Minimum Lethal Dose Research Articles

Wound healing activity of methanolic extract of Turbinella pyrum from Gulf of Mannar, India

Molluscs are a promising source of bioactive substances and are virtually untapped resources of novel compounds. Many novel metabolites with potent pharmacological properties have been discovered in marine organisms in recent years. The present study aimed to investigate the wound healing activity of a methanolic extract of the marine gastropod Turbinella pyrum using an excision and incision wound model. To determine the minimum lethal dose, acute oral toxicity studies were performed as per the Organization for Economic Co-operation and Development (OECD) guidelines. Excision and incision wound models were used to evaluate the wound healing activity of the methanolic extract of T. pyrum by preparing a simple ointment base BP at different concentrations using five groups of Wistar albino rats. Group I (control) received a topical application of the simple ointment BP. Group II (Standard) was treated with topical application of 5% framycetin sulphate cream, and Groups III, IV and V were treated with 5%, 10% and 15% methanolic extracts of T. pyrum. Student's t test was used to analyse the results obtained from the present study, and P<0.05 was considered significant. In the incision wound model, the epithelialization period was also found to be highly significant (P<0.05) in group V (10.77%) when compared to the standard (12.69%). In the excision wound model, the epithelialization period was also found to be highly significant (P<0.05) in group IV (13.17%) and group V (11.38%) when compared to that of the standard (Group II) (14.56%).This finding justifies that the methanolic extract of T. pyrum has properties that render it capable of promoting accelerated wound-healing activity.

ITRAQ-Based Quantitative Proteomic Analysis of Acinetobacter baumannii under Hypoxia and Normoxia Reveals the Role of OmpW as a Virulence Factor.

ABSTRACTAcinetobacter baumannii needs to adapt to hypoxia during infection. Understanding its proteome regulation during infection would allow us to determine new targets to develop novel treatments. iTRAQ proteomic analysis of A549 cell infection by the ATCC 17978 strain was performed. A total of 175 proteins were differentially expressed under hypoxia versus normoxia. We selected the hypoxia-downregulated protein OmpW to analyze its role as a virulence factor. The loss of OmpW decreased the adherence and invasion of A. baumannii in these host cells, without affecting its bacterial growth. Moreover, A549 cell viability with ΔOmpW infection was higher than that with the wild-type strain. ΔOmpW presented less biofilm formation. Finally, the minimum lethal dose required by the ΔOmpW mutant was higher than that of the wild-type strain in a murine peritoneal sepsis model, with lower bacterial loads in tissues and fluids. Therefore, OmpW seems to be a virulence factor necessary for A. baumannii pathogenesis.IMPORTANCE Acinetobacter baumannii causes infections that are very difficult to treat due to the high rate of resistance to most and sometimes all of the antimicrobials used in the clinical setting. There is an important need to develop new strategies to combat A. baumannii infections. One alternative could be blocking specific bacterial virulence factors that this pathogen needs to infect cells. Pathogens modulate their protein expression as a function of the environment, and several studies have reported that hypoxia occurs in a wide range of infections. Therefore, it would be interesting to determine the proteome of A. baumannii under hypoxia in order to find new virulence factors, such as the outer membrane protein OmpW, as potential targets for the design of novel therapies.

Investigation and Identification of Food Poisoning Caused by Clostridium botulinum Type B1 in Shenzhen, China.

Clostridium botulinum produces botulinum neurotoxins (BoNTs), which cause people who ingest them to become seriously ill and sometimes die. In recent years, sporadic food poisoning cases associated with C. botulinum have occurred across the world. In 2016, two men were admitted to our hospital in Shenzhen, China, with foodborne botulism. In this study, we report on these two typical C. botulinum-related food poisoning incidents and the steps taken to identify and characterize the causative pathogen. We characterized the bacterial pathogen isolated from the first patient using cooked meat medium and egg yolk agar bacterial cultures under anaerobic conditions, and morphologically identified the isolate using Gram staining. The in vivo bioassay results in mice showed that the minimum lethal dose of the BoNTs produced by our isolate was 0.001-0.0001 mg/mL (LD50 of the culture was estimated to be 1.5812 mg/kg). Whole genome sequencing (WGS) results showed that the isolate was identified as C. botulinum B1 Okra. The causative strain was successfully isolated from the intestinal lavage fluid collected from the initial patient.

In Vitro Efficacy of Antimicrobial Photodynamic Therapy With Phycocyanin and Diode Laser for the Reduction of Porphyromonas gingivalis.

Introduction: Implant surface disinfection is the most difficult phases in treatment of peri-implantitis. This study sought to assess the efficacy of antimicrobial photodynamic therapy (aPDT) with phycocyanin and diode laser for the reduction of Porphyromonas gingivalis. Methods: In this in vitro, experimental study, first the minimum inhibitory concentration (MIC) of phycocyanin, the sub-lethal exposure time of the diode laser, and the sub-lethal dose of aPDT were determined. The wells containing P. gingivalis suspension were randomly divided into three experimental groups for exposure to ½ MIC phycocyanin for 5 minutes, minimum lethal dose of diode laser (635 nm, 4 minutes), and aPDT with phycocyanin and diode laser. The positive control group was exposed to 0.2% chlorhexidine (CHX) for 5 minutes, and the negative control group received no treatment. The colony count was calculated in the five groups and compared using one-way ANOVA and t test. Results: aPDT with a diode laser (635 nm, 4 minutes) and 125 µg/mL phycocyanin caused a significantly greater reduction in P. gingivalis count (mean reduction of 44.24%) compared with other groups (P<0.0001). Minimum and maximum colony counts were noted in 0.2% CHX and negative control groups respectively. The reduction in the colony count was significant in all experimental groups, compared with the control group (P<0.0001). Conclusion: aPDT with 635-nm diode laser and phycocyanin can significantly decrease P. gingivalis count in vitro. Considering the conservative nature of this modality, it may be used for the decontamination of peri-implant.

A simple method for purification of epsilon toxin of Clostridium perfringens type D for serum neutralization assay

Enterotoxaemia, a disease that affects domestic ruminants, is caused by the epsilon toxin of Clostridium perfringens type D and B. Control and prophylaxis are based on systemic vaccination of small ruminant herds with epsilon toxoid. Purified epsilon toxin is an essential material for vaccine evaluation. It is also necessary for diagnosis of enterotoxaemia disease in the field by in vitro tests including ELISA. The aim of this study was to set up a method for preparation of functional purified epsilon toxin of C. perfringens type D to be used in serum neutralization test. In this study, epsilon toxin was prepared from C. perfringens type D culture precipitated with ammonium sulfate, dialyzed against phosphate buffered saline (PBS) buffer and then, purified using chromatography system. Then, the purified epsilon toxin was detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Toxin function was confirmed by cell culture and minimum lethal dose (MLD) assays. Also rabbits were immunized by vaccine in two turns with a 28-day interval. Then, blood samples were collected, and serum neutralization (SN) test was carried out. Results showed that the purified toxin was suitable for SN assay. Our purification method was simple, fast and cost-effective for preparation of epsilon toxin.

Characteristics of a Bacteriophage, vB_Kox_ZX8, Isolated From Clinical Klebsiella oxytoca and Its Therapeutic Effect on Mice Bacteremia.

Klebsiella oxytoca is an important nosocomial and community-acquired opportunistic pathogenic Klebsiella and has become the second most prevalent strain in the clinic after K. pneumoniae. However, there have been few reports of bacteriophages used for treating K. oxytoca. In this study, a novel bacteriophage, vB_Kox_ZX8, which specifically infects K. oxytoca AD3, was isolated for the first time from human fecal samples. The biological characteristics of vB_Kox_ZX8 showed an incubation period of 10 min, a burst size of 74 PFU/cell, and a stable pH range of 3–11. Genomic bioinformatics studies of vB_Kox_ZX8 showed that it belongs to the genus Przondovirus, subfamily Studiervirinae, family Autographiviridae. The genome of vB_Kox_ZX8 is 39,398 bp in length and contains 46 putative open reading frames encoding functional proteins, such as DNA degradation, packaging, structural, lysin-holin, and hypothetical proteins. We further investigated the efficacy of vB_Kox_ZX8 phage in the treatment of mice with bacteremia caused by K. oxytoca infection. The results showed that vB_Kox_ZX8 (5 × 109 PFU/mouse) injected intraperitoneally alone was metabolized rapidly in BALB/c mice, and no significant side effects were observed in the control and treatment groups. Importantly, intraperitoneal injection with a single dose of phage vB_Kox_ZX8 (5 × 107 PFU/mouse) for 1 h post-infection saved 100% of BALB/c mice from bacteremia induced by intraperitoneal challenge with a minimum lethal dose of K. oxytoca AD3. However, all negative control mice injected with PBS alone died. Owing to its good safety, narrow host infectivity, high lysis efficiency in vitro, and good in vivo therapeutic effect, phage vB_Kox_ZX8 has the potential to be an excellent antibacterial agent for clinical K. oxytoca-caused infections.

Acute toxicity, 28-day repeated-dose toxicity and toxicokinetic study of timosaponin BII in beagle dogs

The safety evaluation of timosaponin BII (TBII) in beagle dogs with toxicokinetic study was performed. For the acute oral toxicity study, the minimum lethal dose (MLD) of TBII was more than 2000 mg/kg and suggested the characteristics of absorption saturation. For the 28-day repeated dose oral toxicity and toxicokinetic studies, there was no significant effect on all test parameters except for prolonged APTT in the 60 and 180 mg/kg groups, which recovered after withdrawal. The increase of drug exposure of 180 mg/kg group was not proportional to the increase of administration dose, showing the characteristics of absorption saturation.

Reduced pathogenicity of velogenic NDV strain AF22420-I via site-directed mutagenesis of V gene

Newcastle disease virus (NDV), an avian paramyxovirus, has the potential to be used as an anti-cancer therapeutic vaccine due to its oncolytic and immunostimulatory activities. The virus can be categorised into three pathotypes: lentogenic, mesogenic, and velogenic; of the three pathotypes, the lentogenic strains such as the La Sota are the preferred pathotype for vaccine development due to their low virulence to birds. On the other hand, the translation of the virus to clinic of the velogenic strain AF2240-I is hindered by its virulence towards birds although it exhibits strong oncolysis with significant outcomes both in vitro and in vivo. This study aims to reduce the pathogenicity of AF2240-I yet retaining the anti-cancer properties of the virus. To achieve this, the V protein that acts as an interferon antagonist was chosen to be mutated. It is a non-structural protein that does not interfere with the binding and infection of the virus; hence, mutation of this virulence factor was deducted to be able to reduce harm to the avian species but retain its anti-cancer properties as much as possible. The V protein, which was produced from the insertion of an additional G into a conserved editing site of the P gene, was mutated by substituting the G nucleotide at position 411 from the start of P gene to a T nucleotide. This mutation will produce a premature stop codon from the V mRNA, resulting in a truncated V protein; but only causes a silent mutation in the P protein. The recombinant virus was recovered by the use of BHK cells stably expressing the phage T7 RNA polymerase. The pathogenicity of the mutated virus was determined in 9- to 11-day-old embryonated SPF chicken eggs. The mean death time (MDT) was determined to be 73.6 hours at the minimal lethal dose of 10-7, resembling to that of a mesogenic strain. The virulence of the mutated virus has been successfully reduced where it could be potentially used as the vector for the development of recombinant oncolytic virus for cancer treatment.

A new practical purification method for type D Clostridium perfringens Epsilon toxin by Size-exclusion chromatography (SEC) and Ultrafiltration (UF)

BACKGROUND: The high potential toxicity of epsilon toxin (Etx) produced by Clostridium perfringens (C. perfringens) type D, has made it the third most lethal clostridial toxin behind botulinum and tetanus, therefore, having a pure and concentrated Etx is very important. OBJECTIVES: The aim of this study was to purify Etx as pure as possible with an applicable, cost-effective, multistep purification protocol with the lowest and shortest time. METHODS: The purification of the Etx was carried out in multiple consecutive steps; ammonium sulfate precipitation, dialysis, size exclusion chromatography by G50, two concentration steps, and ultrafiltration. The Etx activity after different steps was evaluated by the minimum lethal dose (MLD) calculation, according to the standard operating procedure. Toxin quantification was determined using Lowry technique, and its presence and specificity was tracked to identify pure Etx by SDS- PAGE and western blotting. Finally, the purity of Etx was evaluated by capillary electrophoresis. RESULTS: The purified Etx formed a single band of about 32.9 kDa in SDS-PAGE and blotting. The pure Etx concentration was calculated to be 3.9 mg/ml and its MLD value was the dilution of 1/24000 after the ultrafiltration step. The presented purification processes to purify Etx resulted in ~ 87-fold concentration and 88.6% purity. CONCLUSIONS: Due to this high Etx purity, the processes used in this study can provide the technical knowledge of toxin production in a larger industrial scale that can be used in development of clostridial toxoid vaccines, as well as quality control and/or diagnostic tests.

The Inhibitory Efficiencies of Geraniol as an Anti-Inflammatory, Antioxidant, and Antibacterial, Natural Agent Against Methicillin-Resistant Staphylococcus aureus Infection in vivo

IntroductionAntibiotics wee widely used as feed additives in animal husbandry. With the increase of drug resistance of bacteria, there is an urgent need to find alternatives to antibiotics. Clinically, methicillin-resistant Staphylococcus aureus (MRSA) infections account for about 25% to 50% of Staphylococcus aureus infections worldwide. Similarly, it is also one of the pathogens that cause serious animal infections.MethodsWe established a mouse model of systemic infection of MRSA to study the preventive effect of geraniol on MRSA and the immunomodulatory effect of geraniol. The mice in the experiment were injected with geraniol by intramuscular injection and were fed intraperitoneally with minimum lethal dose of MRSA. Then, the survival rate, inflammatory cytokines, oxidative stress factors in serum were measured. These values were used to estimate the bacterial load in different organs and to assess histopathological changes in the lungs, liver and kidneys.ResultsThe above-mentioned two ways of using geraniol could prevent MRSA infection in vivo in mice and showed a significant dose–response relationship. In other words, geraniol significantly decreased the concentrations of inflammatory cytokines and oxidative stress factors in MRSA-infected mice. At the same time, the level of glutathione peroxidase also increased in a dose–proportional relationship. In the group of mice treated with geraniol, their superoxide dismutase levels were significantly higher than those in the vancomycin. After treatment with geraniol, the burden of MRSA decreased. No obvious histopathological abnormalities were found in the liver and kidney of MRSA-infected mice. In addition, geraniol improved the inflammatory changes in the lungs.ConclusionThe results indicated that geraniol was a natural substance that could be used as an anti-inflammatory, antioxidant and antibacterial substance to protect mice from MRSA systemic infection. Generally, the research shows that as a natural medicine, geraniol has broad potential in the development and application of antibiotic substitutes.

Efficacy assessment of a triple anthrax chimeric antigen as a vaccine candidate in guinea pigs: challenge test with Bacillus anthracis 17 JB strain spores

Context Bacillus anthracis secretes a tripartite toxin comprising protective antigen (PA), edema factor (EF), and lethal factor (LF). The human anthrax vaccine is mainly composed of the anthrax protective antigen (PA). Considerable efforts are being directed towards improving the efficacy of vaccines because the use of commercial anthrax vaccines (human/veterinary) is associated with several limitations. Objective In this study, a triple chimeric antigen referred to as ELP (gene accession no: MT590758) comprising highly immunogenic domains of PA, LF, and EF was designed, constructed, and assessed for the immunization capacity against anthrax in a guinea pig model. Materials and methods Immunization was carried out considering antigen titration and immunization protocol. The immunoprotective efficacy of the ELP was evaluated in guinea pigs and compared with the potency of veterinary anthrax vaccine using a challenge test with B. anthracis 17JB strain spores. Results The results demonstrated that the ELP antigen induced strong humoral responses. The T-cell response of the ELP was found to be similar to PA, and showed that the ELP could protect 100%, 100%, 100%, 80% and 60% of the animals from 50, 70, 90, 100 and 120 times the minimum lethal dose (MLD, equal 5 × 105 spore/ml), respectively, which killed control animals within 48 h. Discussion and conclusions It is concluded that the ELP antigen has the necessary requirement for proper immunization against anthrax and it can be used to develop an effective recombinant vaccine candidate against anthrax.

ASSESSMENT OF ACUTE AND SUB CHRONIC TOXICITY OF DEOXYCHOLIC ACID ON ORAL ADMINISTRATION IN RODENTS

Deoxycholic acid (DCA) is one of the secondary bile acids, which are metabolic byproducts of intestinal bacteria and used traditionally to treat immune related disorders. The objective of the present study was to evaluate acute and sub-acute toxicity of DCA in mice and rats after oral administration following OECD test guidelines. Toxicological profile of DCA were evaluated by single-oral toxicity (5, 10, 100 and 200 mg/kg), 14-day repeated oral dose toxicity (1, 2.5 and 5 mg/kg) in mice and 28-days repeated-oral dose toxicity (30, 100, 300 and 1000 mg/kg) in healthy SD rats. Oral single dose of DCA caused decreased body weight and the mice were found lethargic. DCA was lethal at doses ≥ 10 mg/kg and caused mortality/morbidity by 48 hours post dosing. In 14-days repeated oral toxicity study with DCA as oral gavage at 1, 2.5 and 5 mg/kg, no effects on body weight and no treatment related clinical signs were observed. 28-day repeated oral toxicity demonstrates dose-dependent significant increase in WBC (up to 43%), absolute neutrophil (up to 61%) and absolute lymphocyte (up to 42%) counts suggesting an immunostimulatory role on humoral immunity. DCA at doses &gt;1000 mg/kg was toxic and caused anemia. Minimum lethal dose (MLD) and MTD of DCA in mice was estimated to be equal or greater than 10 mg/kg and 5 mg/kg/day, respectively. Oral NOAEL of DCA was30 mg/kg/day in SD rats. DCA did not cause any delayed effects. DCA appears to be safe for human consumption.

New technology to improve the thermal stability of botulinum toxin type D by biomimetic mineralization

The advanced biomimetic mineralization technology was applied to protect the Botulinum neurotoxin type D, and the processing of the mineralization granule of botulinum toxin type D was successfully screened. The loss of activity of the toxin protein at different temperatures and the destructive strength of the gastrointestinal tract against the toxin were determined biologically. The lethal toxicity of the mineralized toxin to wild rodents was determined by median lethal dose. Protective tests at different temperatures showed that the preservation period of botulinum toxin type D mineralized sample 2 was significantly higher than that of the control group at three different temperatures, and its toxicity loss was significantly reduced. The damage intensity of the mineralized toxin to the gastrointestinal contents of plateau zokor and plateau pika was significantly reduced. The minimum lethal doses of the mineralized toxin particles to plateau zokor, plateau pika, and mice were 5200, 8,600,000, and 25,000 MLD/kg. These results showed that biomimetic mineralization could greatly improve the thermal stability of botulinum toxin type D and reduce the damaging effect of the gastrointestinal contents of target animals to botulinum toxin type D. The mineralized toxin could be used to control the population density of urban rodents. This research provides new insights into the protection of toxin protein substances.

Prenylated Trans-Cinnamic Esters and Ethers against Clinical Fusarium spp.: Repositioning of Natural Compounds in Antimicrobial Discovery

Onychomycosis is a common nail infection mainly caused by species belonging to the F. oxysporum, F. solani, and F. fujikuroi species complexes. The aim of this study was to evaluate the in vitro susceptibility of six representative strains of clinically relevant Fusarium spp. toward a set of natural-occurring hydroxycinnamic acids and their derivatives with the purpose to develop naturally occurring products in order to cope with emerging resistance phenomena. By introducing a prenylated chain at one of the hydroxy groups of trans-cinnamic acids 1–3, ten prenylated derivatives (coded 4–13) were preliminarily investigated in solid Fusarium minimal medium (FMM). Minimal inhibitory concentration (MIC) and lethal dose 50 (LD50) values were then determined in liquid FMM for the most active selected antifungal p-coumaric acid 3,3′-dimethyl allyl ester 13, in comparison with the conventional fungicides terbinafine (TRB) and amphotericin B (AmB), through the quantification of the fungal growth. Significant growth inhibition was observed for prenylated derivatives 4–13, evidencing ester 13 as the most active. This compound presented MIC and LD50 values (62–250 µM and 7.8–125 µM, respectively) comparable to those determined for TRB and AmB in the majority of the tested pathogenic strains. The position and size of the prenylated chain and the presence of a free phenol OH group appear crucial for the antifungal activity. This work represents the first report on the activity of prenylated cinnamic esters and ethers against clinical Fusarium spp. and opens new avenues in the development of alternative antifungal compounds based on a drug repositioning strategy.

Susceptibility pattern of Salmonella enterica against commonly prescribed antibiotics, to febrile-pediatric cases, in low-income countries

BackgroundIn most low-income countries, febrile-pediatric-cases are often treated empirically with accessible antibiotics without periodic epidemiological surveillance, susceptibility testing, or minimal lethal dose calculations. With this backdrop, the study was undertaken to evaluate the susceptibility trend of Salmonella enterica against the commonly prescribed antibiotics.MethodsAll isolates of Salmonella enterica were identified by standard protocols of biotyping and serotyping, then tested against antibiotics by the modified Kirby disk-diffusion method. Minimum Inhibitory Concentration (MIC) of isolates was determined by the agar-dilution method and compared with disk diffusion results and on nalidixic-acid sensitive/resistant strains.ResultsAmong 1815 febrile-pediatric patients, 90(4.9%) isolates of Salmonella enterica [serovar: Salmonella Typhi 62(68.8%) and Salmonella Paratyphi A 28(31.1%)] were recovered. The incidence of infection was higher among males, age groups 5 to 9, and patients enrolling in the out-patient department (OPD). On the disk-diffusion test, most isolates were sensitive against first-line drugs i.e.cephalosporins, and macrolides. However, against quinolones, a huge percentile 93.3%, of isolates were resistant [including 58 Typhiand 26 Paratyphiserovar] while nearly 14% were resistant against fluoroquinolones.When MICs breakpoint were adjusted as follows: 4 μg/ml for azithromycin, ≥1 μg/ml for ciprofloxacin, 2 μg/ml for ofloxacin, 8 μg/ml for nalidixic acid, and 1 μg/ml for cefixime, higher sensitivity and specificity achieved. Compared to other tested antibiotics, a low rate of azithromycin resistance was observed. Nevertheless, higher resistance against fluoroquinolones was observed on NARS strain.ConclusionHigher susceptibility of Salmonella enterica to the conventional anti-typhoidal drugs (amoxicillin, chloramphenicol, cotrimoxazole, cephotaxime) advocates for its reconsideration. Although, the lower susceptibility against fluoroquinolones among nalidixic-acid-resistant Salmonella (NARS) strain negates its empirical use among the study age group.

Synthesis and Biological Evaluation of 10-Substituted Camptothecin Derivatives with Improved Water Solubility and Activity.

Despite remarkable clinical achievements, camptothecin (CPT) still suffers from poor solubility and severe toxicity. Therefore, it is necessary to redevelop CPT derivatives as supplementary antitumor agents with good water solubility and small side effects. In this work, 27 camptothecin derivatives were synthesized and screened for their cytotoxicity against A549 (lung) and HCT-116 (colon) cancer cell lines. Among them, compound B7, 7-ethyl-10-(2-oxo-2-(4-methylpiperidin-1-yl)ethoxy)camptothecin,was demonstrated in vitro to be a more potent antitumor agent than SN-38 by comparison of their inhibitory activities against cell proliferation and colony formation and interference effect on process of cell cycle and cell apoptosis. Additionally, a molecular docking model revealed that B7 can interact with the topoisomerase I-DNA complex, and that the solubility of B7 reached 5.73 μg/mL in water. Moreover, B7 significantly inhibited tumor growth in an A549 xenograft model at dosages of 0.4 and 2.0 mg/kg, and exhibited minimum lethal doses comparable to those of irinotecan. These results indicated that B7, with improved solubility, enhanced activity and acceptable acute toxicity, can be used as a lead compound for the development of novel anticancer agents.

Efficacy of dual carbapenem treatment in a murine sepsis model of infection due to carbapenemase-producing Acinetobacter baumannii.

ObjectivesTo evaluate the in vivo efficacy of a dual carbapenem combination containing imipenem plus meropenem against carbapenem-resistant Acinetobacter baumannii producing carbapenemases OXA-23 or OXA-58.MethodsAn experimental model of peritonitis using C57BL/6J female mice was developed and the minimum lethal doses were calculated for infections due to OXA-23 or OXA-58 producers of A. baumannii clinical isolates. The efficacies of the carbapenems in monotherapy and in combination were tested.ResultsMeropenem was better than imipenem in mice infected with either of the carbapenem-resistant A. baumannii (CRAb) strains. The combination of meropenem plus imipenem significantly improved the clearance of CRAbs from spleen compared with non-treated groups. The carbapenem-containing combination was better than imipenem for treating mice infected with both carbapenemase producers. In blood, the carbapenem combination significantly decreased the bacterial load of the OXA-23 producers compared with imipenem or meropenem used in monotherapy.ConclusionsThese results suggest that dual carbapenem combination could be an option for the treatment of infections due to carbapenemase-producing A. baumannii such as OXA-23 and OXA-58 producers.

Toxicological Profile of the Pain-Relieving Antioxidant Compound Thioctic Acid in Its Racemic and Enantiomeric Forms.

Thioctic acid is a multipotent antioxidant compound existing as dextrorotatory (+), eutomer and naturally occurring and levorotatory (−). It has been proven to help fight many pathologies and is sold as racemate. In agreement with studies claiming a greater biopotency of the eutomer compared to the levorotatory compound, we recently preclinically and clinically showed that (+) thioctic acid is a pain-reliever as effective as double-dosed racemate. We investigated acute and subchronical toxicity of (+/−) thioctic acid, (−) thioctic acid, (+) thioctic acid and (+) salt thioctic acid on Sprague–Dawley rats. For acute toxicity, compounds were administered intraperitoneally (i.p.) with a single-injection at 125, 240, 360, 480 µmol/kg, then rodents were tested for motorial coordination and minimum lethal dose (LDmin). A subtoxic dose (360 µmol/kg) was administered i.p. for 15 days and we finally evaluated motorial impairment, glycemia, organ toxicity, and apoptosis state. Acutely administered, the highest doses of all thioctic acid compounds negatively affected motorial ability and (−) thioctic acid LDmin resulted higher than the others. Subchronic administrations caused overall body weight loss, motorial impairment, mass loss in some organs. (+/−) and (−) thioctic acid injections enhanced caspase-3 activity in some organs, (−) enantiomer-treated animals displayed more marked organ toxicity signs. Together with our previous study on the biologic role of enantiomers, these data suggest a therapeutic use of (+) enantiomer-based formulations, thus lowering dose and toxicity without affecting the positive effects brought by the drug.

The possible mechanisms of copper resistance in the pathogen Pseudomonas syringae pathovars in stone fruit trees

One of the most common diseases affecting Stone-fruits is Bacterial Canker, caused by Pseudomonas syringae pv. syringae (Pss) and morsprunorum (Psm). In this study, Pss and Psm were isolated from stone-fruit trees from the Western Aegean region (WA) and Lake Van Basin (LVB) of Turkey, where pesticide usage is high and low, respectively. We aimed to determine copper resistance levels, the involved resistance mechanisms and the reactions to copper pesticides of these isolates. The minimum inhibitory concentration (MIC) and lethal dose 50 (LD50) of the isolates were detected using CuSO4 and other Cu-pesticides. To determine the mechanisms of copper resistance, the presence of copA and cusA genes was investigated. The modified Fe/Cu Blue-CAS Agar media were used to investigate the relationship between the isolates’ copper resistance and each isolate’s ability to produce siderophores. The highest MIC value was 2 mM in CuSO4. The tolerance levels of the isolates from the WA and LVB regions were 29% and 47% susceptible, 29% and 47% low resistance and 42% and 6% resistant, respectively. The most successful pesticide was (CuSO4 + Ca (OH)2) + mancozeb. While none of the isolates had the cusA gene, four isolates had the copA gene, which was proven to be plasmid-borne. Differences between copA gene sequences were detected and were determined to be not related to the pathovars. The amount of siderophore produced against copper in Ps pathovars affected seems to be related with the resistance level. Also, Ps pathovars were able to tolerate copper at doses as high as 1.1 mM by producing siderophores, and at doses of 1.7 mM and above through the copA gene.

F10, a new camptothecin derivative, was identified as a new orally–bioavailable, potent antitumor agent

Topoisomerases are interesting targets for drug discovery. In the present study, we attached saturated carbon atoms to the 10-position of camptothecin and synthesized 10 new camptothecin derivatives from 10-HCPT or SN-38. The activities of new compounds were evaluated both in vitro and in vivo. The most promising compound F10, 7-ethyl-10-(2-oxo-2-(piperidin-1-yl)ethoxy)camptothecin, inhibited cancer cells growth with the IC50 of 0.002, 0.003, 0.011 and 0.081 μM on Raji, HCT116, A549 and Lovo cells, respectively. Meanwhile, oral administration of F10 remarkably suppressed the HCT116-xenograft tumor growth in the nude-mice model at the dosage of 0.5, 2.0 and 8.0 mg/kg in vivo. Intraperitoneal administration of F10 can completely inhibit Raji-xenograft tumor growth in established NPG mouse model at 2.0 and 4.0 mg/kg. In addition, the minimum lethal doses of F10 and SN-38 in mice by intravenous administration were 80 and 40 mg/kg (or 0.155, 0.102 mmol/kg), respectively. The solubility of F10 reached 9.86 μg/mL in a buffer solution of pH 4.5. The oral bioavailability of F10 achieved 22.4% in mice. The molecular docking model revealed that F10 can interact with topoisomerase I-DNA complex. Our findings indicate that F10 is a new orally-oavailable antitumor agent with potent anticancer effect. Furthermore, attaching a ring hydrophobic moiety to the 10-position of camptothecin provides a favorable approach in the optimization of camptothecin.