Mucosal Vaccination Strategies against Clostridioides difficile Infection.

Clostridioides difficile infection (CDI) presents a major public health threat by causing frequently recurrent, life-threatening cases of diarrhea and intestinal inflammation. The ability of C. difficile to express antibiotic resistance and to form long-lasting spores makes the pathogen particularly challenging to eradicate from healthcare settings, raising the need for preventative measures to curb the spread of CDI. Since C. difficile utilizes the fecal-oral route of transmission, a mucosal vaccine could be a particularly promising strategy by generating strong IgA and IgG responses that prevent colonization and disease. This mini-review summarizes the progress toward mucosal vaccines against C. difficile toxins, cell-surface components, and spore proteins. By assessing the strengths and weaknesses of particular antigens, as well as methods for delivering these antigens to mucosal sites, we hope to guide future research toward an effective mucosal vaccine against CDI.

Draft Genome Sequences and Genome Characterization of Three Toxigenic and Two Nontoxigenic Clostridioides difficile Clinical Isolates from Florida, USA.

Draft genome sequences of five Clostridioides difficile clinical isolates were obtained in Florida, USA. Three isolates, designated TGH29 (sequence type 1 [ST1]/clade 2), TGH79 (ST11/clade 5), and TGH91 (ST35/clade 1), contained toxin-encoding genes. The two nontoxigenic strains were classified as TGH114 (ST109/clade 4) and TGH132 (ST15/clade 1). Antimicrobial resistance determinants and plasmids were detected and putative prophages predicted in some isolates.

Recombinant Fusion Protein Vaccine Containing Clostridioides difficile FliC and FliD Protects Mice against C. difficile Infection.

Bacterial flagella are involved in infection through their roles in host cell adhesion, cell invasion, auto-agglutination, colonization, the formation of biofilms, and the regulation and secretion of nonflagellar bacterial proteins that are involved in the virulence process. In this study, we constructed a fusion protein vaccine (FliCD) containing the Clostridioides difficile flagellar proteins FliC and FliD. The immunization of mice with FliCD induced potent IgG and IgA antibody responses against FliCD, protected mice against C. difficile infection (CDI), and decreased the C. difficile spore and toxin levels in the feces after infection. Additionally, the anti-FliCD serum inhibited the binding of C. difficile vegetative cells to HCT8 cells. These results suggest that FliCD may represent an effective vaccine candidate against CDI.

Genomic and Phenotypic Characterization of the Nontoxigenic Clostridioides difficile Strain CCUG37785 and Demonstration of Its Therapeutic Potential for the Prevention of C. difficile Infection.

Symptoms of Clostridioides difficile infection (CDI) are attributed largely to two toxins, TcdA and TcdB. About 17-23% of C. difficile isolates produce binary toxin, which enhances C. difficile pathogenesis. Previously, we engineered the nontoxigenic C. difficile strain CCUG37785 (designated as CCUG37785) to express immunogenic fragments of TcdA and TcdB as an oral mucosal CDI vaccine candidate. In this study, we performed genomic and phenotypic analyses of CCUG37785 and evaluated its potential use for preventing and treating CDI. Whole genome sequencing showed that CCUG37785 is ribotype ST3 and lacks toxin genes. Comparative analyses of PaLoc and CdtLoc loci of CCUG37785 revealed 115-bp and 68-bp conserved fragments in these regions, respectively. Phenotypic comparisons between CCUG37785 and C. difficile R20291 (an epidemic hypervirulent BI/NAPI/027 strain, designated as R20291) found that CCUG37785 exhibited significantly higher adhesion and sporulation, significantly lower spore germination and biofilm formation, and comparable motility to R20291. We also showed that oral inoculation of CCUG37785 spores prior to infection with R20291 spores provided mice almost full protection against developing CDI. However, oral inoculation of CCUG37785 spores after infection with R20291 spores only provided minor protection against CDI. Further analysis showed that mice pretreated with CCUG37785 spores secreted significantly less R20291 spores, while mice treated with CCUG37785 spores after infection with R20291 secreted a comparable amount of R20291 spores to mice infected with R20291 spores only. Our data both highlight the potential use of CCUG37785 for the prevention of primary and recurrent CDI in humans and support its use as an oral mucosal vaccine carrier against CDI. IMPORTANCE Clostridioides difficile infection (CDI) symptoms range from diarrhea to intestinal inflammation/lesion and death and are mainly caused by two exotoxins, TcdA and TcdB. Active vaccination provides the attractive opportunity to prevent CDI and recurrence. No vaccine against CDI is currently licensed. Tremendous efforts have been devoted to developing vaccines targeting both toxins. However, ideally, vaccines should target both toxins and C. difficile cells/spores that transmit the disease and cause recurrence. Furthermore, C. difficile is an enteric pathogen, and mucosal/oral immunization would be particularly useful to protect the host against CDI considering that the gut is the main site of disease onset and progression. Data in our current study not only highlight the potential use of CCUG37785 to prevent primary and recurrent CDI in humans but also further support its use as an oral mucosal vaccine carrier against CDI.

Clostridioides difficile phage biology and application.

Clostridium difficile, now reclassified as Clostridioides difficile, is the causative agent of C. difficile infections (CDI). CDI is particularly challenging in healthcare settings because highly resistant spores of the bacterium can persist in the environment, making it difficult to curb outbreaks. Dysbiosis of the microbiota caused by the use of antibiotics is the primary factor that allows C. difficile to colonize the gut and cause diarrhea and colitis. For this reason, antibiotics targeting C. difficile can be ineffective at preventing recurrent episodes because they exacerbate and prolong dysbiosis. The emergence of antibiotic resistance in C. difficile also presents a significant threat. The diverse array of bacteriophages (phages) that infect C. difficile could offer new treatment strategies and greater insight into the biology of the pathogen. In this review, we summarize the current knowledge regarding C. difficile phages and discuss what is understood about their lifestyles and genomics. Then, we examine how phage infection modifies bacterial gene expression and pathogenicity. Finally, we discuss the potential clinical applications of C. difficile phages such as whole phage therapy and phage-derived products, and we highlight the most promising strategies for further development.

Point-of-care detection, characterization, and removal of chocolate bloom using a handheld Raman spectrometer.

Chocolate bloom is an off-white coating on the surface of chocolate products due to the altered distribution of the ingredients. Bloom reduces the shelf-life of chocolate and affects its visual and tactile quality, all of which are serious concerns for chocolate manufacturers and consumers. The automated, rapid, and noninvasive point-of-care detection of chocolate bloom has been an essential but challenging problem. The ability to detect and characterize chocolate bloom using portable laser spectroscopy could be used to develop in-situ quality control sensors. In this work, a handheld Raman spectrometer was used to detect chocolate bloom. Raman spectra acquired from bloomed HERSHEY'S milk chocolate, Hawaiian Host milk chocolate covered macadamia nuts, and Babayevsky Russian dark chocolate were used to characterize the type of bloom. The 1064 nm laser beam of the handheld Raman instrument was used to partially remove the fat bloom of the dark chocolate and to induce sugar bloom on the milk chocolate. The handheld Raman approach has a high potential for industrial and consumer applications for the on-site chemical analysis of chocolate bloom and as an alternative laser-based chocolate decoration.