Antibacterial activity in three Chaetoceros microalgae species cultures by using antibiotics

Diatoms, such as Chaetoceros, grow in a mutualistic relationship with bacteria. However, in some cases, it is necessary to grow them in bacteria-free cultures. To reduce bacterial load, antibiotics are used, and on certain occasions it is necessary to use a mixture with more than one antibiotic. This work aimed to obtain a quick and effective protocol to reduce the bacterial load and evaluate the response of three Chaetoceros species with aquacultural importance. Single and mix antibiotics were used. Microalgal and bacterial growth was measured. The growth parameters for diatoms showed that the significantly highest cell concentration was for C. muelleri (3.15 x106 cells mL-1) and the lowest values to C. calcitrans (2.98 x106 cells mL-1). The significantly highest growth rate was for C. calcitrans (0.77 divisions per day), and the lowest values for Chaetoceros sp. (0.60 divisions per day). The growth parameters for heterotrophic bacteria showed that the significantly highest bacterial load was for Chaetoceros sp. (19.16 x106 CFU (Colony-Forming Units) mL-1) and the lowest values were for C. calcitrans (12.23 x106 CFU mL-1). The growth rate of the heterotrophic bacteria present in Chaetoceros cultures was similar among the three studied species. Streptomycin® and sulfate G41® produced a partial reduction of bacterial load. The most effective treatment for all three species was the use of an antibiotic mix composed of ampicillin® (250 μg mL-1), kanamycin® (200 μg mL-1), neomycin® (50 μg mL-1), and streptomycin® (100 μg mL-1) for three days. The mix prepared with the highest antibiotic concentration produced a reduction of bacteria (100%) for three days; however, it also induced a significant reduction of the growth of the three Chaetoceros species.

In Vitro Synergistic Antimicrobial Activity of Combinations of Meropenem, Colistin, Tigecycline, Rifampin, and Ceftolozane/Tazobactam Against Carbapenem-Resistant Acinetobacter Baumannii

The aim of this study was to investigate the in vitro activity of various antimicrobial combinations against carbapenem-resistant Acinetobacter baumannii (CRAB) isolates producing OXA-23 carbapenemases.In vitro activity of six two-drug combinations against CRAB isolates collected from patients with CRAB bacteremia was evaluated using the checkerboard method and time-kill assay [0.5 ×, 1 ×, 2 × minimum inhibitory concentrations (MIC)], to identify potential synergistic and bactericidal two-drug combinations against CRAB isolates, using meropenem, colistin, tigecycline, rifampin, and ceftolozane/tazobactam. All 10 CRAB isolates in our study carried the OXA-58-type and OXA-23-type carbapenem-hydrolyzing oxacillinase. The colistin-ceftolozane/tazobactam combination demonstrated a synergistic effect in both the time-kill assay (using an antibiotic concentration of 1 × MIC) and the checkerboard method, while simultaneously showing a bactericidal effect in the time-kill assay. For all 10 CRAB isolates, time-kill curves showed a significant synergistic bactericidal activity of the colistin-ceftolozane/tazobactam combination at 0.5 × MIC. Overall, there is substantial discordance of synergistic activity between the checkerboard microdilution and time-kill assay (with a concordance of 35%). Our study demonstrated that the two-drug combinations of colistin and ceftolozane/tazobactam can be a potential alternative for treating CRAB infections. The effect of these antibiotic combinations should be evaluated through clinical trials.

Environmental Biofilms as Reservoirs for Antimicrobial Resistance

Characterizing the response of microbial communities to a range of antibiotic concentrations is one of the strategies used to understand the impact of antibiotic resistance. Many studies have described the occurrence and prevalence of antibiotic resistance in microbial communities from reservoirs such as hospitals, sewage, and farm feedlots, where bacteria are often exposed to high and/or constant concentrations of antibiotics. Outside of these sources, antibiotics generally occur at lower, sub-minimum inhibitory concentrations (sub-MICs). The constant exposure to low concentrations of antibiotics may serve as a chemical “cue” that drives development of antibiotic resistance. Low concentrations of antibiotics have not yet been broadly described in reservoirs outside of the aforementioned environments, nor is the transfer and dissemination of antibiotic resistant bacteria and genes within natural microbial communities fully understood. This review will thus focus on low antibiotic-concentration environmental reservoirs and mechanisms that are important in the dissemination of antibiotic resistance to help identify key knowledge gaps concerning the environmental resistome.

Conceptual model of adding antibiotics to dialysate fluid during renal replacement therapy

Studies have shown significant variability in antibiotic trough concentrations in critically ill patients receiving renal replacement therapy (RRT). The purpose of this study was to assess whether adding beta-lactam antibiotics to dialysate solution can maintain stable antibiotic concentrations during RRT in experimental conditions. A single compartment model reflecting the patient was constructed and connected to the RRT machine. Dialysate fluid was prepared in three different concentrations of meropenem (0 mg/L; 16 mg/L; 64 mg/L). For each dialysate concentration various combinations of dialysate and blood flow rates were tested by taking different samples. Meropenem concentration in all samples was calculated using spectrophotometry method. Constructed experimental model results suggest that decrease in blood meropenem concentration can be up to 35.6%. Moreover, experimental data showed that antibiotic loss during RRT can be minimized and stable plasma antibiotic concentration can be achieved with the use of a 16 mg/L Meropenem dialysate solution. Furthermore, increasing meropenem concentration up to 64 mg/L is associated with an increase antibiotic concentration up to 18.7–78.8%. Administration of antibiotics to dialysate solutions may be an effective method of ensuring a constant concentration of antibiotics in the blood of critically ill patients receiving RRT.

Exploration of the Pharmacodynamics for Pseudomonas aeruginosa Biofilm Eradication by Tobramycin

Pseudomonas aeruginosa is a Gram-negative, opportunistic pathogen which is involved in numerous infections. It is of growing concern within the field of antibiotic resistant and tolerance and often exhibits multi-drug resistance. Previous studies have shown the emergence of antibiotic resistant and tolerant variants within the zone of clearance of a biofilm lawn after exposure to aminoglycosides. As concerning as the tolerant variant emergence is, there was also a zone of killing (ZOK) immediately surrounding the antibiotic source from which no detectable bacteria emerged or were cultured. In this study, the ZOK was analyzed using both in vitro and in silico methods to determine if there was a consistent antibiotic concentration versus time constraint (area under the curve, (AUC)) which is able to completely kill all bacteria in the lawn biofilms in our in vitro model. Our studies revealed that by achieving an average AUC of 4,372.5 μg*hr/mL, complete eradication of biofilms grown on both agar and hydroxyapatite was possible. These findings show that appropriate antibiotic concentrations and treatment duration may be able to treat antibiotic resistant and tolerant biofilm infections.

Investigation of Antibiotic Release from Bone Allograft in an Experiment on Rabbits

BACKGROUND: The treatment of chronic osteomyelitis, despite the use of new methods, is still an urgent problem. Local use of antibacterial drugs in combination with systemic antibiotic therapy has become popular in recent decades. Autologous bone grafts are considered ideal for bone defects filling. Different methods of allograft preparation may have differences in the rate and duration of antibiotic release. Moreover, it can affect the effectiveness of microbial agent eradication. The study analyzed the differences in the release of gentamicin from different types of allografts in dynamics and methods of preparation: «PerOssal» medium, whole bone allograft soaked in antibiotic, whole bone allograft, welded with an antibiotic, and perforated bone allograft soaked in an antibiotic solution. AIM: The objective of the study was to study the stability of antibiotic release and to determine the effectiveness of local transport systems. Evaluation of the difference in gentamicin release from different types of allografts in dynamics and methods of preparation had been realized: “PerOssal” medium, whole bone allograft soaked in antibiotic, whole bone allograft welded with an antibiotic, and perforated bone allograft, soaked in antibiotic solution. MATERIALS AND METHODS: The research was conducted between September 2020 and March 2021. The experiments were performed on 120 laboratory rabbits (weight – 3000–3500 g, age – 6–8 months), which were divided into four groups (30 animals in each group). Group 1 consisted of animals treated with “PerOssal.” The whole bone allograft soaked in an antibiotic was used in the treatment of animals of Group 2. The whole bone allograft, welded with an antibiotic, was used in the treatment of animals of Group 3. Perforated bone allograft soaked in an antibiotic was used in Group 4. Osteomyelitis of the proximal femur was formed in experimental animals. RESULTS AND DISCUSSION: Statistically insignificant decrease in the concentration of gentamicin was observed by the 7th day in all experimental groups. In rabbits whose bone defect was filled with a whole bone allograft welded with antibiotic and perforated bone allograft impregnated with an antibiotic (Groups 3 and 4), the most stable concentration of gentamicin was noted throughout the study period. Statistically significant differences were revealed between the experimental groups in relation to the dynamics of changes in the concentration of gentamicin in blood plasma. It was found that the group using the biodegradable material “PerOssal” on the 1st day showed a high concentration of the antibiotic in the blood plasma. However, by the 2nd day, a lower concentration of the antibiotic was recorded compared to all comparison groups of the bone allograft. CONCLUSIONS: The results of the analysis of the dynamics of gentamicin concentration may indicate significant differences between the methods of graft preparation, especially in the relationship with antibiotic release into the blood plasma. The most stable antibiotic concentration was registered in the groups of animals that underwent the filling of bone defect using a whole bone allograft welded with an antibiotic and a perforated bone allograft impregnated with antibiotic. A significant decrease of gentamicin concentration in the femur homogenate by the 7th day after transplantation was observed when using a whole bone allograft impregnated with an antibiotic. At the same time, a stable concentration of the antibiotic in the blood plasma was registered. The highest initial antibiotic concentration in the homogenate with a gradual decrease over 7 days was observed when using the antibiotic-impregnated biodegradable material “PerOssal.”

Impact of PrsA on membrane lipid composition during daptomycin-resistance-mediated β-lactam sensitization in clinical MRSA strains

Background The cyclic anionic lipopeptide daptomycin is used in the treatment of severe infections caused by Gram-positive pathogens, including MRSA. Daptomycin resistance, although rare, often results in treatment failure. Paradoxically, in MRSA, daptomycin resistance is usually accompanied by a concomitant decrease in β-lactam resistance in what is known as the ‘see-saw effect’. This resensitization is extensively used for the treatment of MRSA infections, by combining daptomycin and a β-lactam antibiotic, such as oxacillin. Objectives We aimed: (i) to investigate the combined effects of daptomycin and oxacillin on the lipid composition of the cellular membrane of both daptomycin-resistant and -susceptible MRSA strains; and (ii) to assess the involvement of the post-translocational protein PrsA, which plays an important role in oxacillin resistance in MRSA, in membrane lipid composition and remodelling during daptomycin resistance/β-lactam sensitization. Results The combination of microbiological and biochemical studies, with fluorescence microscopy using lipid probes, showed that the lipid composition and surface charge of the daptomycin-resistant cells exposed to daptomycin/oxacillin were dependent on antibiotic concentration and directly associated with PrsA, which influenced cardiolipin remodelling/relocation. Conclusions Our findings show that PrsA, in addition to its post-transcriptional role in the maturation of PBP 2a, is a key mediator of cell membrane remodelling connected to the see-saw effect and may have a key role in the resensitization of daptomycin-resistant strains to β-lactams, such as oxacillin.

Express determination of cephalexin

Test means based on immobilized Fehling’s reagent for the determination of cephalexin have been obtained. Methods for visual and colorimetric assessment of antibiotic concentration using a smartphone camera have been developed. The optimal conditions for the indicator reaction have been selected with varying heating time and temperature. For a visual semi-quantitative assessment of the content of cephalexin, a color scale has been obtained. The metrological characteristics of the test method have been determined: the range of the determined contents is 0.5–16 mg/ml, the unreliability interval is 0.1–0.4 mg/ml, and the detection limit is 0.4 mg / ml. For the colorimetric assessment of the concentration of cephalexin, a linear dependence of the intensity of the Blue channel on the logarithm of the concentration of cephalexin (у = –96x + 144, r 2 = 0.99) has been constructed, and linear dependences of the area (у = –36986x + 62458, r 2 = 0.96) and perimeter (у = –270x + 786, r 2 = 0.93) from the logarithm of the concentration of cephalexin have been obtained. The range of the determined contents was 0.1–16 mg/ml, the lower limit was 0.1 mg / ml. The verification of the correctness of the developed test methods was carried out by the «introduced-found» method (Sr  0.13).

Streptococcal and enterococcal endocarditis: time for individualized antibiotherapy?

Recommendations for the treatment of streptococcal and enterococcal endocarditis are based on old efficacy studies, but the starting doses have never been reassessed and are associated with significant adverse events. Based on data from other serious infections, we suggest that maintaining a concentration of β-lactams higher than 4–6 times the responsible bacteria MIC 100% of the time in the heart of the vegetation would be a pertinent therapeutic objective. The data point to a diffusion gradient of β-lactams in the vegetation. Yet, so far as is known, the ratio of antibiotic concentration at steady state between plasma and vegetation cannot be completely determined. Answering this crucial question would make it possible for each patient to have a targeted β-lactam plasma concentration, according to the MIC for the responsible bacteria. This would lead the way to personalized antibiotherapy and allow a safe switch to oral medication.

Antibiotic Breakdown by Susceptible Bacteria Enhances the Establishment of β-Lactam Resistant Mutants

For a better understanding of the evolution of antibiotic resistance, it is imperative to study the factors that determine the initial establishment of mutant resistance alleles. In addition to the antibiotic concentration, the establishment of resistance alleles may be affected by interactions with the surrounding susceptible cells from which they derive, for instance via the release of nutrients or removal of the antibiotic. Here, we investigate the effects of social interactions with surrounding susceptible cells on the establishment of Escherichia coli mutants with increasing β-lactamase activity (i.e., the capacity to hydrolyze β-lactam antibiotics) from single cells under the exposure of the antibiotic cefotaxime (CTX) on agar plates. We find that relatively susceptible cells, expressing a β-lactamase with very low antibiotic-hydrolyzing activity, increase the probability of mutant cells to survive and outgrow into colonies due to the active breakdown of the antibiotic. However, the rate of breakdown by the susceptible strain is much higher than expected based on its low enzymatic activity. A detailed theoretical model suggests that this observation may be explained by cell filamentation causing delayed lysis. While susceptible cells may hamper the spread of higher-resistant β-lactamase mutants at relatively high frequencies, our findings show that they promote their initial establishment.