imperative to better characterize the intricate interplay between C. difficile microbial factors, host immunologic signatures, and clinical features that are associated with adverse outcomes of severe CDI. In this narrative review, we discuss the implications of C. difficile genetics and virulence factors in the molecular epidemiology of CDI, and the utility of early biomarkers in predicting the clinical trajectory of patients at risk of developing severe CDI. Furthermore, we identify associations between host immune factors and CDI outcomes in both animal models and human studies. Next, we highlight clinical factors including renal dysfunction, aging, blood biomarkers, level of care, and chronic illnesses that can affect severe CDI diagnosis and outcome. Finally, we present our perspectives on two specific treatments pertinent to patient outcomes: metronidazole administration and surgery. Together, this review explores the various venues of CDI research and highlights the importance of integrating microbial, host, and clinical data to help clinicians make optimal treatment decisions based on accurate prediction of disease progression.

scholarly journals Biofilm regulation in Clostridioides difficile: Novel systems linked to hypervirulence

2021 ◽  
Vol 17 (9) ◽  
pp. e1009817
Author(s):  
Megan G. Taggart ◽  
William J. Snelling ◽  
Patrick J. Naughton ◽  
Roberto M. La Ragione ◽  
James S. G. Dooley ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Gastrointestinal Disease ◽  
Regulatory System ◽  
Disease Recurrence ◽  
The United States ◽  
Extracellular Dna ◽  
Published Data ◽  
Bacterial Cells ◽  
Healthcare Associated Infection ◽  
Clostridioides Difficile

Clostridiodes difficile (C. difficile) was ranked an “urgent threat” by the Centers for Disease Control and Prevention (CDC) in 2019. C. difficile infection (CDI) is the most common healthcare-associated infection (HAI) in the United States of America as well as the leading cause of antibiotic-associated gastrointestinal disease. C. difficile is a gram-positive, rod-shaped, spore-forming, anaerobic bacterium that causes infection of the epithelial lining of the gut. CDI occurs most commonly after disruption of the human gut microflora following the prolonged use of broad-spectrum antibiotics. However, the recurrent nature of this disease has led to the hypothesis that biofilm formation may play a role in its pathogenesis. Biofilms are sessile communities of bacteria protected from extracellular stresses by a matrix of self-produced proteins, polysaccharides, and extracellular DNA. Biofilm regulation in C. difficile is still incompletely understood, and its role in disease recurrence has yet to be fully elucidated. However, many factors have been found to influence biofilm formation in C. difficile, including motility, adhesion, and hydrophobicity of the bacterial cells. Small changes in one of these systems can greatly influence biofilm formation. Therefore, the biofilm regulatory system would need to coordinate all these systems to create optimal biofilm-forming physiology under appropriate environmental conditions. The coordination of these systems is complex and multifactorial, and any analysis must take into consideration the influences of the stress response, quorum sensing (QS), and gene regulation by second messenger molecule cyclic diguanosine monophosphate (c-di-GMP). However, the differences in biofilm-forming ability between C. difficile strains such as 630 and the “hypervirulent” strain, R20291, make it difficult to assign a “one size fits all” mechanism to biofilm regulation in C. difficile. This review seeks to consolidate published data regarding the regulation of C. difficile biofilms in order to identify gaps in knowledge and propose directions for future study.

scholarly journals Genomic analysis of Clostridioides difficile in two regions reveals a diversity of strains and limited transmission

2020 ◽  
Vol 41 (S1) ◽  
pp. s237-s238
Author(s):  
Nicole Pecora ◽  
Stacy Holzbauer ◽  
Xiong Wang ◽  
Yu Gu ◽  
Trupti Hatwar ◽  
...  
Keyword(s):  
New York ◽  
Genomic Analysis ◽  
Illumina Miseq ◽  
The United States ◽  
Snp Analysis ◽  
Whole Genome ◽  
Emerging Infections ◽  
Clostridioides Difficile ◽  
Healthcare Associated ◽  
Incident Cases

Background: The epidemic NAP1/027 Clostridioides difficile strain (MLST1, ST1) that emerged in the mid-2000 is on the decline. The current distribution of C. difficile strain types and their transmission dynamics are poorly defined. We performed whole-genome sequencing (WGS) of C. difficile isolates in 2 regions to identify the predominant multilocus sequence types (MLSTs) in community- and healthcare-associated cases and potential transmission between cases using whole-genome single-nucleotide polymorphism (SNP) analysis. Methods: Isolates were collected through the CDC Emerging Infections Program population-based surveillance for C. difficile infections (CDI) for 3 months between 2016 and 2017 in 5 Minnesota counties and 1 New York county. Isolates were limited to incident cases (CDI in a county resident with no positive C. difficile test in the preceding 8 weeks). Cases were classified as healthcare associated (HA-CDI) or community associated (CA-CDI) based on healthcare exposures as previously described. WGS was performed on an Illumina Miseq. The CFSAN (FDA) pipeline was used to compute whole-genome SNPs, SPAdes was used for assembly, and MLST was assigned according to www.pubmlst.org. Results: Of 431 isolates, 269 originated from New York and 162 from Minnesota; 203 cases were classified as CA-CDI and 221 as HA-CDI. The proportion of CA-CDI cases was higher in Minnesota than in New York: 62% vs 38%. The predominant MLSTs across both sites were ST42 (9%), ST8 (8%), and ST2 (8%). MLSTs more frequently encountered in HA-CDI than CA-CDI included ST1 (note that this ST includes PCR Ribotype 027; 76% HA-CDI), ST53 (84% HA-CDI), and ST43 (80% HA-CDI). In contrast, ST110 (63% CA-CDI) and ST3 (67% CA-CDI) were more commonly isolated from CA-CDI cases. ST1 accounted for 7.6% of circulating strains and was more common in New York than Minnesota (10% vs 3%) and was concentrated among New York HA-CDI cases. Also, 412 isolates (1 per patient) were included in the final whole-genome SNP analysis. Of these, only 12 pairs were separated by 0–3 SNPs, indicating potential transmission, and most involved HA-CDI cases. ST1, ST17, and ST46 accounted for 8 of 12 pairs, with ST17 and ST46 potentially forming small clusters. Conclusions: This analysis provides a snapshot of the current genomic epidemiology of C. difficile across 2 geographically and epidemiologically distinct regions of the United States and supports other studies suggesting that the role of direct transmission in the spread of CDI may be limited.Funding: NoneDisclosures: None

scholarly journals 2404. Molecular Epidemiology of Clostridioides difficile in the United States, 2017

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S830-S830 ◽  
Author(s):  
Ashley Paulick ◽  
Michelle Adamczyk ◽  
Lauren C Korhonen ◽  
Alice Guh ◽  
Amy Gargis ◽  
...  
Keyword(s):  
United States ◽  
Molecular Epidemiology ◽  
The United States ◽  
Emerging Infections ◽  
Convenience Sample ◽  
Clostridioides Difficile ◽  
Positive Stool ◽  
Control And Prevention ◽  
Clostridioides Difficile Infection ◽  
Healthcare Associated

Abstract Background In 2009, the Centers for Disease Control and Prevention (CDC) implemented Clostridioides difficile infection (CDI) surveillance through the Emerging Infections Program (EIP) to monitor the incidence and evolving epidemiology of CDI in the United States. Since 2012, ribotypes (RTs) 027, 106, 002, 014, and 020 have constituted the top five strain types among both US community- and healthcare-associated isolates. Here we describe the changes in molecular epidemiology of C. difficile isolates collected in the United States in 2017. Methods In 2017, CDI surveillance was conducted at 10 EIP sites (CA, CO, CT, GA, MD, MN, NM, NY, OR, and TN). A convenience sample of clinical laboratories across EIP sites submitted C. difficile-positive stool specimens to the MN Department of Health Public Health Laboratory and Hines VA Hospital (IL) for culture. Isolates were forwarded to CDC and characterized by capillary-based PCR-ribotyping and PCR detection of tcdA, tcdB, cdtA, cdtB, and deletions in tcdC. Results In 2017, 1,051 C. difficile isolates were submitted; the total number of isolates received from each site ranged from 11 to 286 with a median of 85.5. In total, 143 RTs were observed, with the majority of isolates harboring toxin genes tcdA and tcdB (95%) and a wild-type tcdC sequence (71%). Among 556 healthcare-associated isolates, RT 027 was the most prevalent and the top RT at 5 sites (CA, GA, MD, NM, TN). Ribotype 106 was the most prevalent among 495 community-associated CA isolates and the top RT at 6 sites (CO, CT, GA, MD, MN, TN). Ribotype 027 significantly decreased from 2012 to 2017 among both healthcare-associated (21% vs 15%; p = 0.02) and community-associated isolates (17% vs 6%; P < 0.0001). Among healthcare-associated isolates, RT 076, which was observed in 8 EIP sites, increased from 2% in 2016 to 5% in 2017 (p = 0.05) and replaced RT 020 as one of the top 5 healthcare-associated RTs in 2017. Conclusion Despite an overall decline since 2012, RT 027 remained the most prevalent RT among healthcare-associated isolates submitted in 2017. The increased frequency of RT 076 among healthcare-associated isolates submitted in 2017 highlights the evolving molecular epidemiology of C. difficile and the need for continued surveillance to monitor potential emerging strains. Disclosures All authors: No reported disclosures.

scholarly journals Epidemiology of community-acquired and recurrent Clostridioides difficile infection

2021 ◽  
Vol 14 ◽  
pp. 175628482110162
Author(s):  
Yichun Fu ◽  
Yuying Luo ◽  
Ari M Grinspan
Keyword(s):  
Large Scale ◽  
Infection Prevention ◽  
The United States ◽  
Future Research ◽  
Clostridioides Difficile ◽  
Epidemiologic Surveillance ◽  
Scale Population ◽  
Clostridioides Difficile Infection ◽  
Healthcare Associated ◽  
Hospital Acquired

Clostridioides difficile infection is a leading cause of healthcare-associated infections with significant morbidity and mortality. For the past decade, the bulk of infection prevention and epidemiologic surveillance efforts have been directed toward mitigating hospital-acquired C. difficile. However, the incidence of community-associated infection is on the rise. Patients with community-associated C. difficile tend to be younger and have lower mortality rate. Rates of recurrent C. difficile infection overall have decreased in the United States, but future research and public health endeavors are needed to standardize and improve disease detection, stratify risk factors in large-scale population studies, and to identify regional and local variations in strain types, reservoirs and transmission routes to help characterize and combat the changing epidemiology of C. difficile.

scholarly journals Spo0A suppresses sin locus expression in Clostridioides difficile

2020 ◽  
Author(s):  
Babita Adhikari Dhungel ◽  
Revathi Govind
Keyword(s):  
Diarrheal Disease ◽  
Toxin Production ◽  
The United States ◽  
Upstream Region ◽  
Pcr Analysis ◽  
Bacterial Cells ◽  
Master Regulator ◽  
Clostridioides Difficile ◽  
Severity Of The Disease

AbstractClostridioides difficile is the leading cause of nosocomial infection and is the causative agent of antibiotic-associated diarrhea. The severity of the disease is directly associated with the production of toxins, and spores are responsible for the transmission and persistence of the organism. Previously we characterized sin locus regulators SinR and SinR’, where SinR is the regulator of toxin production and sporulation, while the SinR’ acting as its antagonist. In Bacillus subtilis, Spo0A, the master regulator of sporulation, regulates SinR, by regulating the expression of its antagonist sinI. However, the role of Spo0A in the expression of sinR and sinR’ in C. difficile is not yet reported. In this study, we tested spo0A mutants in three different C. difficile strains R20291, UK1, and JIR8094, to understand the role of Spo0A in sin locus expression. Western blot analysis revealed that spo0A mutants had increased SinR levels. The qRT-PCR analysis for its expression further supported this data. By carrying out genetic and biochemical assays, we have shown that Spo0A can bind to the upstream region of this locus to regulates its expression. This study provides vital information that Spo0A regulates sin locus, which controls critical pathogenic traits such as sporulation, toxin production, and motility in C. difficile.IMPORTANCEClostridioides difficile is the leading cause of antibiotic-associated diarrheal disease in the United States. During infection, C. difficile spores germinate, and the vegetative bacterial cells produce toxins that damage host tissue. In C. difficile, sin locus is known to regulate both sporulation and toxin production. In this study, we have shown that Spo0A, the master regulator of sporulation to control the sin locus expression. We performed various genetic and biochemical experiments to show that Spo0A directly regulates the expression of this locus by binding to its upstream DNA region. This observation adds new detail to the gene regulatory network that connects sporulation and toxin production in this pathogen.

scholarly journals Clostridioides difficile Spores: Bile Acid Sensors and Trojan Horses of Transmission

2020 ◽  
Vol 33 (02) ◽  
pp. 058-066
Author(s):  
Aimee Shen
Keyword(s):  
Disease Transmission ◽  
Disease Recurrence ◽  
The United States ◽  
Life Forms ◽  
Obligate Anaerobe ◽  
Infectious Particle ◽  
Clostridioides Difficile ◽  
Healthcare Associated ◽  
High Treatment ◽  
Annual Treatment Cost

AbstractThe Gram-positive, spore-forming bacterium, Clostridioides difficile is the leading cause of healthcare-associated infections in the United States, although it also causes a significant number of community-acquired infections. C. difficile infections, which range in severity from mild diarrhea to toxic megacolon, cost more to treat than matched infections, with an annual treatment cost of approximately $6 billion for almost half-a-million infections. These high–treatment costs are due to the high rates of C. difficile disease recurrence (>20%) and necessity for special disinfection measures. These complications arise in part because C. difficile makes metabolically dormant spores, which are the major infectious particle of this obligate anaerobe. These seemingly inanimate life forms are inert to antibiotics, resistant to commonly used disinfectants, readily disseminated, and capable of surviving in the environment for a long period of time. However, upon sensing specific bile salts in the vertebrate gut, C. difficile spores transform back into the vegetative cells that are responsible for causing disease. This review discusses how spores are ideal vectors for disease transmission and how antibiotics modulate this process. We also describe the resistance properties of spores and how they create challenges eradicating spores, as well as promote their spread. Lastly, environmental reservoirs of C. difficile spores and strategies for destroying them particularly in health care environments will be discussed.

scholarly journals Effectiveness of Twenty Germicides Against Five Strains of C. difficile spores, With and Without Calf Serum, at Several Exposure Times

2020 ◽  
Vol 41 (S1) ◽  
pp. s204-s206
Author(s):  
William Rutala ◽  
Maria Gergen ◽  
Dale Gerding ◽  
David Jay Weber
Keyword(s):  
Fetal Calf Serum ◽  
Calf Serum ◽  
The United States ◽  
Log10 Reduction ◽  
Clostridioides Difficile ◽  
Transmission Mechanisms ◽  
Environmental Surfaces ◽  
High Concentrations ◽  
High Level ◽  
Healthcare Associated

Background:Clostridioides difficile is a major cause of antibiotic-associated colitis and the most common healthcare-associated pathogen in the United States. Interrupting the known transmission mechanisms of C. difficile in hospitals requires appropriate hand hygiene, disinfection of potentially contaminated surfaces, and patient equipment. However, only limited data are available on the effectiveness of germicides against various strains of C. difficile, with and without fetal calf serum, and at multiple exposure times. For this reason, we undertook the following evaluation to determine the effectiveness of germicides. Methods: The effectiveness of the sporicidal activity of the germicides against 5 strains of C. difficile was evaluated using a quantitative carrier test, a standard of ASTM International developed by Sattar et al. In this protocol, metal carriers (1 cm diameter 0.7 mm thick) were inoculated with 10 L spore suspension, containing ~103 or 106 C. difficile spores, and we then exposed them to 50 L germicide for 1, 5, 10, or 20 minutes. The following C. difficile strains were used in these studies: ATCC strains 9689; J9; BI-9; 630; and CF-4. To determine whether C. difficile spore susceptibility was similar to other spores, we also tested Bacillus atrophaeus spores, ATCC strain 19659. Fetal calf serum (FCS) was used to simulate organic matter. Results: In general, high-level disinfectants (eg, OPA, glutaraldehyde), chemical sterilants (eg, peracetic acid), and high concentrations of chlorine (>5,000 ppm) were generally sporicidal (>3 log10 reduction) in 5–10 minutes (and sometimes 1 minute). This level of sporicidal activity was demonstrated for the various strains of C. difficile spores and B. atrophaeus spores (Table 1). There did not appear to be any significant differences in inactivation of C. difficile spores (BI-9 strain) in the presence or absence of FCS (Table 2). Discussion: The sporicidal activity of disinfectants is critical because such formulations are routinely used to eliminate the risk associated with noncritical and semicritical instruments and environmental surfaces. Our data suggest that immersion in most (but not all) high-level disinfectants for 10 minutes is likely to be successful in eradicating C. difficile spores (>4 log10 reduction) from semicritical equipment (eg, endoscopes). Additionally, high concentrations of chlorine and some high-level disinfectants will kill C. difficile spores in 1 or 2 minutes.Funding: NoneDisclosures: Drs. Rutala and Weber are consultants to PDI (Professional Disposable International)

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