Drop Plate Method Research Articles

A New Guanidine-Core Small-Molecule Compound as a Potential Antimicrobial Agent against Resistant Bacterial Strains.

The guanidine core has been one of the most studied functional groups in medicinal chemistry, and guanylation reactions are powerful tools for synthesizing this kind of compound. In this study, a series of five guanidine-core small molecules were obtained through guanylation reactions. These compounds were then evaluated against three different strains of Escherichia coli, one collection strain from the American Type Culture Collection (ATCC) of E. coli ATCC 35218, and two clinical extended-spectrum beta-lactamase (ESBL)-producing E. coli isolates (ESBL1 and ESBL2). Moreover, three different strains of Pseudomonas aeruginosa were studied, one collection strain of P. aeruginosa ATCC 27853, and two clinical multidrug-resistant isolates (PA24 and PA35). Among Gram-positive strains, three different strains of Staphylococcus aureus, one collection strain of S. aureus ATCC 29213, and two clinical methicillin-resistant S. aureus (MRSA1 and MRSA2) were evaluated. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) experiments were reported, and the drop plate (DP) method was used to determine the number of viable suspended bacteria in a known beaker volume. The results from this assessment suggest that guanidine-core small molecules hold promise as therapeutic alternatives for treating infections caused by clinical Gram-negative and Gram-positive bacteria, highlighting the need for further studies to explore their potential. The results from this assessment suggest that the chemical structure of CAPP4 might serve as the basis for designing more active guanidine-based antimicrobial compounds, highlighting the need for further studies to explore their potential.

Niosome gels encapsulate green mangosteen peel extract (Garcinia mangostana L.) as an anti-acne-inducing bacterial and anti-inflammatory activity

This research aims to develop a niosome gel from green mangosteen peel extract (Garcinia mangostana L.) by maceration with an ethanol solvent. The result of the yield percentage was 14.35 ± 0.90. The result of analyzing the phytochemicals by high-performance liquid chromatography (HPLC) was that tannin and xanthone were equal to 0.7483 ± 0.0825 mg per 100 mg of extract and 0.02964 ± 0.0088 mg per 100 mg of extract, respectively. The results of the determination of the effect include anti-inflammatory and anti-bacterial (Cutibacterium acnes) as anti-inflammatory with nitric oxide from LPS-induced macrophage cells up to a maximum equal to 29.10 ± 4.78% as a concentration at 1 mg mL–1 as acetonide can inhibit nitric oxide equal to 33.12 ± 3.62% as a concentration at 1 mg mL–1. and anti-bacterial (Cutibacterium acnes) by broth microdilution and drop plate methods, it was found that clindamycin has a minimum inhibitory concentration (MIC) and a minimum bactericidal concentration (MBC) of 0.08 μg mL–1 and 0.31 μg mL–1, respectively. The development of niosomes consisting of cholesterol, tween 60, and mangosteen peel extract at 1% w w–1 and 2% w w–1 in every formula showed good stability. The reduction of particle size in the formula by an ultrasonic bath at 30 minutes and measurement of particle size with a transmission electron microscope (TEM) were found to be equal to 1,265 and 123 – 219 nm, respectively. The polydispersity index (PDI) was in the range of 0.1 – 0.2, and the zeta potential value was in the range of –26.15 to –28.61 mV. The result of hydration and trans epidermal water loss (TEWL) was found to be that after 4 weeks of use, the formula containing the niosomes of mangosteen peel extract concentrated at 2% w w–1 maximizes skin moisture. It has a value of 346 ± 39.27 and has the least surface water loss. GRAPHICAL ABSTRACT HIGHLIGHTS The green mangosteen peel extract (Garcinia mangostana L.) can inhibiting nitric oxide production from LPS-induced macrophage cells up to 29.10 ± 4.78% at a concentration of 1 mg ml-1, the lowest inhibitory (MIC) and kill (MBC) concentration of C. acnes bacteria less than 0.24 mg ml-1. The particle size of niosomes encapsulate extracts before reducing particle size equal to 1,265 nm. and after reducing particle size in the range of 123 - 219 nm. The skin hydration value after using product for 4 weeks as niosome gel encapsulate extract 2%w w-1 of extract was maximum moisture and minimal water loss.

The Effects of Grape Seed Oligomeric Proanthocyanidin and Nisin on Dental Pulp Stem Cells.

This study aimed to evaluate the biological effects of "proanthocyanidin" (PA), and "nisin" (Ni), on dental pulp stem cells (DPSCs) and LPS-induced DPSCs as well as their antimicrobial effects against S. aureus and E. coli. After characterization of DPSCs, cytotoxicity of PA and Ni on DPSCs were evaluated using a water-soluble tetrazolium salt (WST-1). The cytokines and chemokines released by DPSCs and the expression levels of IL-6, IL-8, and TNF alpha were detected with human Cytokine Array C5 and enzyme-linked immunosorbent assay (ELİSA), respectively. The antibacterial activities of PA and Ni were tested using the drop plate method. PA at 75 μg/ml increased cell viability, decreased TNF-α expression of DPSCs, did not show any cytotoxic effects on LPS-induced DPSCs, and also showed a tendency to decrease TNF-α expression. PA at 75 μg/ml exhibited higher expressions of TIMP-2, OPG, IL-7, and IL-8 in LPS-induced DPSCs compared to DPSCs. Ni at 100 μg/ml decreased TNF-α expression in DPSCs with no cytotoxic effects. It provided increased cell viability and a downregulation trend of TNF-α expression in LPS-induced DPSCs. Both Ni and PA provided strong antibacterial effects against S. aureus. Ni at 200μg/ml had strong antibacterial effects against E. coli without affecting negatively the viability of both DPSCs and LPS-induced DPSCs and showed anti-inflammatory activity by decreasing TNF-α expression. PA provided strong antibacterial effects against E. coli at 200 μg/ml but affected DPSCs viability negatively. PA and Ni at specific concentrations exhibited immunomodulatory activity on DPSCs and LPS-induced DPSCs without any cytotoxic effects and strong antibacterial effects on S. aureus.

Effect of oregano essential oil against selective microorganisms in raw mutton

The current study was conducted to ascertain the impact of various concentrations of oregano essential oil on the microbial population in raw mutton. The experiment was conducted using a complete randomized design (CRD) with six (6) treatments as T0 , T1 , T2 , T3 , T4 , T5 and three (3) replications. The drop plate method was utilized to detect the presence of certain microorganisms. The outcomes showed that adding oregano oil reduced the microbial population in samples. Colony-forming units gradually decreased where T5 being the lowest number of colony-forming units. There was a highly significant difference (p<0.0001) in the colony-forming unit of Staphylococcus epidermidis and Proteus vulgaris. For Klebsiella pneumonia, there was a significant reduction (p<0.01) of the microbes among samples. On the other hand, Staphylococcus epidermidis significantly outperformed Proteus vulgaris in terms of growth inhibition. In light of the findings, it is possible to conclude that the addition of oregano essential oil at a 200 µl level reduces the microbial population of raw mutton.

Efficacy of green tea extract on Staphylococcus aureus and Klebsiella pneumonia of raw chevon

The efficacy of green tea extract with different strengths on selective microorganisms of raw chevon was investigated in this study. The experiment was conducted using completely randomized design (CRD) six treatments (T0 =0 ml of green tea; T1 =1 ml of green tea; T2 =2 ml of green tea; T3 =3 ml of green tea; T4 =4 ml of green tea and T5=5 ml of green tea) having three replications. Drop plate method was used for analyzing antimicrobial activity in this experiment. The outcome illustrated a declining microbial population in samples while incorporating green tea with colony-forming units steadily reducing from T0 to T5. There was a significant effect (p<0.0001) of green tea on the number of E. coli and Staphylococcus epidermidis. The minimum and maximum population of TVC of E. coli, in the treated sample were 6×105 CFU/ml in T5 and 45×105 CFU/ml in T0 respectively while for Staphylococcus epidermidis, it was 4×105 CFU/ml in T5 and 10×105 CFU/ml in T0 respectively. Green tea extracts showed equal growth suppression and extremely significant differences (p<0.0001) in their antimicrobial activity against both E. coli and Staphylococcus epidermidis. Given the results, it is possible to draw the conclusion that adding green tea extract at a level of 5 ml to raw chevon will diminish its microbial population. For Staphylococcus aureus and Klebsiella pneumonia were significant reduction (p<0.0001) of the microbes among samples treated with green tea extracts.

Investigating the Ability of Edwardsiella ictaluri and Flavobacterium covae to Persist within Commercial Catfish Pond Sediments under Laboratory Conditions.

Two prevalent bacterial diseases in catfish aquaculture are enteric septicemia of catfish and columnaris disease caused by Edwardsiella ictaluri and Flavobacterium covae, respectively. Chronic and recurring outbreaks of these bacterial pathogens result in significant economic losses for producers annually. Determining if these pathogens can persist within sediments of commercial ponds is paramount. Experimental persistence trials (PT) were conducted to evaluate the persistence of E. ictaluri and F. covae in pond sediments. Twelve test chambers containing 120 g of sterilized sediment from four commercial catfish ponds were inoculated with either E. ictaluri (S97-773) or F. covae (ALG-00-530) and filled with 8 L of disinfected water. At 1, 2, 4-, 6-, 8-, and 15-days post-inoculation, 1 g of sediment was removed, and colony-forming units (CFU) were enumerated on selective media using 6 × 6 drop plate methods. E. ictaluri population peaked on Day 3 at 6.4 ± 0.5 log10 CFU g-1. Correlation analysis revealed no correlation between the sediment physicochemical parameters and E. ictaluri log10 CFU g-1. However, no viable F. covae colonies were recovered after two PT attempts. Future studies to improve understanding of E. ictaluri pathogenesis and persistence, and potential F. covae persistence in pond bottom sediments are needed.

Study on the Efficacy of Green Tea Extract on Staphylococcus aureus and Klebsiella pneumonia of Raw Chicken Meat

We investigated the efficacy of green tea extract on Staphylococcus aureus and Klebsiella pneumonia of raw chicken meat. The experiment was conducted using complete randomized design (CRD) having six (6) treatments with three (3) replications. The germicidal activity was determined using the drop plate method. The results indicated that the addition of green tea extract (GTE) decreased microbial population in raw chicken meat. The lowest colony forming unit/ml (CFU ml-1) was found in T5 in 10-3 dilution factor (DF). There was a highly significant difference (p<0.001) in CFU ml-1of Staphylococcus aureus and Klebsiella pneumonia among different treatments i.e., GTE has significant effect (p<0.001) on Klebsiella pneumoniae and Staphylococcus aureus. The maximum number of Staphylococcus aureus in raw chicken meat was 14×105 CFU ml-1in T0 using TVC, while minimum was 4×104 CFU ml-1 in T5. Similar trend was also found in Klebsiella pneumoniae. GTE also showed highly significant (p<0.001) germicidal efficacy on both of Staphylococcus aureus and Klebsiell. The GTE also showed significant (p<0.01) germicidal efficacy on Staphylococcus epidermidis, Salmonella spp., Shigella spp. and E. coli. Considering the results obtained, it might be concluded that addition of 5 ml GTE reduce microbial population of raw chicken.

The clinical effectiveness of probiotics and autoprobiotics in treatment of <i>Helicobacter pylori</i>-associated dyspepsia

The aim of our study was to evaluate the clinical performance of a monotherapy by Enterococcus faecium-based probiotics and indigenous autoprobiotics against H. pylori associated dyspepsia.
 Materials and methods. There were examined 95 patients with dyspepsia. The entire patient cohort underwent clinical evaluation including filling out the questionnaire to assess dyspepsia symptoms before and after treatment, gastric endoscopy as well as gastric multi-focal biopsy (gastric body and gastric antrum) and verification of H. pylori infection with the three clinical laboratory methods (biochemical, bacteriological and molecular detection). An antagonistic in vitro activity of probiotics against H. pylori was detected by drop plate method for probiotic strains Enterococcus faecium SF68 and Bifidobacterium bifidum (Bifiform), Enterococcus faecium L3 (Laminolact), and autoprobiotic strains combined with indigenous Enterococcus faecium. To examine an antagonistic activity of probiotics and autoprobiotics in clinical trials, we used a starter culture based on the Enterococcus faecium L3 strain and an autoprobiotic based on indigenous Enterococcus faecium. The probiotic or autoprobiotic were administered orally to patients with gastritis twice a day at dose of 50 ml (8.0 lgCFU/ml) for 20 days. H. pylori eradication was assessed by stool antigen test 1.52 months after the end of treatment.
 Results. Initially the H. pylori infection was confirmed with 49.4% of patients. The sensitivity of H. pylori to the probiotics was detected in 81% of individuals for indigenous Enterococci (the autoprobiotic), 76% for Laminolact, and in 62% for Bifiform. 22 patients with previous history of allergic reactions to antibiotics used in routine H. pylori eradication regimens were divided in two cohorts. One cohort (10 patients) received the autoprobiotic only, another cohort (12 patients) received only probiotic. Monotherapy with autoprobiotic resulted in 100% H. pylori eradication, single-agent therapy with probiotic led to 60% eradication of H. pylori. Dyspepsia symptoms were completely resolved in both groups of patients.
 Conclusion. Our research demonstrated the sensitivity of examined H. pylori strains to be similar for traditional eradication treatment agents (antibiotics) and the proposed intervention agents (probiotics and autoprobiotics). An autoprobiotic monotherapy with indigenous enterococci led to higher levels of H. pylori eradication than with E. faecium L3-based probiotic agent. Our work demonstrated advantage for application of probiotics in patients with antibiotic allergies or other obstacles for the standard eradication therapy. Nonetheless, further investigation to better understand underlying mechanisms of action, as well as larger observational and randomized studies, are necessary to determine the scope of therapeutic application for probiotics and autoprobitics to eradicate H. pylori infection.

Probiotic viability, pH and lactic acid concentration of opened commercial probiotic dairy drinks stored at different temperatures and durations

BackgroundThe commercial cultured milk drinks contain either single or mixed probiotic species and supply in different serving sizes. It is known that different combinations of probiotics might provide the various products’ quality in terms of nutritional value during their manufacturing process. However, a lack of information about probiotic viability and physicochemical properties of the opened fermented products for continuous fermentation leads to the driving force in conducting this study. Therefore, four locally available cultured milk drinks (branded Y, F, N and V) with 20 bottles each were aseptically transferred into their respective sterile containers and stored at 4 °C, 25 °C and − 20 °C for 1–13 days. Then, the viable cells were quantified using the drop plate method on de Man, Rogosa and Sharpe (MRS) agar. The pH change was investigated using the calibrated pH meter, and the Enzytec D-/L-Lactic acid kit determined the content of D-lactic acid via spectrophotometer. Eventually, the data were analysed using the statistical tool.ResultsThe viability of probiotics in brands Y and V was significantly increased even when stored at − 20 °C and 4 °C with at least 1 log CFU/mL increment. The proliferation of probiotics was moderately influenced by the pH of the opened cultured milk. High content of D-lactate was found in Y- and F-branded products after 13 days of storage. The Y-branded cultured milk drink had the highest content of D-lactate with 0.52 g/L and 0.40 g/L when stored for 13 days at room temperature and 4 °C, respectively.ConclusionsThis study sheds light on the necessity to elucidate the properties of opened probiotic beverages over time, especially when bottled in large quantities. This allows some improvement steps.

What Is the Duration of Irrigation? An InVitro Study of the Minimum Exposure Time to Eradicate Bacteria With Irrigation Solutions.

Antiseptic irrigation solutions are commonly used by arthroplasty surgeons to reduce intraoperative bacterial colonization with the goal of reducing postoperative infections in the setting of primary total joint arthroplasty. Currently, the minimum irrigation time to eliminate common microbes implicated in periprosthetic joint infection is unknown. We sought to determine the minimum effective exposure time required to prevent growth of Staphylococcus aureus, Staphylococcus epidermidis, and Cutibacterium acnes with common antiseptic solutions. S aureus, S epidermidis, and C acnes cultures were treated with povidone-iodine (0.35%), chlorhexidine (0.05%), sodium hypochlorite (0.5%), polyhexamethylene biguanide, and an acetic acid-based solution for 15, 30, 60, 90, and 120seconds in triplicate. Bacterial growth was quantified using the drop plate method. Failure to eliminate all bacteria was considered "not effective" at that time point. Povidone-iodine 0.35% (Betadine), sodium hypochlorite 0.5% (HySept), and acetic acid (Bactisure) eradicated all bacterial growth after 90seconds of treatment, and as low as 15seconds in S aureus and C acnes (Betadine) or S epidermidis (Bactisure). Polyhexamethylene biguanide (Prontosan) required 90seconds for elimination of S aureus and S epidermidis, and 120seconds for C acnes. Chlorhexidine 0.05% (Irrisept) did eliminate S epidermidis at 120seconds but did not effectively eradicate S aureus or C acnes. All tested antiseptic solutions demonstrated successful eradication of all bacterial growth in under 2minutes of treatment time except chlorhexidine. Povidone-iodine may require the shortest duration of treatment time to successfully eradicate common bacteria.

ANTIMICROBIAL EFFICACY OF PAPAYA SEED ETHANOLIC EXTRACT AGAINST SALMONELLA TYPHI THAT CAUSES TYPHOID FEVER

Salmonella typhi is a bacteria that causes typhoid . In Indonesia, the disease is an endemic throughout the year. Papaya that is often found to have efficacy as an antibacterial agent. The research is aimed to detect the antibacterial ability of ethanol extract papaya seeds to Salmonella typhi . Extraction method was done by soaking using 70 % ethyl alcohol solvent then agar diffusion method was used to find out Minimum inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC) in antibacterial testing. The statistics will be examined through analysis of variance (one way ANOVA with p-value (α) set at 0.05). Liquid dilution method is utilized to assess Minimum inhibitory concentration while drop plate method is made to determine Minimum bactericidal concentration. The research discovered that ethanol extract of papaya seeds contain secondary metabolite compounds such as : phenolics, flavonoids, alkaloids, saponins, tannins and terpenoids at 100 % concentration was effective to inhibit Salmonella typhi growth with an area of 6,86 mm inhibition zone which is categorized as moderate. In this study, Ciprofloxacin was used for positive control and distilled water for negative control. The percentage of sample concentrations used are: 3%, 6%, 12%, 25%, 50% and also 100%. The research used Ciprofloxacin The ethanol extract of papaya seeds was not found to be an effective bactericidal agent against Salmonella typhi.

Growth of Lactobacillus casei FNCC0900 in Media Based Umbi Porang Plant (Amorphophallus muelleri BI.)

Porang tuber (Amorphophallus muellerii BI.) Is a type of tuber that has a high enough glucomannan content of 67%. Glucomannan is very difficult to digest by humans directly so it takes the role of probiotics. L. casei bacteria FNCC0900 as a probiotic agent capable of utilizing glucomannan as a carbon source for growth. The purpose of this study was to determine the growth pattern and changes in environmental factors, namely the pH value of the probiotic bacteria L. casei FNCC0900 growth medium. The parameters in this study consisted of the highest cell density, generation time and pH value changes in Glucose Yeast Peptone Liquid Media, Porang Boiled Water Media and Porang Flour Liquid Media using the drop plate method which had 4 repeated calculations. Porang Boiled Water Liquid Media has a faster log phase period with a higher cell density than Porang Flour Liquid Media, but the shortest generation time is found in Porang Flour Liquid Media with the highest number of generations. L. casei FNCC0900 bacteria are more able to reduce the pH of Glucose Yeast Peptone Liquid Media compared to porang tuber-based media, so in this case L. casei FNCC0900 can be stated to be able to grow on porang tuber-based media with growth patterns, generation time, cell density and pH value. which varies.

The drop plate method as an alternative for Azospirillum spp. viable cell enumeration within the consensus protocol of the REDCAI network

Quality evaluation of commercial inoculants is essential to warrant an adequate crop response to inoculation within a biosecurity framework. In this sense, this work is aimed at standardizing and validating the drop plate method for the enumeration of Azospirillum viable cells as an alternative to the spread plate technique, which is currently proposed in the consensus protocol of the REDCAI network. Between 14 and 25 private and public laboratories participated in three independent trials. We obtained consistent and robust results that allowed to confirm that both techniques are equivalent, concluding that the drop plate method is an alternative enumeration technique that is adequate to be included in the abovementioned consensus protocol.

The Roles of Escherichia coli cyaA / crp Genes in Metal Stress

Adenosine 3 ′, 5′-monophosphate (cAMP) is an important signaling molecule. CRP, the receptor protein of cAMP, acts as the 'main' regulator for transcription factors. The CRP-cAMP complex directly controls at least 500 promoters in Escherichia coli. In this study, the roles of cyaA and crp genes in E. coli BW25113 strain under metal stress were investigated. The minimal inhibition concentration (MIC) and minimal cidal concentration (MCC) of 5 different metals (Zn, Ni, Co, Cd and Cu) on Escherichia coli BW25113 wild type, cyaA and crp mutant cells were determined. In addition, the effect of these metals on the survival of E. coli cyaA / crp mutants was determined by growth and drop plate method. According to E. coli BW25113 wild type, cyaA mutant strain was observed sensitivity in all metals except copper, whereas resistance was observed in crp mutant strain only to zinc metal. The roles of the cyaA and crp genes in metal stress were confirmed by completing the genes on the plasmid. As a result, the roles of cyaA and crp genes in metal resistance were revealed in this study.

Assessing the surface free energy of modified asphalt binder with image processing technique

When the asphalt binder does not comply with the required specification for pavement, the suggested remedy is the implication of modifiers to enhance the quality. In the present investigation, asphalt cement binder was modified with varying percentages of two types of modifiers, Sulfur (3, 5, 7, and 10) % and Carbon Black (5, 10, and 15) % by weight of asphalt cement. The image processing technique was implemented to evaluate the surface free energy components (dynamic and static contact angles) of the treated asphalt cement by implementing the sessile drop and Wilhelmy plate techniques. The aim of this investigation is to assess how the image processing can detect the variation in the adhesive properties through the assessment of surface free energy of the asphalt cement due to the addition of the modifiers. For the comparison process, the surface free energy components of the control asphalt are also determined. Considering the test result obtained, it was concluded that Carbon black is more appropriate additive, it gives an ultimate total surface free energy increment of (7.7 and 6) % as compared with (2.8 and 3.1) % for sulfur when Sessile drop and Wilhelmy plate methods are implemented respectively when compared with the control binder.

Role of Phenotypic Switching in Stability and Persistence of Pseudomonas aeruginosa Biofilms

Objectives: Objective: This study was designed to explore the role of different phenotypes of P. aeruginosa in the development, stability and persistence of biofilm. Methods: A total of seventeen (17) waterborne biofilm producing strains of P. aeruginosa were studied. These isolates were identified on the basis of typical phenotypic characters and the tube method was used for the study of biofilms. Population and phenotypic variance were studied by the drop plate method. The hydrophobicity of strains was evaluated by the bacterial adhesion to apolar solvent test. Results: Study showed that the subject isolates of P. aeruginosa adopted a biofilm life style after 36 h of incubation at 35 °C. After 24 h the adhesion started, but it was reversible and easily dispersed by simple washing. However, after 36 h the irreversible adhesion was noticed. The biofilm consortia harbor three different phenotypes: i. wild types, showed typical P. aeruginosa characters on Cetrimide agar; ii. Slow growers, showed poor pigmentation and take >36 h for colony development, and iii. Small colony variants (SCVs) are metabolically inactive and producing pinpointed non pigmented colonies. Comparative analysis showed that these phenotypes i.e. SCVs were highly hydrophobic and persistent in biofilm consortia due to the production of excessive amounts of exopolysaccharides. Conclusions: This study showed that phenotypic heterogeneity is a characteristic feature of P. aeruginosa biofilms and all of these phenotypes have a major role in stability and persistence of biofilm consortia. J Microbiol Infect Dis 2020; 10(1):10-17.

Complement C9 binding site and the anti-microbial activity of caprine vitronectin are localized in close proximity in the N-terminal region of the protein

Vitronectin (Vn) is a ligand for complement C9 and modulates its activity that favors bacterial growth and survival. At the same time, the anti-microbial activity of the heparin-binding region of human Vn has been documented. To understand these diverse and opposite functions of the protein, we have analyzed the interaction of caprine Vn with C9 in the homologous system. In a previous study, the C9 binding activity was mapped to the N-fragment of the caprine Vn (N-Vn), representing the first 200 amino acids. Interestingly, this fragment also inhibited bacterial growth. In this study, we have generated four sub-fragments of N-Vn and analyzed C9 binding by ELISA, blot overlay, surface plasmon resonance and circular dichroism spectroscopy. These sub-fragments were also tested for antimicrobial activity against E. coli and S. aureus by drop plate method and analyzing cell death by flow cytometry. Results of these analyses together with previous data suggest that in addition to the second RGD motif (106–108 amino acids), the first 47 residues are also required for C9 binding. The anti-microbial tests employed indicate that the growth inhibitory property is contributed by 101–150 residues of Vn. These results provide an initial insight into two diverse Vn functions.

Evaluation of growth and motility in non-photosynthetic Azospirillum brasilense exposed to red, blue, and white light.

Azospirillum brasilense is a non-photosynthetic rhizobacterium that promotes the growth of plants. In this work, we evaluated the effects of different light qualities on the growth, viability, and motility in combination to other culture conditions such as temperature or composition of the culture medium. Exponential cultures of A. brasilense Az39 were inoculated by drop-plate method on nutritionally rich (LB) or chemically defined (MMAB) media in the presence or absence of Congo Red indicator (CR) and exposed continuously to white light (WL), blue light (BL), and red light (RL), or maintained in dark conditions (control). The exposure to BL or WL inhibited growth, mostly in LB medium at 36°C. By contrast, the exposure to RL showed a similar behavior to the control. Swimming motility was inhibited by exposure to WL and BL, while exposure to RL caused only a slight reduction. The effects of WL and BL on plant growth-promoting rhizobacteria should be considered in the future as deleterious factors that could be manipulated to improve the functionality of foliar inoculants, as well as the bacterial effects on the leaf after inoculation.

Azalomycin F5a Eradicates Staphylococcus aureus Biofilm by Rapidly Penetrating and Subsequently Inducing Cell Lysis.

Antimicrobial resistance has emerged as a serious threat to public health. Bacterial biofilm, as a natural lifestyle, is a major contributor to resistance to antimicrobials. Azalomycin F5a, a natural guanidine-containing polyhydroxy macrolide, has remarkable activities against Gram-positive bacteria, including Staphylococcus aureus, a major causative agent of hospital-acquired infections. To further evaluate its potential to be developed as a new antimicrobial agent, its influence on S. aureus biofilm formation was evaluated using the crystal violet method, and then its eradication effect against mature biofilms was determined by confocal laser scanning microscopy, the drop plate method, and regrowth experiments. The results showed that azalomycin F5a could significantly inhibit S. aureus biofilm formation, and such effects were concentration dependent. In addition, it can also eradicate S. aureus mature biofilms with the minimum biofilm eradication concentration of 32.0 μg/mL. As extracellular deoxyribonucleic acid (eDNA) plays important roles in the structural integrity of bacterial biofilm, its influence on the eDNA release in S. aureus biofilm was further analyzed using gel electrophoresis. Combined with our previous works, these results indicate that azalomycin F5a could rapidly penetrate biofilm and causes damages to the cell membrane, leading to an increase in DNase release and eventually eradicating S. aureus biofilm.

A New Antibiotic-Loaded Sol-Gel Can Prevent Bacterial Prosthetic Joint Infection: From in vitro Studies to an in vivo Model.

The aim of this study was to evaluate the effect of a moxifloxacin-loaded organic-inorganic sol-gel with different antibiotic concentration in the in vitro biofilm development and treatment against Staphylococcus aureus, S. epidermidis, and Escherichia coli, cytotoxicity and cell proliferation of MC3T3-E1 osteoblasts; and its efficacy in preventing the prosthetic joint infection (PJI) caused by clinical strains of S. aureus and E. coli using an in vivo murine model. Three bacterial strains, S. epidermidis ATCC 35984, S. aureus 15981, and, E. coli ATCC 25922, were used for microbiological studies. Biofilm formation was induced using tryptic-soy supplemented with glucose for 24 h, and then, adhered and planktonic bacteria were estimated using drop plate method and absorbance, respectively. A 24-h-mature biofilm of each species growth in a 96-well plate was treated for 24 h using a MBECTM biofilm Incubator lid with pegs coated with the different types of sol-gel, after incubation, biofilm viability was estimated using alamrBlue. MC3T3-E1 cellular cytotoxicity and proliferation were evaluated using CytoTox 96 Non-Radioactive Cytotoxicity Assay and alamarBlue, respectively. The microbiological studies showed that sol-gel coatings inhibited the biofilm development and treated to a mature biofilm of three evaluated bacterial species. The cell studies showed that the sol-gel both with and without moxifloxacin were non-cytotoxic and that cell proliferation was inversely proportional to the antibiotic concentration containing by sol-gel. In the in vivo study, mice weight increased over time, except in the E. coli-infected group without coating. The most frequent symptoms associated with infection were limping and piloerection; these symptoms were more frequent in infected groups with non-coated implants than infected groups with coated implants. The response of moxifloxacin-loaded sol-gel to infection was either total or completely absent. No differences in bone mineral density were observed between groups with coated and non-coated implants and macrophage presence lightly increased in the bone grown directly in contact with the antibiotic-loaded sol-gel. In conclusion, moxifloxacin-loaded sol-gel coating is capable of preventing PJI caused by both Gram-positive and Gram-negative species.