Antimicrobial resistance, particularly in healthcare-associated infections (HAIs), highlights the need for more effective and sustainable disinfection strategies. This study aimed to evaluated the bactericidal effects of two conventional biocides-benzalkonium chloride (BAC) and peracetic acid (PAA)-and two phytochemicals-salicylic acid (SAL) and eugenol (EUG)-againstEscherichia coli and Staphylococcus aureus, focusing on potential synergistic interactions. Antimicrobial efficacy was assessed through standardized dose- and time-response assays (EN 1276) and kinetic modeling using the Chick-Watson and Weibull equations. BAC and PAA achieved complete bacterial inactivation at 3 mg/L and 1 mg/L, respectively, within 5-15 min, confirming rapid and potent activity. Conversely, SAL and EUG required substantially higher concentrations (500-1700 mg/L) to achieve total loss of culturability. Chick-Watson modeling demonstrated high disinfection rate coefficients for BAC and PAA, while SAL and EUG exhibited markedly lower values. Synergy testing revealed a strong interaction between BAC and EUG, with fractional bactericidal concentration indices of 0.350 for E. coli and 0.309 for S. aureus, whereas other combinations were additive. Weibull modeling further indicated that bacterial tolerance was dependent on compound type and concentration, with S. aureus generally more susceptible than E. coli. These findings collectively confirm the enhanced efficacy of selected biocide/phytochemical combinations, allowing lower concentrations and promoting more sustainable antimicrobial practices.

Managing bacterial infections and the spread of microbial resistance is one of the most critical and complex tasks of modern healthcare infrastructures. Antiseptics and disinfectants such as biocides play a significant role in controlling microbial resistance by reducing the microbial load on surfaces, skin, and environments, thereby limiting the opportunity for pathogens to proliferate and develop resistance. Herein, we tested the different interactions of quaternary ammonium compound (QAC)-based biocide compositions in pursuit of a better antimicrobial performance. An extensive microbiological analysis was conducted for 12 selected compositions of various combinations of mono-QACs, bis-QACs, and alcohols on 17 strains of bacteria of the ESKAPEE group and fungi, including 11 clinical highly resistant varieties, highlighting synergistic or additive dynamics. The evaluation showed noticeable improvements in activity, with up to 16-fold MBC and 32-fold MBEC reductions for alcohol-based compositions of lead QAC. Moreover, synergistic interactions were detected and confirmed via an optimized checkerboard assay for pyridinium QAC combinations against planktonic Gram-positive S. aureus with a fractional inhibitory concentration index (FICI) and fractional bactericidal concentration index (FBCI) of 0.39-0.5 and Gram-negative A. baumannii biofilms. The studied biocides demonstrated the long-term preservation of antimicrobial efficiency without resistance development during a 40-day period and do not induce QAC-associated cross-resistance for four commercially available antibiotics with similar mechanisms of action.

Susceptibility testing of clinical multidrug-resistant (MDR) and reference P. aeruginosa strains was performed using the standard twofold serial dilution method. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of antiseptics were determined. MIC and MBC values were also interpreted as the bacteriostatic index of antiseptic activity (BSIAA) and the bactericidal index of antiseptic activity (BCIAA). The ability of strains to form biofilms, the inhibition of biofilm formation, and the destruction of mature biofilms under the influence of bacteriostatic, bactericidal, and ½ of the initial antiseptic concentration were modeled using Christensen’s test. Antiseptics from the detergent group, decamethoxine (0.1 and 0.02%) and polyhexanide (0.1%), demonstrated the highest antimicrobial activity. Their bacteriostatic concentrations were 63.2 ± 5.2 μg/mL and 68.7 ± 4.2 μg/mL, respectively. The ranking of antiseptics by bacteriostatic efficacy was: decamethoxine > polyhexanide > octenidine > miramistin > chlorhexidine. The highest BSIAA values were observed for povidone-iodine 10%, decamethoxine 0.1%, octenidine 0.1%, and polyhexanide 0.1%. The highest bactericidal IAA values were found for povidone-iodine 10%, decamethoxine 0.1%, octenidine 0.1%, and polyhexanide 0.1%. Miramistin 0.01% was deemed insufficiently effective. Polyhexanide exhibited the highest bactericidal activity, with a BCIAA to BSIAA ratio of 0.88. For all other antiseptics, this ratio ranged from 0.5 to 0.6. All tested strains exhibited a high capacity for biofilm formation. All antiseptics significantly inhibited biofilm formation. Octenidine had the strongest effect on immature biofilms, reducing their formation by 28.5% (p < 0.0001). The MICs of most antiseptics stimulated mature biofilm development. The bacteriostatic concentration of octenidine led to the eradication of biofilm by 4.7% (p < 0.001) compared to the control. The MBC of most antiseptics (except chlorhexidine) eradicated mature biofilms by 4–30.6%, whereas chlorhexidine stimulated mature biofilm growth by 17.9%. All antiseptics, at half their initial concentration, partially eradicated MDR Pseudomonas biofilms by 11.3–42.4%. Analysing the effect of octenidine at different concentrations and stages of biofilm formation highlights its strong activity against P. aeruginosa biofilms. Our findings underscore the importance of carefully monitoring P. aeruginosa isolates for antiseptic susceptibility. This approach can help prevent the development of selective conditions that promote resistant microorganisms and limit their spread.

Carbon-based materials have emerged as promising biomaterials due to their biocompatibility and inherent antibacterial properties. Carbyne, a unique allotrope of carbon, characterized by sp-hybridized carbons forming alternating single and triple bonds, exhibits exceptional toughness. Herein, we explore the potential of carbyne-enriched plasma coatings for antibacterial applications in conjunction with micro- and nano-textured polymeric surfaces. We investigate and characterize carbyne-enriched plasma coatings onto superhydrophilic or superhydrophobic poly (methyl methacrylate) (PMMA) plasma micro-nanotextured surfaces. Our analysis evaluates the wetting properties and durability of these surfaces, particularly in liquid immersion conditions. The integration of carbyne-enriched plasma coatings serves a dual purpose: it enhances the chemical bactericidal action and protects surface micro-nanostructures from deformation due to capillary forces thanks to the material’s innate toughness. The results show that the micro-nanotextured and carbyne-enriched coated PMMA surfaces exhibit a significant bactericidal activity as expressed by a bactericidal index of approximately 50%, owing to the combined effect of both the surface topography and the plasma-deposited carbyne coating.

Enterococcus faecalis is strongly associated with secondary/persistent root canal infections, being the most prevalent bacterium in cases of apical periodontitis in previously treated teeth. This study was elaborated to evaluate the anti-planktonic, antibiofilm, and synergistic effects of Nasturtium officinale and Psidium guajava hydroethanolic extracts against standard and clinical strains of E. faecalis. Firstly, the N. officinale extract was prepared from watercress leaves, and P. guajava extract was prepared from guava tree leaf shoots. Then, the content of soluble solids was quantified in both. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of the isolated N. officinale and P. guajava extracts for each bacterial strain were determined using the broth microdilution method, following the Clinical and Laboratory Standards Institute (CLSI) guideline M7-A9. The MTT assay was used to evaluate the antibiofilm activity, and the fractional bactericidal concentration index (FBCI) was utilized to evaluate the synergistic effect of the N. officinale and P. guajava extracts using the checkerboard technique. Again, the MTT assay was used to evaluate the antibiofilm activity of the combined extracts this time. The data were subjected to statistical analysis using ANOVA and Tukey’s test, with a significance level of p ≤ 0.05. It was found that the soluble solid content of N. officinale was 50 mg/mL, and of P. guajava was 33.5 mg/mL. The MBC value of N. officinale was 12.5 mg/mL, and of P. guajava was 0.52 mg/mL against all the tested strains of E. faecalis. The combined 0.1 mg/mL N. officinale + 0.1 mg/mL P. guajava, and 0.1 mg/mL N. officinale + 0.5 mg/mL P. guajava hydroethanolic extracts effectively reduced the biofilm formation of the standard and clinical strain 4 of E. faecalis. Therefore, these combined extracts may be considered as endodontic irrigants in future studies.

The misuse of antibiotics in livestock farming has led to the emergence of antibiotic-resistant bacteria, which pose a serious threat to global animal and human health. Essential oils extracted from the leaves of Ocimum basilicum, Perilla frutescens and Mentha spicata contain bioactive compounds with bactericidal properties. This study aimed to evaluate the bactericidal activity of Ocimum basilicum, Perilla frutescens and Mentha spicata essential oils and their combination with the antibiotic norfloxacin, against antibiotic-resistant Salmonella spp. isolated from diseased chickens. The antibiotic susceptibility testing of the isolated bacteria was conducted using the disc diffusion method. The bactericidal efficacy of essential oils and antibiotics was assessed by determining the minimum bactericidal concentration. The interaction between essential oils and antibiotics was analysed using the fractional bactericidal concentration index through the microdilution chequerboardtechnique. Salmonella typhimurium and Salmonella enteritidis were recovered from the organs of infected chickens. Isolated Salmonella spp. displayed significant resistance to amoxicillin, ampicillin, streptomycin and cefuroxime. The essential oils of Ocimum basilicum and Mentha spicata demonstrated similar minimum bactericidal concentration (MBC) values of 8,000 µg/mL, while the essential oil of Perilla frutescens showed a higher MBC value of 10,000 µg/mL. Analysis of the interaction between these essential oils and norfloxacin indicated that both Ocimum basilicum and Mentha spicata exhibited a synergistic effect, whereas Perilla frutescens exhibited an additive effect when combined with norfloxacin in eradicating Salmonella spp. The study concluded that combining essential oils from three herbs with the antibiotic norfloxacin was highly effective in fighting antibiotic-resistant Salmonella spp. isolated from infected chickens.

This in vitro experimental study examines and evaluates the antimicrobial and synergistic effects of the ethanolic extract of six plants: Biancaea sappan (L.) Tod, Bauhinia malabarica Roxb, Carthamus tinctorius L., Derris scandens (Roxb.) Benth, Hibiscus sabdariffa L., and Piper nigrum L. against common microbial species representing gram-positive, gram-negative bacteria, and fungi, consisting of Staphylococcus aureus, Escherichia coli, Staphylococcus epidermidis, Pseudomonas aeruginosa, and Candida albicans. The plants were extracted using 90% ethanol. According to the standard method of agar diffusion assay, the micro-dilution method for minimal inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined. This study found that among the six plants, only B. sappan and B. malabarica exhibited moderate inhibitory effects against S. aureus and S. epidermidis. B. sappan had MIC values of 250 µg/ mL and 125 µg/ mL, respectively, and B. malabarica showed MIC values of 62.50 µg/ mL and 31.25 µg/ mL, respectively. The synergistic effects of a combination of B. sappan and B. malabarica extracts at a ratio of 25:75 were analyzed, and it was found that the combination inhibited S. aureus and S. epidermidis with MIC values of 250 µg/ mL and 125 µg/ mL, respectively. The fractional inhibitory concentration index (FICI) and the fractional bactericidal concentration index (FBCI) indicated antagonistic or synergistic effects of the combination, with FICI and FBCI values of 2.5–5.0 for both B. sappan and B. malabarica extracts in the 25:75 mixture. In conclusion, single plant ethanolic extracts of B. sappan and B. malabarica possess potent antimicrobial activity to varying degrees. However, the antimicrobial potency of the 25:75 ratio mixture of these extracts was shown to decrease against the same organisms, with in vitro antimicrobial activity and antagonistic effects observed only against the tested gram-positive bacteria.

IntroductionDiabetes mellitus is a chronic metabolic disorder characterized by persistent hyperglycemia due to impaired insulin production or utilization, leading to severe health complications. Diabetic foot ulcers (DFUs) represent a major complication, often exacerbated by polymicrobial infections involving Staphylococcus aureus and Acinetobacter baumannii. These pathogens, notorious for their resistance to antibiotics, complicate treatment efforts, especially due to biofilm formation, which enhances bacterial survival and resistance. This study explores the synergistic effects of combining gentamicin, imipenem, and fucoidan, a sulfated polysaccharide with antimicrobial properties, against both planktonic and biofilm forms of S. aureus and A. baumannii.MethodsIsolates of S. aureus and A. baumannii were collected from DFUs and genetically confirmed. Methicillin resistance in S. aureus was identified through disk diffusion and PCR. Biofilm formation, including dual-species biofilms, was analyzed using the microtiter plate method. The antimicrobial efficacy of gentamicin, imipenem, and fucoidan was assessed by determining the minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), minimum biofilm inhibitory concentration (MBIC), and minimum biofilm eradication concentration (MBEC). Synergistic interactions were evaluated using the fractional inhibitory concentration index (FICi) and fractional bactericidal concentration index (FBCi). The expression of biofilm-associated genes (icaA in S. aureus and bap in A. baumannii) was analyzed, and the cytotoxicity of fucoidan was assessed.ResultsThe study revealed that 77.4% of S. aureus and all A. baumannii isolates showed multidrug resistance. Among 837 tested conditions for dual-species biofilm formation, 72 resulted in strong biofilm formation and 67 in moderate biofilm formation. The geometric mean MIC values for gentamicin were 12.2 µg/mL for S. aureus, 22.62 µg/mL for A. baumannii, and 5.87 µg/mL for their co-culture; for imipenem, they were 19.84, 9.18, and 3.70 µg/mL, respectively, and for fucoidan, 48.50, 31.20, and 19.65 µg/mL, respectively. The MBC values for gentamicin were 119.42, 128, and 11.75 µg/mL; for imipenem, they were 48.50, 14.92, and 8 µg/mL; and for fucoidan, they were 88.37, 62.62, and 42.48 µg/mL. The MBIC values were 55.71, 119.42, and 18.66 µg/mL for gentamicin; 68.59, 48.50, and 25.39 µg/mL for imipenem; and 153.89, 101.49, and 53.53 µg/mL for fucoidan. The MBEC values were 315.17, 362.03, and 59.25 µg/mL for gentamicin; 207.93, 157.58, and 74.65 µg/mL for imipenem; and 353.55, 189.46, and 99.19 µg/mL for fucoidan. When cultured in planktonic form, the geometric mean FICi and FBCi values indicated additive effects, while co-culture showed FICi values of ≤ 0.5, suggesting a synergistic interaction. Treatment with gentamicin and fucoidan led to significant downregulation of the icaA and bap genes in both single-species and dual-species biofilms of S. aureus and A. baumannii. The reductions in gene expression were more pronounced in dual-species biofilms compared to single-species biofilms. Additionally, treatment with imipenem and fucoidan also resulted in significant downregulation of these genes in both biofilm types. Cytotoxicity assessments indicated that higher concentrations of fucoidan were toxic, yet no harmful effects were noted at the optimal synergistic concentrations used with antibiotics.ConclusionIn our investigation, we found that combining gentamicin, imipenem, and fucoidan had a synergistic effect on dual-species biofilms of S. aureus and A. baumannii, suggesting potential benefits for treating such infections effectively. This underscores the importance of understanding microbial interactions, antibiotic susceptibility, and biofilm formation in DFUs.

The treatment of acute and chronic wounds with critical colonization by wound pathogens remains a serious problem, which is becoming more and more threatening every year due to the global problem of antibiotic resistance. Aim - to study the antimicrobial activity of modern antiseptic Polyhexanide and antimicrobial biomaterials based on it against most common pathogens of wound infections. Materials and methods. Antimicrobial activity of polyhexanide solution were studied against MLS -resistant clinical isolates of S. aureus, MDR- strains of E. coli, clinical XDR-isolates of K. pneumoniae, P. aeruginosa, and A. baumannii as well as referent strains S. aureus ATCC 25923, E. coli ATCC 25922, K. pneumoniae ATCC 708603, P. aeruginosa ATCC 27853, A. baumannii ATCC ВАА-747. MIC and MBC of polyhexanide were detected by standard dilution method; bactericidal index of antiseptic activity (BC IAA) for 0.1% polyhexamethylen-biguanide solution for wound irrigation (PHMB) was calculated. The bioactivity of wound antimicrobial dressings with polyhexamethylenguanidyn-hydrochoridi (PHMB-HCl) and PHMB was determined by standard disk diffusion method; growth inhibition zones (GIZ) were measured and compared. Results. MLS-resistant strains of S. aureus were the most susceptible to the action of the antiseptic: the MIC and MBC of PHMB were on average 22.69±4.6 μg/ml and 40.55±8.57 μg/ml, respectively. The MIC values against S. aureus was lower than those against E. coli by 1.97 times, against K. pneumoniae – by 1.78 times, against A. baumannii – by 2.97, against P. aeruginosa – 3.9 times. The MBC of PHMB against clinical strains of S. aureus had the lowest values and differed significantly from those against E. coli by 2.21 times, K. pneumoniae by 1.99 times, A. baumannii by 3.4 times, P. aeruginosa by 4.52 times. The bactericidal index of antiseptic activity (BC IAA) of PHMB had the highest values in relation to clinical strains of S. aureus (BC IAA=24.7), K. pneumoniae (BC IAA=12.4) and E. coli (BC IAA=11.2). Wound dressings PHMB-HCl and PHMB demonstrated high antimicrobial properties against reference and clinical strains of S. aureus, E. coli, K. pneumoniae. Reference and clinical strains of P. aeruginosa were the least susceptible to PHMB-based wound dressings. Conclusions. Polyhexanide antiseptic solution for wound irrigation and polyhexanide-containing dressings with PHMB and PHMB-HCl have the high antimicrobial properties in relation to the most common types of wound pathogens. No conflict of interests was declared by the authors.

Background: Due to the rapid emergence of extensively drug-resistant strains of Acinetobacter baumannii worldwide, there is a necessity for greater consideration of the role of preventive vaccines in combating these pathogens.Objective: The study aims to assess the effectiveness of a heat-inactivated whole-cell vaccine against A. baumannii.Methods: Eleven Swiss albino mice were divided into Experimental (n=4), Placebo-controlled (n=4), and Negative Control (n=3) Groups. Clinically isolated extensively drug-resistant A. baumannii species emulsified wih Complete Freund’s adjuvant was used for intramuscular vaccination in the Experimental group in three different phases on 14-day intervals whereas the Placebo-controlled group received phosphatebuffered saline emulsified with CFA in the same schedule and Negative Control group was kept unimmunized. Serum was collected from the tail blood of each mouse on the 10th day after each immunization, by cardiac puncture on the 14th day after the lethal dose from the experimental group, and after the death of the placebo-controlled group.Results: Mice inoculated with heatinactivated whole-cell vaccine survived and showed a higher neutralizing (IgG) antibody response after 2nd (&gt;7-fold titer) and 3rd inoculation (&gt;12-fold titer) whereas all mice from the Placebocontrolled group and Negative Control group did not survive after lethal challenge with different extensively drug-resistant strain of A. baumannii. Sera from the experimental group of mice collected after 3rd inoculation resulted in a higher serum bactericidal killing index after three hours of incubation.Conclusion: These results suggest intramuscular vaccination with heat-inactivated whole-cell vaccine survival after extensively drug-resistant A. baumannii infection.International Journal of Human and Health Sciences Vol. 08 No. 03 Jul’24 Page: 275-282

Antimicrobial tolerance is a significant concern in the food industry, as it poses risks to food safety and public health. To overcome this challenge, synergistic combinations of antimicrobials have emerged as a potential solution. In this study, the combinations of two essential oil constituents (EOCs), namely carvacrol (CAR) and eugenol (EUG), with the quaternary ammonium compounds (QACs) benzalkonium chloride (BAC) and didecyldimethylammonium chloride (DDAC) were evaluated for their antimicrobial effects against Escherichia coli and Bacillus cereus, two common foodborne bacteria. The checkerboard assay was employed to determine the fractional inhibitory concentration index (FICI) and the fractional bactericidal concentration index (FBCI), indicating the presence of bactericidal, but not bacteriostatic, synergy in all QAC-EOC combinations. Bactericidal synergism was clearly supported by Bliss independence analysis. The bactericidal activity of the promising synergistic combinations was further validated by time-kill curves, achieving a >4-log10 reduction of initial bacterial load, which is significant compared to typical industry standards. The combinations containing DDAC showed the highest efficiency, resulting in the eradication of bacterial population in less than 2-4 h. These findings emphasize the importance of considering both bacteriostatic and bactericidal effects when evaluating antimicrobial combinations and the potential of EOC-QAC combinations for sanitization and disinfection in the food industry.

BackgroundThe emergence and rapid spread of multi-drug resistant (MDR) bacterial strains, such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant S. aureus (VRSA), have posed a significant challenge to the medical community due to their ability to form biofilm and develop resistance to common antibiotics. Traditional antibiotics that were once effective in treating bacterial infections are now becoming increasingly ineffective, leading to severe consequences for patient outcomes. This concerning situation has called for urgent research to explore alternative treatment strategies. Recent studies have shown that antimicrobial peptides (AMPs) hold promise as effective agents against biofilm-associated drug-resistant infections as well as to enhance the efficacy of conventional antibiotics. Accordingly, we aimed to investigate the antimicrobial and antibiofilm effects of melittin AMP, both alone and in combination with penicillin and oxacillin, against biofilm-forming MDR-MRSA and -VRSA.MethodsIn this study, we investigated the kinetics of biofilm formation and assessed various parameters related to the antimicrobial and antibiofilm efficacy of melittin and antibiotics, both alone and in combination, against MDR-MRSA and -VRSA. The antimicrobial parameters included the Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC), Fractional Inhibitory Concentration Index (FICi), Fractional Bactericidal Concentration Index (FBCi), and the antibiofilm activity of melittin and antibiotics indicated by the Minimum Biofilm Inhibitory Concentration (MBIC), Minimal Biofilm Eradication Concentration (MBEC), Fractional Biofilm Inhibitory Concentration Index (FBICi), and Fractional Biofilm Eradication Concentration Index (FBECi).ResultsThe MIC results showed that all S. aureus isolates were resistant to penicillin (≥0.25 μg/mL), and 66% of isolates were resistant to oxacillin. The geometric means of the MIC values for penicillin, oxacillin, and melittin were 19.02, 16, and 1.62 μg/ml, respectively, and the geometric means of the MBC values for penicillin, oxacillin, and melittin were 107.63, 49.35, and 5.45 μg/ml, respectively. The study revealed that the combination indexes of melittin-penicillin and melittin-oxacillin, as determined by FIC values against all isolates, were 0.37 and 0.03, respectively. Additionally, melittin-penicillin and melittin-oxacillin exhibited combination indexes based on FBC values against all isolates at 1.145 and 0.711, respectively. Besides, melittin inhibited the biofilm formation of all S. aureus isolates, with MBIC values ranging from 10 to 1.25 μg/mL, and MBEC values ranging from 40 to 10 μg/mL. Generally, the combination indexes of melittin-penicillin and melittin-oxacillin, determined using FBIC values against all isolates, were 0.23 and 0.177, respectively. Moreover, melittin-penicillin and melittin-oxacillin typically had combination indexes based on FBEC values against all isolates at 5 and 2.97, respectively.ConclusionIn conclusion, our study provides evidence that melittin is effective against both planktonik and biofilm forms of MRSA and VRSA and exhibits significant synergistic effects when combined with antibiotics. These results suggest that melittin and antibiotics could be a potential candidate for further investigation for in vivo infections caused by MDR S. aureus. Furthermore, melittin has the potential to restore the efficacy of penicillin and oxacillin antibiotics in the treatment of MDR infections. Applying AMPs, like melittin, to revive beta-lactam antibiotics against MRSA and VRSA is an innovative approach against antibiotic-resistant bacteria. Further research is needed to optimize dosage and understand melittin mechanism and interactions with beta-lactam antibiotics for successful clinical applications.

The necessity for the investigation of novel approaches and strategies for the treatment of multidrug-resistant E. coli related infections becomes more and more essential. Purpose - to investigate and compare the level of antimicrobial and anti-biofilm activity of antiseptic preparations against MDR clinical isolates of E. coli. Materials and methods. In vitro effectiveness of modern antiseptics; octenidine 0.1% (OCT), polyhexanide 0.1% (PHMB), chlorhexidine 0.5% (CHG), miramistin 0.01% (MRM), decamethoxine 0.1% 0.02% (DCM), povidone-iodine 10% (PVP-I), was determined against forty-six polyresistant clinical strains of E. coli. MIC, MBC were found by standard methods, the value of which was interpreted as a bacteriostatic and bactericidal index of antiseptic activity (BS IAA and BC IAA). The effect of antiseptics on the immature biofilm was modelled using the Christensen test. Results. MIC and MBC values were the lowest in DCM and OCT. The highest values of the antiseptic activity index (IAA&gt;4) were determined for the antiseptics PHMB 0.1%, OCT 0.1% and DCM 0.1%. It was found that the feasibility of using MRM at a concentration of 0.01% is questionable as the BS IAA is above the threshold value, while the BC IAA is not. The effectiveness of PVP-I 1% against MDR E. coli was found insufficient. Sub-bacteriostatic concentrations of DCM, CHG, and PHMB reliably inhibited the formation of E. coli biofilms within 24 hours. MRM and PVP-I in sub-bacteriostatic concentrations stimulated biofilm formation. Conclusions. Based on the analysis of all conducted studies, 0.1% and 0.02% DCM, 0.05% CHG, 0.1% OCT, 0.1% PHMB, 10% and 2% PVP-I are the most active against MDR clinical isolates of E. coli. The research was carried out in accordance with the principles of the Helsinki Declaration. The study protocol was approved by the Local Ethics Committee of the participating institution. The informed consent of the patient was obtained for conducting the studies. No conflict of interests was declared by the authors.

The outbreaks caused by Vibrio spp. are a notable threat to the potential growth of the economy of penaeid culture, which is still controlled by the administration of antibiotics. At first, the infected group was subjected to phenotypic bacteriological examination with subsequent molecular identification via 16S rRNA gene sequencing, which confirmed four strains of Vibrio spp., V. atlanticus, V. natriegens, V. alginolyticus, and V. harveyi, from moribund-infected shrimp during mortality events in an Egyptian hatchery. To better understand the defense mechanism of the most effective antibiotic against Vibrio strains, the immune responses were compared and evaluated in infected Litopenaeus vannamei broodstock after being fed 5 mg kg−1 of florfenicol antibiotic, which was first determined through in vitro antibiogram tests. Therefore, our study aimed to determine the immune response of L. vannamei during Vibrio spp. infection in Egyptian hatcheries and after antibiotic medication. The parameters assessed were the total and differential hemocyte count (THC), granular cells (GC), semi-granular cells (SGC), and hyaline cells (HC). As well as the metabolic and immune enzymes: alanine aminotransferases (ALT), aspartate aminotransferases (AST), alkaline phosphatase (ALP), acid phosphatase (ACP), and lysozyme activity; an antioxidant index, such as superoxide dismutase (SOD) and glutathione (GSH); a phagocytic assay; changes in reactive oxygen species (ROS); and bactericidal activity in the hemolymph of the control, infected, and treated groups. Further evaluation of the mRNA expression levels of the prophenoloxidase (LvproPO), toll-like receptor 1 (LvToll1), and haemocyanin (LvHc) genes were performed in the hepatopancreas of the same groups. A significant drop in the THC, GC, SGC, and HC counts, as well as lysozyme and bactericidal activities, phagocytic assay, ROS, SOD, and GSH index, were represented in infected shrimp compared to control shrimp; however, a marked increase in the activity of ALT, AST, ALP, and ACP was observed. These activities were significantly restored in the treated shrimp compared to the infected shrimp. Nevertheless, no significant changes were noted in the transcriptional levels of the LvproPO and LvToll1 genes in the treated shrimp when compared to the infected shrimp; however, a significant suppression of the LvHc gene was noted. Our study aimed to determine the immune response of L. vannamei during Vibrio spp. infection in Egyptian hatcheries and after antibiotic medication. We concluded that florfenicol in medicated feed could be effective in controlling vibriosis and ameliorating the immune response of shrimp.

Antibiotic resistance is increasing alarmingly in all parts of the world. Cinnamomum camphora (Linn.) Presl (C. camphora) is one of the earliest herbal remedies still in use today in traditional medicine. This study aimed to analyze the component of C. camphora grown widelyin Saudi Arabia (Qassim region) using GC-MS. Also, this study evaluates the in vitro antibacterial properties of C. camphora against certain clinical bacteria obtained from hospitals, including multi-drug resistant pathogens. Leaves of C. camphora tree were collected and essential oil was extracted for this study. The extract was subjected to GC-MS analysis. Eight clinical antibiotic-resistant pathogens were used in this study for the following assays: antibiotics susceptibility assay, determination of minimum inhibitory concentration (MIC), determination of minimum bactericidal concentration (MBC) and MIC index (MBC/MIC). The results show that the main components of the essential oil (EO) from the leaves ofC. camphorawere Eucalyptol. The EO had good antibacterial activity against eight clinical antibiotic-resistant pathogens, namely, Pseudomonas aeruginosa, Acinetobacter baumannii (two strains), Klebsiella pneumonia (two strains), Escherichia coli (one strain), Staphylococcus aureus (two strains). These findings may lead to a more complete knowledge of this aromatic plant's antibacterial action against antibiotic-resistant pathogens (in vitro).

With the increasing bacterial resistance to traditional antibiotics, there is an urgent need for the development of alternative drugs or adjuvants of antibiotics to enhance antibacterial efficiency. The combination of antimicrobial peptides (AMPs) and traditional antibiotics is a potential alternative to enhance antibacterial efficiency. In this study, we investigated the synergistic bactericidal effect of AMPs, including chicken (CATH-1,−2,−3, and -B1), mice (CRAMP), and porcine (PMAP-36 and PR-39) in combination with conventional antibiotics containing ampicillin, tetracycline, gentamicin, and erythromycin against Staphylococcus aureus, Salmonella enteritidis, and Escherichia coli. The results showed that the minimum bactericidal concentration (MBC) of CATH-1,−3 and PMAP-36 was lower than 10 μM, indicating that these three AMPs had good bacterial activity against S. aureus, S. enteritidis, and E. coli. Then, the synergistic antibacterial activity of AMPs and antibiotics combination was determined by the fractional bactericidal concentration index (FBCI). The results showed that the FBCI of AMPs (CATH-1,−3 and PMAP-36) and erythromycin was lower than 0.5 against bacterial pathogens, demonstrating that they had a synergistic bactericidal effect. Furthermore, the time-killing kinetics of AMPs (CATH-1,−3 and PMAP-36) in combination with erythromycin showed that they had a continuous killing effect on bacteria within 3 h. Notably, the combination showed lower hemolytic activity and cytotoxicity to mammal cells compared to erythromycin and peptide alone treatment. In addition, the antibacterial mechanism of CATH-1 and erythromycin combination against E. coli was studied. The results of the scanning electron microscope showed that CATH-1 enhanced the antibacterial activity of erythromycin by increasing the permeability of bacterial cell membrane. Moreover, the results of bacterial migration movement showed that the combination of CATH-1 and erythromycin significantly inhibits the migration of E. coli. Finally, drug resistance analysis was performed and the results showed that CATH-1 delayed the emergence of E. coli resistance to erythromycin. In conclusion, the combination of CATH-1 and erythromycin has synergistic antibacterial activity and reduces the emergence of bacterial drug resistance. Our study provides valuable information to develop AMPs as potential substitutes or adjuvants for traditional antibiotics.

Methicillin-resistant Staphylococcus epidermidis (MRSE) strains are increasingly emerging as serious pathogens because they can be resistant to many antibiotics called multidrug resistance (MDR) that limit the therapeutic options. In the case of vancomycin- and rifampin-resistant MDR-MRSE, the physicians are not allowed to increase the doses of antibiotics because of severe toxicity. Accordingly, we investigated the synergistic activity of melittin antimicrobial peptide with vancomycin and rifampin against vancomycin-resistant, and rifampin-resistant MDR-MRSE isolates. Minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), fractional inhibitory concentration index (FICi), and fractional bactericidal concentration index (FBCi) of antimicrobial agents against isolates were determined. Coagulate activities and serum and salt stability as well as melittin cytotoxicity on the human embryonic kidney (HEK) 293 cells and human red blood cells (RBCs) at their synergistic concentrations. MIC and MBC values for melittin were in the range of 0.312–2.5 and 0.312–5, respectively. Results also showed that the interaction of melittin with drugs was highly synergistic in which the geometric means of FICi and FBCi were < 0.5. Induced synergism led to a decrease in melittin, rifampin, and vancomycin concentrations by 8–1,020, 2–16, and 4–16-folds, respectively. This phenomenon caused a reduction in melittin toxicity by which the synergistic concentration of melittin needed to kill bacteria did not show cytotoxicity and hemolytic activity. Besides, no coagulation activity was found for the synergistic and alone concentrations of melittin in both Prothrombin Time (PT) and Partial Thromboplastin Time (PTT). Interestingly, the antibacterial activity of melittin in Mueller Hinton Broth (MHB) containing human serum did no significant differences between MIC and MBC values of melittin in MHB and MHB containing 10% human serum. The present findings showed that the therapeutic index of melittin was improved by 32.08- and 12.82-folds when combined with vancomycin and rifampin, respectively. Taken together, the obtained data show that melittin alone was effective against MDR-MRSE isolates and this antimicrobial peptide showed highly synergistic effects with vancomycin and rifampin without causing toxicity. Therefore, the combination of melittin and traditional antibiotics could be a promising strategy for the treatment of infections caused by MDR-MRSE.

Assessment of the efficacy of clofazimine alone and in combination with primary agents against Mycobacterium tuberculosis in vitro

The therapeutic virtues of honey no longer need to be proven. Honey, which is rich in nutrients, is an excellent nutritional food because of its many properties; however, honey has been diverted from this primary function and used in clinical research. Evidence has shown that honey still possesses unknown properties and some of these aspects have never been addressed. In this work, two bioactive compounds found in honey (methylglyoxal and antimicrobial peptides) were evaluated for their anti-Bacillus subtilis activity with particular attention to their dilution factor. Although this bacterial strain does not possess an indigenous virulence factor gene, it becomes virulent by transferring plasmids with B. thuringiensis or expression of toxins from Bordetella pertussis. As is known, methylglyoxal is a toxic electrophile present in many eukaryotic and prokaryotic cells, which is generated by enzymatic and non-enzymatic reactions. Its overexpression successfully kills bacteria by inducing membrane disruption. Also, AMPs show potent inhibitory action against Gram-positive bacteria. Because of the lack of information concerning the main ingredients of honey, the microencapsulation process was used. Both methylglyoxal (MGO) and peptide-loaded liposomes were synthesized, characterized and compared to their free forms. The liposomal formulations contained a mixture of eggPC, cholesterol, and octadecylamine and their particle sizes were measured and their encapsulation efficacy calculated. The results revealed that Algerian multifloral white honey contained higher levels of MGO compared to manuka honey, which prevented bacterial growth and free MGO was relatively less effective. In fact, MGO killed BS in the loaded form with the same bacteriostatic and bactericidal index. However, the action of AMPs was different. Indeed, the investigation into the reactivity of MGO in the solvent indicated that regardless of the level of water added, honey is active at a fixed dilution. This data introduces the notion of dilution and abolishes the concept of concentration. Moreover, the synergistic antibacterial effect of the compounds in honey was diminished by the matrix effect. The degree of liposome-bacteria-fusion and the delay effect observed could be explain by both the composition and nature of the lipids used. Finally, this study reinforces the idea that under certain conditions, the metalloproteinases in honey produce AMPs.

Staphylococcus saprophyticus is frequently involved in various difficult-to-treat infections due to the formation of biofilms. To identify useful antibiofilm strategies, this study explored the efficacy and mechanism of baicalin in enhancing the ability of azithromycin against multidrug-resistant Staphylococcus saprophyticus-Liu-2016-Liyang, China-francolin (MDRSS) biofilms in vitro and in vivo. When azithromycin was used in combination with baicalin, the minimum inhibitory concentration in biofilm (MICB) for azithromycin decreased 4- to 512-fold. Compared with the azithromycin and baicalin groups, the combination of azithromycin and baicalin could not reduce the biofilm biomass, but the dispersion rates of biofilm were decreased and the bactericidal ability was increased. Furthermore, the relative transcript levels of WalK/R system-related genes were upregulated by the addition of baicalin or azithromycin plus baicalin compared with that of the azithromycin and blank control groups. The strong correlation relationship between the WalK/R system and the bactericidal index demonstrated that baicalin enhanced the bactericidal effect of azithromycin on MDRSS biofilms by modulating the WalK/R system. In the mouse cutaneous infection model, the combination of azithromycin and baicalin succeeded in eradicating MDRSS and decreasing pathological injuries. This study indicated that baicalin has the potential to be an adjuvant to enhance the antimicrobial activity of azithromycin against MDRSS in the biofilm form by modulating the WalK/R system.