Antibacterial and Antibiofilm Activity of Clerodendron Cyrtophyllum Turcz. Ethanolic Extracts against Staphylococcus aureus from Bovine Mastitis

Staphylococcus aureus (SA) is representative of gram-positive bacteria. Sanguinarine chloride hydrate (SGCH) is the hydrochloride form of sanguinarine (SG), one of the main extracts of Macleaya cordata (M. cordata). There are few reports on its antibacterial mechanism against SA. Therefore, in this study, we investigated the in vitro antibacterial activity and mechanism of SGCH against SA. The inhibitory zone, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) were measured, and the bactericidal activity curve was plotted. In addition, the micromorphology, alkaline phosphatase (AKP) activity, Na+K+, Ca2+Mg2+-adenosine triphosphate (ATP) activity, intracellular reactive oxygen species (ROS), and fluorescein diacetate (FDA) were observed and detected. The results showed that the inhibitory zone of SGCH against SA was judged as medium-sensitive; the MIC and MBC were 128 and 256 μg/mL, respectively; in the bactericidal activity curve, SGCH with 8 × MIC could completely kill SA within 24 h. SGCH was able to interfere with the integrity and permeability of the SA cell wall and membrane, as confirmed by the scanning electron microscopy (SEM) images, the increase in extracellular AKP and Na+ K+, Ca2+ Mg2+-ATP activities as well as the fluorescein diacetate (FDA) staining experiment results. Moreover, a high concentration of SGCH could induce SA to produce large amounts of ROS. In summary, these findings revealed that SGCH has a preferable antibacterial effect on SA, providing an experimental and theoretical basis for using SG as an antibiotic substitute in animal husbandry and for the clinical control and treatment of diseases caused by SA.

Dental caries results from the colonization of oral microorganisms, such as Streptococcus mutans (S. mutans), and their adherence to teeth is the initial step in this process. Therefore, blocking the attachment to the enamel surface would lead to inhibition of biofilm formation. We investigated the antibacterial, antibiofilm, and anti-attachment effects of nano-liposomal herbal extracts, including Shirazi thyme (Zataria multiflora; Z. multiflora), Summer savory (Satureja hortensis; S. hortensis), and Red Cabbage (Brassica oleracea var. capitata f. rubra; BRSOR), against biofilm formation and adherence of S. mutans. S. mutans strain ATCC 35,668 was cultured, and aqueous extracts were prepared. An antibacterial assessment was conducted using the minimum inhibitory concentration (MIC) method by the broth microdilution method. Nanoliposomes of Z. multiflora (Z-NLP) and S. hortensis (S-NLP) were prepared and characterized. Anti-biofilm assay was performed using the minimum biofilm inhibitory concentration (MBIC) test. Finally, the effects of nanoliposomal herbal extracts were evaluated against S. mutans attachments to glass and enamel surfaces. The aqueous extract of BRSOR did not exhibit an antibacterial effect, and therefore, it was abandoned from further consideration in the work. No significant difference was observed between the herbal extracts and the control groups in MIC. Extract-loaded nanoliposomes demonstrated a significantly higher anti-biofilm effect compared to aqueous extracts. Among the study groups, only Z-NLP had a similar attachment level to enamel with Chlorhexidine 0.2%. According to the results, Z-NLP showed a significant effect on the attachment of S. mutans to enamel, thereby inhibiting biofilm formation.

The role of Streptococcus mutans in the initiation of caries is related to its acidogenicity, aciduricity, and polysaccharides extracellular layer production by glucosyltransferases in dental biofilms. Therefore, inhibition of glucosyltransferase activity impairs the virulence of cariogenic biofilms, which can be used to prevent dental caries. We evaluated the anti-bacterial, anti-biofilm, and anti-glucosyltransferases effects of nanoliposomal herbal aqueous extracts of Liquorice (Glycyrrhiza glabra; G. glabra), Ginger (Zingiber officinale; Z. officinale), Pomegranate (Punica granatum; P. granatum), and Rose (Rosa damascene; R. damascene) via minimum bactericidal concentration and minimum inhibitory concentration against Streptococcus mutans strain ATCC 35,668. An anti-biofilm assay was performed using a minimum biofilm inhibitory concentration test. Among herbs, only P. granatum showed an antibacterial effect. Therefore, a nanoliposomal formulation of P. granatum was developed and characterized. Its effect on S.mutans glucosyltransferases was assessed by measuring glucan amount. The nanoliposomal formulation of P.granatum showed a significantly higher anti-biofilm effect than P. granatum aqueous extract. Their similar potential in blocking glucosyltransferases showed that the nanoliposomal formulation of P.granatum blocked other pathways rather than blocking glucosyltransferases for its anti-biofilm effect. Collectively, the nanoliposomal formulation of P.granatum, due to its anti- Streptococcus mutans characteristics, would be a production which open a new horizon for the oral pharmaceutical industry.

Phenotypic and genotypic characterization of antimicrobial resistance profiles in Streptococcus dysgalactiae isolated from bovine clinical mastitis in 5 provinces of China

Skin, our first interface to the external environment, is subjected to oxidative stress caused by a variety of factors such as solar ultraviolet, infrared and visible light, environmental pollution, including ozone and particulate matters, and psychological stress. Excessive reactive species, including reactive oxygen species and reactive nitrogen species, exacerbate skin pigmentation and aging, which further lead to skin tone unevenness, pigmentary disorder, skin roughness and wrinkles. Besides these, skin microbiota are also a very important factor ensuring the proper functions of skin. While environmental factors such as UV and pollutants impact skin microbiota compositions, skin dysbiosis results in various skin conditions. In this review, we summarize the generation of oxidative stress from exogenous and endogenous sources. We further introduce current knowledge on the possible roles of oxidative stress in skin pigmentation and aging, specifically with emphasis on oxidative stress and skin pigmentation. Meanwhile, we summarize the science and rationale of using three well-known antioxidants, namely vitamin C, resveratrol and ferulic acid, in the treatment of hyperpigmentation. Finally, we discuss the strategy for preventing oxidative stress-induced skin pigmentation and aging.

Chlamydomonas reinhardi, cultured under normal growth conditions, secreted significant amounts of protein and carbohydrates but not lipids or nucleic acids. A fivefold increase in light intensity led to a tenfold increase in secreted protein and carbohydrate. Among the proteins secreted was acid phosphatase with a pH optimum at 4.8 like the enzyme in the cells. Phosphorus depleted algae grown on minimal orthophosphate contained and secreted both acid and alkaline phosphatase. The pH optimum of the intracellular alkaline phosphatase was 9.2. When phosphorus-depleted cells were grown with increasing orthophosphate, intra- and extracellular alkaline phosphatase was almost completely repressed and intra- and extracellular acid phosphatase was partially repressed. Extracellular acid and alkaline phosphatase increased with the age of the culture. Electrophoresis indicated only one acid and one alkaline phosphatase in phosphorus-satisfied and phosphorus-depleted cells. Chlamydomonas cells suspended in an inorganic salt solution secreted only acid phosphatase; the absence of any extr-cellular cytoplasmic marker enzyme indicated that there was little, if any, autolysis to account for the extracellular acid enzyme. Phosphorus-depleted cells were able to grow on organic phosphates as the sole source of orthophosphate. Ribose-5-phosphate was the best for cell multiplication, and its utility was shown to be due to the cell's ability to use the ribose as well as the orthophosphatase for cell multiplication.

Pseudomonas aeruginosa is one of the causative pathogens of bovine mastitis. Most of P. aeruginosa cells can form biofilm, thereby reducing antibiotic efficacy which has become a significant public health challenge nowadays. In the present study, biofilm formation of 50 isolates was assessed. The inhibitory effect of three designed antimicrobial peptides (FASK, YDVD, WSF) on P. aeruginosa planktonic growth and biofilms was evaluated. The minimum inhibitory concentration (MIC) of all the peptides under study was 1600 µg/mL. The synthetic compounds had a significant inhibitory effect at concentrations of 1/2 MIC (800 µg/mL) and 1/4 MIC (400 µg/mL) on biofilm formation of these isolates, and showed anti-biofilm activity at a lower concentration than MIC. Antibacterial peptides FASK, YDVD with 70% antibiofilm effect and WSF with 60% effect prevented the formation of biofilm by mastitis isolates at a concentration of 1/2 MIC. The peptides of this study are promising candidates for inhibiting P. aeruginosa biofilm formation.

Background and Aim:Bovine mastitis has a negative impact on animals, and improper antibiotic use has caused an increase in bacterial resistance. Therefore, medicinal plants could serve as an alternative treatment for this condition. Polyphenols have potential as antibiotic agents. Oak bark has long been used as a medicine and has shown antibacterial effects. Moreover, research on heather plant demonstrated that it has antibacterial properties. This study aimed to assess the antibacterial effects of oak (Quercus robur) bark and heather (Calluna vulgaris L.) herb extracts against common bovine mastitis pathogens.Materials and Methods:Dried oak bark and heather herb were used to prepare extracts using 30%, 50%, and 70% ethanol and acetone as solvents. Their polyphenol content was determined using the Folin–Ciocalteu method. Bovine mastitis-inducing clinical isolates of Escherichia coli, Streptococcus agalactiae, Streptococcus uberis, Serratia liquefaciens, Staphylococcus aureus, and reference cultures of S. aureus and E. coli were used for antibacterial tests. All extracts were screened through a disk diffusion test to ascertain their antibacterial effects, and the minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were determined for the most effective extracts.Results:Oak bark extracts had variable antibacterial effects against S. aureus and Streptococcus strains, but no statistically significant difference was observed in activity against E. coli. The disk diffusion test showed that the oak bark extracts obtained using acetone and ethanol at 30% yielded the best results. However, the 70% acetone oak extract alone affected all types of bacteria. Further antibacterial tests of 70% acetone and 30% ethanol oak extracts revealed that the lowest MIC and MBC scores were against S. aureus strains and E. coli reference cultures. Conversely, the heather herb extracts exhibited broader activity against all types of bacteria, although better results were observed against Gram-positive bacteria. There was also a negative correlation between solvent concentration and antibacterial effect (p < 0.05, r = −0.507). The highest inhibition zone scores and broadest spectrum were observed in samples prepared in 30% ethanol. There was no statistically significant correlation between the phenolic content of plants and their antibacterial effects.Conclusion:Oak bark and heather extracts could be used as potential antibacterial agents against bovine mastitis pathogens.

Antibiotics represent the first line therapy for bovine mastitis. However, the increasing prevalence of multidrug-resistant organisms (MDROs) highlights the need for alternative therapeutic approaches. This study evaluated the antimicrobial and antibiofilm activities of Eucalyptus globulus leaf extract (EGL-L), ursolic acid (UA) and asiatic acid (AA) against Staphylococcus aureus (SA), Streptococcus uberis (SU), Streptococcus agalactiae (SAG), and Enterococcus spp. (EN) isolated from bovine mastitis, 39.7% of which were MDROs. The minimal inhibitory concentration (MIC) assay demonstrated that all the compounds exhibited antimicrobial activity against the tested bacteria, including MDROs. However, EGL-L was less effective (p < 0.001) than UA or AA against field strains. UA was more effective against SAG and SU compared to SA (p < 0.001), whereas AA was more effective against SU than SA (p < 0.001). Conversely, EGL-L exhibited similar inhibitory effects on all bacteria. The biofilm-forming ability of the bacterial strains was also assessed, and the minimal biofilm inhibitory concentrations (MBICs) of the compounds were evaluated for moderate and strong biofilm producers. None of the compounds were able to completely inhibit biofilm formation. However, MBIC80 values within the tested concentration range were achieved for 15 out of 32 strains with EGL-L and for 27 out of 32 strains with UA and AA. These findings highlight a promising alternative to conventional antimicrobials for AA and UA, showing potential for topical intramammary use for the control and prevention of bovine mastitis, especially because of their efficacy against biofilm formation. Future research should focus on toxicity assessments and formulation development for potential topical administration.

Surgical site infections (SSIs) represent the most common nosocomial infections, and surgical sutures are optimal surfaces for bacterial adhesion and biofilm formation. Staphylococcus spp., Enterococcus spp., and Escherichia coli are the most commonly isolated microorganisms. The aim of this research was to evaluate the antibiofilm activity of a medical device (MD) containing TIAB, which is a silver-nanotech patented product. The antibacterial effect was evaluated against Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212, and E. coli ATCC 25922 by assessing the minimum inhibitory concentration (MIC) by the Alamar Blue® (AB) assay. The antibiofilm effect was determined by evaluation of the minimum biofilm inhibitory concentration (MBIC) and colony-forming unit (CFU) count. Subsequently, the MD was applied on sutures exposed to the bacterial species. The antimicrobial and antibiofilm effects were evaluated by the agar diffusion test method, confocal laser scanning microscopy (CLSM), and scanning electron microscopy (SEM). The MIC was determined for S. aureus and E. faecalis at 2 mg/mL, while the MBIC was 1.5 mg/mL for S. aureus and 1 mg/mL for E. faecalis. The formation of an inhibition zone around three different treated sutures confirmed the antimicrobial activity, while the SEM and CLSM analysis performed on the MD-treated sutures underlined the presence of a few adhesive cells, which were for the most part dead. The MD showed antimicrobial and antibiofilm activities versus S. aureus and E. faecalis, but a lower efficacy against E. coli. Surgical sutures coated with the MD have the potential to reduce SSIs as well as the risk of biofilm formation post-surgery.

Toona sinensis is a traditional plant used in food and medicine, but no reports exist on its antibacterial and antibiofilm effects against Listeria monocytogenes. This study analyzed the chemical components of the ethyl acetate extract of T. sinensis and explored its antibacterial, antibiofilm, and antioxidant effects. (1) A total of 58 compounds were detected, primarily including 12 flavonoids, 10 fatty acids, 6 organic acids, and 3 alkaloids. Linoleic acid, succinic acid, and quercetin may contribute to antibacterial and antibiofilm properties, while polyphenols and flavonoids enhance antioxidant activity. (2) The minimum inhibitory concentration (MIC) of the extract against L. monocytogenes ATCC 19112 was 100µg/mL. Growth curves showed complete inhibition at 2 MIC, with live/dead staining and scanning electron microscope images confirming bacterial cell destruction at this concentration. (3) Crystal violet staining showed that at 0.5, 1, and 2 MIC, biofilm reduction of L. monocytogenes ATCC 19112 was 62.75, 82.66, and 85.50%, respectively, which was further confirmed by fluorescein isothiocyanate staining. (4) Proteomic analysis suggested the antibacterial mechanism involves disruptions in nucleic acid, protein, carbon, and energy metabolism, as well as cell wall synthesis. (5) At a concentration of 125µg/mL, the T. sinensis extract exhibited 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging efficiency of 93.98% ± 1.56% and 2,2'-azino-bis (ABTS) free radical scavenging efficiency of 99.85% ± 0.08%. These results demonstrate the potential of T. sinensis extract as an antibacterial and antioxidant agent. This is the first report on the antibacterial and antibiofilm effects of T. sinensis extract against L. monocytogenes.

We compared the values of the minimum inhibitory concentration (MIC) and mutant prevention concentration (MPC) values ​​of three antimicrobial agents for 72 bovine isolates ofPasteurella multocida, 80 swine isolates ofP. multocida, 80 bovine isolates ofEscherichia coli, 80 swine isolates ofE. coli, and 80 isolates ofStaphylococcus aureusfrom bovine mastitis. The ratio of MIC90​​/MPC90which limited mutant selection window (MSW) was ≤ 0.12/4 mg/l for enrofloxacin, 0.5/≥ 64 mg/l for florfenicol and 4/≥ 128 mg/l for tulathromycin in bovineP. multocidaisolates, ≤ 0.12/2 mg/l for enrofloxacin, 0.5/≥ 64 mg/l for florfenicol and 4/≥ 128 mg/l for tulathromycin in swineP. multocidaisolates, 1/16 mg/l for enrofloxacin, 8/≥ 64 mg/l for florfenicol and 8/≥ 128 mg/l for tulathromycin in bovineE. coliisolates, 0.5/16 mg/l for enrofloxacin, ≥ 64/≥ 64 mg/l for florfenicol and 8/≥ 128 mg/l for tulathromycin in swineE. coliisolates, and 0.25/16 mg/l for enrofloxacin, 4/≥ 64 mg/l for florfenicol and 4/≥ 128 mg/l for tulathromycin inS. aureusisolates. These findings indicate that the dosage of antimicrobial agents to achieve serum concentration equal to or higher than MPC could reduce selection of resistant bacterial subpopulation.

The exposure of cells to ultraviolet B radiation (UVB) can induce the production of reactive oxygen species (ROS) which damage cellular components. Free radical scavengers and antioxidants can interfere with the production of ROS. We studied cytotoxicity, intracellular ROS levels, lipid peroxidation, antioxidant status and oxidative DNA damage in cultured human skin dermal fibroblast adult cells (HDFa) exposed to UVB in the presence of sesamol, a natural phenolic compound. The levels of cytotoxicity, intracellular ROS, lipid peroxidation, oxidative DNA damage and apoptotic morphological changes were significantly increased in UVB irradiated HDFa cells. We also observed that the activities of enzymatic antioxidants (superoxide dismutase, catalase and glutathione peroxidase) and the levels of non-enzymatic antioxidant status (GSH) were significantly decreased in UVB irradiated cells. On the other hand, sesamol pretreatment significantly decreased cytotoxicity, intracellular ROS, lipid peroxidation, oxidative DNA damage and apoptotic morphological changes in sesamol-pretreated and UVB-irradiated HDFa cells. We have also observed increased enzymatic and non-enzymatic antioxidants status in sesamol plus UVB-irradiated cells. Among the different doses tested, 80 μM of sesamol shows maximum protection for UVB-induced oxidative damage. In conclusion, UVB-induced ROS formation, cell fatality, lipid peroxidation, antioxidant depletion and oxidative DNA damage in HDFa cells is inhibited by sesamol, which, probably through its ROS scavenging activity.

Antimicrobial mechanism of recombinant enterocin CHQS on Listeriamonocytogenes and its application on pasteurized milk

Tuberculosis (TB) remains a major threat to human health. Due to the prevalence of drug-resistant Mycobacterium tuberculosis (Mtb), it is urgent to discover drugs with new mechanisms of action (MOA) to ensure effectiveness against strains that are resistant to existing TB drugs. Cynoglossum lanceolatum Forsk was used to treat TB in Traditional Chinese Medicine. In this article, shikonin, the anti-Mtb active component, was obtained from the whole herb extract of C. lanceolatum by bioassay-guided isolation. Using the microplate alamar blue assay (MABA), the minimum inhibitory concentration (MIC) of shikonin against Mtb was determined to be 128 μg/mL. In order to obtain a more efficient anti-Mtb molecule, (E)-1-(6-bromo-2,3-dihydrochromen-4-ylidene)thiosemicarbazide was synthesized based on the scaffold of shikonin, which exhibited potent activity against Mtb (MIC=4 μg/mL). These results highlight that both naphthalene-1,4-dione and chroman-4-one are pharmacophores with activities against Mtb. To investigate a plausible mechanism of action, the molecular docking was firstly performed against catalase-peroxidase enzyme (KatG) of Mtb using AutoDock 4 software. The results demonstrated that both shikonin and (E)-1-(6-bromo-2,3-dihydrochromen-4-ylidene)thiosemicarbazide could bind to the active site of Mtb KatG. KatG enzyme activity and intracellular reactive oxygen species (ROS) levels in Mtb cells were then measured by ultraviolet spectrophotometric method and fluorescence microplate reader assay, respectively. The experiments confirmed that above compounds could inhibit the catalytic activity of Mtb KatG, and cause the ROS accumulation in Mtb cells. Therefore, inhibition of KatG may be a novel mechanism of action for these two compounds to fight against Mtb.