BACKGROUND: Only a few studies dealt with the occurrence of endospore-forming clostridia in the microbiota of infants without obvious health complications. METHODS: A methodology pipeline was developed to determine the occurrence of endospore formers in infant feces. Twenty-four fecal samples (FS) were collected from one infant in monthly intervals and were subjected to variable chemical and heat treatment in combination with culture-dependent analysis. Isolates were identified by MALDI-TOF mass spectrometry, 16S rRNA gene sequencing, and characterized with biochemical assays. RESULTS: More than 800 isolates were obtained, and a total of 21 Eubacteriales taxa belonging to the Clostridiaceae, Lachnospiraceae, Oscillospiraceae, and Peptostreptococcaceae families were detected. Clostridium perfringens, C. paraputrificum, C. tertium, C. symbiosum, C. butyricum, and C. ramosum were the most frequently identified species compared to the rarely detected Enterocloster bolteae, C. baratii, and C. jeddahense. Furthermore, the methodology enabled the subsequent cultivation of less frequently detectable gut taxa such as Flavonifractor plautii, Intestinibacter bartlettii, Eisenbergiella tayi, and Eubacterium tenue. The isolates showed phenotypic variability regarding enzymatic activity, fermentation profiles, and butyrate production. CONCLUSIONS: Taken together, this approach suggests and challenges a cultivation-based pipeline that allows the investigation of the population of endospore formers in complex ecosystems such as the human gastrointestinal tract.
- MeSH
- Clostridium * genetika MeSH
- feces mikrobiologie MeSH
- Firmicutes genetika MeSH
- kojenec MeSH
- lidé MeSH
- mikrobiota * MeSH
- RNA ribozomální 16S genetika MeSH
- Check Tag
- kojenec MeSH
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
A total of, 78 Clostridium septicum (CLSE) isolates were screened for genes encoding: α-toxin, flagellin, and resistance to vancomycin (VANg). The isolates were also tested for their ability to form biofilm and their antibiotic susceptibility. All isolates were positive for α-toxin and flagellin genes. However, only 19 isolates (24.3%) showed prevalence for VANg. We observed the strongest capacity to form a biofilm (100%) in isolates from patients with oncologic or septic and febrile diagnoses. This percentage was also very high in patients with colitis and gastrointestinal hemorrhage (72.7%). No less than 43 isolates showed antibiotic resistance, and 21 were multidrug-resistant (MDR). Interestingly, our studies showed a correlation between antibiotic resistance and biofilm formation. A statistically significant difference was observed between biofilm-forming MDR isolates and those with low/no biofilm-forming ability. However, the most impressive observation was the correlation with mortality rate. While the overall mortality rate for CLSE infections was 16.7% (13/78), the mortality rate for patients infected with MDR isolates forming biofilm moderately or strongly reached 38.1% (8/21). This number increased even further when only infections with the biofilm-forming VANg-positive isolates were considered (61.5%; 8/13). Therefore, the ability of a VANg-positive CLSE isolate to form a biofilm has been suggested as a biomarker of poor prognosis.
This article describes the characterization and application of collagenase-based chitosan nanofiber membranes with rat burns. Electrospun chitosan nanofibers were functionalized with clostridial collagenase using carbodiimide chemistry. The immobilized collagenase was characterized by enzyme activity, kinetic constants, and dry storage stability measurements using a Pz-peptide substrate. The apparent kinetic constants KM and Vmax of immobilized collagenase showed a high affinity for the peptide substrate compared to the free enzyme. Drying of chitosan membranes with immobilized collagenase ensured 98 % stability of enzyme activity after rehydration. The effect of collagenase immobilized on chitosan nanofibers on the burn of the rat model was compared with a control treatment with chitosan nanofibers. The healing of the wound with both materials was terminated after 30 days at the same time, although the collagenase wound healed more rapidly during healing. The scar area size after the application of collagenase-containing chitosan nanofiber membranes was 31.6 % smaller than when only chitosan nanofibers were used.
- MeSH
- chitosan terapeutické užití MeSH
- Clostridium histolyticum MeSH
- enzymy imobilizované MeSH
- hojení ran * účinky léků MeSH
- krysa rodu rattus MeSH
- kůže zranění MeSH
- mikrobiální kolagenasa * metabolismus terapeutické užití MeSH
- nanovlákna terapeutické užití MeSH
- pilotní projekty MeSH
- rány a poranění farmakoterapie patologie MeSH
- stabilita enzymů MeSH
- výsledek terapie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
The main bottleneck in the return of industrial butanol production from renewable feedstock through acetone-butanol-ethanol (ABE) fermentation by clostridia, such as Clostridium beijerinckii, is the low final butanol concentration. The problem is caused by the high toxicity of butanol to the production cells, and therefore, understanding the mechanisms by which clostridia react to butanol shock is of key importance. Detailed analyses of transcriptome data that were obtained after butanol shock and their comparison with data from standard ABE fermentation have resulted in new findings, while confirmed expected population responses. Although butanol shock resulted in upregulation of heat shock protein genes, their regulation is different than was assumed based on standard ABE fermentation transcriptome data. While glucose uptake, glycolysis, and acidogenesis genes were downregulated after butanol shock, solventogenesis genes were upregulated. Cyclopropanation of fatty acids and formation of plasmalogens seem to be significant processes involved in cell membrane stabilization in the presence of butanol. Surprisingly, one of the three identified Agr quorum-sensing system genes was upregulated. Upregulation of several putative butanol efflux pumps was described after butanol addition and a large putative polyketide gene cluster was found, the transcription of which seemed to depend on the concentration of butanol.
- MeSH
- biologický transport genetika MeSH
- bioreaktory mikrobiologie MeSH
- buněčná membrána metabolismus MeSH
- butanoly toxicita MeSH
- Clostridium beijerinckii účinky léků genetika metabolismus MeSH
- fyziologický stres genetika MeSH
- glukosa metabolismus MeSH
- glykolýza genetika fyziologie MeSH
- mastné kyseliny metabolismus MeSH
- plasmalogeny biosyntéza MeSH
- proteiny tepelného šoku metabolismus MeSH
- quorum sensing genetika MeSH
- stanovení celkové genové exprese MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Clostridium diolis DSM 15410 is a type strain of solventogenic clostridium capable of conducting isopropanol-butanol-ethanol fermentation. By studying its growth on different carbohydrates, we verified its ability to utilize glycerol and produce 1,3-propanediol and discovered its ability to produced isopropanol. Complete genome sequencing showed that its genome is a single circular chromosome and belongs to the cluster I (sensu scricto) of the genus Clostridium. By cultivation analysis we highlighted its specific behavior in comparison to two selected closely related strains. Despite the fact that several CRISPR loci were found, 16 putative prophages showed the ability to receive foreign DNA. Thus, the strain has the necessary features for future engineering of its 1,3-propanediol biosynthetic pathway and for the possible industrial utilization in the production of biofuels.
A fatal case of 67-year-old female with metastatic breast cancer on chemotherapy complicated with febrile neutropenia, colitis and sepsis due to Clostridium septicum is presented. Important clinical symptoms, laboratory and radiology findings together with therapy and outcome of neuropenic colitis are also discussed.
- MeSH
- bakteriemie * diagnóza MeSH
- Clostridium septicum * MeSH
- klostridiové infekce * komplikace diagnóza MeSH
- kolitida * diagnóza etiologie MeSH
- lidé MeSH
- nádory prsu * komplikace MeSH
- senioři MeSH
- Check Tag
- lidé MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
Clostridial collagenases are essential biotechnological tissue dissociation agents owing to their ability to cleave different types of collagen. Standardization of collagenase-based protocols has been hampered by impurities in products manufactured from Clostridium histolyticum. To enhance the purification process, we produced recombinant collagenase classes G and H, taking advantage of the Escherichia coli expression system. The respective gene sequences were derived from C. histolyticum and modified by addition of a C-terminal polyhistidine tag. Harvested bacteria were lysed and the collagenase protein was affinity purified using a His-tag column. The purity, identity, integrity of the eluted collagenases G and H were determined by SDS electrophoresis and Western blot. The proteolytic activity of the collagenase G and H blend (rColGH) was determined by the standard FALGPA assay. The tissue dissociation activity was verified using a standardized method for isolation of rat pancreatic islets. Biocompatibility of the blend was validated by a standardized viability assay on the isolated islets. Two batches of rColGH were produced and compared to a commercially available collagenase. Based on our results, we conclude that rColGH is a functional and non-toxic novel recombinant collagenase worth further characterization and blend optimization in order to make it a competitive commercial product.
- MeSH
- Clostridium MeSH
- kolagenasy * MeSH
- krysa rodu rattus MeSH
- Langerhansovy ostrůvky * MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Commensal bacterium Clostridium paraputrificum J4 produces several extracellular chitinolytic enzymes including a 62 kDa chitinase Chit62J4 active toward 4-nitrophenyl N,N'-diacetyl-β-d-chitobioside (pNGG). We characterized the crude enzyme from bacterial culture fluid, recombinant enzyme rChit62J4, and its catalytic domain rChit62J4cat. This major chitinase, securing nutrition of the bacterium in the human intestinal tract when supplied with chitin, has a pH optimum of 5.5 and processes pNGG with Km = 0.24 mM and kcat = 30.0 s-1. Sequence comparison of the amino acid sequence of Chit62J4, determined during bacterial genome sequencing, characterizes the enzyme as a family 18 glycosyl hydrolase with a four-domain structure. The catalytic domain has the typical TIM barrel structure and the accessory domains-2x Fn3/Big3 and a carbohydrate binding module-that likely supports enzyme activity on chitin fibers. The catalytic domain is highly homologous to a single-domain chitinase of Bacillus cereus NCTU2. However, the catalytic profiles significantly differ between the two enzymes despite almost identical catalytic sites. The shift of pI and pH optimum of the commensal enzyme toward acidic values compared to the soil bacterium is the likely environmental adaptation that provides C. paraputrificum J4 a competitive advantage over other commensal bacteria.
- MeSH
- bakteriální proteiny genetika metabolismus MeSH
- chitin metabolismus MeSH
- chitinasy chemie genetika metabolismus MeSH
- Clostridium růst a vývoj izolace a purifikace metabolismus MeSH
- katalytická doména MeSH
- koncentrace vodíkových iontů MeSH
- lidé MeSH
- rekombinantní proteiny genetika metabolismus MeSH
- střevní mikroflóra MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Clostridium perfringens forms biofilms and spores that are a source of food contamination. In this study, the antibacterial activities of Lactobacillus plantarum culture supernatants (LP-S), LP-S fractions, and the plant-derived compound epigallocatechin gallate (EG) were evaluated. Specifically, their effects on the viability and biofilm-forming ability of C. perfringens were assessed. Moreover, the expression of quorum sensing-regulated genes associated with the pathogenesis of this microorganism and that of genes involved in biofilm formation was also investigated. The results showed that both EG and the LP-S exerted bactericidal activity against all C. perfringens strains tested. The minimal bactericidal concentration (MBC) of EG was 75 µg/mL for all strains but ranged from 61 to 121 µg of total protein per mL for LP-S. EG exerted only minor effects on biofilm formation, whereas LP-S, particularly its 10 and 30 K fractions, significantly reduced the biofilm-forming ability of all the strains. The antibiofilm activity of LP-S was lost following preincubation with proteases, suggesting that it was mediated by a proteinaceous molecule. The treatment of C. perfringens with either EG or LP-S did not change the transcript levels of two CpAL (C. perfringens quorum-sensing Agr-like system)-related genes, agrB and agrD, which are known to be involved in the regulation of biofilms, suggesting that LP-S exerted its biofilm inhibitory activity downstream of CpAL signaling. In summary, we demonstrated the bactericidal activity of EG and LP-S against C. perfringens and antibiofilm activity of LP-S at a subinhibitory dose. Our results suggested that these compounds can be further explored for food safety applications to control agents such as C. perfringens.
Clostridium ventriculi (syn. Sarcina ventriculi) is a Gram-positive opportunistic pathogen with sarcina morphology. In the case of gastrointestinal disorders, the treatment is often empirical. Due to the common occurrence in primates and the potential risk of dysbiosis; the antibiotic susceptibility screening of C. ventriculi strains isolated from guenon monkeys and crested gibbons to 58 antibiotics was performed to reduce potentially ineffective antibiotic use in case of disease. Isolates were found to be susceptible to the majority of the tested antibiotics, mainly to (fluoro)quinolones, macrolides, penicillins, and tetracyclines. The susceptibility profiles were similar despite the hosts. Tested strains showed also natural resistance to a few antibiotics on the genus level. Detected in vitro antibiotic efficiency is consistent with documented human treatment cases.
- MeSH
- antibakteriální látky terapeutické užití MeSH
- Clostridium účinky léků genetika MeSH
- genetická variace MeSH
- genotyp MeSH
- Haplorrhini mikrobiologie MeSH
- Hylobates mikrobiologie MeSH
- klostridiové infekce diagnóza farmakoterapie MeSH
- primáti mikrobiologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
