Search Results (318)

Background/Objectives: Necrotic enteritis (NE), induced by Clostridium perfringens, is responsible for significant economic losses in the poultry industry worldwide. The growing restrictions on antibiotic use have driven the search for alternative strategies for disease control. The purpose of this study is to isolate and characterize lytic phages targeting multidrug-resistant C. perfringens type G recovered from chickens with NE. Methods: C. perfringens was isolated from chickens with NE using a culture method with selective TSC agar. Bacterial identification was carried out using biochemical tests and PCR. C. perfringens isolates were toxinotyped by PCR. Antibiotic susceptibility test was performed using the agar dilution method. Bacteriophages were isolated from chicken intestine samples collected from wet markets using the double-layer agar technique. Phage isolates were characterized by host range, one-step growth, stability, and whole genome sequencing. The efficacy of phage CPP8 in controlling multidrug-resistant C. perfringens type G was evaluated in GAM broth. Results: In this study, 16 C. perfringens strains were isolated from 100 chickens suspected of NE. Among these isolates, 10 (62.5%) belonged to type G, while the remaining 6 (37.5%) were type A. A total of 11 phages capable of lysing C. perfringens type G were isolated from the chicken intestine. Among them, phage CPP8 has the widest host range, short latent period, large burst size, and high stability. Moreover, the genome of CPP8 lacked genes related to antibiotic resistance, toxins, virulence factors, or lysogeny. Treatment with CPP8 resulted in a significant reduction in viable counts of C. perfringens at 37 °C. Conclusions: Our findings highlight phage CPP8 as a promising candidate for bio-control of multidrug-resistant C. perfringens type G. Full article

Silage is a fundamental component of cattle feed, and its microbiological quality is critical for animal health and human safety. Improper ensiling conditions, such as oxygen exposure or inadequate acidification, can promote the growth of pathogens like Listeria monocytogenes, Clostridium botulinum, and Bacillus cereus. This study aimed to evaluate the microbial status of maize silages and identify pre-ensiling factors influencing its hygienic safety. Over a four-year period, 406 silage samples were collected from cattle farms across Poland. The research evaluated general hygiene indicators and screened for specific pathogens using standard culture methods, polymerase chain reaction toxotyping, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The impact of agricultural practices, including soil quality, organic fertilization, and microbial inoculation, was also analyzed. The analysis revealed that 32.1% of silages fell outside the reference pH range, indicating potential aerobic instability. While Salmonella and Campylobacter were not detected, Clostridium spp. were highly prevalent (81.0%), and C. perfringens was confirmed in 24.9% of samples. Listeria species occurred in 2.9% of silages, with L. innocua being the most frequent isolate. Statistical analysis showed that organic fertilization was significantly linked to specific C. perfringens toxotypes, though it did not increase the overall microbial burden. Conversely, microbial inoculation generally reduced the counts of several undesirable bacteria, although these differences were not statistically significant across all parameters. High pH values and significant contamination with Clostridium, B. cereus, and fungi remain critical challenges for silage safety. The results underscore the necessity for improved agricultural practices—specifically the minimization of soil and manure contamination during harvest—and the broader adoption of microbial inoculation to ensure the microbiological stability of fermented forage. Full article

Increased demands for protein have led to a search for alternatives to traditional protein sources like soy and animal protein. Black Soldier Fly Larvae can be reared on many feedstocks to produce a high-quality nutrient source for livestock and pets. These insects contain biologically meaningful compounds like antimicrobial peptides, lauric acid, and chitin. This combination of compounds highlights the need to investigate BSFL as a functional ingredient in broilers. This study examined the impact of BSFL inclusion on broiler performance with and without a subclinical Necrotic Enteritis (NE) challenge over two experiments. In both experiments, diets included 0%, 2.5%, or 5.0% BSFL from 0–42 d. During Experiment 2, birds were given a live coccidiosis vaccine at hatch and challenged with C. perfringens at 19, 20, and 21 d. Primary variables include growth performance, lesion scores, and NE-specific mortality. A BSFL inclusion of 2.5% and 5.0% significantly improved feed conversion ratio in both experiments (p < 0.001); 5% BSFL inclusion also significantly improved body weight gain in both experiments (p = 0.014, p = 0.023, respectively). Overall, results indicate that BSFL is an effective and safe feed ingredient option for commercial broiler production, providing biologically relevant improvements in performance with and without disease pressure. Full article

Necrotizing enteritis (NE) is an important intestinal disease threatening the poultry farming industry, and the ban on antibiotic growth promoters has created an urgent demand for safe and effective NE vaccines. Recombinant attenuated Salmonella vectors (RASVs) administered orally can induce mucosal immune responses against delivered antigens, thus showing great potential to elicit protective immunity against NE. The EntB protein is a newly discovered putative enterotoxin of Clostridium perfringens (C. perfringens). Bioinformatic predictions in this study revealed that EntB contains nineteen potential antigenic epitopes, two functional domains (NlpC and YgiM), and interacts with ten proteins, supporting its potential as a target antigen for NE vaccines. To optimize the immunogenicity of EntB-based vaccines, we constructed a novel recombinant attenuated Salmonella Enteritidis (S. Enteritidis) vector rSC0169 harboring a rhamnose-regulated delayed attenuation system, which was then used to deliver EntB to generate the recombinant strain rSC0169(pS-EntB). This system enhanced the immunogenicity of the Salmonella vector rSC0169 and further elicited robust mucosal immune responses against EntB, as well as humoral and cellular immune responses. Compared with the control strain rSC0169(pS0018), rSC0169(pS-EntB) candidate vaccine strain significantly alleviated NE symptoms, increased the intestinal villus height/crypt depth (VH/CD) ratio, upregulated tight junction protein expression, and reduced excessive pro-inflammatory cytokine production. In conclusion, this study provides a promising NE candidate vaccine and offers a valuable strategy for developing vaccines against other intestinal bacterial diseases. Full article

The study aimed to determine the effect of newly developed multi-strain synbiotics on the hygienic quality of feed for pigs and pork edible raw materials originating from animals supplemented with synbiotics. The trial was conducted on 54 feed samples and 54 pigs. Animals were divided into six groups corresponding to three synbiotic preparations (A, B, C), two positive controls with probiotics (D, E), and a negative control (K) with no feed additive. The supplementation of the sow’s basal diet with the feed additive started 10 days before farrowing and continued for 38 days, covering the lactation period. The diet of piglets from groups A-E was supplemented with the feed additive starting from two weeks of age until slaughter at 24 weeks of age. Feed and animal raw materials were tested for the presence of Salmonella spp., Campylobacter spp., L. monocytogenes, Clostridium spp., C. perfringens, C. botulinum, and the count of AMB, TPC, fungi, ASFB, C. perfringens, Enterobacteriaceae family, E. coli, presumptive B. cereus, CoPS, HS, LAB, yeast probiotic strains, and Enterococcus spp. Statistically significant differences were found between individual groups in the count of C. perfringens, AMB, TPC, Enterococcus spp., and LAB in all feeds tested. Moreover, synbiotics A, B, and C lowered the count of AMB, TPC, and LAB, and synbiotics A and C decreased Enterobacteriaceae family contamination in both total raw materials as well as raw materials of fatteners. The higher effectiveness of synbiotics was associated with a higher number of probiotic strains in one preparation. Our study found that dietary supplementation of synbiotics alters the occurrence of C. perfringens in feed and enhances the hygienic quality of edible pork raw materials. Full article

Chrysomya megacephala is a synanthropic fly with a high potential to act as a mechanical vector of pathogenic bacteria, surpassing Musca domestica in both bacterial load and diversity. Native to Asia and Africa, it has become a cosmopolitan species, successfully adapting to a wide range of environments, including natural ecosystems. In Colombia, studies on its role as a vector are limited and have largely relied on traditional culturing methods. This study aimed to characterize the pathogenic bacterial microbiota associated with C. megacephala using 16S rRNA gene sequencing in urban, rural, and forest settings of a coastal tourist city. Flies were collected using Van Someren Rydon traps with attractants and sterile materials. Bacterial identification was performed through Oxford Nanopore MinION sequencing (Manufactured by Oxford Nanopore Technologies, Oxford, UK). A total of 49 bacterial species were identified, with urban environments showing the highest taxonomic richness. The forest environment was characterized by a highly dominant community structure, led by Vagococcus carniphilus. Notably, 20 bacterial species of public health relevance were detected, including Clostridium botulinum, Clostridium perfringens, Ignatzschineria ureiclastica, Escherichia coli, and Streptococcus agalactiae. These findings indicate that bacterial community composition varies by environment and underscore the potential role of C. megacephala as a mechanical vector, highlighting the importance of surveillance for its public health implications. Full article

Tetanus toxin evaluation has traditionally relied on mouse LD₅₀ bioassays, which require extensive animal use and time, necessitating development of alternative methods in accordance with 3R principles (Replacement, Reduction, and Refinement). We developed and validated a sandwich enzyme-linked immunosorbent assay (ELISA) as an alternative to animal testing for evaluating tetanus toxin biological activity using 18 environmental and clinical isolates of Clostridium tetani, complemented by an immunochromatographic (IC) assay for rapid screening. The ELISA demonstrated excellent analytical performance with a lower limit of quantification of 2.4 ng/mL (equivalent to 85.4 LD₅₀/mL), favorable linearity (R² = 0.999), precision (CV < 1.7–8.2%), and specificity (<1% cross-reactivity with C. septicum, C. novyi, and C. perfringens). Correlation analysis between ELISA relative potency and observed minimum lethal dose values revealed a robust positive correlation (r = 0.974). Both parallel line assay and single-point quantification methods showed strong correlations with mouse bioactivity measurements (r = 0.998). The IC assay successfully detected all isolates within 15 min. The measurement range of 2.4–45.6 ng/mL effectively covered diverse toxin-production capabilities spanning a 600-fold concentration range. This validated ELISA and IC assay combination provides a reliable, rapid alternative to animal experimentation for tetanus toxin evaluation. Full article

This study aimed to investigate the impacts of chestnut tannin (CT) on growth performance, immune response, and intestinal health of broilers challenged with necrotic enteritis (NE) through the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING)-ferroptosis pathway. A total of 240 one-day-old male Cobb 500 broilers (44.54 ± 0.51 g) were randomly divided into four groups, including a Control group, NE group, 500 mg/kg CT group (L-CT), and 1000 mg/kg CT group (H-CT), with six replicates per group and ten broilers per replicate. Sporulated coccidia oocysts on day 14 and Clostridium perfringens solution from days 19 to 21 were given to all broilers except the Control group through oral administration to establish the NE infection model. The results demonstrated that dietary supplementation with CT improved (p < 0.05) growth performance, intestinal morphology, and intestinal mucosal barrier function of broilers challenged with NE. CT supplementation decreased (p < 0.05) interleukin (IL)-1β, IL-6, type I interferon, interferon-γ, and tumor necrosis factor-α concentrations and increased (p < 0.05) IL-10 concentration in the jejunal mucosa. Furthermore, CT supplementation decreased (p < 0.05) Fe²⁺ concentration, malondialdehyde concentration, mitochondrial DNA level, and mitochondrial reactive oxygen species level in the jejunal mucosa. Broilers under NE challenge had upregulated (p < 0.05) jejunal protein expression of cGAS, STING, phospho-TANK-binding kinase 1, phospho-interferon regulatory factor 7, phospho-nuclear factor kappa B, ferroptosis suppressor protein 1, prostaglandin-endoperoxide synthase 2, acyl-CoA synthetase long-chain family member 4, WD repeat domain phosphoinositide-interacting protein 2, nuclear receptor co activator factor 4 and autophagy related protein 5 and downregulated (p < 0.05) glutathione peroxidase 4, ferritin heavy chain 1, ferritin light chain and ferroportin 1 compared with the Control group, while the supplementation of CT reversed these effects. In conclusion, CT improved intestinal inflammatory damage of broilers challenged with NE by inhibiting the cGAS-STING-ferroptosis pathway, which was more effective at a dose of 1000 mg/kg in this study. Full article

The scientific understanding of cetacean pathology has advanced significantly in recent decades. However, data concerning the health status of members of the family Kogiidae remains scarce. This study presents a comprehensive pathological assessment and determination of causes of death in 45 stranded kogiids, comprising 35 Kogia breviceps and 10 K. sima, along the coasts of the Canary Islands between 1999 and 2018. Causes of death (CD) were classified as natural (30/45; 66.6%) or anthropogenic (11/45; 24.4%), while the cause remained undetermined in four cases (9%). Among natural causes, the most prevalent etiologies included trauma (13/30; 43.3%), infectious (7/30; 23.3%), parasitic (5/30; 16.6%), cardiomyopathy (3/30; 10%), malnutrition (1/30; 3.3%), and fetal distress (1/30; 3.3%). Anthropogenic causes were vessel collisions (7/11; 63.6%), interactions with fishing activities (2/11; 18.2%), and foreign body-associated pathology (2/11; 18.2%). Notably, intra- and interspecific traumatic interactions were frequently identified in this cohort, and variable degrees of gross and histologic cardiomyopathic changes were observed in 68.9% of individuals (31/45) without evident sex bias. Septicemia caused by Clostridium perfringens and C. tertium was confirmed in three cases. Additionally, cervical gill slit adenitis by Crassicauda sp. (15/45; 33%) and parasitic gastritis by Anisakis sp. (27/45; 60%) were frequent findings. These findings provide novel insights into the pathologic spectrum affecting kogiids and enhance the current understanding of their health status, with implications for diagnostic protocols, stranding response, and conservation strategies. Full article

This study investigates the elaboration, structural characteristics, and antibacterial performance of silver-based nanoparticles obtained via a hydrolytic chemical route, with and without assistance from ultrasound and microwave irradiation. Two silver nitrate precursor concentrations (1 M and 2 M) were employed to evaluate the influence of synthesis conditions on phase composition, morphology, and antimicrobial efficiency. The obtained powders were characterized by ATR-FTIR, X-ray diffraction (XRD), and scanning electron microscopy (SEM). XRD analysis revealed that drying at 120 °C led to oxide-rich systems dominated by Ag₂O, with minor contributions from metallic Ag and carbonate species, while calcination at 550 °C resulted in complete phase transformation into highly crystalline metallic silver. SEM observations demonstrated that precursor concentration and synthesis assistance strongly affect particle size, aggregation degree, and surface morphology. Ultrasound- and microwave-assisted synthesis promoted finer crystallite sizes and more homogeneous particle distributions compared to non-assisted routes. The antibacterial activity was evaluated against Escherichia coli (Gram-negative) and Clostridium perfringens (Gram-positive, anaerobic, spore-forming). Oxide-rich samples, particularly Ox.Ag/2 M, exhibited rapid and complete bacterial inactivation within 30 min, while metallic silver samples showed time-dependent antibacterial behavior, achieving full inhibition after 4 h. The results demonstrate that antibacterial efficiency is governed by a synergistic interplay between silver oxidation state, nanoscale morphology, and surface reactivity. These findings highlight the potential of tailored silver-based nanomaterials as effective antibacterial materials for biomedical, food safety, and environmental applications. Full article

Despite increasing interest in probiotics as antibiotic alternatives in swine production, few studies have directly compared the functional efficacy of different commercial probiotic formulations under controlled conditions. We conducted an in vitro study using porcine intestinal epithelial (IPEC-J2) and macrophage-like (3D4/21) cell models to compare the efficacy of three commercial probiotic consortia (C1: three strains of Bacillus velezensis; C2: B. licheniformis + B. subtilis; C3: Clostridium butyricum). Treatments were evaluated for their ability to inhibit pathogenic Escherichia coli, Clostridium perfringens, and Salmonella spp., enhance epithelial barrier integrity, and modulate immune responses. Experimental endpoints included pathogen inhibition assays, adhesion to IPEC-J2 cells, transepithelial electrical resistance (TEER), tight junction protein expression, and cytokine profiling via RT-qPCR and proteomics. Data were analyzed using the Kruskal–Wallis test with false discovery rate (FDR) control at 5%. C1 cell-free supernatant (CFS) strongly inhibited pathogen growth (84.8 ± 5.3% inhibition of ETEC F4⁺F18⁻ vs. medium control; p < 0.05), whereas C2 had no effect, and C3 inhibited only one isolate. The coculture of IPEC-J2 cells with C1 CFS increased the expression of TJ proteins ZO-1, MUC13, and MUC20 (+12.9–46.6% vs. control; p < 0.001) and anti-inflammatory TGF-β; reduced pro-inflammatory IL-6 in LPS-stimulated 3D4/21 cells. In comparison, C2 and C3 showed minimal impact on epithelial barrier integrity and immune modulation, as indicated by negligible changes in TEER values, tight junction protein expression (ZO-1, MUC13, MUC20), and cytokine profiles relative to the control. In conclusion, C1 demonstrated greater in vitro efficacy than C2 (B. licheniformis + B. subtilis) and C3 (Clostridium butyricum), including pathogen inhibition assays, epithelial adhesion, TEER measurements, and cytokine modulation, suggesting its potential as a leading candidate for functional probiotic applications. Full article

Clostridium perfringens is one of the main causes of death in poultry with no vaccines approved for poultry at present. The appropriate adjuvant is critical for the development of vaccines in C. perfringens in poultry. Here, we utilized Levilactobacillus brevis for high-yielding selenium biotransformation and demonstrated that heat-inactivated nano-selenium Lactobacillus (HiSeL) is a safe, efficient, and chemically stable selenium immunopotentiator for C. perfringens vaccines. We evaluated the effectiveness of HiSeL as an immune adjuvant to modulate the efficacy of multi-epitope vaccine in mice. Subcutaneous immunization mice with HiSeL promoted high levels of specific IgG, modulated cytokine secretion, downregulated stress-related gene expression, and provided 100% protection against lethal challenge with C. perfringens. Surprisingly, we found that HiSeL can quickly and effectively induce SIgA production even by subcutaneous immunization. Transcriptome sequencing revealed the pivotal role of TGF-β and NF-κB signaling pathways in IgA immune responses in mice immunized with the HiSeL-adjuvanted multi-epitope vaccine. Collectively, our study provides proof-of-concept evidence that HiSeL functions as a potent adjuvant candidate for the multi-epitope vaccine in a murine model, offering new insights into the development of engineered postbiotic-based adjuvants. Full article

C. perfringens is a Gram-positive, anaerobic, spore-forming bacterium that causes serious diseases in humans and animals. In this study, C. perfringens was isolated from the intestinal content of an alpaca, cultured, and then identified using laboratory methods including Gram staining, biochemical tests, and PCR for 16S rRNA of six toxins. Furthermore, the pathogenicity of different strains was analyzed in mice. The results showed that C. perfringens was identified as type A and caused severe pathology of the spleen, lungs, and duodenum in mice through CD4⁺ and CD8⁺ T cells. Also, the mRNA expression levels of ZO-1 and Occludin were further quantified by qRT-PCR with normalization to β-actin, which showed decreased expression levels in the duodenum of mice in the gavage group compared to those in the NC groups, with significant differences (n = 3; * p < 0.05, ** p < 0.01). The results could inform the development of drugs and vaccines resistant to C. perfringens in alpaca. Full article

Background: Clostridium perfringens is an infectious agent of concern in wild/farmed shellfish. Hence, this study assessed shellfish-borne Clostridium perfringens (ShbCp) prevalence. Methods: A total of 1469 ShbCp from 2336 shellfish were modelled using hierarchical generalized linear and 1000-permutation-based-mixed-effects, meta-regression models. Results: The overall ShbCp prevalence was 54.12% (19.73–84.99) with a 32.02% (14.52–56.64) toxigenic rate and a higher estimate in 2020–2025 (41.01%, 17.00–70.23) versus 1970–2019 (20.01%, 4.49–57.08). Culture media significantly affect ShbCp recovery, with cooked meat medium and thioglycollate medium registering higher estimates (77% and 25.15%, respectively) than selective agars (<7%). The molluscans had a higher ShbCp rate (60.68%) than crustaceans (1.57%) and cephalopods (0.14%); oysters (85.97%) than mussels (71.81%), clams (50.38%), slug/snails (48.23%), scallops (16.24%), crabs (11.91%), shrimps (1.05%), and squids (0.42%); and Crassostrea gigas (89.27%) versus Ruditapes philippinarum (45.92%) versus Mytilus galloprovincialis (30.14%). ShbCp differed significantly by nations but not by continent with Spain (87.79%) having the highest rate, then China (47.01%), Japan (43.91%), the USA (10.44%), and Greece (0.00%); South America (51.36%), then Asia (44.77%), Europe (21.97%), and North America (10.44%). Sample size, growth medium, nation, and shellfish class significantly explained 27.58%, 72.30%, 67.52%, and 28.51% ($R^2$) variance in ShbCp prevalence, respectively. Conclusions: The present study estimated a high ShbCp prevalence, suggesting a significant public health risk. It recommends that C. perfringens should be incorporated as a supplemental indicator into shellfish safety/shellfish water quality monitoring alongside traditional indicators. Also, geographical data gaps from Africa, Latin America, the Middle East, and Oceania underline the need for national and global monitoring attention and priority on C. perfringens in shellfish/shellfish beds. Full article

Clostridium perfringens is an opportunistic Gram-positive bacterium that causes necrotic enteritis and other severe infections in animals, as well as food poisoning in humans. In this study, we introduce a framework consisting of rpoB sequence typing (RSTing) and network analysis to investigate the evolutionary trajectories of C. perfringens. By analyzing 319 rpoB sequences—300 from public databases and 19 newly sequenced isolates from chicken and cattle sources—we identified 84 rpoB sequence types (RSTs). Among them, the early emerging RST 1-1 was the most prevalent (21.3%), while the putative ancestral type, RST 0, was the fifth most common (4.7%). The high RST diversity and the predominance of RST 1-1, mainly from chickens, suggest that chickens may serve as an important reservoir. By integrating virulence gene profiling, MLST, and comparative genomics, we separated identical RSTs into distinct genotypes and uncovered genomic evidence of possible interspecies transmission between chickens and cattle, two major food-producing species. These findings indicate that RSTing provides a useful complementary approach to investigating the evolutionary and epidemiological dynamics of C. perfringens. Full article