Lung microdialysis and in vivo PK/PD integration of cefquinome against Actinobacillus pleuropneumoniae in a porcine experimental lung infection model.

This study aim to explore the application of microdialysis in pharmacokinetic (PK) and pharmacodynamic (PD) integration of cefquinome against Actinobacillus pleuropneumoniae in a porcine experimental lung infection model. The model was established via intratracheal inoculation where average bacterial counts (CFU) in the lungs of infected pigs reached 6.57 log10 CFU/g after 3 h. The PK profiles of unbound cefquinome in lung dialysates were determined following intramuscular injection of single doses of 0.125, 0.25, 0.5, 1, 2, and 4 mg/kg. Lung dialysate samples were collected using microdialysis at a flow rate of 1.5 µL/min until 24 h. The PD studies were conducted over 24 h based on 10 intermittent dosing regimens and total daily doses ranged from 0.25 to 4 mg/kg and dosage intervals included 12 and 24 h. The lung tissue was collected after 24 h of treatment and homogenized for bacterial counts. The relationships between PK/PD parameters derived from lung dialysates and drug efficacy were analyzed using an inhibitory sigmoid Emax model. The percentage of time the free drug concentration exceeded the minimum inhibitory concentration (%fT > MIC) was the PK/PD index best describing the antimicrobial activity (R2 = 0.96) in the porcine experimental infection model. The %fT > MIC values required to achieve net bacterial stasis, 1, 2 and 3 log10 CFU/g reductions in the lung were 22.45, 28.86, 37.62, and 56.46%, respectively. Cefquinome exhibited time-dependent characteristics against A. pleuropneumoniae in vivo. These results provide valuable insights into the application of microdialysis in PK/PD integration model studies and optima regimen of cefquinome for the treatment of porcine respiratory diseases caused by A. pleuropneumoniae.

Pharmacokinetics and relative bioavailability study of two cefquinome sulfate intramammary infusions in cow milk.

In this study, two intramammary infusions of cefquinome sulfate were investigated for pharmacokinetics and bioavailability. Twelve lactating cows for each group were administered an effective dose of 75 mg/gland for cefquinome, with milk samples collected at various time intervals. The concentrations of cefquinome in milk at different times were determined by the UPLC-MS/MS method. Analyses of noncompartmental pharmacokinetics were conducted on the concentration of cefquinome in milk. Mean pharmacokinetic parameters of group A and group B following intramammary administration were as follows: AUClast 300558.57 ± 25052.78 ng/mL and 266551.3 ± 50654.85 ng/mL, Cmax 51786.35 ± 11948.4 ng/mL and 59763.7 ± 8403.2 ng/mL, T1/2 5.69 ± 0.62 h and 5.25 ± 1.62 h, MRT 7.43 ± 0.79 h and 4.8 ± 0.78 h, respectively. Pharmacokinetic experiments showed that the relative bioavailability of group B was 88.69% that of group A. From our findings, group B (3 g: 75 mg) shows a quicker drug elimination process than group A (8 g: 75 mg), which suggests that the withdrawal period for the new formulation may be shorter.

PK-PD integration of enrofloxacin and cefquinome alone and in combination against Klebsiella pneumoniae using an in vitro dynamic model.

Introduction: Klebsiella pneumoniae is classified as a critical pathogen in both animals and humans and infections can be fatal in chickens resulting in substantial economic losses. However, the misuse of antibiotics can also lead to drug resistance and a potential transmission chain between animals and humans. Three K. pneumoniae strains with different susceptibility phenotypes were chosen to study the pharmacokinetic/pharmacodynamic (PK/PD) integration of enrofloxacin (ENR) and cefquinome (CEQ) alone and in combination. Results: Checkerboard assay results indicated that the combination treatment for type strain ATCC 700603 was synergistic effect with a fractional inhibitory concentration index (FICI) of ≤0.5. The other two clinical strains demonstrated an additive effect (FICI >0.5 to ≤1). Furthermore, static time-kill curves indicated that enrofloxacin and cefquinome added singly were effective in killing K. pneumoniae at concentrations of >2 MIC and ≥1 MIC, respectively. Additionally, the combination of enrofloxacin and cefquinome led to an enhanced antibacterial activity of cefquinome. The dynamic time-kill curves indicated that enrofloxacin and cefquinome had bactericidal and bacteriostatic activities, respectively at ≥1.5 mg/L (single-dose) and 4 mg/L (8 h split-dose) causing a decrease in bacterial counts of ≥4.45 and >2 log10 CFU/mL. Enrofloxacin possessed no bacteriostatic effects against K. pneumoniae at a constant concentration of 1× MIC. Cefquinome used in combination with 1× MIC enrofloxacin exhibited bactericidal activity at ≥4 mg/L (12 h split-dose) with reductions of ≥3.65 log10 CFU/mL. The PK/PD parameters were also analyzed to determine the concentration and duration of the drugs needed to reduce bacteria by 3 log10 CFU/mL. For enrofloxacin alone, the AUC24h/MIC was 23.29 h and the Cmax/MIC was 3.18. For cefquinome alone, the %T > MIC was 48.66 and when used in combination with enrofloxacin was 18.04. The combined use of cefquinome and enrofloxacin can increase the antibacterial activity of cefquinome against K. pneumoniae under a 12-h split-dose regimen regardless of individual drug susceptibility. Discussion: The static and dynamic time-kill curves indicated that enrofloxacin exhibited concentration-dependent activity, while cefquinome exhibited time-dependent activity. In the in vitro dynamic model, enrofloxacin alone exhibited better antimicrobial effects against K. pneumoniae compared to cefquinome alone. However, the antibacterial effect of cefquinome can be enhanced by combining it with enrofloxacin. These findings suggest a potentially effective approach for combating K. pneumoniae infections.

Detection of Antibiotic Resistance in Feline-Origin ESBL Escherichia coli from Different Areas of China and the Resistance Elimination of Garlic Oil to Cefquinome on ESBL E. coli.

The development of drug-resistance in the opportunistic pathogen Escherichia coli has become a global public health concern. Due to the share of similar flora between pets and their owners, the detection of pet-origin antibiotic-resistant E. coli is necessary. This study aimed to detect the prevalence of feline-origin ESBL E. coli in China and to explore the resistance elimination effect of garlic oil to cefquinome on ESBL E. coli. Cat fecal samples were collected from animal hospitals. The E. coli isolates were separated and purified by indicator media and polymerase chain reaction (PCR). ESBL genes were detected by PCR and Sanger sequencing. The MICs were determined. The synergistic effect of garlic oil and cefquinome against ESBL E. coli was investigated by checkerboard assays, time-kill and growth curves, drug-resistance curves, PI and NPN staining, and a scanning electronic microscope. A total of 80 E. coli strains were isolated from 101 fecal samples. The rate of ESBL E. coli was 52.5% (42/80). The prevailing ESBL genotypes in China were CTX-M-1, CTX-M-14, and TEM-116. In ESBL E. coli, garlic oil increased the susceptibility to cefquinome with FICIs from 0.2 to 0.7 and enhanced the killing effect of cefquinome with membrane destruction. Resistance to cefquinome decreased with treatment of garlic oil after 15 generations. Our study indicates that ESBL E. coli has been detected in cats kept as pets. The sensitivity of ESBL E. coli to cefquinome was enhanced by garlic oil, indicating that garlic oil may be a potential antibiotic enhancer.

Elimination of Cefquinome Sulfate Residue in Cow's Milk after Intrauterine Infusion.

As set in the maximum residue limit regulations of the European Commission, this study aimed to obtain the residual parameters in milk with optimized UPLC-MS/MS conditions and to determine the conclusive drug withdrawal period to ensure food safety. In this research, an ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed to study cefquinome sulfate's residue elimination in milk and to calculate cefquinome's withdrawal period. Twelve healthy cows free of endometritis were selected for the experiment. Before using the drug, the vaginal orifice and perineum of each cow was disinfected. One dose of intrauterine perfusion was used for each cow, followed by an additional dose after 72 h. Before administration and 12 h, 18 h, 24 h, 36 h, 42 h, 48 h, 60 h, 66 h, 72 h, 84 h, 90 h, and 96 h after the last dose, milk (10 mL) was gathered from each cow's teat and pooled. For the measurement of cefquinome concentrations in milk, UPLC-MS/MS was performed. A calibration curve was generated using linear regression as follows: Y = 250.86X - 102.29, with a correlation coefficient of 0.9996; the limits of detection and the limits of quantitation were 0.1 µg·kg-1 and 0.2 µg·kg-1, respectively. The average recovery of cefquinome was 88.60 ± 16.33% at 0.2 µg·kg-1, 100.95 ± 2.54% at 10 µg·kg-1, and 97.29 ± 1.77% at 50 µg·kg-1. For 5 consecutive days at the three spiking levels, the intra and inter-day relative standard deviations (RSD) were 1.28%-13.73% and 1.81%-18.44%, respectively; the residual amount of cefquinome was less than the maximum residue limit of 20 µg·kg-1, 36 h after administration; and the residual amount was less than the limit of detection (0.1 µg·kg-1) 48 h after administration. The withdrawal time of cefquinome in cow's milk was 39.8 h, as calculated using WTM1.4 software. In terms of clinical practical use, the withdrawal period of milk was temporarily set at 48 h after the administration of the cefquinome sulfate uterus injection to cows, in accordance with the recommended dose and course.

Low temperature synthesis of carbon dots in microfluidic chip and their application for sensing cefquinome residues in milk.

In this study, the N-doped carbon dots were continuously synthesized by a facile microfluidic strategy at 90 °C, and their quantum yields reached 19.2%. The characteristics of the obtained carbon dots could be real-time monitored in order to synthesize carbon dots with specific properties. By incorporating the carbon dots into a well-established enzymatic cascade amplification system, an inner filter effect-based fluorescence immunoassay was set up for ultrasensitive detection of cefquinome residues in milk samples. The developed fluorescence immunoassay provided a low detection limit of 0.78 ng/mL, which satisfied the maximum residue limit set by authorities. The fluorescence immunoassay had an 50% inhibition concentration of 0.19 ng/mL against cefquinome and showed a good linear relationship from 0.013 ng/mL to 1.52 ng/mL. While, the average recovery values ranged from 77.8% to 107.8% in spiked milk samples, with relative standard deviations ranging from 6.8% to 10.9%. Compared with conventional methods, the microfluidic chip was more flexible on carbon dots synthesis and the developed fluorescence immunoassay was more sensitive and eco-friendlier for ultra-trace cefquinome residue analysis.

Pharmacokinetics of cefquinome in rainbow trout (Oncorhynchus mykiss) after intravascular, intraperitoneal, and oral administrations.

This study aimed to determine the pharmacokinetics and bioavailability of cefquinome in rainbow trout (Oncorhynchus mykiss) following intravascular (IV), intraperitoneal (IP), and oral (PO) administrations at 14 ± 1°C. In this study, three hundred and six clinically healthy rainbow trout (110-140 g) were used. The fish received single IV, IP, and PO injections of cefquinome at 10 mg/kg dose. The plasma concentrations of cefquinome were measured using HPLC-UV and were evaluated using non-compartmental analysis. Cefquinome was measured up to 96 h for PO route and 144 h for IV and IP routes in plasma. Following IV administration, t1/2ʎz , ClT , and Vdss were 18.85 h, 0.037 L/h/kg, and 0.84 L/kg, respectively. The Cmax of IP and PO routes was 9.75 and 1.64 µg/ml, respectively. The bioavailability following IP and PO administrations was 59.46% and 12.33%, respectively. Cefquinome at 10 mg/kg dose may maintain T > MIC above 40% at 72 and 96 h intervals, respectively, following the IP and IV routes for bacteria with MIC values of ≤2 µg/ml and at 24 h intervals following the PO route for bacteria with MIC value of ≤0.75 µg/ml. However, further studies are needed to determine in vitro and in vivo antibacterial efficacy and multiple dosage regimens of cefquinome against pathogens isolated from rainbow trout.

Evaluation of cefquinome's efficacy in controlling avian colibacillosis and detection of its residues using high performance liquid chromatography (HPLC).

This study aimed to evaluate the efficacy of cefquinome in treatment and controlling of Escherichia coli experimentally infected broiler chickens, in addition of detection of its residues using High performance liquid chromatography (HPLC). In this study, 150 one-day old Cobb broiler chicks were used. On the 14th day chicks experimentally infected and divided into 3 equal groups (50 each); control group (G1) non-infected, non-treated, (G2) infected with E. coli O78 non treated, (G3) infected with E. coli O78 , cefquinome treated. Cefquinome was administrated 5th day post infection, intramuscularly by a dose of (2 mg/ kg b w.t) for 3 consecutive days. Experimental E. coli infection in broilers induced weakness, loss of appetite, depression, cough and watery diarrhea in addition to a recorded mortality (30%) with reduction in growth performance, erythrogram, total proteins, albumin, antioxidants and haemagglutination inhibition (HI) titers. In addition, a significant increase in feed conversion rate (FCR), leukocytic count, liver enzymes, kidney functions, total globulins, malondialdehyde, nitric oxide and lysozyme activity. Treatment with cefquinome led to decreased mortality rate, improvement in clinical signs, growth performance and modulated most of these altered parameters. Cefquinome's residues was not detected in breast muscles 3rd day and liver and kidneys 7th days post treatment. Therefore, it's recommended that cefquinome is a good choice for controlling of colibacillosis in broilers and its withdrawal time 3 days in breast muscles and 7 days in liver and kidney post treatment.

Cefquinome Sulfate Oily Nanosuspension Designed for Improving its Bioavailability in the Treatment of Veterinary Infections.

Introduction: Cefquinome sulfate (CS) is the first fourth-generation antibiotic for animals, which has a wide antibacterial spectrum, strong antibacterial activity and low drug resistance. However, it is accompanied by problems of poor therapeutic efficacy. In this context, the use of nanosuspensions have been found to be an attractive strategy. The main objective of this work is to develop a new oily nanosuspension to improve bioavailability and stability of CS formulations. Methods: After screening the formulations, cefquinome sulfate oily nanosuspension (CS-NSP) was prepared by mortar grinding, using propylene glycol dicaprolate/dicaprate (Labrafac™ PG) as oil medium and caprylocaproyl polyoxyl-8 glycerides (Labrasol®) as stabilizer. The properties of CS-NSP were investigated by testing its physicochemical characteristics, stability, in vitro release, hemolysis, and muscle irritation. The in vivo pharmacokinetics of CS-NSP was studied using rats. Results: Results show that CS-NSP presents suitable stability, physicochemical properties and safety. Moreover, a rapid release and high bioavailability of CS-NSP have also been verified in the study. Pharmacokinetic experiments in vivo showed that the bioavailability of CS-NSP was about 1.6 times that of commercial cefquinome sulfate injection (CS-INJ, Chuangdao®) (p<0.01). These advantages of CS-NSP were carried out by small particle size and low viscosity, being associated with the use of Labrafac PG and stabilizer Labrasol. Conclusion: The results proved that the new preparation is safe and effective and is expected to become a promising veterinary nanodelivery system.

Ex Vivo Pharmacokinetics and Pharmacodynamics Modeling and Optimal Regimens Evaluation of Cefquinome Against Bovine Mastitis Caused by Staphylococcus aureus.

Cefquinome, the fourth-generation cephalosporin applied solely for veterinary medicine, is commonly used for bovine mastitis caused by Staphylococcus aureus. The present study aims to establish an optimal dose and provide a PK/PD Cutoff value (COPD) for cefquinome against S. aureus based on ex vivo pharmacokinetics and pharmacodynamics (PK/PD) integration. This study investigated the pharmacokinetics (PK) of cefquinome when administered as three consecutive intramammary (IMM) doses of cefquinome in three healthy dairy cows at 75 mg/gland. Drug concentration was determined by HPLC-MS/MS assay. The ex vivo pharmacodynamics (PD) of cefquinome were evaluated by using a milk sample from a PK experiment. The relationship between the AUC/ MIC of cefquinome and bacterial loading reduction was simulated using a Sigmoid Emax model. The cefquinome concentration in milk attained a maximum level of 1.55 ± 0.21 mg/mL at 1.8 h after the third administration. The mean value of the area under the concentration-time curve (AUC0-24) was 26.12 ± 2.42 mg·h/mL after the third administration. The elimination half-life was 10.6 h. For PD profile, the MICs of cefquinome in milk were 2-4 times higher than those in the broth. In vitro time-killing curve shows that initial bacterial concentration has a huge impact on antibacterial effect on three strains. The antibacterial effect was weakened with the initial bacterial concentration increasing from 106 to 108 CFU/mL. The AUC0-24h/MIC index correlated well with ex vivo efficacy both for the initial inoculum of 106 CFU/mL and 108 CFU/mL (R 2 > 0.84). According to the inhibitory sigmoid Emax model analysis, the PK/PD surrogate (AUC0-24/MIC) values were 8,638, 1,397, and 3,851 for bactericidal effect (E = -3) with an initial inoculum of 106 CFU/mL, while the corresponding values were 12,266, 2,295, and 5,337, respectively, with the initial inoculum of 108 CFU/mL. The ex vivo PK/PD based population dose prediction indicated a target attainment rate (TAR) of 90% of 55 mg/gland/12 h. The COPD for cefquinome against S. aureus was 2 µg/mL under the recommended dose of 55 mg/gland/12 h. However, it should be validated in clinical practice in future investigations. These results contribute to the rational use of cefquinome for mastitis treatment in clinical veterinary medicine.

PK/PD integration and pharmacodynamic cutoff of cefquinome against cow mastitis due to Escherichia coli.

Cefquinome is the fourth generation of cephalosporin approved solely in animal usage. In order to slow down the resistance development of E. coli to cefquinome, and to protect and maintain the effectiveness of cefquinome, an ex vivo PK/PD modeling of cefquinome against E. coli in cows after intramammary infusion administration was conducted. The epidemiologic cutoff (ECOFF) and pharmacodynamic cutoff (COPD) of cefquinome against E. coli in lactation cows after intramammary infusion administration were recommended. The MICs of cefquinome against 1073 clinical E. coli isolates ranged from 0.015 to >64 µg/ml, and the ECOFF was defined as 0.125 µg/ml. The pharmacokinetic results showed that cefquinome maintained high concentration in milk for a long period with the T1/2ß of 10.60 h after intramammary infusion in dairy cows. The drug concentration in skimmed milk was still as high as 0.15 mg/ml after 48 h. Cefquinome displayed bacterial killing effect at 2× MIC with the initial inoculum of 106  cfu/ml in vitro; however, the same effect was attained with a concentration as high as 32× MIC with the initial inoculum of 108  cfu/ml both in artificial medium and in skimmed milk. The initial inoculum is an important factor on time-killing curve accounting for weakened killing pattern of cefquinome. The AUC0-24 h /MIC index correlated well with ex vivo efficacy. The AUC0-24 h /MIC values for bactericidal effect were 50, 016, and 67,644, respectively, for initial inoculum of 106 and 108  cfu/ml, indicating the bacterial loading or the severity of infection would infect the PK/PD modeling results. The ex vivo PK/PD-based population dose prediction indicated a target attainment rate (TAR) at the existing daily dose (75 mg/udder) of 84.77% against E. coli. Thus, it was recommended as rational dosage. The COPD of cefquinome against E. coli was determined as 8 µg/ml at the dose of 75 mg/udder. The derived ECOFF, COPD, together with ex vivo PK/PD-based population dose prediction served as important steps in the establishment of optimum dose regimen and provided a useful interpretative criterion to categorize the antimicrobial susceptibility testing results of cefquinome.

Kill Rate and Evaluation of Ex Vivo PK/PD Integration of Cefquinome Against Actinobacillus pleuropneumoniae.

We wished to study the detailed and precise antibacterial activity of cefquinome against Actinobacillus pleuropneumoniae (APP) in vitro and ex vivo. We analyzed the relationships between kill rate and cefquinome concentration in broth and between pharmacokinetic/pharmacodynamic (PK/PD) parameters and antibacterial effect in serum and tissue cage fluid (TCF) of piglets. Cefquinome exhibited time-dependent antibacterial activity against APP according to the kill rate. The maximum kill rate was 0.48 log10 CFU/mL/h at the 0-9-h period in broth. In the ex vivo PK/PD study, the maximum concentration (Cmax), time to reach the maximum concentration (Tmax), terminal half-life (T1/2ß), and area under the concentration time curve (AUCinfinity) were 5.65 µg/ml, 0.58 h, 2.24 h, and 18.48 µg·h/ml in serum and 1.13 µg/ml, 2.60 h, 12.22 h, and 20.83 µg·h/ml in TCF, respectively. The values of area under the curve during 24 h/minimum inhibitory concentration (AUC24h/MIC) for bacteriostatic, bactericidal, and bacterial eradication effects were 18.94, 246.8, and 1013.23 h in serum and 4.20, 65.81, and 391.35 h in TCF, respectively. Our findings will provide a valuable basis for optimization of dosage regimens when applying cefquinome to treat APP infection.

Disposition of Cefquinome in Turkeys (Meleagris gallopavo) Following Intravenous and Intramuscular Administration.

The bioavailability and pharmacokinetics in turkeys of cefquinome (CFQ), a broad-spectrum 4th-generation cephalosporin antibiotic, were explored after a single injection of 2 mg/kg body weight by intravenous (IV) and intramuscular (IM) routes. In a crossover design and 3-weeks washout interval, seven turkeys were assigned for this objective. Blood samples were collected prior to and at various time intervals following each administration. The concentration of CFQ in plasma was measured using HPLC with a UV detector set at 266 nm. For pharmacokinetic analysis, non-compartmental methods have been applied. Following IV administration, the elimination half-life (t1/2ʎz), distribution volume at steady state (Vdss), and total body clearance (Cltot) of CFQ were 1.55 h, 0.54 L/kg, and 0.32 L/h/kg, respectively. Following the IM administration, CFQ was speedily absorbed with an absorption half-life (t1/2ab) of 0.25 h, a maximum plasma concentration (Cmax) of 2.71 µg/mL, attained (Tmax) at 0.56 h. The bioavailability (F) and in vitro plasma protein binding of CFQ were 95.56% and 11.5%, respectively. Results indicated that CFQ was speedily absorbed with a considerable bioavailability after IM administration. In conclusion, CFQ has a favorable disposition in turkeys that can guide to estimate optimum dosage regimes and eventually lead to its usage to eradicate turkey's susceptible bacterial infections.

Antibacterial pathway of cefquinome against Staphylococcus aureus based on label-free quantitative proteomics analysis.

Cefquinome (CEQ) is a novel ß-lactam antibiotic that exhibits excellent antibacterial activity against Staphylococcus aureus. However, the bacterial protein targets of CEQ are unclear. To evaluate the relationship between the pharmacokinetic/pharmacodynamic (PK/PD) parameters of CEQ and strains with varying degrees of resistance and to elucidate bacterial protein responses to CEQ treatment, label-free quantitative proteomics analysis was conducted. The sensitive S. aureus ATCC6538 and the resistant 2MIC and 8MIC were tested for differentially expressed proteins. An in vitro model was treated with different concentrations of CEQ (3, 5, or 10 µg/ml) with different terminal half-lives (2.5 or 5 h) at different intervals (12 or 24 h). Differentially expressed proteins were evaluated using Gene Ontology analysis followed by KEGG pathway enrichment analysis and STRING network analysis. RT-qPCR was performed to validate the differentially expressed proteins at the molecular level. The results showed that the degree of resistance increased in a cumulative manner and increased gradually with the extension of administration time. The resistant strain would not have appeared in the model only if %T > mutant prevention concentration ≥ 50%. The expression of 45 proteins significantly changed following CEQ treatment, among which 42 proteins were obviously upregulated and 3 were downregulated. GO analysis revealed that the differentially expressed proteins were mainly present on cells and the cell membrane, participated in metabolic and intracellular processes, and had catalytic and binding activities. The RPSO, SDHB, CITZ, ADK, and SAOUHSC 00113 genes in S. aureus may play important roles in the development of resistance to CEQ. These results provided important reference candidate proteins as targets for overcoming S. aureus resistance to CEQ.

RNA-seq-based transcriptome analysis of a cefquinome-treated, highly resistant, and virulent MRSA strain.

The emergence and dissemination of methicillin-resistant Staphylococcus aureus (MRSA) strains of animal origin that are resistant to several antibiotics is of great concern. Cefquinome is a fourth-generation cephalosporin developed specifically for veterinary use. The mechanism of MRSA resistance to cefquinome is still not established. Therefore, we designed this study to evaluate the effect of cefquinome on the transcriptome of MRSA1679a, a strain that was isolated from a chicken. The transcriptome analysis indicated that multiple efflux pumps (QacA, NorB, Bcr, and ABCb) were upregulated in MRSA1679a as a resistance mechanism to expel cefquinome. Additionally, penicillin-binding protein 1A was overexpressed, which conferred resistance to cefquinome, a ß-lactam antibiotic. Adhesion and the biofilm-forming capacity of the MRSA strain was also enhanced in addition to overexpression of many stress-related genes. Genes related to carbohydrate metabolism, secretion systems, and transport activity were also significantly upregulated in MRSA1679a. In conclusion, global transcription was triggered to overcome the stress induced by cefquinome, and the MRSA1679a showed a great genetic potential to survive in this challenging environment. This study provides a profound understanding of MRSA1679a as a potentially important pathogen and identifies key resistance characteristics of MRSA against cefquinome. Studies should be aimed to demonstrate multidrug resistance mechanisms of virulent strains by exposing to different antibiotic combinations.

Determination of Susceptibility Breakpoint for Cefquinome against Streptococcus suis in Pigs.

Streptococcus suis (S. suis), a zoonotic pathogen, causes severe diseases in both pigs and human beings. Cefquinome can display excellent antibacterial activity against gram-negative and gram-positive bacteria. The aim of this study was to derive an optimal dosage of cefquinome against S. suis with a pharmacokinetic/pharmacodynamic (PK/PD) integration model in the target infection site and to investigate the cutoffs monitoring the changes of resistance. The minimum inhibitory concentration (MIC) distribution of cefquinome against 342 S. suis strains was determined. MIC50 and MIC90 were 0.06 and 0.25 µg/mL, respectively. The wild-type cutoff was calculated as 1 µg/mL. A two-compartmental model was applied to calculate the main pharmacokinetic parameters after 2 mg/kg cefquinome administered intramuscularly. An optimized dosage regimen of 3.08 mg/kg for 2-log10 CFU reduction was proposed by ex vivo PK/PD model of infected swine. The pharmacokinetic-pharmacodynamic cutoff was calculated as 0.06 µg/mL based on PK/PD targets. Based on the clinical effectiveness study of pathogenic MIC isolates, the clinical cutoff was calculated as 0.5 µg/mL. A clinical breakpoint was proposed as 1 µg/mL. In conclusion, the results offer a reference for determining susceptibility breakpoint of cefquinome against S. suis and avoiding resistance emergence by following the optimal dosage regimen.

Development and Validation of Liquid Chromatography-Tandem Mass Spectrometry Methods for the Quantification of Cefquinome, Ceftiofur, and Desfuroylceftiofuracetamide in Porcine Feces with Emphasis on Analyte Stability.

Cefquinome and ceftiofur are ß-lactam antibiotics used for the treatment of bacterial infections in swine. Although these antimicrobials are administered intramuscularly, the exposure of the gut microbiota to these cephalosporins is not well described. This exposure can contribute to the emergence and spread of antimicrobials in the environment and to the possible spread of antimicrobial resistance genes. To assess the impact of drug administration on the intestinal excretion of these antimicrobials it is essential to measure the amounts of native compound and metabolites in feces. Two (ultra)-high-performance liquid chromatography-tandem mass spectrometry ((U)HPLC-MS/MS) methods were developed and validated, one for the determination of cefquinome and ceftiofur and the other for the determination of ceftiofur residues, measured as desfuroylceftiofuracetamide, in porcine feces. The matrix-based calibration curve was linear from 5 ng g-1 to 1000 ng g-1 for cefquinome (correlation coefficient (r) = 0.9990 ± 0.0007; goodness of fit (gof) = 3.70 ± 1.43) and ceftiofur (r = 0.9979 ± 0.0009; gof = 5.51 ± 1.14) and quadratic from 30 ng g-1 to 2000 ng g-1 for desfuroylceftiofuracetamide (r = 0.9960 ± 0.0020; gof = 7.31 ± 1.76). The within-day and between-day precision and accuracy fell within the specified ranges. Since ß-lactam antibiotics are known to be unstable in feces, additional experiments were conducted to adjust the sampling protocol in order to minimize the impact of the matrix constituents on the stability of the analytes. Immediately after sampling, 500 µL of an 8 µg mL-1 tazobactam solution in water was added to 0.5 g feces, to reduce the degradation in matrix.

Selection of ESBL-producing Escherichia coli in the gut of calves experimentally fed with milk containing antibiotic residues.

In the bovine sector, the spread of Enterobacterales producing extended-spectrum and AmpC ß-lactamases (ESBL/AmpC) mostly concerns veal calves, and the use of waste milk containing antibiotic residues has been recurrently incriminated. In this study, calves were experimentally fed with milk containing either 2,000 µg/L or 20,000 µg/L of the critically important antibiotic cefquinome. The total counts of enterobacterales and ESBL-producing E. coli were monitored using non-selective and selective media. Our data highlighted the important combination of two main factors (cefquinome exposure and initial ESBL colonization level) in the ESBL selection and amplification process in the gut of calves. Results also proved the dose-independent effect of cefquinome administration on the selection and amplification of ESBL-producing E. coli. Finally, the blaCTX-M-1/IncI1 ST3 plasmid was systematically recovered after cefquinome exposure, highlighting its epidemic success. Altogether, this work is one of the rare experimental studies providing quantitative information on the impact of waste milk containing antimicrobials on the ESBL load in calves' microbiota, and the first one using cefquinome. These data emphasise the need for global guidelines on the use of waste milk on dairy farms in order to decrease the antimicrobial resistance burden in this sector.

Evidence for Establishing the Clinical Breakpoint of Cefquinome against Haemophilus Parasuis in China.

Haemophilus parasuis can cause high morbidity and mortality in swine. Cefquinome possesses excellent antibacterial activity against pathogens causing diseases of the respiratory tract. This study aimed to establish the clinical breakpoint (CBP) of cefquinome against H. parasuis and to monitor the resistance change. Referring to the minimum inhibitory concentration (MIC) distribution of cefquinome against 131 H. parasuis isolates, the MIC50 and MIC90 were determined to be 0.125 and 1 µg/mL, respectively. And the epidemiological cutoff (ECOFF) value was 1 µg/mL. HPS42 was selected as a representative strain for the pharmacodynamic (PD) experiment, pharmacokinetic (PK) experiment and clinical experiments. The PK/PD index values, area under concentration-time curve (AUC)/MIC, of the bacteriostatic, bactericidal, and bacterial elimination effects were 23, 41, and 51 h, respectively. The PK/PD cutoff was calculated as 0.125 µg/mL by Monte Carlo simulation (MCS), and the clinical cutoff was 0.25-4 µg/mL by WindoW. Combing these three values, the CBP of cefquinome against H. parasuis was found to be 1 µg/mL. In conclusion, this was the first study to integrate various cutoffs to establish the CBP in the laboratory. It is helpful to distinguish wild type H. parasuis and reduce the probability of treatment failure.

Comparative pharmacokinetics and pharmacokinetic/pharmacodynamic analysis by nonlinear mixed-effects modeling of cefquinome in nonpregnant, pregnant, and lactating goats after intravenous and intramuscular administration.

Cefquinome is a fourth-generation cephalosporin that is used empirically in goats. Different physiologic factors like pregnancy or lactation could determine the pharmacokinetic behavior of drugs in the organism. The objectives of this study are to (a) compare the pharmacokinetics of cefquinome after intravenous and intramuscular administration in adult nonpregnant (n = 6), pregnant (n = 6), and lactating goats (n = 6), at a dose of 2 mg/kg, with rich sampling by nonlinear mixed-effects modeling, (b) conduct a pharmacokinetic/pharmacodynamic analysis to evaluate the efficacy of the recommended posology in goats with different physiological states, and (c) determine the optimal posology that achieve a PTA value ≥ 90%, taking into account a T > MIC ≥ 60% of a MIC value ≤ 0.25 µg/ml, in the different subpopulations of goats for both routes. Gestation significantly increased Ka and V1, while reduced F0, Cl, and Q. On the other hand, lactation significantly increased V1 and reduced Tk0. Cefquinome concentrations achieved in placental cotyledon, amniotic fluid, and fetal serum indicate a minimal penetration across the placental barrier. Moreover, milk penetration of cefquinome was minimal. The total body clearance of cefquinome for goats was 0.29 L kg-1  hr-1 , that is apparently higher than the reported for cows (0.13 L kg-1  hr-1 ) and pigs (0.16 L kg-1  hr-1 ). So, the optimal dose regimen for cefquinome after intravenous and intramuscular administration required higher dose and frequency of administration compared with recommendations for cows or pigs. Therefore, 2 mg kg-1  8 hr-1 and 5 mg kg-1  12 hr-1 could be used for IV and IM routes, respectively, for the treatment of respiratory infections caused by P. multocida and M. haemolytica, but only 5 mg kg-1  12 hr-1 by both routes should be recommended for Escherichia coli infections.