Search Results (274)

Invasive infections with rare yeasts are increasing worldwide and are associated with higher mortality rates due to their resistance to antifungal drugs. Accurate antifungal susceptibility testing (AFST) is crucial for proper management of rare yeast infections. We performed AFST of 74 Candida kefyr isolates by Etest, EUCAST-based MICRONAUT-AM assay (MCN-AM) and reference Clinical and Laboratory Standards Institute broth microdilution method (CLSI). Essential agreement (EA, ±1 two-fold dilution), categorical agreement (CA), major errors (MEs) and very-major errors (VmEs) were determined using epidemiological cut-off values of ≤1.0 µg/mL, ≤0.03 µg/mL, ≤0.5 µg/mL and ≤1 µg/mL, defining wild-type isolates for fluconazole, voriconazole, micafungin and amphotericin B (AMB), respectively. Results for AMB susceptibility were correlated with ERG2/ERG3 mutations and total-cell sterols. CA of ≥97% was recorded between any two methods while EA varied between 72 and 82%, 87 and 92%, and 49 and 76% for fluconazole, voriconazole and micafungin, respectively. For AMB, CAs between CLSI and Etest; CLSI and MCN-AM; MCN-AM and Etest were 95% (4 ME, 0 VmE), 96% (3 ME, 0 VmE) and 99%, respectively, while EA varied from 32% to 69%. Non-synonymous ERG2/ERG3 mutations and no ergosterol were found in seven of eight isolates of non-wild types for AMB by Etest. Our data show that Etest, CLSI and MCN-AM methods are suitable for AFST of C. kefyr for fluconazole, voriconazole and micafungin. Excellent CAs for AMB between Etest and MCN-AM with concordant sterol profiles but not with CLSI suggest that Etest is also an excellent alternative for the detection of C. kefyr isolates with reduced susceptibility to AMB. Full article

Background: The effect of antibiotics can be severely affected by external factors. Combining the oxidative impact of photodynamic therapy with antibiotics is largely unexplored, which may result in positive results with great impact on clinical applications. In particular, that can be relevant in the case of antibiotic resistance. Objectives: In this study, we examined the effects of aPDT using the photosensitizers (PSs), methylene blue (MB) or Photodithazine (PDZ), both alone and in combination with the antibiotics ciprofloxacin (CIP), gentamicin (GEN), and ceftriaxone (CEF), against the Gram-negative bacterium Klebsiella pneumoniae. Methods: A standard suspension of K. pneumoniae was subjected to PDT with varying doses of MB and PDZ solutions, using a 75 mW/cm² LED emitting at 660 nm with an energy of 15 J/cm². The MICs of CIP, GEN, and CEF were determined using the broth dilution method. We also tested the photosensitizers MB or PDZ as potentiating agents for synergistic combinations with antibiotics CIP, GEN, and CEF against K. pneumoniae. Results: The results showed that MB was more effective in inhibiting survival and killing K. pneumoniae compared to PDZ. The tested antibiotics CIP, GEN, and CEF suppressed bacterial growth (as shown by reduced MIC values) and effectively killed K. pneumoniae (reduced Log CFU/mL). While antibiotic treatment or aPDT alone showed a moderate effect (1 Log₁₀ to 2 Log₁₀ CFU reduction) on killing K. pneumoniae, the combination therapy significantly increased bacterial death, resulting in a ≥3 Log₁₀ to 6 Log₁₀ CFU reduction. Conclusions: Our study indicates that pre-treating bacteria with PDT makes them more susceptible to antibiotics and could serve as an alternative for treating local infections caused by resistant bacteria or even reduce the required antibiotic dosage. This work explores numerous possible combinations of PDT and antibiotics, emphasizing their interdependence in controlling infections and the unique properties each PS-antibiotic combination offers. Clinical application for the combination is a promising reality since both are individually already adopted in clinical use. Full article

Background/objectives: Proteus mirabilis is a frequent causative agent of urinary and wound infections in both community and hospital settings. It develops resistance to expanded-spectrum cephalosporins (ESCs) due to the production of extended-spectrum β-lactamases (ESBLs) or plasmid-mediated AmpC β-lactamases (p-AmpCs). Recently, carbapenem-resistant isolates of P. mirabilis emerged due to the production of carbapenemases, mostly belonging to Ambler classes B and D. Here, we report an outbreak of infections due to carbapenem-resistant P. mirabilis that were observed in a psychiatric hospital in Zagreb, Croatia. The characteristics of ESBL and carbapenemase-producing P. mirabilis isolates, associated with an outbreak, were analyzed. Materials and methods: The antibiotic susceptibility testing was performed by the disk-diffusion and broth dilution methods. The double-disk synergy test (DDST) and inhibitor-based test with clavulanic and phenylboronic acid were applied to screen for ESBLs and p-AmpCs, respectively. Carbapenemases were screened by the modified Hodge test (MHT), while carbapenem hydrolysis was investigated by the carbapenem inactivation method (CIM) and EDTA-carbapenem-inactivation method (eCIM). The nature of the ESBLs, carbapenemases, and fluoroquinolone-resistance determinants was investigated by PCR. Plasmids were characterized by PCR-based replicon typing (PBRT). Selected isolates were subjected to molecular characterization of the resistome by an Inter-Array Genotyping Kit CarbaResisit and whole-genome sequencing (WGS). Results: In total, 20 isolates were collected and analyzed. All isolates exhibited resistance to amoxicillin alone and when combined with clavulanic acid, cefuroxime, cefotaxime, ceftriaxone, cefepime, imipenem, ceftazidime–avibactam, ceftolozane–tazobactam, gentamicin, amikacin, and ciprofloxacin. There was uniform susceptibility to ertapenem, meropenem, and cefiderocol. The DDST and combined disk test with clavulanic acid were positive, indicating the production of an ESBL. The MHT was negative in all except one isolate, while the CIM showed moderate sensitivity, but only with imipenem as the indicator disk. Furthermore, eCIM tested positive in all of the CIM-positive isolates, consistent with a metallo-β-lactamase (MBL). PCR and sequencing of the selected amplicons identified VIM-1 and VIM-4. The Inter-Array Genotyping Kit CarbaResist and WGS identified β-lactam resistance genes bla_VIM, bla_CTX-M-15, and bla_TEM genes; aminoglycoside resistance genes aac(3)-IId, aph(6)-Id, aph(3″)-Ib, aadA1, armA, and aac(6′)-IIc; as well as resistance genes for sulphonamides sul1 and sul2, trimethoprim dfr1, chloramphenicol cat, and tetracycline tet(J). Conclusions: This study revealed an epidemic spread of carbapenemase-producing P. mirabilis in two wards in a psychiatric hospital. Due to the extensively resistant phenotype (XDR), therapeutic options were limited. This is the first report of carbapenemase-producing P. mirabilis in Croatia. Full article

Background/Objectives: The rapid emergence of antibiotic-resistant Escherichia coli in food and clinical environments necessitates new, clean-label antimicrobials. This study assessed eight Greek native essential oils—oregano, thyme, dittany, rosemary, peppermint, lavender, cistus and helichrysum—for activity against six genetically and phenotypically diverse E. coli strains (reference, pNorm, mecA, mcr-1, blaOXA and O157:H7). We aimed to identify oils with broad-spectrum efficacy and clarify the chemical constituents responsible. Methods: Disk-diffusion assays measured inhibition zones at dilutions from 50% to 1.56% (v/v). MIC and MBC values were determined by broth microdilution. GC–MS profiling identified dominant components, and Spearman rank-order correlations (ρ) linked composition to activity. Shapiro–Wilk tests (W = 0.706–0.913, p ≤ 0.002) indicated non-normal data, so strain comparisons used Kruskal–Wallis one-way ANOVA with Dunn’s post hoc and Bonferroni correction. Results: Oregano, thyme and dittany oils—rich in carvacrol and thymol—exhibited the strongest activity, with MIC/MBC ≤ 0.0625% (v/v) against all strains and inhibition zones > 25 mm at 50%. No strain-specific differences were detected (H = 0.30–3.85; p = 0.998–0.571; p_adj = 1.000). Spearman correlations confirmed that carvacrol and thymol content strongly predicted efficacy (ρ = 0.527–0.881, p < 0.001). Oils dominated by non-phenolic terpenes (rosemary, peppermint, lavender, cistus, helichrysum) showed minimal or no activity. Conclusions: Phenolic-rich EOs maintain potent, strain-independent antimicrobial effects—including against multidrug-resistant and O157:H7 strains—via a multi-target mode that overcomes classical resistance. Their low-dose efficacy and GRAS status support their use as clean-label food preservatives or adjuncts to antibiotics or bacteriophages to combat antimicrobial resistance. Full article

Background/Objectives:Titanium implant surface modifications enhance osseointegration and prevent microbial colonization, improving implant longevity. Antimicrobial coatings, particularly cerium- and bismuth-doped hydroxyapatite (CeHAp and BiHAp), have gained attention for reducing infection-related complications. This study evaluates the antimicrobial activity of CeHAp and BiHAp coatings on CpTi compared to untreated CpTi in artificial saliva at pH levels of 4.5, 6.5, and 8. Methods: Antibacterial efficacy against Staphylococcus aureus (S. aureus), Escherichia coli (E. coli), and Candida albicans (C. albicans) was assessed using the broth dilution method. Titanium rods coated with test compounds were incubated in inoculated nutrient broth, and microbial inhibition was determined via optical density at 600 nm. A statistical analysis was performed using the Kruskal–Wallis ANOVA test, the median and Interquartile Range were determined for the variables, and a Dwass–Steel–Critchlow–Fligner intergroup pairwise comparison was conducted. Results: The results showed that both the CeHAp and BiHAp coatings demonstrated significant antimicrobial activity against S. aureus (OD = 0.01) at pH 6.5, which was more pronounced than the activity observed against E. coli (OD = 0.05), with the difference being statistically significant (p = 0.001). The least antimicrobial activity was observed against C. albicans (0.21) at pH 8 (p = 0.001). Conclusion: These findings highlight the pH-dependent effectiveness of BiHAp and CeHAp coatings in inhibiting microbial growth. Their application on titanium implants may enhance antimicrobial properties, contributing to improved dental implant success and broader biomedical applications. Full article

Excessive application of phosphate fertilizers exacerbates water pollution, while the low phosphorus availability in acidic soils results in diminished phosphorus utilization efficiency of crops. This study conducted a maize pot experiment to investigate the effects of soybean fermentation broth value-added phosphorus fertilizer (SFB-VAPF) on soil phosphorus availability and microbial communities in acidic lateritic red soils during the 31-day seedling stage to determine its growth promotion efficacy. Conducted in Guangzhou, China, under greenhouse conditions, the experimental design comprised 11 treatments: CK (no fertilizer), treatments with P alone at two levels (0.05 and 0.15 g·kg⁻¹), and eight SFB-VAPF treatments combining each P level with four dilutions of soybean fermentation broth (SFB; 100-, 300-, 500-, and 700-fold dilutions). Each treatment had five replications. Application of SFB-VAPF significantly improved the soil chemical attributes, enzyme activities, and promoted maize growth and nutrient accumulation. Compared to the high-P treatments (0.15 g·kg⁻¹ P), low-P SFB-VAPF demonstrated superior enhancement of the soil organic matter (SOM), available nutrients, maize biomass, and nutrient accumulation. The treatment combining 0.05 g·kg⁻¹ P and 100-fold diluted SFB significantly increased the acid phosphatase activity (ACP) by 28.01% and the AP content by 69.63%, while achieving the highest maize biomass. Although SFB-VAPF application reduced the microbial species richness, the combinations of low P with high SFB and high P with low SFB enhanced both the community structural diversity and distribution evenness. SFB-VAPF application reduced the abundance of Alphaproteobacteria, while the Gammaproteobacteria abundance significantly increased in the low-P SFB-VAPF groups. The microbial beta diversity analysis demonstrated that combining 0.05 g·kg⁻¹ P with SFB significantly altered the microbial community structure. The key driving factors included soil EC and SOM, AP, Al-P, and Fe-P contents, with AP content exerting an extremely significant influence on the bacterial community composition and structure (p ≤ 0.001). This study demonstrates that SFB-VAPF enhances soil phosphorus availability, and improves the structural diversity and distribution evenness of microbial communities, thereby promoting crop growth. Critically, SFB synergistically enhances the efficiency of low-concentration phosphorus fertilizers. Full article

Microbial detection in milk is crucial for food safety and quality, as beneficial and harmful microorganisms can affect consumer health and dairy product integrity. Identifying and quantifying these microorganisms helps prevent contamination and spoilage. The study employs advanced molecular techniques to detect and quantify the genomic DNA for the target hydrolytic enzyme coding genes lipA and aprX based on the multi-align sequence conserved region, specific primer pair, and hydrolysis probes designed using the singleplex qPCR and multiplex qPCR. Cultured isolates and artificially contaminated sterilized ultra-high-temperature (UHT) milk were analyzed for their specificity, cross-reactivity, and sensitivity. The finding indicated that strains with lipA and aprX genes were amplified while the other strains were not amplified. This indicated that the designed primer pairs/probes were very specific to the target gene of interest. The specificity of each design primer pair was checked using SYBR Green qPCR using 16 different isolate strains from the milk sample. The quantification specificity of each strain target gene was deemed to be with a mean Ct value for positive pseudomonas strain > 16.98 ± 1.76 (p < 0.0001), non-pseudomonas positive strain ≥ 27.47 ± 1.25 (p < 0.0001), no Ct for the negative control and molecular grade water. The sensitivity limit of detection (LOD) analyzed based on culture broth and milk sample was >10⁵ and >10⁴ in PCR amplification while it was >10⁴ and >10³ in real-time qPCR, respectively. At the same time, the correlation regression coefficient of the standard curve based on the pure culture cell DNA as the DNA concentration serially diluted (20 ng/µL to 0.0002 ng/µL) was obtained in multiplex without interference and cross-reactivity, yielding R² ≥ 0.9908 slope (−3.2591) and intercepting with a value of 37, where the efficiency reached the level of 95–102% sensitivity reached up to 0.0002 ng/µL concentration of DNA, and sensitivity of microbial load was up to 1.2 × 10² CFU/mL. Therefore, multiplex TaqMan qPCR simultaneous amplification was considered the best method developed for the detection of the lipA and aprX genes in a single tube. This will result in developing future simultaneous (three- to four-gene) detection of spoilage psychrotrophic bacteria in raw milk. Full article

The genus Terminalia has a long history of use in traditional medicine to treat various diseases, including bacterial infections. We previously reported a metabolomic analysis using liquid chromatography–mass spectrometry of selected Australian Terminalia spp. and highlighted numerous flavonoids that may contribute to the antimicrobial activities of those plants. This study examines the antibacterial activities of fifteen flavonoids found in Terminalia spp. against a range of gastrointestinal pathogens using broth dilution assays. Flavonoids were also combined with six different classes of conventional antibiotics to investigate interactions. The efflux pump inhibitory activity of the flavonoid was evaluated using ethidium bromide accumulation and efflux assays. Toxicities were assessed via human dermal fibroblast cell line assays. Fisetin, hispidulin, isoorientin, orientin, rutin, and vitexin showed noteworthy growth inhibitory activity (MIC values 62.5–250 µg/mL). Isoorientin and orientin were most potent against Bacillus cereus and Alcaligenes faecalis, displaying MIC values of 62.5 µg/mL against both bacteria. All flavonoids except genistein, isorhamnetin, kaempferol, luteolin, taxifolin, and vitexin were nontoxic in human dermal fibroblast (HDF) cell proliferation assays. When individual flavonoids were combined with selected antibiotics, some potentiated the activity of these antibiotics. Two synergistic, eighteen additive and thirty-one non-interactive interactions were observed. The synergistic interactions were all observed in combination with orientin. Notably, orientin exhibited efflux pump inhibitory effects at concentrations from 15.26 µg/mL to 125 µg/mL. The findings reported herein indicate that the selected flavonoids have the potential for addressing bacterial antibiotic resistance and highlight the need for further study. Full article

Arthrospira platensis (spirulina) is a cyanobacterium containing various phytochemical compounds associated with various antioxidant, antimicrobial, antiviral, anticancer, anti-inflammatory, and immune-promoting properties. The efficacy of ethanolic and methanolic crude extracts of A. platensis regarding antibacterial, antioxidant, and anticancer effects was determined in this study. The ethanolic extract showed the highest antioxidant activity by 8.96 ± 0.84 mg gallic acid equivalent per gram of extract (mg GAE/g extract), 53.03 ± 4.21 mg trolox equivalent antioxidant capacity per gram of extract (mg TEAC/g extract), and 48.06 ± 0.78 mg TEAC/g extract as determined by DPPH, ABTS, FRAP assays, respectively. Moreover, the ethanolic extract showed the highest total phenolic and flavonoid compound contents by 38.79 ± 1.61 mg GAE/g extract and 27.50 ± 0.53 mg of quercetin equivalent per gram of extract (mg QE/g extract). Gallic acid and quercetin in the extracts were also determined by HPLC. The antibacterial activity was evaluated by agar well diffusion and broth dilution methods on skin pathogenic bacteria, including Staphylococcus aureus, Staphylococcus epidermidis, methicillin-resistant S. aureus (MRSA), Micrococcus luteus, Pseudomonas aeruginosa, and Cutibacterium acnes. The inhibition zone of A. platensis extracts ranges from 9.67 ± 0.58 to 12.50 ± 0.50 mm. In addition, MIC and MBC values ranged from 31.25 to 125 mg/mL. The inhibition of biofilm formation and biofilm eradication by A. platensis ethanolic extract was 87.18% and 99.77%, as determined by the crystal violet staining assay. Furthermore, the anticancer activity of A. platensis on A375 human melanoma cells was examined. The ethanolic and methanolic extracts induced DNA apoptosis through both intrinsic and extrinsic pathways by upregulating the expression of caspase-3, caspase-8, and caspase-9. These findings suggested that A. platensis demonstrated promising antioxidant, antibacterial, and anticancer activities, emphasizing its potential as a natural therapeutic agent for the management of pathological conditions. Full article

Background/Objectives: Proteus mirabilis is a frequent causative agent of urinary tract and wound infections in community and hospital settings. It develops resistance to expanded-spectrum cephalosporins (ESC) due to the production of extended-spectrum β-lactamases (ESBLs) or plasmid-mediated AmpC β-lactamases (p-AmpC). Here, we report the characteristics of ESBLs and p-AmpC β-lactamases encountered among hospital and community isolates of P. mirabilis in two hospitals and the community settings in Zagreb, Croatia. Methods: Antibiotic susceptibility testing was performed using disk-diffusion and broth dilution methods. The double-disk-synergy test (DDST) and inhibitor-based test with clavulanic and cloxacillin were applied to screen for ESBLs and p-AmpC, respectively. PCR investigated the nature of ESBL, carbapenemases, and fluoroquinolone resistance determinants. Selected strains were subjected to molecular analysis of resistance traits by the Inter-Array CarbaResist Kit and whole-genome sequencing (WGS). Results: In total, 39 isolates were analyzed. Twenty-two isolates phenotypically tested positive for p-AmpC and seventeen for ESBLs. AmpC-producing organisms exhibited uniform resistance to amoxicillin-clavulanate, ESC, ciprofloxacin, and sulphamethoxazole-trimethoprim, and uniform susceptibility to carbapenems and piperacillin-tazobactam and all harbored bla_CMY-16 genes. ESBL-positive isolates demonstrated resistance to amoxicillin-clavulanate, cefuroxime, cefotaxime, ceftriaxone, and ciprofloxacin but variable susceptibility to cefepime and aminoglycosides. They possessed bla_CTX-M genes that belong to cluster 1 (n = 5) or 9 (n = 12), with CTX-M-14 and CTX-M-65 as the dominant allelic variants. Conclusions: The study demonstrated the presence of CTX-M ESBL and CMY-16 p-AmpC among hospital and community-acquired isolates. AmpC-producing isolates showed uniform resistance patterns, whereas ESBL-positive strains had variable degrees of susceptibility/resistance to non-β-lactam antibiotics, resulting in more diverse susceptibility patterns. The study found an accumulation of various resistance determinants among hospital and outpatient isolates, mandating improvement in detecting β-lactamases during routine laboratory work. Full article

Objective: Fungal corneal infections are challenging to treat due to delayed diagnostic procedures, bacterial co-infections, and limited antifungal efficacy. This study investigates the therapeutic potential of ultraviolet C (UVC) light alone and combined with antifungal drugs. Methods: A subsurface infection model was developed in semi-solid agar droplets, with Candida albicans cells or Aspergillus brasiliensis spores inoculated into 0.75% w/v yeast peptone dextrose (YPD) agar in a 96-well microplate (5 µL per well). Two treatment groups were tested: (1) UVC exposure (265 nm, 1.93 mW/cm²) for durations of 0 s, 5 s, 10 s, 15 s, 30 s, 60 s, or 120 s, and (2) UVC combined with antifungal drugs (Amphotericin B and Natamycin) at their minimum inhibitory concentrations (MICs), determined in YPD broth. After treatment, agar droplets were homogenized, diluted, and plated for microbial enumeration. The most effective UVC doses were further tested in an ex vivo C. albicans porcine keratitis model, where the corneal epithelium was debrided, infected with C. albicans, and exposed to UVC. Corneas were then homogenized and plated to evaluate treatment efficacy. Results: UVC exposure of ≥15 s inhibited C. albicans, and ≥10 s inhibited A. brasiliensis (all p < 0.05). The broth MICs were 0.1875 µg/mL for Amphotericin B against C. albicans, 6.25 µg/mL against A. brasiliensis, and 0.78125 µg/mL for Natamycin against C. albicans, 7.8125 µg/mL against A. brasiliensis. The broth MIC did not eradicate fungi in the subsurface model. Combined treatments enhanced inhibition (all p < 0.05), with 30 s UVC + amphotericin B for C. albicans (p = 0.0218) and 30 s UVC + natamycin for A. brasiliensis (p = 0.0017). Ex vivo, 15 s and 30 s UVC inhibited growth (p = 0.0476), but no differences were seen between groups (all p > 0.05). Conclusion: UVC demonstrated strong antifungal efficacy, with supplementary benefits from combining UVC with low doses of antifungal drugs. Full article

Fungal endophytes of medicinal plants produce diverse secondary metabolites and extracellular enzymes with therapeutic and biotechnological potential. However, the biological and biotechnological potential of fungal endophytes from South African medicinal plants remain relatively underexplored. In this study, the antimicrobial, antioxidant, anti-inflammatory and extracellular enzymatic capabilities of five fungal endophytes previously isolated from the leaves of Kirkia acuminata Oliv. were investigated. Sequencing of the internal transcribed spacer (ITS) regions revealed that the isolates belonged to the genera Setosphaeria, Diaporthe and Corynespora. The broth micro-dilution assay and the Folin–Ciocalteau reagent method were used to assess the antibacterial activity and the total phenolic content (TPC) of the fungal endophytes’ ethyl acetate crude extracts (CEs), respectively. The antioxidant activity was assessed using the ferric reducing antioxidant power (FRAP) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assays. The influence of the CE of the Setosphaeria rostrata KaL-4 on the viability and LPS-induced interleukin-6 (IL-6) production in Raw 264.7 macrophages was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and an ELISA, respectively. The ability of the isolates to produce extracellular proteases, laccases and peroxidases was also determined. The CEs displayed antimicrobial activity with MICs ranging from 0.63 to 1.25 mg/mL and reducing power and scavenging activity ranging from 40% to 18% and from 60% to 48%, respectively. The S. rostrata KaL-4 CE possessed the highest TPC and demonstrated dose-dependent cytotoxicity. The CE further demonstrated a significant reduction in IL-6 production at a concentration of 0.75 µg/mL. Only one isolate demonstrated the ability to produce proteases with an enzymatic index (EI) of 0.66, while laccases (EI range of 0.14 to 1.15) and peroxidases were produced by all of the isolates. These findings suggest that fungal endophytes from South African medicinal plants are promising sources of bioactive compounds and industry-significant extracellular enzymes. Full article

Phyllanthus emblica Linn. (commonly known as Amla or Indian Gooseberry) is commonly used in Ayurvedic medicine to treat respiratory infections, skin disorders, and gastrointestinal issues. The fruit contains an abundance of polyphenols, which contribute to its strong antioxidant properties. The antibacterial activity of fruit extracts derived from P. emblica against Escherichia coli, Staphylococcus aureus, and Klebsiella pneumoniae was determined along with the antibiotic-resistant variants extended-spectrum β-lactamase (ESBL) E. coli, methicillin-resistant S. aureus (MRSA), and ESBL K. pneumoniae. Disc diffusion and broth dilution assays were conducted to assess the activity of aqueous, methanolic, and ethyl acetate extracts, with large zones of inhibition of up to 15 mm on agar observed for S. aureus and MRSA. Minimum inhibitory concentration (MIC) values ranging from 158 to 1725 µg/mL were calculated. The aqueous and methanolic extracts of P. emblica were less active against E. coli, ESBL E. coli, K. pneumoniae, and ESBL K. pneumoniae, with the only noteworthy MIC (633 µg/mL) observed for the aqueous extract against K. pneumoniae. Interestingly, a lack of inhibition was observed on agar for any of the extracts against these bacteria. Liquid chromatography–mass spectrometry (LC-MS) analysis identified several notable flavonoids, phenolic acids, terpenoids, and tannins. Notably, Artemia nauplii bioassays indicated that all extracts were nontoxic. The antibacterial activity and absence of toxicity in P. emblica extracts suggest their potential as candidates for antibiotic development, highlighting the need for further mechanistic and phytochemical investigations. Full article

Background: Combination therapy with antibiotics and phages has been suggested to increase the antibacterial activity of both antibiotics and phages. We tested the in vitro activity of five antibiotics belonging to different classes in combination with lytic bacteriophages against multidrug-resistant metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa isolates. Material/Methods: A total of 10 non-repetitive well-characterized MBL-producing P. aeruginosa isolates (5 NDM, 5 VIM) co-resistant to aminoglycosides and quinolones were used. Phage–antibiotic interactions were assessed using an ISO-20776-based broth microdilution checkerboard assay in 96-well microtitration plates. Two-fold dilutions of colistin (8–0.125 mg/L), ciprofloxacin, meropenem, aztreonam, and amikacin (256–4 mg/L) were combined with ten-fold dilutions of five different phages (5 × 10⁹–5 × 10⁰ PFU/mL) belonging to Pakpunavirus, Phikzvirus, Pbunavirus, and Phikmvvirus genus. Plates were incubated at 35 ± 2 °C for 24 h, and the minimum inhibitory concentration of antibiotics (MIC_A) and phages (MIC_P) were determined as the lowest drug and phage concentration, resulting in <10% growth based on photometric reading at 550 nm. Interactions were assessed based on the fractional inhibitory concentration index (FICi) of three independent replicates and clinical relevance based on the reversal of phenotypic resistance. The statistical significance of each drug alone and in combination with phages was assessed using GraphPad Prism 8.0. Results: Synergistic and additive interactions were found for 60–80% of isolates for all drugs. FICis were statistically significantly lower than 0.5 for colistin (p = 0.005), ciprofloxacin (p = 0.02), meropenem (p = 0.003), and amikacin (p = 0.002). Interactions were found at clinically achievable concentrations for colistin, meropenem, and amikacin, and a reversal of phenotypic resistance was observed for most strains (63–64%) for amikacin and meropenem. Antagonism was found for few isolates with all antibiotics tested. Phage vB_PaerM_AttikonH10 and vB_PaerP_AttikonH4 belonging to Phikzvirus and Phikmvvirus genus, respectively, showed either synergistic (FICi ≤ 0.35) or additive effects with most antibiotics tested. Conclusions: Synergy was observed for most drugs and phages with amikacin, showing strong synergy and reversal of phenotypic resistance against most isolates. Taking into account the wide utility of jumbo phages obtained, the findings of vB_PaerM_AttikonH10 in combination with different classes of antibiotics can enhance the activity of currently ineffective antibiotics against MBL-producing P. aeruginosa isolates. Full article

Background/Objectives: The rise in antibiotic-resistant ESKAPE pathogens, which are responsible for severe and hard-to-treat infections, highlights the urgent need for alternative therapeutic agents. While species in the Leonotis genus have demonstrated antimicrobial potential, limited research exists on Leonotis ocymifolia. This study evaluated the phytochemical profiles and antioxidant, antibacterial, and antibiofilm activities of L. ocymifolia leaf and stem extracts. Methods: Acidified acetone and hexane were used for extraction, followed by liquid–liquid fractionation with dichloromethane (DCM), ethyl acetate, and butanol. Phytochemicals were profiled using thin-layer chromatography (TLC), while polyphenolic content and antioxidant activity were determined using colorimetric and DPPH assays, respectively. Antibacterial activity was assessed via bioautography and micro-broth dilution assays. Antibiofilm activities were evaluated using crystal violet staining, and metabolic activity was assessed using tetrazolium salt as a cell viability indicator. Results: Ethyl acetate fractions had the highest phenolic (98.15 ± 9.63 mg GAE/g) and tannin contents (108.28 ± 8.78 mg GAE/g), with strong DPPH scavenging activity (79–90% at 250 µg/mL). DCM extracts had potent antibacterial activity, with a minimum inhibitory concentration (MIC) of 0.31–0.625 mg/mL against Pseudomonas aeruginosa, Escherichia coli, and Klebsiella pneumoniae. Antibiofilm assays revealed over 50% inhibition across biofilm formation phases, with DCM leaf extracts disrupting biofilms by inhibiting microbial metabolism. Conclusions: This study highlights L. ocymifolia as a promising source of bioactive compounds with significant antioxidant and antibacterial properties. The DCM and ethyl acetate extracts demonstrated high polyphenol content and effective biofilm inhibition. Further studies are warranted to isolate bioactive compounds and elucidate their mechanisms of action. Full article