Search Results (94)

Avian pathogenic Escherichia coli (APEC) is highly infective in poultry, causing significant economic losses to the poultry industry. As an extraintestinal pathogenic strain, adherence is a critical step in the infection. The functions of several adhesins, including type I, P, and Curli fimbriae, have been extensively studied. However, the roles of other adhesins, like Sfm, remain largely unexplored. Sfm is widely present in E. coli. Although the Sfm cluster is an ortholog of the fim gene cluster of Salmonella type I fimbriae, the biological function of Sfm in APEC has not yet been elucidated. To investigate whether Sfm in APEC CE129 plays a role in virulence, in this study, we constructed recombinant strains by expressing Sfm in the fimbriae-deficient strain SE5000. Additionally, a CE129 sfmA mutant strain was constructed. The resulting changes in adherence, biofilm formation, resistance to macrophage phagocytosis, and resistance to serum bactericidal ability were observed. The adherence ability of CE129ΔsfmA was reduced by 41%. HD-11 cells demonstrated a 30% increase in the phagocytosis of CE129ΔsfmA, and a 50% reduction in SE5000 (pBR322-sfm). The sfm deletion mutant showed a 23.9% reduction in the resistance to serum bactericidal ability, while SE5000 (pBR322-sfm) displayed a 32% increase. SE5000 (pBR322-sfm) exhibited a 34% increase in biofilm formation, and CE129ΔsfmA demonstrated a 21% decrease. Real-time PCR was employed to examine the impact of Sfm deletion on the transcription level of key virulence factors (fimA, fliC, papC, tsh, ompA, and iss). The results indicated that Sfm in CE129 is closely associated with bacterial adherence and survivability, contributing to biofilm formation and influencing the expression of key virulence factors. This study yields initial insight into the functional roles of Sfm in APEC and provides a foundation for the effective control of E. coli in the poultry industry. Full article

Background: The combined diphtheria–tetanus–acellular pertussis (three-component), Haemophilus influenzae type b (Hib, conjugate), and ACYW135 meningococcal (conjugate) vaccine (DTaP-Hib-MCV4) offers a promising alternative to single-component vaccines, potentially simplifying immunization schedules and improving vaccination coverage. Methods: We evaluated the safety, immunogenicity, and protective efficacy of DTaP-Hib-MCV4 in animal models. Acute and long-term toxicity studies were conducted in Sprague-Dawley (SD) rats with equal numbers of male and female animals. Immunogenicity was assessed in female NIH mice and SD rats using a three-dose regimen at 14-day intervals. Orbital blood was collected 14 days post-immunization to measure IgG titers against pertussis, diphtheria, tetanus, Hib, and meningococcal antigens. The protective efficacy was determined using potency tests for the pertussis, diphtheria, and tetanus components; passive protection studies for Hib; and serum bactericidal antibody (SBA) titers against A/C/Y/W135 meningococcal serogroups. Results: Acute and repeated-dose toxicity studies in SD rats showed no signs of abnormal toxicity or irritation at either high (three doses/rat) or low (one dose/rat) doses levels. The no-observed-adverse-effect level (NOAEL) for DTaP-Hib-MCV4 was established at three doses/rat after 8 weeks of repeated intramuscular administration and a 4-week recovery period. Specific IgG antibodies against all the vaccine components were detected in animal sera at both one and three doses/rat, with no evidence of immunotoxicity. Following two-dose primary immunization in murine models, the combined vaccine elicited robust antigen-specific antibody responses, with geometric mean titers (GMTs) as follows: 1,280,000 for pertussis toxin (PT); 761,093 for filamentous hemagglutinin (FHA); 1,159,326 for pertactin (PRN); 1,659,955 for diphtheria toxoid (DT); 1,522,185 for tetanus toxoid (TT); 99 for Haemophilus influenzae type b (Hib); and 25,600, 33,199, 8300, and 9051 for serogroups A, C, Y, and W135 of Neisseria meningitidis, respectively. In the rat models, three-dose primary immunization also elicited robust antigen-specific antibody responses. Protection studies demonstrated efficacy against pertussis, tetanus toxin, and diphtheria toxin challenges. In the Hib challenge study, none of the 10 animals given anti-DTaP-Hib-MCV4 antiserum developed bacteremia after the live Hib challenge (vs. 5814/0.1 mL in the negative control, p < 0.001). In addition, the SBA titers against meningococcal serogroups exceeded the protective threshold (≥1:8) in 92.2% of the immunized mice and 100% of the immunized rats. Crucially, the combined vaccine induced potent immune responses and protective efficacy, with antibody levels and protection against each component antigen comparable to or greater than those of the individual components: DTaP, Hib, and MCV4. Conclusions: These findings demonstrate that the DTaP-Hib-MCV4 combined vaccine is both safe and immunogenic, supporting its potential as a viable alternative to individual vaccines. This combined vaccine may streamline immunization programs and enhance vaccination coverage. Full article

Carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp), a pathogen causing severe nosocomial infections and high mortality rates, is increasingly becoming a serious global public health threat. Capsular polysaccharide (CPS), a major virulence factor of hvKp, can be enzymatically degraded by bacteriophage-derived depolymerases. However, to our knowledge, depolymerases targeting K. pneumoniae K54-type strains have rarely been identified. Here, we identified and characterized three novel capsule depolymerases, Dep_C, Dep_Y, and Dep_Z, derived from three different K. pneumoniae phages, which retained robust activity across a broad pH range (pH 3.0–12.0) and demonstrated thermal stability up to 50 °C. These depolymerases could efficiently digest the CPS of K. pneumoniae K54-serotype strains, significantly inhibit biofilm formation, and remove their mature biofilms. Although no bactericidal activity was detected, these depolymerases rendered host bacteria susceptible to serum complement-mediated killing. We further demonstrate that Dep_C, Dep_Y, and Dep_Z can effectively and significantly prolong the survival time of mice in a pneumonia model infected with K54-type K. pneumoniae and reduce the colonization and virulence of the bacteria in the mice. These findings indicate that depolymerases Dep_C, Dep_Y, and Dep_Z could increase bacterial susceptibility to host immune responses of hvKp to the host through their degradation effect on the CPS. In conclusion, our study demonstrates that the three capsule depolymerases are promising antivirulent agents to combat CR-hvKp infections. Full article

Background/Objectives: Accurate detection of periprosthetic joint infection (PJI) in patients with inflammatory arthritis (IA), including rheumatoid arthritis (RA), remains challenging due to overlapping inflammatory parameters and the influence of immunosuppressive regimens. Methods: A narrative review was conducted using PubMed/MEDLINE (2010–2025). Search terms included “periprosthetic joint infection”, “inflammatory arthritis”, “rheumatoid arthritis”, “diagnosis”, “biomarkers”, “synovial fluid”, and “immunosuppression”. Eventually, 50 studies were included. Results: IA patients diagnosed with PJI are more frequently younger, female, and present with a higher burden of comorbidities and an increased rate of false-positive histological findings and culture-negative infections. Standard biomarkers, such as serum C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), as well as synovial fluid white blood cell count and polymorphonuclear leukocyte percentage, have a low to moderate value for diagnosing PJI in patients with IA. Optimal thresholds for these tests differ from those recommended by the Musculoskeletal Infection Society (MSIS). Alpha-defensin has demonstrated superior diagnostic performance among synovial fluid biomarkers included in MSIS criteria. Novel markers, such as serum bactericidal permeability-increasing protein (BPI) and neutrophil elastase-2 (ELA-2), as well as synovial C-reactive protein and calprotectin, along with molecular techniques like polymerase chain reaction (PCR), are showing increasing potential. Conclusions: Disease and treatment-related confounders hinder PJI diagnosis in IA. Adjusted thresholds and IA-specific approaches are needed. Further research should validate emerging biomarkers, among which BPI, ELA-2, and synovial CRP show the greatest diagnostic potential and guide perioperative immunosuppressive strategies. Full article

This study investigates the health benefits of supplementing Asian seabass diets with hot water crude extract from the sea lettuce Ulva rigida (Ur-HWCE). The extract’s proximate composition consists of 57.63% carbohydrates, 6.75% protein, 31.96% ash, and 6.01% sulfate polysaccharides, as confirmed by FTIR spectrum analysis. It also exhibits significant antioxidant properties, including total antioxidants, ABTS, DPPH, and reducing power. The study involved four groups fed Ur-HWCE at 0.5, 1.0, and 5 g/kg compared to a control group, with feed prepared daily and given twice at 5% of body weight for 4 weeks. Ur-HWCE supplementation did not negatively impact growth performance. It significantly upregulated insulin-like growth factor 1 (igf1) in the brain and liver, enhancing growth processes. Ur-HWCE reduced oxidative stress markers, such as malondialdehyde (MDA). Enhanced immune responses were observed, including increased bactericidal activity, serum IgM levels, and the upregulation of immune-related genes (dcs, c3, ighm, lyz, il8, il10). Gut microbiota analyses showed increased beneficial aerobic and natural probiotic Bacillus spp., particularly Bacillus amyloliquefaciens, enhancing gut health by reducing pathogenic bacteria. Blood biochemical parameters remained stable, and no histopathological alterations were found in the liver and intestine tissues, confirming the supplement’s safety. Fish fed with Ur-HWCE showed significantly higher survival rates and relative percent survival (RPS) against Vibrio vulnificus AAHM-VV2312 compared to the control group, demonstrating improved disease resistance. The study concludes that Ur-HWCE is a promising dietary supplement for enhancing the health, growth, and disease resistance of Asian seabass, supporting its potential in sustainable aquaculture practices. Full article

Avian pathogenic Escherichia coli (APEC) causes bloodstream infections mainly by resisting the bactericidal action of host serum. Although various protein and polysaccharide factors involved in serum resistance have been identified, the role of small non-coding RNA (sRNA) in serum resistance has rarely been studied. The sRNA RyhB contributes to serum resistance in APEC, but the regulation mechanism of RyhB to serum resistance-related targets remains unknown. Here, we studied the regulatory mechanism of RyhB on capsule synthesis and how RyhB regulates serum resistance, macrophage phagocytosis resistance, and pathogenicity to natural hosts by regulating capsule synthesis. The results showed that RyhB upregulates capsular synthesis by interacting with the promoter regions of the capsule gene cluster and activating the translation of the capsule. The deletion of ryhB and/or neu reduced the ability of resistance to serum, macrophage phagocytosis, and pathogenicity of APEC in ducks. It can be concluded that RyhB directly upregulates the expression of capsular gene cluster and capsular synthesis and then indirectly promotes resistance to serum and macrophage phagocytosis and pathogenicity to ducks. Full article

Cefquinome is used to treat septicemia caused by Escherichia coli (E. coli) and respiratory infections caused by Streptococcus equi subsp. zooepidemicus in foals. However, studies reporting the use of cefquinome to target E. coli as pathogens of sepsis are lacking. Therefore, this study aimed to determine the optimal dosage regimen for cefquinome against E. coli using a PK/PD model. After the administration of 1 mg/kg cefquinome (intramuscularly or intravenously), blood samples were collected at different time points to determine the serum concentration of cefquinome via HPLC. The pharmacokinetic parameters were evaluated via NCA (WinNonlin 5.2.1 software). The main pharmacokinetic parameters of cefquinome in foals were as follows: after intravenous administration, the elimination half-life (T_1/2β) was 2.35 h, the area under the curve (AUC_0–last) was 12.33 μg·h/mL, the mean residence time (MRT_0–last) was 2.67 h, and the clearance rate (CL) was 0.09 L/h/kg. After intramuscular administration, the peak concentration (C_max) was 0.89 μg/mL, the time to reach the maximum serum concentration (T_max) was 2.16 h, T_1/2β was 4.16 h, AUC_0–last was 5.41 μg·h/mL, MRT_0–last was 4.92 h, CL was 0.15 L/h/kg, and the absolute bioavailability (F) was 43.86%. An inhibitory sigmoid Emax model was used to integrate the PK/PD indices with ex vivo antimicrobial effects to identify pharmacodynamic targets (PDTs). According to the dose calculation formula, the doses of intramuscularly administered cefquinome required to achieve bacteriostatic effects, bactericidal effects, and bactericidal elimination were 1.10, 1.66, and 2.28 mg/kg, respectively. However, further studies are warranted to verify the therapeutic efficacy of cefquinome in clinical settings. Full article

Serum bactericidal assay (SBA) is a functional assay that evaluates infection- and vaccine-induced neutralizing antibodies representing the serological correlate of protection against Neisseria meningitidis. However, it is time consuming due to its readout using the enumeration of colony-forming units (CFUs), making this conventional SBA (C-SBA) difficult for large-scale use. We developed a new SBA method that takes advantage of a bioluminescence N. meningitidis serogroup B (BioLux-SBA). The assay development steps involved the human complement source validation, the setup of the optimal incubation time, and the assessment of intra-day and inter-day variability. BioLux-SBA was then compared to C-SBA using a serum collection of Norman children vaccinated in 2011 with MenBvac, an OMV meningococcal vaccine. While a conventional approach requests 48 h of work to test 24 sera per day, BioLux-SBA takes only 5 h to test 96 sera per day. The SBA titers (n = 10) correlated with R² of 0.98 (p-value < 0.0001). The deposition of terminal complement components (C5b-C9) measured by flow cytometry on the bacterial surface well correlated with BioLux SBA titers. This high-throughput method to evaluate the immunogenicity of meningococcal vaccines appears to be a reliable method for an OMV meningococcal B vaccine and requires further assessment in other laboratories and against other meningococcal vaccines. Full article

Today, most anti-acne treatments employ topical and systemic antibiotics such as erythromycin and clindamycin, which induce cutaneous dysbiosis with adverse side effects to the skin’s normal microbiota, consequently leading to the emergence of antimicrobial resistance. In our quest to discover natural anti-acne bioactives as alternatives, we undertook a research program with the aim to identify a new blend of active ingredients based on the monoterpene phenol moiety. Within this program, we evaluated the in vitro anti-acne efficacy of thymol, Curcuma turmerones and their patented combination “Acnocure” in a cosmetic formulation. The minimum inhibitory concentration (MIC) of Acnocure against C. acnes (ATCC 6919), S. aureus (ATCC 6538), S. epidermidis (ATCC 12228) and C. freneyi (CIP 52.16) was determined to be 0.32, 0.26, 0.47 and 0.11 mg/mL, respectively. In the time-kill curve study against C. acnes, Acnocure, containing thymol 0.25% and 0.1% Curcuma turmerone as well as thymol 0.1% and 0.1% Curcuma turmerone in a cosmetic simplex formulation, demonstrated rapid bactericidal activity with a 4.7 log reduction at pH 5.5, occurring within just two hours of the study and lasting for over 24 h. The killing efficacy was similar to our cosmetic reference benchmark, Effaclar DUO serum, used in the same study. Additionally, thymol, Curcuma turmerones and Acnocure were evaluated in an anti-inflammatory efficacy assay in lipopolysaccharide (LPS)-primed U937 macrophages model and demonstrated moderate inhibition of interleukin-1β (IL-1β) at 100 µg/mL and significant inhibition of prostaglandin E-2 (PGE-2) at 1 µg/mL, respectively. Further evidence gathered on thymol and Curcuma turmerones in an IL-1α-stimulated dermal fibroblast model showed >90% inhibition of PGE-2 release between 2 µg/mL and 30 µg/mL concentrations. These promising results position Acnocure as a natural alternative for the replacement of synthetic corticosteroids and antibiotics with potent anti-acne skincare properties. Full article

To investigate the anti-bacterial and anti-inflammatory properties of sophoridine and elucidate its mechanism of action, we carried out both in vitro and in vivo experiments. Multiple bacterial strains were utilized to determine the effective concentration of sophoridine in antibacterial and bactericidal assays. Subsequently, LPS-stimulated RAW264.7 cells and E. coli-challenged BALB/c mice models were employed to evaluate the production of inflammatory cytokines. Our results showed that sophoridine concentrations exceeding 5.12 mg/mL significantly inhibited cell viability, while 0.32 mg/mL of sophoridine demonstrated the optimal anti-inflammatory activity at 12 h. In E. coli-induced diarrheal mice, doses of 15, 30, and 60 mg/kg BW of sophoridine alleviated fecal occult blood and exhibited anti-inflammatory effects by reducing the level of serum TNF-α, IL-1β, and IL-6 levels, increasing serum IL-10, and inhibiting leucocyte infiltration in the duodenum. Notably, 15 mg/kg BW of sophoridine effectively decreased the mRNA and protein expression of NF-κB p65. These findings suggest that sophoridine has promising potential for the treatment of diarrhea through its anti-inflammatory effects mediated by the inhibition of NF-κB activation. Full article

The antimicrobial peptide P6.2 was previously de novo designed as an alpha helix cationic amphipathic molecule. In previous work, we have shown that this peptide displayed significant antimicrobial activity against both Gram-positive (Staphylococcus aureus) and Gram-negative (Pseudomonas aeruginosa) bacteria. However, while P6.2 lacked biofilm-inhibiting properties against the P. aeruginosa strain PA01, it displayed anti-inflammatory effects in a murine acute lung infection model challenged with this pathogen. In this work, the peptide P6.2 antimicrobial activity and its possible synergy with meropenem were evaluated both in vitro and in vivo using a Galleria mellonella infection model against a carbapenem-resistant KPC-producing clinical isolate of P. aeruginosa. Firstly, the cytotoxic effect of the peptide on A549 and RAW264.7 cell lines was assayed, showing no cytotoxicity at 64 µg/mL and below. Then, the MIC (minimal inhibitory concentration) and bactericidal effect against the carbapenemase-producing strain P. aeruginosa M13513 strain were determined. P6.2 showed a MIC between 32 and 64 µg/mL, and a rapid bactericidal activity against this strain (less than 45 min). The peptide stability at different temperatures and in bovine serum at 37 °C was also analyzed, showing good stability and almost no degradation after 15 min of incubation at 100 °C or 24 h at 37 °C in serum, respectively. The antibiofilm activity was also evaluated, and although the peptide did not show biofilm inhibitory activity, it did demonstrate biofilm disruptive activity, together with bactericidal activity inside the pre-formed biofilm. The possible synergistic effect with the carbapenem meropenem was then analyzed in vitro by killing kinetics, revealing a synergistic interaction between P6.2 and the antibiotic against this strain. Finally, P6.2 was evaluated in vivo in the Galleria mellonella larvae infection model. Interestingly, in G. mellonella, P6.2 alone did not completely clear the infection caused by P. aeruginosa M13513. However, when combined with meropenem, P6.2 demonstrated a synergistic effect, leading to increased survival rates in infected larvae. The results presented here highlight the potential that this peptide displays when used in combination with carbapenems against a clinically relevant KPC-producing P. aeruginosa. Full article

Seeds are a major source of contamination by foodborne pathogens such as Salmonella typhimurium, Escherichia coli, and Listeria monocytogenes, significantly increasing the risk of foodborne diseases associated with fresh produce like sprouts. In this study, we described novel endolysins and the engineered variants that exhibited potent bactericidal activity against these pathogens. These endolysins demonstrated strong bactericidal effects independently of outer membrane permeabilizers, effectively killing S. typhimurium, E. coli, and L. monocytogenes to undetectable levels (>4-log kill) at concentrations as low as 12.5 μg/mL. The enzymes retained their activity in complex environments, such as a wide range of temperatures (4–100 °C), pH values (4–10), serum concentrations (0–50%), and sodium chloride concentrations (0–500 mM). Furthermore, their rapid bactericidal kinetics, excellent storage stability (>18 months), and broad-spectrum antimicrobial activity enhanced their potential for application. These endolysins remained effective against stationary-phase bacteria and biofilm-forming bacteria, achieving more than 99% biofilm eradication at 200 μg/mL. Notably, at concentrations as low as 50 μg/mL, these enzymes completely decontaminated foodborne pathogens in a mung bean seed model contaminated with 4–5 log CFU of bacteria. This study is the first to report the successful use of lysins to control both Gram-negative and Gram-positive pathogens on mung bean seeds. Full article

Background: Typhoid and paratyphoid fevers represent a global health burden, especially in Southern Asia, exacerbated by the increase in antimicrobial resistance. While vaccines against Salmonella Typhi have been successfully introduced, a vaccine against S. Paratyphi A is not available, yet. Efforts to develop an effective vaccine targeting both Salmonella serovars are currently ongoing. GVGH is developing a bivalent vaccine constituted by the Vi-CRM₁₉₇ typhoid conjugate vaccine (TCV), and the Salmonella Paratyphi A O-antigen (O:2), also conjugated to the CRM₁₉₇ carrier protein (O:2-CRM₁₉₇). In this work we have characterized a panel of S. Paratyphi A clinical isolates from endemic regions, differing in terms of their O:2 structural features. Methods: Rabbits were immunized with the S. Paratyphi A component of the vaccine candidate and the resulting sera were tested for their ability to bind and kill the isolates using flow cytometry and luminescence-based serum bactericidal assay (L-SBA). Results: The O:2-CRM₁₉₇ glycoconjugate induced a functional immune response in rabbits, effectively binding and killing a diverse panel of clinical isolates. The sera demonstrated bactericidal activity independent of the O:2 structural variations, including differences in O-acetylation and glucosylation levels. Additionally, the study found that the O:2-CRM₁₉₇ conjugate’s adsorption to Alhydrogel did not significantly impact its immunogenicity or bactericidal efficacy. Conclusions: The O:2-CRM₁₉₇ component of the bivalent vaccine candidate shows promise in providing broad protection against S. Paratyphi A isolates, regardless of their O-antigen structural variations. The ongoing clinical studies on human sera are expected to confirm these results. Full article

Bacterial killing is important for recovering from infection. Pasteurella multocida is a key bacterial pathogen causing swine respiratory disease and is associated with substantial mortality. Antimicrobial therapy remains an important therapeutic intervention for treating infected animals. Pradofloxacin (fluoroquinolone) is the most recently approved antimicrobial agent for treating pigs with swine respiratory disease. We compared in vitro killing of swine P. multocida strains by pradofloxacin in comparison to ceftiofur, enrofloxacin, florfenicol, marbofloxacin, tildipirosin, tilmicosin, and tulathromycin over a range of bacterial densities and four clinically relevant drug concentrations. Pradofloxacin killed 92–96.9% of cells across 10⁶–10⁸ cfu/mL densities at the mutant prevention drug concentration following 2–24 h of drug exposure, 96.9–98.9% of cells across 10⁶–10⁹ cfu/mL at the maximum serum drug concentration following 30 min of drug exposure, increasing to 99.9–100% kill following 12–24 h of drug exposure. At the maximum tissue drug concentration and against bacterial densities of 10⁶–10⁹ cfu/mL, pradofloxacin killed 91.3–99.8% of cells following 2 h of drug exposure, which increased to 99.9–100% kill following 12–24 h of drug exposure. Pradofloxacin was rapidly bactericidal across a range of bacterial densities and at clinically relevant drug concentrations. Pradofloxacin will be an important antibiotic for treating pigs with swine respiratory disease and where clinically indicated. Full article

Shigellosis represents a significant global health concern particularly affecting children under 5 years in low- and middle-income countries (LMICs) and is associated with stunting and antimicrobial resistance. There is a critical need for an effective vaccine offering broad protection against the different Shigella serotypes. A correlate of protection has not yet been established but there is a general consensus about the relevant role of anti-O-Antigen-specific IgG and its functionality evaluated by the Serum Bactericidal Assay (SBA). This study aims to characterize a high-throughput luminescence-based SBA (L-SBA) against seven widespread Shigella serotypes. The assay was previously developed and characterized for S. sonnei and S. flexneri 1b, 2a, and 3a and has now been refined and extended to an additional five serotypes (S. flexneri 4a, 5b, 6, X, and Y). The characterization of the assay with human sera confirmed the repeatability, intermediate precision, and linearity of the assays; both homologous and heterologous specificity were verified as well; finally, limit of detection and quantification were established for all assays. Moreover, different sources of baby rabbit complement showed to have no impact on L-SBA output. The results obtained confirm the possibility of extending the L-SBA to multiple Shigella serotypes, thus enabling analysis of the functional response induced by natural exposure to Shigella in epidemiological studies and the ability of candidate vaccines to elicit cross-functional antibodies able to kill a broad panel of prevalent Shigella serotypes in a complement-mediated fashion. Full article