Bioisosteres at C9 of 2-Deoxy-2,3-didehydro-N-acetyl Neuraminic Acid Identify Selective Inhibitors of NEU3.

Human neuraminidases play critical roles in many physiological and pathological processes. Humans have four isoenzymes of NEU, making selective inhibitors important tools to investigate the function of individual isoenzymes. A typical scaffold for NEU inhibitors is 2-deoxy-2,3-didehydro-N-acetylneuraminic acid (DANA) where C9 modifications can be critical for potency and selectivity against human NEU. To design improved DANA analogues, we generated a library of compounds with either a short alkyl chain or a biphenyl substituent linked to the C9 position through one of six amide bioisosteres. Bioisostere linkers included triazole, urea, thiourea, carbamate, thiocarbamate, and sulfonamide groups. Within this library, we identified a C9 biphenyl carbamate derivative (963) that showed high selectivity and potency for NEU3 (Ki = 0.12 ± 0.01 µM). In contrast, NEU1 and NEU4 isoenzymes preferred amide and triazole linkers, respectively. Finally, analogues with urea, sulfonamide, and amide linkers showed enhanced inhibitory activity for a bacterial NEU, NanI from Clostridium perfringens.

The bacterial profile and antibiotic susceptibility in skin and soft tissue infections at a tertiary care hospital of Quetta, Pakistan.

Objectives: To determine the bacterial profile and antibiotic susceptibility in skin and soft tissue infections among patients in a tertiary care setting. METHODS: The cross-sectional cohort study was conducted at the Centre for Advanced Studies in Vaccinology and Biotechnology, University of Balochistan, Quetta, Pakistan, from June 2021 to May 2022, and comprised bacteriainfected skin samples that were collected from the Bolan Medical Complex Hospital, Quetta, and the Sandeman Provincial Hospital, Quetta. The swab samples were immediately cultured, and positive samples were evaluated for biochemical tests, antibiotic susceptibility test and polymerase chain reaction. Data was analysed using SPSS 22. RESULTS: Of the 800 samples, 598(74.7%) tested positive for pathogenic bacteria. Staphylococcus aureus accounted for 316(39.5%) infections, followed by clostridium perfringens 18.96(2.37%), escherichia coli 120(15.12%), pseudomonas aeruginosa 98(12.25%) and klebsiella pneumoniae 44(5.5%). Among all the infected samples, 380(47.5%) belonged to males, 218(27.25%) to patients aged 5-20 years, 448(56%) to the uneducated subjects, and 462(57.87%) to patients having lower socioeconomic status. Pseudomonas aeruginosa showed the highest level of resistance against all antibiotics. Conclusion: Regular surveillance and proper use of antibiotics should be encouraged in hospitals to limit the spread of antibiotic resistance against pathogenic bacteria.

Emergence of genetic diversity and multi-drug resistant Clostridium perfringens from wild birds.

BACKGROUND: Clostridium perfringens (C. perfringens) is an important zoonotic microorganism that can cause animal and human infections, however information about the prevalence status in wild birds of this pathogenic bacterium is currently limited. RESULT: In this study, 57 strains of C. perfringens were isolated from 328 fecal samples of wild birds. All the isolates were identified as type A and 70.18% of the isolates carried the cpb2 gene. Antimicrobial susceptibility testing showed that and 22.80% of the isolates were classified as multidrug-resistant strains. The MLST analysis of the 57 isolates from wild birds was categorized into 55 different sequence types (STs) and clustered into eight clonal complexes (CCs) with an average of 20.1 alleles and the Simpson Diversity index (Ds) of 0.9812, and revealed a high level of genetic diversity within the C. perfringens populations. Interestingly, the isolates from swan goose were clustered in the same CC while isolates from other bird species were more scattered suggesting that a potential difference in genetic diversity among the C. perfringens populations associated with different bird species. CONCLUSION: C. perfringens exhibits a wide range of host adaptations, varying degrees of antimicrobial resistance, and a high degree of genetic diversity in wild birds. Understanding the prevalence, toxin type, antimicrobial resistance, and genetic diversity of C. perfringens in wildlife populations is essential for developing effective strategies for disease control and management.

Inhibition of Clostridium perfringens and Bacillus cereus by Dry Vinegar and Cultured Sugar Vinegar During Extended Cooling of Uncured Beef and Poultry Products.

The 2021 FSIS Stabilization Guidelines for Meat and Poultry Products (Appendix B) Option 1.2 limits Phase 1 cooling from 48.8 to 26.7 °C in uncured meats to 1 h. However, this time restriction is impractical to achieve in large-diameter whole-muscle products. The objective of this study was to compare the inhibitory effect of commercial dry vinegars (DVs) and cultured sugar-vinegar blends (CSVs) on Clostridium perfringens and Bacillus cereus in uncured beef and poultry products during extended cooling. Treatments (beef: 72-73% moisture, pH 6.2-6.3, 0.85-0.95% NaCl; turkey: 76-77% moisture, pH 6.5-6.7, 1.3-1.6% NaCl) included Controls without antimicrobials, and four DV and four CSV, each tested at 0.75 and 1.25%. Batches were inoculated with 2.5-log C. perfringens or B. cereus spores, vacuum-packaged, and cooked to 73 °C. Packages were cooled from 48.8 to 27 °C (Phase 1) in 3, 4, or 5 h; Phase 2 (27-12.8 °C) and Phase 3 (12.8-4 °C) were standardized for 5-h cooling each. Pathogens were enumerated on selective agar in triplicate samples assayed at precook, postcook, and at the end of Phase 1, 2, and 3 cooling. Experiments were conducted twice. B. cereus did not grow (<0.5-log increase) in any treatment when Phase 1 cooling was extended to 5 h. C. perfringens grew rapidly (2.5 to >4.5 log) in Control treatments when Phase 1 cooling was extended to ≥3 h. All 1.25% DV ingredients limited C. perfringens growth to ≤1-log when Phase 1 cooling was extended to 3 h but supported a >1-log increase when Phase 1 cooling was extended to 5 h. All 1.25% CSV inhibited growth under 3-h Phase 1 cooling; 1.25% CSV-A and ≥0.75% CSV-D inhibited growth in turkey during 5-h Phase 1 cooling, but inhibition with 1.25% CSV-C was inconsistent in beef. This study revealed that formulating uncured meats with 1.25% DV or certain CSV can extend Phase 1 cooling to 3 h. Although all ingredients inhibited growth when used at 0.75% or greater compared to a control, greater variability of inhibition was observed among CSV than for DV.

Screening and selection of eubiotic compounds possessing immunomodulatory and anti-Clostridium perfringens properties.

Eubiotics are water and/or feed additives used in poultry to promote gut health and control enteric burden of pathogens, including Clostridium perfringens. While several eubiotic compounds (ECs) are being introduced commercially, it is essential to devise an in vitro model to screen these compounds to assess their immunomodulatory and antimicrobial properties prior to their testing in vivo. A chicken macrophage cell-line (MQ-NCSU) was used to develop an in vitro model to screen the immunological and anti-C. perfringens properties of 10 ECs: monobutyrin, monolaurin, calcium butyrate, tributyrin, carvacrol, curcumin, green tea extract, rosemary extract, monomyristate, and tartaric acid. An optimal concentration for each EC was selected by measuring the effect on viability of MQ-NCSU cells. Cells were then treated with ECs for 6, 12, and 24 h. and expression of interferon-gamma (IFNγ), interleukin (IL)-1ß, IL-6, IL-10, transforming growth factor-beta (TGFß) and cluster of differentiation (CD40) genes, as well as major histocompatibility complex (MHC)-II protein were evaluated. At 6 h post-stimulation, monobutyrin, calcium butyrate, and green tea extract treatments induced a significant downregulation of IFNγ, IL-6, or IL-1ß gene transcription and MHC-II expression, while the IL-10 or TGFß gene expression in these treatments as well as those receiving rosemary extract and tartaric acid was significantly upregulated, when compared to control, suggesting immunomodulatory properties of these ECs. Finally, pretreatment of macrophages with these selected 5 ECs for 24 h followed by C. perfringens infection showed that monobutyrin, green tea extract, rosemary extract, and calcium butyrate treatments can inhibit bacterial growth significantly at 12 and/or 24 h post-infection, when compared to the control. Collectively, our findings show that ECs possessing immunomodulatory and anti-C. perfringens properties can be selected using an in vitro avian macrophage cell-based model so that such ECs can further be tested in vivo for their disease prevention efficacy.

High intestinal carriage of Clostridium perfringens in healthy individuals and ICU patients in Hangzhou, China.

Clostridium perfringens has emerged as a growing public health concern due to its ability to cause various infections and its increasing resistance to antibiotics. To assess its current epidemiology in clinical settings, we conducted a survey involving 426 healthy individuals and 273 ICU inpatients at a provincial hospital in China. Our findings revealed a high prevalence of C. perfringens in healthy individuals (45.77%, 95% CI: 41.0%-50.6%) and ICU patients (12.82%, 95% CI: 9.1%-17.4%). The identified 220 C. perfringens isolates displayed substantial resistance to erythromycin (57.9%), clindamycin (50.7%), and tetracycline (32.0%), primarily attributed to the presence of erm(Q) (54.4%), lnu(P) (13.8%), tetB(P) (83.6%), and tetA(P) (66.7%). Notably, C. perfringens isolates from this particular hospital demonstrated a high degree of sequence type diversity and phylogenic variation, suggesting that the potential risk of infection primarily arises from the bacteria's gut colonization rather than clonal transmissions within the clinical environment. This study provides an updated analysis of the current epidemiology of C. perfringens in healthy individuals and ICU patients in China and emphasizes the need to optimize intervention strategies against its public health threat. IMPORTANCE: Clostridium perfringens is a bacterium of growing public health concern due to its ability to cause infections and its increasing resistance to antibiotics. Understanding its epidemiology in clinical settings is essential for intervention strategies. This study surveyed healthy individuals and ICU inpatients in a provincial hospital in China. It found a high prevalence of C. perfringens, indicating infection risk. The isolates also showed significant antibiotic resistance. Importantly, the study revealed diverse sequence types and phylogenetic variation, suggesting infection risk from intestinal colonization rather than clonal transmission in hospitals. This analysis emphasizes the need to optimize intervention strategies against this public health threat.

Genomic analysis of Clostridium perfringens type D isolates from goat farms.

C. perfringens type D strains are the leading cause of enterotoxaemia in ruminants such as goats, sheep, and cattle. However, there has been no prior research on the genomic characteristics of C. perfringens type D strains from various regions in China. Here, we investigated the antibiotic resistance, genomic characteristics, and phylogenetic relationship of C. perfringens type D isolates recovered from goat farms in Shaanxi, Gansu, and Ningxia provinces. The antibiotic resistance test indicated that the isolates displayed high minimum inhibitory concentration (MIC) values to sulfafurazole, whereas the other antibiotics tested, such as penicillin, enrofloxacin, and florfenicol, worked well on them. Additionally, only tetracycline resistance genes [tetA(P) and tetB(P)] were identified from the isolates. A collective of 13 toxin genes, including etx and cpe were detected among the isolates. Sequence comparison revealed that the etx and cpe genes shared high sequence identities, and they could coexist on a pCW3-like plasmid, representing a potential risk to both animal breeding and public health. Phylogenetic analysis using core genome multi-locus sequence typing (cgMLST) and core genome single nucleotide polymorphisms (SNPs) revealed the close genetic relationship and potential regional/transregional transmission of the C. perfringens type D isolates in Shaanxi and Gansu provinces. Furthermore, pan-genomic analysis suggested the functional differences at the protein-coding gene level, although isolates from the same source shared a close genetic relationship. In conclusion, this study indicated the antibiotic resistance, virulence markers, potential transregional transmission, and genomic diversity of C. perfringens type D strains from various regions in China, which could provide references for the prevention of C. perfringens foodborne diseases and further research.

Research Note: Bovine lactoferrin in chickens: an investigation into its viability as an antibiotic alternative.

Finding effective antibiotic alternatives is crucial to managing the re-emerging health risk of Clostridium perfringens (CP) type A/G-induced avian necrotic enteritis (NE), a disease that has regained prominence in the wake of governmental restrictions on antibiotic use in poultry. Known for its antimicrobial and immunomodulatory effects, the use of bovine lactoferrin (bLF) in chickens is yet to be fully explored. In this study, we hypothesized that bLF can accumulate in the small intestines of healthy chickens through gavage and intramuscular supplementation and serves as a potential antibiotic alternative. Immunohistochemistry located bLF in various layers of the small intestines and ELISA testing confirmed its accumulation. Surprisingly, sham-treated chickens also showed the presence of bLF, prompting a western blotting analysis that dismissed the notion of cross-reactivity between bLF and the avian protein ovotransferrin. Although the significance of the route of administration remains inconclusive, this study supports the hypothesis that bLF is a promising and safe antibiotic alternative with demonstrated resistance to the degradative environment of the chicken intestines. Further studies are needed to determine its beneficial pharmacological effects in CP-infected chickens.

Phytochemical composition and In-vitro activity of ethanolic extract of Eucalyptus globulus leaves against multidrug resistant poultry pathogens.

Aim of the present study was to determine the In-vitro antibacterial activity of ethanolic extract of E. globulus leaves against common multidrug resistant poultry pathogens. Phytochemical analysis through HPLC revealed that kaempeferol (7.315min) followed by querecetin (6.655min) and myrecetin (3.655min). Percent area of kaempeferol (6826.88%) was highest, followed by myrecetin (5516.22%) and querecetin (163.748%). Phytochemical investigation of ethanolic extract of E. globulus leaves through GCMS showed highest retention time (min) α-pinene (20.43) and α-terpineol (20.15) accompanied by spathulenol (11.97), piperitone (11.04). The ethanolic extracts of E. globulus leaves showed a highest zone of inhibition against S. pullorum SP6; 20.64± 2.08, E. coli SE 12; 19.75± 2.83, C. perfringens type A (CPM38-01); 19.46± 2.02. The highest level of MIC of E. globulus noted were against S. gallinarum S22; 133.37±53.294, S. gallinarum S1; 130.20±45.10, S. gallinarum S4; 129.47±24.182, S. gallinarum S3; 126.83±72.392. In conclusion, the study confirmed that the ethanolic extract of E. globulus is composed of active ingredients having antibacterial activity and can be referred as an alternate to antibiotics.

Activity of ethanolic extract of Eucalyptus globulus leaves against multi drug resistant poultry pathogens in broiler chicks.

The present study was designed to evaluate the antimicrobial activity of E. globulus leaves in broiler chicks. Total (n=255) day-old chicks were segregated into five groups i.e. Pathogenic E. coli, S. pullorum, S. gallinarum and C. perfringens type A and control negative group. Each bacterial challenged (1x 107 CFU) group was divided into control positive, antibiotic, probiotic and E. globulus group. Experimental birds were exposed to E. coli, S. pullorum, S. gallinarum and C. perfringens type A at different ages. At 35th day of experiment the log reduction for each group was determined. The highest log reduction in E. coli and C. perfringens Type A colonies count were found in E. globulus (3.26) (2.33) treated group followed by antibiotic (2.85) (1.59) and probiotic (2.84) (1.50) respectively. The log reduction in S. pullorum colonies count was highest in E. globulus (2.50) followed by probiotic (2.24) and antibiotic (2.16). The S. gallinarum colonies count log reduction was found highest for antibiotic (2.84) followed by probiotic (2.48) and E. globulus group. The results of in-vivo experiment revealed that ethanolic extract of E. globulus has antibacterial activity and it can be used as a replacement to low level of antibiotics added in poultry feed.

The Impact of Berberine on Intestinal Morphology, Microbes, and Immune Function of Broilers in Response to Necrotic Enteritis Challenge.

The objective of this study was to explore the therapeutic effects of berberine on necrotic enteritis (NE) in broilers caused by Clostridium perfringens. A total of 240 1-day-old Arbor Acres chicks were divided into four groups, as negative controls (NC), positive controls (PC), berberine- (BER-) treated, or lincomycin- (LMY-) treated groups. Broilers were challenged with C. perfringens at 15-21 days of age, followed by BER or LMY supplied in drinking water for 7 days. Experimental results showed that C. perfringens infection significantly decreased growth performance and increased intestinal necrosis index and the number of C. perfringens present to 6.45 Log10CFU/g (P < 0.001). Proinflammatory cytokines in the ileum were significantly increased, but the expression of ileal tight junction proteins occludin and claudin-1 was significantly reduced. Both BER and LMY ameliorated some of these observations. Compared with the PC group, the number of C. perfringens in the cecum was significantly decreased following treatment (P < 0.001), and growth performance and small intestine morphology were similar to those of the NC group (P > 0.05). IL-1ß, IL-6, and TNF-α levels as well as occludin and claudin-1 expression were also significantly improved (P < 0.05). BER has the potential to replace antibiotics for NE caused by C. perfringens.

Prevalence and characterization of Clostridium perfringens isolated from different chicken farms in China.

Clostridium perfringens (C. perfringens) is a common pathogenic microorganism present in nature, which can cause animal and human diseases, such as necrotizing enteritis (NE) in poultry. Little is known about the current prevalence status of C. perfringens from poultry farms of different types and regions in China. From December 2018 to August 2019, we investigated the prevalence, genotype distribution and drug resistance of C. perfringens from Guangdong, Pingyin, Tai'an and Weifang. A total of 622 samples were collected and processed for C. perfringens isolation, among which 239 (38.42%) samples were determined to be positive for C. perfringens. A total of 312 isolates of C. perfringens were recovered (1-5 strains were isolated for each positive sample), and 98.72% of the isolates were identified as type A, while the others were type F. Antimicrobial susceptibility testing revealed that 47.71% of the isolates were resistant to at least five classes of commonly used antibiotics. Multilocus sequence typing (MLST) showed that 74 representative isolates were divided into 63 sequence types (STs), and the Simpson's diversity index (Ds) of the STs for the five farms was 0.9799. 37.84% of the isolates were classified into seven clonal complexes (CC1-CC7), and the isolates from the same farm were more concentrated in the minimum spanning tree. In addition, some cloaca isolates and feed isolates were distributed in the same ST or CC; this result indicates that the C. perfringens in chicken can come from the environment (feed etc.).

A study on fungal defensin against multidrug-resistant Clostridium perfringens and its treatment on infected poultry.

In the present study, we aimed to investigate the antibacterial activity and mechanisms of plectasin-derived peptide NZ2114 in vitro and its therapeutic effects in vivo on broilers challenged with Clostridium perfringens. In vitro assay showed that NZ2114 had potent (minimal inhibitory concentration, 0.91 µM) and rapid antibacterial activity (99.9% reduction within 2 h), and the dual antibacterial mechanisms (including interfering with the cell membrane and intracellular DNA) against C. perfringens CVCC 2030. In vivo study, NZ2114 tended to increase linearly and quadratically the average daily gain as NZ2114 level increased and was the highest at 20 mg/L. NZ2114 at 10 ~ 40 mg/L dramatically reduced jejunal lesion score. Besides, the levels of IL-6, TNF-α, and IL-1ß tended to downregulate linearly and quadratically as the NZ2114 level increased and were all the lowest at the dose of 20 mg/L. NZ2114 significantly upregulated those levels of IgA, IgG, IgM, and sIgA with a linear and quadratic dose effect, with the highest IgA, IgG, IgM, and sIgA at 20 mg/L. Finally, NZ2114 tended to linearly and quadratically increase the numerical value of crypt depth, with the lowest value at 40 mg/L. Lincomycin only dramatically reduced the jejunal lesion score and increased the numerical value of crypt depth. These results indicate that NZ2114 has the potential as a new alternative to antibiotics for the treatment of C. perfringens-induced necrotic enteritis infection.Key points• NZ2114 could kill C. perfringens by dual antibacterial mechanisms• Broiler necrotic enteritis model induced by C. perfringens was established• NZ2114 treatment could ameliorate C. perfringens-induced necrotic enteritis.

Assessment of Immune Response and Efficacy of Essential Oils Application on Controlling Necrotic Enteritis Induced by Clostridium perfringens in Broiler Chickens.

Necrotic enteritis (NE) caused by Clostridium perfringens is one of the most important enteric diseases in poultry. The antibacterial activity of two different essential oil (EO) blends against C. perfringens was investigated both in vitro and in vivo. Additionally, the immunological response to EO treatment was assessed. In the in vitro study, the antibacterial activity of EO formulas and commonly used antibiotics was evaluated against C. perfringens using disk diffusion assay, minimum inhibitory concentration (MIC) assay, and minimum bactericidal concentration (MBC) assay. In the in vivo study, NE experimental infection was performed on 440 Ross broiler chicks at 19 days of age for 4 continuous days. The chicks were treated with either EOs or amoxicillin at 22 days of age for 5 continuous days. One day after the end of treatment, the birds' performance was evaluated by calculating the feed conversion ratio. Serum samples from 120 birds were collected to measure the levels of IL-1ß, IFN-γ, IL-8, IL-10, and IL-17. After that, all birds were slaughtered, and their small intestines were subjected to gross and histopathological evaluation. In addition, bacterial counts in the small intestines were evaluated. In the in vitro study, EOs showed higher antimicrobial activities in comparison with antibiotics against C. perfringens. In the in vivo study, birds treated with EOs showed a significant decrease in bacterial counts, a significant decrease in intestinal lesions, and a significant improvement in performance compared with untreated birds (p < 0.05). Moreover, treating birds with EOs directed the immune system toward an anti-inflammatory pathway. None of the treated birds died due to NE compared with the 10% mortality rate in untreated birds. In conclusion, EOs might be an effective and safe alternative to antibiotics in the treatment of chicken NE.

The Biological Activity of Monarda didyma L. Essential Oil and Its Effect as a Diet Supplement in Mice and Broiler Chicken.

The use of growth-promoting antibiotics in livestock faces increasing scrutiny and opposition due to concerns about the increased occurrence of antibiotic-resistant bacteria. Alternative solutions are being sought, and plants of Lamiaceae may provide an alternative to synthetic antibiotics in animal nutrition. In this study, we extracted essential oil from Monarda didyma, a member of the Lamiaceae family. We examined the chemical composition of the essential oil and then evaluated the antibacterial, antioxidant, and anti-inflammatory activities of M. didyma essential oil and its main compounds in vitro. We then evaluated the effectiveness of M. didyma essential oil in regard to growth performance, feed efficiency, and mortality in both mice and broilers. Carvacrol (49.03%) was the dominant compound in the essential oil extracts. M. didyma essential oil demonstrated antibacterial properties against Escherichia coli (MIC = 87 µg·mL-1), Staphylococcus aureus (MIC = 47 µg·mL-1), and Clostridium perfringens (MIC = 35 µg·mL-1). Supplementing the diet of mice with essential oil at a concentration of 0.1% significantly increased body weight (+5.4%) and feed efficiency (+18.85%). In broilers, M. didyma essential oil significantly improved body weight gain (2.64%). Our results suggest that adding M. didyma essential oil to the diet of broilers offers a potential substitute for antibiotic growth promoters.

Epigallocatechin gallate and Lactobacillus plantarum culture supernatants exert bactericidal activity and reduce biofilm formation in Clostridium perfringens.

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.

Bacteriocin-Like Inhibitory Substance (BLIS) Activity of Enterococcus faecium DB1 Against Biofilm Formation by Clostridium perfringens.

The antibiofilm effect of bacteriocin-like inhibitory substance (BLIS) from Enterococcus faecium DB1 against Clostridium perfringens was investigated in the present study. BLIS of E. faecium DB1 significantly reduced biofilm formation by C. perfringens in a dose-dependent manner for 24 and 48 h. In particular, treatment with BLIS of E. faecium DB1 significantly inhibited biofilm formation by C. perfringens on chicken meat and stainless steel coupon surfaces. Moreover, BLIS of E. faecium DB1 decreased the viability of C. perfringens biofilm and planktonic cells, indicating that the reduction of biofilm formation by C. perfringens might be achieved by killing the bacterial cells. Taken together, the present results suggest that BLIS of E. faecium DB1 can be a promising antibiofilm agent to eradicate C. perfringens.

Selection and application of natural antimicrobials to control Clostridium perfringens in sous-vide chicken breasts inhibition of C. perfringens in sous-vide chicken.

Current consumer preferences for both clean label food ingredients and convenience-based foods has provided a unique opportunity to explore the application of novel natural food preservatives in sous vide products. The anaerobic environment and relatively low thermal processing of the sous vide process creates a favorable environment for the survival, germination, and outgrowth of spore-forming bacterium Clostridium perfringens. The aim of this study was to identify effective novel natural ingredient formulations against C. perfringens and apply them within a vacuum-sealed sous vide chicken model exposed to abusive storage and chilling conditions. Among six commercial vinegar-based formulations, liquid vinegar with citrus extract (CE; 1.0%) and with lemon juice concentrate (LJC; 1.5%) were identified as the most effective at inhibiting three individual C. perfringens strains. Both reduced viable cell counts by 5 log CFU/mL (P < 0.05), whereas reductions in spore counts ranged from 2 to 4 log CFU/mL depending on formulation and concentration used. Once incorporated to chicken meat 1.0% CE and 1.5% LJC before sous-vide cooking, completely inhibited the growth of mixed C. perfringens strains (P < 0.05) during storage for 16 days at 12 and 16 °C. Exponential cooling from 54 to 4 °C was performed for 18 h to imitate abusive storage conditions. CE and LJC at 3.0% inhibited growth and reduced counts by 3.4 and 2.9 log CFU/g compared to respective controls. Treatments CE and LJC could be implemented within the formulation of a sous vide chicken product to provide an effective protection against C. perfringens meeting clean label expectations.

The Multifunctional Sactipeptide Ruminococcin C1 Displays Potent Antibacterial Activity In Vivo as Well as Other Beneficial Properties for Human Health.

The world is on the verge of a major antibiotic crisis as the emergence of resistant bacteria is increasing, and very few novel molecules have been discovered since the 1960s. In this context, scientists have been exploring alternatives to conventional antibiotics, such as ribosomally synthesized and post-translationally modified peptides (RiPPs). Interestingly, the highly potent in vitro antibacterial activity and safety of ruminococcin C1, a recently discovered RiPP belonging to the sactipeptide subclass, has been demonstrated. The present results show that ruminococcin C1 is efficient at curing infection and at protecting challenged mice from Clostridium perfringens with a lower dose than the conventional antibiotic vancomycin. Moreover, antimicrobial peptide (AMP) is also effective against this pathogen in the complex microbial community of the gut environment, with a selective impact on a few bacterial genera, while maintaining a global homeostasis of the microbiome. In addition, ruminococcin C1 exhibits other biological activities that could be beneficial for human health, as well as other fields of applications. Overall, this study, by using an in vivo infection approach, confirms the antimicrobial clinical potential and highlights the multiple functional properties of ruminococcin C1, thus extending its therapeutic interest.

Sequencing of Clostridium perfringens toxin genes (cpa, etx, iap) from Iraqi hospitals and detection by PCR of the genes encoding resistance to metronidazole, tetracycline, and clindamycin.

PURPOSE: This cross sectional study was designed to investigate the molecular detection of C. perfringens toxins (alpha, epsilon, iota) and antibiotics resistance genes, as well as sequencing of their toxin genes. METHODS: Different wound swabs were taken from 140 patients. PCR was applied for detection of clostridial toxins; alpha toxin (cpa) gene, epsilon toxin (etx) gene, and iota toxin (iap) gene. Metronidazole nim gene, tetracycline resistance genes; (tetQ, tetM, tetB, and tetW) and clindamycin resistance genes erm (A,B), were used for genotypic detection of antibiotic resistance. RESULTS: Out of 140 clinical samples collected, 7 isolates were detected using specific primer 16S-23S intergenic rRNA spacer gene of C. perfringens. Results showed presence of alpha toxin (cpa) genes in all clostridial isolates, Epsilon toxin (etx) genes in 2/7(28.4%), and iota toxin (iap) genes in 2/7 (28.4%). Results of antibiotic resistance genes showed that all isolates were not able to produce nim, tet W and Q, tet B, erm (A), erm (B) genes except for tetM. Gene sequence analysis for cpa gene showed that there were 3 mutations in the sample of this gene, the results of etx gene when had been sent 2 samples from this gene. The first sample showed that there was one mutation and the results of iap gene showed that there were no mutation in 2 samples of the iap gene. The samples had been showed identical 100%. CONCLUSION: The DNA sequence analysis of bacterial genome revealed several important feature including that may allow to confirm new the presence of toxin and to identify new or mutant toxins that may be missed by diagnostic PCR specifically target toxin encoding genes.