Emergence Of Hypervirulent Strains Research Articles

Characterization of community-acquired Clostridioides difficile strains in Israel, 2020-2022.

The prevalence of community-acquired Clostridioides difficile infection (CA-CDI) has been rising, due to changes in antibiotics prescribing practices, emergence of hypervirulent strains and improved diagnostics. This study explored CA-CDI epidemiology by examining strain diversity and virulence factors of CA-CDI isolates collected across several geographical regions in Israel. Stool samples of 126 CA-CDI patients were subjected to PCR and an immunoassay to identify toxin genes and proteins, respectively. Toxin loci PaLoc and PaCdt were detected by whole-genome sequencing (WGS). Biofilm production was assessed by crystal violet-based assay. Minimum inhibitory concentration was determined using the Etest technique or agar dilution. WGS and multi-locus sequence typing (MLST) were used to classify strains and investigate genetic diversity. Sequence types (ST) 2 (17, 13.5%), ST42 (13, 10.3%), ST104 (10, 8%) and ST11 (9, 7.1%) were the most common. All (117, 92.8%) but ST11 belonged to Clade 1. No associations were found between ST and gender, geographic area or antibiotic susceptibility. Although all strains harbored toxins genes, 34 (27%) produced toxin A only, and 54 (42.9%) strains produced toxin B only; 38 (30.2%) produced both toxins. Most isolates were biofilm-producers (118, 93.6%), primarily weak producers (83/118, 70.3%). ST was significantly associated with both biofilm and toxin production. C. difficile isolates in Israel community exhibit high ST diversity, with no dominant strain. Other factors may influence the clinical outcomes of CDI such as toxin production, antibiotic resistance and biofilm production. Further studies are needed to better understand the dynamics and influence of these factors on CA-CDI.

Klebsiella pneumoniae survives on surfaces as a dry biofilm

BackgroundDry surface biofilms (DSB) are widespread in healthcare settings presenting a challenge to cleaning and disinfection. Klebsiella pneumoniae has been a focus of attention due to antibiotic resistance and the emergence of hypervirulent strains. Few studies have demonstrated K pneumoniae survival on surfaces following desiccation. MethodsDSB were formed over 12 days. Bacterial culturability and transfer were investigated following DSB incubation up to 4 weeks. Bacterial viability in DSB was investigated with live/dead staining using flow cytometry. ResultsK pneumoniae formed mature DSB. After 2 and 4 weeks of incubation, transfer from DSB was low (<55%) and reduced further (<21%) following wiping. Culturability at 2 and 4 weeks varied although viability remained high indicating viable but non culturable state (VBNC). DiscussionK pneumoniae was removed from surfaces by mechanical wiping as shown with DSB of other species. Although culturability was reduced over time, bacteria remained viable up to 4 weeks incubation, proving the need for robust cleaning regimens. ConclusionsThis is the first study confirming K pneumoniae survival on dry surfaces as a DSB. The presence of VBNC bacteria indicated that K pneumoniae can for extended periods, raising questions about its persistence on surfaces.

Effect of ZnO Nanoparticles as Antimicrobial on Multidrug Resistance Klebsiella Pneumonia: A Review

Klebsiella pneumoniae (K. pneumoniae)is a common microorganism for different infections, including respiratory, urinary tract, and biliary diseases, where immunocompromised individuals are directly affected by this bacterium. However, with the emergence of hypervirulent strains, individuals,whether healthy or immunocompetent, are equally susceptible to K. pneumoniae infections. This problem is diagnosed by the wide spreading of multidrug-resistant bacteria in the environment with multiple resistance mechanisms, which are the main challenges for an effective treatment. This put physicians in an inflexible confusion because of the limitation of medical treatment options. Among many remedy substances, zinc oxide (ZnO) nanoparticles (NPs) showed remarkable antibacterial properties versus many Gram-positive and Gram-negative bacteria. This work focuses on presenting the influence of these NPs on the expression of genes in charged in generating the K. pneumoniae capsule. Furthermore, perspectives for applying ZnO NPs in clinical practices are also discussed. As the in vivo studies show a powerful impact of ZnO on bacteria, it is anticipated that this method of treatment will be utilized by hospitals.

P26 Phenotypic and genotypic characterization of antimicrobial resistance in Clostridioides difficile in North Wales

Background Clostridioides difficile poses a major infection control challenge and remains a huge burden to our healthcare system. The emergence of hypervirulent strains of C. difficile has highlighted the need for monitoring antimicrobial resistance (AMR) and continual epidemiological vigilance. Agar dilution is the current gold standard method for antimicrobial susceptibility testing of C. difficile and the method routinely utilized in the Public Health Wales (PHW) UK Anaerobe Reference Unit (UKARU) at the University Hospital of Wales, Cardiff. However, this method is laborious and time consuming. Meanwhile, the emergence of WGS introduces the possibility of acquiring rapid genotypic information in terms of AMR.MethodsIn this research project, isolates obtained from faecal samples submitted across the Betsi Cadwalader health board (BCUHB) that were either GDH or PCR positive were sequenced as part of the C. difficile Genomic Sequencing and Typing (DIGEST) pilot project performed at the PHW Pathogen Genomics Unit (PenGU) in Cardiff.ResultsResults highlights include the finding that 93% of the isolates analysed for the presence of AMR genes were found to have the cdeA gene associated with conferring resistance to fluoroquinolones such as ciprofloxacin. Other common genes observed included blaCDD-1 and blaCDD-2, associated with β-lactam resistance.ConclusionsWhile further work must be done to link genotypic characterization with phenotype, this study provides a valuable insight into AMR mechanisms in C. difficile and may in future become an important tool providing early indication of shift in C. difficile epidemiology or the emergence of a new hypervirulent and MDR strain.

Clostridioides (Clostridium) difficile isolated from paediatric patients in Western Australia 2019–2020

Less is understood about the epidemiology of Clostridioides difficile infection (CDI) in children compared to adults, and its impact is complicated by variations in the natural development of infection in paediatric patients. The interplay of rising CDI incidence in hospitalised paediatric patients, emergence of hypervirulent strains and community associated CDI (CA-CDI) in the past decade is a potential threat in both hospital and community settings. Research in Australia regarding paediatric CDI is limited. Here, we report the molecular characterisation of C.difficile isolated from paediatric patients at a tertiary hospital in Perth, Western Australia. A total of 427 stool samples was collected from patients aged from <1 to 17 years being investigated for diarrhoea from July 2019 to June 2020. Stool specimens were cultured and isolates of C.difficile characterised by ribotyping and toxin gene profiling. Clostridioides difficile was recovered from 84/427 (19.7%) samples tested. The most prevalent PCR ribotypes (RTs) were RT 002 (12.4%), a toxigenic strain, and RT 009 (15.7%), a non-toxigenic strain. Interestingly, C.difficile RT 078 and RT 017, strains that are not endemic in Australia, were isolated from a 1- and 4-year-old child, respectively. Clostridioides difficile RT 106, a strain of emerging importance in Australia, was recovered from two cases (5.3%). Resistance to metronidazole, fidaxomicin, amoxicillin, rifaximin and meropenem was not detected, however, 45 isolates (50.6%) showed resistance to at least one agent, and multidrug resistance was observed in 13.3% of the resistant isolates (6/45). This study provides a baseline for future surveillance of paediatric CDI in Australia. Given that young children can be asymptomatically colonised with toxigenic C.difficile strains, they represent a potential reservoir of strains causing CDI in adults.

Clostridium Difficile Infection in Inflammatory Bowel Disease Patients.

The rising incidence of Clostridium difficile infection (CDI) in the general population has been recognized by health care organizations worldwide. The emergence of hypervirulent strains has made CDI more challenging to understand and treat. Inflammatory bowel disease (IBD) patients are at higher risk of infection, including CDI. A diagnostic approach for recurrent CDI has yet to be validated, particularly for IBD patients. Enzyme immunoassay (EIA) for toxins A and B, as well as glutamate dehydrogenase EIA, are both rapid testing options for the identification of CDI. Without a high index of suspicion, it is challenging to initially differentiate CDI from an IBD flare based on clinical evaluation alone. Here, we provide an up-to-date review on CDI in IBD patients. When caring for an IBD patient with suspected CDI, it is appropriate to empirically treat the presumed infection while awaiting further test results. Treatment with vancomycin or fidaxomicin, but not oral metronidazole, has been advocated by an expert review from the clinical practice update committee of the American Gastroenterology Association. Recurrent CDI is more common in IBD patients compared to non-IBD patients (32% versus 24%), thus more aggressive treatment is recommended for IBD patients along with early consideration of fecal microbiota transplant. Although the use of infliximab during CDI has been debated, clinical experience exists supporting its use in an IBD flare, even with active CDI when needed.

Epitopes identified in GAPDH from Clostridium difficile recognized as common antigens with potential autoimmunizing properties

Clostridium difficile (CD) infections are a growing threat due to the strain resistance to antibiotic treatment and the emergence of hypervirulent strains. One solution to this problem is the search for new vaccine antigens, preferably surface-localized that will be recognized by antibodies at an early stage of colonization. The purpose of the study was to assess the usefulness of novel immunoreactive surface proteins (epitopes) as potential vaccine antigens. Such approach might be tough to pursue since pathogens have acquired strategies to subvert adaptive immune response to produce humoral response against non-essential proteins for their survival. In this study CD surface proteins were isolated, immunoreactive proteins identified and mapped to select potential epitopes. The results of the study exclude the use of CD glyceraldehyde 3-phosphate dehydrogenase as a vaccine antigen, especially as a whole protein. Sequences P9 (201AAGNIVPNTTGAAKAI218) and P10 (224KGKLDGAAQRVPVVTG241) recognized by patients sera are conserved and widespread among CD strains. They show cross-reactivity with sera of people suffering from other bacterial infections and are recognized by sera of autoimmune disease patients. Our study documents that special care in analyzing the sequence of new epitope should be taken to avoid side effects prior to consider it as a vaccine antigen.

Population genomic footprints of host adaptation, introgression and recombination in coffee leaf rust.

Coffee leaf rust, caused by Hemileia vastatrix (Hv), represents the biggest threat to coffee production worldwide and ranks amongst the most serious fungal diseases in history. Despite a recent series of outbreaks and emergence of hypervirulent strains, the population evolutionary history and potential of this pathogen remain poorly understood. To address this issue, we used restriction site-associated DNA sequencing (RADseq) to generate ∼19000 single nucleotide polymorphisms (SNPs) across a worldwide collection of 37 Hv samples. Contrary to the long-standing idea that Hv represents a genetically unstructured and cosmopolitan species, our results reveal the existence of a cryptic species complex with marked host tropism. Using phylogenetic and pathological data, we show that one of these lineages (C3) infects almost exclusively the most economically valuable coffee species (tetraploids that include Coffea arabica and interspecific hybrids), whereas the other lineages (C1 and C2) are severely maladapted to these hosts, but successfully infect diploid coffee species. Population dynamic analyses suggest that the C3 group may be a recent 'domesticated' lineage that emerged via host shift from diploid coffee hosts. We also found evidence of recombination occurring within this group, which could explain the high pace of pathotype emergence despite the low genetic variation. Moreover, genomic footprints of introgression between the C3 and C2 groups were discovered and raise the possibility that virulence factors may be quickly exchanged between groups with different pathogenic abilities. This work advances our understanding of the evolutionary strategies used by plant pathogens in agro-ecosystems with direct and far-reaching implications for disease control.

Clostridium difficile flagella induce a pro-inflammatory response in intestinal epithelium of mice in cooperation with toxins

Clostridium difficile is the most important enteropathogen involved in gut nosocomial post-antibiotic infections. The emergence of hypervirulent strains has contributed to increased mortality and morbidity of CDI. The C. difficile toxins contribute directly to CDI-associated lesions of the gut, but other bacterial factors are needed for the bacteria to adhere and colonize the intestinal epithelium. The C. difficile flagella, which confer motility and chemotaxis for successful intestinal colonization, could play an additional role in bacterial pathogenesis by contributing to the inflammatory response of the host and mucosal injury. Indeed, by activating the TLR5, flagella can elicit activation of the MAPK and NF-κB cascades of cell signaling, leading to the secretion of pro-inflammatory cytokines. In the current study, we demonstrate, by using an animal model of CDI, a synergic effect of flagella and toxins in eliciting an inflammatory mucosal response. In this model, the absence of flagella dramatically decreases the degree of mucosal inflammation in mice and the sole presence of toxins without flagella was not enough to elicit epithelial lesions. These results highlight the important role of C. difficile flagella in eliciting mucosal lesions as long as the toxins exert their action on the epithelium.

Development of a Novel Vaccine Containing Binary Toxin for the Prevention of Clostridium difficile Disease with Enhanced Efficacy against NAP1 Strains.

Clostridium difficile infections (CDI) are a leading cause of nosocomial diarrhea in the developed world. The main virulence factors of the bacterium are the large clostridial toxins (LCTs), TcdA and TcdB, which are largely responsible for the symptoms of the disease. Recent outbreaks of CDI have been associated with the emergence of hypervirulent strains, such as NAP1/BI/027, many strains of which also produce a third toxin, binary toxin (CDTa and CDTb). These hypervirulent strains have been associated with increased morbidity and higher mortality. Here we present pre-clinical data describing a novel tetravalent vaccine composed of attenuated forms of TcdA, TcdB and binary toxin components CDTa and CDTb. We demonstrate, using the Syrian golden hamster model of CDI, that the inclusion of binary toxin components CDTa and CDTb significantly improves the efficacy of the vaccine against challenge with NAP1 strains in comparison to vaccines containing only TcdA and TcdB antigens, while providing comparable efficacy against challenge with the prototypic, non-epidemic strain VPI10463. This combination vaccine elicits high neutralizing antibody titers against TcdA, TcdB and binary toxin in both hamsters and rhesus macaques. Finally we present data that binary toxin alone can act as a virulence factor in animal models. Taken together, these data strongly support the inclusion of binary toxin in a vaccine against CDI to provide enhanced protection from epidemic strains of C. difficile.

Fidaxomicin--the new drug for Clostridium difficile infection.

Clostridium difficile is one of the many aetiological agents of antibiotic associated diarrhoea and is implicated in 15-25 per cent of the cases. The organism is also involved in the exacearbation of inflammatory bowel disease and extracolonic manifestations. Due to increase in the incidence of C. difficile infection (CDI), emergence of hypervirulent strains, and increased frequency of recurrence, the clinical management of the disease has become important. The management of CDI is based on disease severity, and current antibiotic treatment options are limited to vancomycin or metronidazole in the developing countries. this review article briefly describes important aspects of CDI, and the new drug, fidaxomicin, for its treatment. Fidaxomicin is particularly active against C. difficile and acts by inhibition of RNA synthesis. Clinical trials done to compare the efficacy and safety of fidaxomicin with that of vancomycin in treating CDI concluded that fidaxomicin was non-inferior to vancomycin for treatment of CDI and that there was a significant reduction in recurrences. The bactericidal properties of fidaxomicin make it an ideal alternative for CDI treatment. However, fidaxomicin use should be considered taking into account the potential benefits of the drug, along with the medical requirements of the patient, the risks of treatment and the high cost of fidaxomicin compared to other treatment regimens.

Characterization of a multidrug resistant C. difficile meat isolate

Clostridium difficile is a pathogen of significant public health concern causing a life-threatening, toxin-mediated enteric disease in humans. The incidence and severity of the disease associated with C. difficile have increased in the US with the emergence of hypervirulent strains and community associated outbreaks. The detection of genotypically similar and identical C. difficile strains implicated from human infections in foods and food animals indicates the potential role of food as a source of community associated C. difficile disease. One hundred samples each of ground beef, pork and chicken obtained from geographically distant grocery stores in Connecticut were tested for C. difficile. Positive isolates were characterized by ribotyping, antibiotic susceptibility, toxin production and whole genome sequencing. Of the 300 meat samples, only two pork samples tested positive for C. difficile indicating a very low prevalence of C. difficile in meat. The isolates were non toxigenic; however, genome characterization revealed the presence of several antibiotic resistance genes and mobile elements that can potentially contribute to generation of multidrug resistant toxigenic C. difficile by horizontal gene transfer. Further studies are warranted to investigate potential food-borne transmission of the meat isolates and development of multi-drug resistance in these strains.

Fidaxomicin: A new fight against clostridium difficile-associated diarrhea

Clostridium difficile-associated diarrhea (CDAD) occurs commonly as a side effect of broad-spectrum antibiotics. Drugs approved for CDAD are only two of them, metronidazole and vancomycin. Most promising drug for treatment of CDAD is vancomycin, but due to emergence of hypervirulent strains, outcomes have become poor. Fidaxomicin has being approved lately for treatment of CDAD, which has shown good results as compared to vancomycin. Fidaxomicin inhibits protein synthesis by inhibiting transcription carried out by sigma subunit of RNA polymerase. Fidaxomicin has been seen to be a well tolerated, with minimal adverse effects. Clinical trials has showed that fidaxomicin has similar results to vancomycin in showing clinical response, whereas it was superior to vancomycin in having sustained clinical effect, in CDAD.

SP14-2 Clinical implications and microbiological characteristics of bacteremia caused by Streptococcus bovis group

to invasive disease and rheumatic heart disease and mainly amongst the young and elderly populations. Population-based studies have described the epidemiology and clinical features of GAS diseases ranging from classic sore throat to severe life-threatening infections, such as septicaemia, streptococcal toxic shock syndrome (STSS) and necrotising fasciitis. GAS epidemiology varies with time and is dependent upon geographic location and socio-economic conditions. Timely monitoring of incidence, mortality and microbiological characteristics is essential to identify changes in disease patterns and emergence of hypervirulent strains, providingopportunities for alerts to be cascaded to frontline medical staff to facilitate early diagnosis, prompt initiation of life-saving therapy and inform the development of guidelines for control and management. The organism possesses numerous cell surface proteins that play a key role in host–bacteria relationships such as virulence and or adherence and form the basis of the GAS typing scheme. These proteins also represent choice candidates for vaccine developments. M-protein is a surface protein encoded by the emm gene which acts as a major virulence factor. Emm -typing is the molecular gold standard and there are currently more than 200 emm types described worldwide. More recently the application of novel comparative genomics has contributed towards our knowledge of the mechanisms underlying GAS pathogenesis. The information gained from current whole genome sequencing studies could translate into improved diagnostics; identification of novel molecular markers and potential predictors for the iGAS disease process and new targets for therapeutic drugs and vaccines.

Preventing downstream Clostridium difficile infections with upstream antibiotic management

Clostridium difficile infection (CDI) remains a devastating cause of hospital-acquired diarrhea. Treatment modalities have centered traditionally on two antibiotics, metronidazole and oral vancomycin. Both drugs, however, have been associated with variable relapse rates up to 20%. Fidaxomicin, a new oral agent with targeted C. difficile activity, may reduce the chance of relapse, but has not yet entered mainstream clinical practice. CDI is associated with significant morbidity and mortality. In the past decade, the emergence of hypervirulent strains has led to medical management failures and the increased need for surgical intervention.Control of the disease requires excellent infection prevention practices, yet can remain a difficult operational challenge. Selective pressure of antibiotic therapy can increase or lessen the risk depending on the agent used. We believe that antibiotic selection for the treatment of patients with any infectious disease must account for the possibility of subsequent severe CDI. We posit ‘upstream’ antibiotic selection will prevent ‘downstream’ CDI and potentially ameliorate deficiencies in infection prevention practices. Formal studies evaluating such an endpoint would be useful in this era of dangerous CDI.

Update on Clostridium difficile infection

This review summarizes the most recent epidemiological data and advances in research into the pathogenesis, diagnosis and treatment of Clostridium difficile infection (CDI). The epidemiology of CDI has changed with the emergence of hypervirulent strains. CDI rates have increased in the community, in children and in patients with inflammatory bowel disease. Although the North American pulsed-field gel electrophoresis type 1, restriction endonuclease analysis group BI, PCR ribotype 027 (NAP1/BI/027) strain remains prevalent in North America, surveillance suggests that it is decreasing in Europe. A similar strain, PCR ribotype 078, is emerging which is associated with community-associated CDI and has been isolated in animals and food products. The Society for Healthcare Epidemiology of America and the Infectious Diseases Society of America have published new guidelines on the epidemiology, diagnosis, treatment, infection control and environmental management of C. difficile. Several novel therapies for CDI are at different stages of development. There have been promising trial results with fidaxomicin, a novel antibiotic for the treatment of CDI and monoclonal antibodies against toxins A and B, which have been shown to significantly reduce CDI recurrence rates. Major advances have been made in our understanding of the spread and pathogenesis of C. difficile and new treatment options are becoming available.

Clostridium difficile infection in cancer patients and hematopoietic stem cell transplant recipients

Clostridium difficile has become the most common bacterial cause of nosocomial diarrhea. High rates of C. difficile infection (CDI) coupled with increasing morbidity and mortality attributed to CDI have sparked a renewed interest in this disease. Emergence of hypervirulent strains, rising rates of severe and recurrent infection and associated infection control challenges, and diagnostic and therapeutic dilemmas are major issues in the non-oncology population. Scant data on CDI exist in the cancer/transplant population. The purpose of this article is to describe the epidemiology, pathogenesis and management of CDI in patients receiving cancer chemotherapeutic agents, and in hematopoietic stem cell transplant recipients.