BACKGROUND: Colibacillosis in broiler chickens is associated with economic loss and localized or systemic infection. Usually, the last resort is antibacterial therapy. Insight into the disease pathogenesis, host responses and plausible immunomodulatory effects of the antibacterials is important in choosing antibacterial agent and optimization of the treatment. Selected responses of broiler chickens experimentally infected with Escherichia coli (E. coli) and also those treated with florfenicol are evaluated in this study. Chickens (n = 70, 5 weeks old) were randomly assigned to four groups. The control groups included normal control (NC) and intratracheal infection control (ITC) (received sterile bacterial medium). The experimental groups consisted of intratracheal infection (IT) that received bacterial suspension and intratracheal infection with florfenicol administration (ITF) group.
RESULTS: Florfenicol reversed the decreased albumin/globulin ratio to the level of control groups (p > 0.05). Serum interleukin 10 (IL-10) and interferon-gamma (IFN-γ) concentrations decreased in IT birds as compared to NC group. Florfenicol decreased the serum interleukin 6 (IL-6) concentration as compared to IT group. Milder signs of inflammation, septicemia, and left shift were observed in the leukogram of the ITF group. Florfenicol decreased the severity of histopathological lesions in lungs and liver. Depletion of lymphoid tissue was detected in spleen, thymus and bursa of IT group but was absent in ITF birds. The number of colony forming units of E. coli in liver samples of ITF group was only slightly lower than IT birds.
CONCLUSIONS: Experimental E. coli infection of chickens by intratracheal route is associated with remarkable inflammatory responses as shown by changes in biochemical and hematological parameters. Histopathological lesions in lymphoid organs (especially in the spleen) were also prominent. Florfenicol has positive immunomodulatory effects and improves many of the lesions before the full manifestation of its antibacterial effects. These effects of florfenicol should be considered in pharmacotherapy decision-making process.

Antibiotic-induced dysbiosis in the fish gut causes significant adverse effects. We use fecal microbiota transplantation (FMT) to accelerate the restoration of florfenicol-perturbed intestinal microbiota in koi carp, identifying key bacterial populations and metabolites involved in the recovery process through microbiome and metabolome analyses. We demonstrate that florfenicol disrupts intestinal microbiota, reducing beneficial genera such as Lactobacillus, Bifidobacterium, Bacteroides, Romboutsia, and Faecalibacterium, and causing mucosal injuries. Key metabolites, including aromatic amino acids and glutathione-related compounds, are diminished. We show that FMT effectively restores microbial populations, repairs intestinal damage, and normalizes critical metabolites, while natural recovery is less effective. Spearman correlation analyses reveal strong associations between the identified bacterial genera and the levels of aromatic amino acids and glutathione-related metabolites. This study underscores the potential of FMT to counteract antibiotic-induced dysbiosis and maintain fish intestinal health. The restored microbiota and normalized metabolites provide a basis for developing personalized probiotic therapies for fish.

Knowledge on the characteristics of different infection models of avian colibacillosis is mandatory for performing reliable experimental studies. This study compares the characteristics of two infection models of colibacillosis (by intratracheal and subcutaneous routes) in broilers. Broilers (125), 35 days old, were randomly allocated to four experimental (n = 20) and three control (n = 15) groups: 1) negative control; 2) intratracheal infection control; 3) subcutaneous infection control; 4) intratracheal infection (IT); 5) intratracheal infection with florfenicol administration (ITF); 6) subcutaneous infection (SC); and 7) subcutaneous infection with florfenicol administration (SCF). Clinical signs and mortalities were recorded, and gross examination of carcasses was performed. On days 3 and 5 postchallenge (PC), 5 birds/group were euthanatized. Despite similar onset and type of clinical signs and mortality of birds in the IT and SC groups, especially on early days PC, birds in the SC group had lower body weight than IT birds. Lung injuries were more prominent in the IT group than the SC group. Birds of the SC group showed cellulitis and severely injured kidneys in contrast to the IT group. On day 3 PC, the number of positive heart blood samples for bacterial growth in the ITF group was half that of the IT group, whereas SC and SCF birds had similar positive numbers. Infection by the subcutaneous route results in more severe disease. The intratracheal route produces results more similar to colibacillosis under field conditions with regard to clinical signs, pathogenesis, and response to therapy, albeit it is more difficult to perform than a subcutaneous challenge.
Un estudio comparativo sobre dos modelos de infección de colibacilosis en pollos de engorde: Características clínicas, patogénesis y respuesta a la terapia. El conocimiento de las características de los diferentes modelos de infección de colibacilosis aviar es necesario para realizar estudios experimentales confiables. Este estudio compara las características de dos modelos de infección de colibacilosis (por vía intratraqueal y por vía subcutánea) en pollos de engorde. Pollos de engorde (n = 125), de 35 días de edad, se asignaron aleatoriamente a cuatro grupos experimentales (n = 20) y tres controles (n = 15): 1) control negativo (NC), y grupos 2 y 3) controles de infección intratraqueal o subcutánea (ITC o SCC), 4) infección intratraqueal (IT), 5) infección intratraqueal con administración de florfenicol (ITF), 6) infección subcutánea (SC), y 7) infección subcutánea con administración de florfenicol (SCF). Se registraron los signos clínicos y la mortalidad, y se realizó un examen macroscópico de las canales. En los días tres y cinco posteriores a la exposición (PC), se sacrificaron 5 aves por grupo. A pesar de que fueron similares el inicio y tipo de signos clínicos y la mortalidad en las aves de los grupos inoculados por infección intratraqueal y subcutánea, especialmente en los primeros días después del desafío, las aves de los grupos inoculado por vía subcutánea tenían un peso corporal más bajo que las aves inoculadas por la vía intratraqueal. Las lesiones pulmonares fueron más prominentes en el grupo inoculado por vía intratraqueal que en el grupo inoculado por vía subcutánea. Las aves del grupo inoculado por vía subcutánea mostraron celulitis y riñones severamente lesionados en contraste con el grupo inoculado por vía intratraqueal. En el día tres después del desafío, el número de muestras de sangre del corazón positivas para crecimiento bacteriano en el grupo con infección intratraqueal y con administración de florfenicol fue la mitad que en el grupo inoculado intratraquealmente, mientras que las aves inoculadas por vía subcutánea e inoculadas subcutáneamente con administración de florfenicol tuvieron números positivos similares. En conclusión, la infección por vía subcutánea resulta en una enfermedad más severa. La vía intratraqueal produce resultados más similares a la colibacilosis en condiciones de campo con respecto a los signos clínicos, la patogenia y la respuesta a la terapia, aunque es más difícil de realizar que un desafío subcutáneo.

The Clinical and Laboratory Standards Institute has published epidemiological cut-off values for susceptibility data generated at 22°°C and read after 44-48 h for florfenicol, oxolinic acid and oxytetracycline against Aeromonas salmonicida. The cut-off values for the minimum inhibitory concentration (MIC) and disc diffusion were derived from data obtained by 1 laboratory and 2 laboratories respectively. The present work reports the generation of susceptibility data from additional laboratories and the calculation of provisional cut-off values from aggregations of these data with previously published data. With respect to MIC data, the provisional cut-off values, derived from aggregations of the data from 4 laboratories, were ≤4 µg ml-1 for florfenicol, ≤0.0625 µg ml-1 for oxolinic acid and ≤1 µg ml-1 for oxytetracycline. For disc diffusion data, the provisional cut-off values derived from aggregations of the data from 5 laboratories were ≥30 mm for florfenicol, ≥32 mm for oxolinic acid and ≥25 mm for oxytetracycline. In addition, a cut-off value of ≥29 mm for ampicillin was derived from the aggregation of data from 4 laboratories.

Actinobacillus pleuropneumoniae is responsible for porcine pleuropneumonia, a highly contagious lung infection. The control of this respiratory disease remains heavily reliant on antibiotics, with phenicols being one of the primary classes of antibiotics used in pig farming. In the present study, we describe three isolates (B2278, B2176 and B2177) of A. pleuropneumoniae resistant to florfenicol attributed to the presence of the floR gene, which were obtained from two pig farms in Italy. Florfenicol susceptibility tests indicated that B2176 exhibited an intermediate susceptibility profile, while B2177 and B2278 were resistant. All three isolates belonged to serovar 6 and tested positive for the presence of the floR gene. Whole genome sequencing analysis revealed that isolates B2176, B2177 and B2278 harbored genes encoding the toxins ApxII and ApxIII, characteristic of strains with moderate virulence. Moreover, phylogenetic analysis demonstrated that these isolates were closely related, with single nucleotide polymorphisms (SNPs) ranging from 8 to 19. The floR gene was located on a novel 5588 bp plasmid, designated as pAp-floR. BLASTN analysis showed that the pAp-floR plasmid had high nucleotide identity (99 %) and coverage (60 %) with the pMVSCS1 plasmid (5621 bp) from Mannheimia varigena MVSCS1 of porcine origin. Additionally, at least under laboratory conditions, pAp-floR was stably maintained even in the absence of direct selective pressure, suggesting that it does not impose a fitness cost. Our study underscores the necessity of monitoring the spread of florfenicol-resistant A. pleuropneumoniae isolates in the coming years.

Florfenicol (F), an antimicrobial agent exclusive to veterinary use within the chloramphenicol class, is extensively applied as a broad-spectrum remedy for animal diseases. Despite its efficacy, concerns arise over potential deleterious residues in animal-derived edibles, posing threats to human health. This study pioneers an innovative approach, introducing a quantum dot fluorescence-based immunoassay (FLISA) for the meticulous detection of F residues in animal-derived foods and feeds. This method demonstrates heightened sensitivity, with a detection limit of 0.3 ng/mL and a quantitative detection range of 0.6-30.4 ng/mL. Method validation, applied to diverse food sources, yields recoveries from 90.4 % to 109.7 %, featuring RSDs within 1.3 % to 8.7 %, the results showed high consistency with the national standard HPLC-MS/MS detection method. These findings underscore the method\'s accuracy and precision, positioning it as a promising tool for swift and reliable F residue detection, with substantial implications for fortifying food safety monitoring.

Veterinary antibiotics have become an emerging pollutant in water and wastewater sources due to excess usage, toxicity and resistance to traditional water and wastewater treatment. The present study explored the degradation of a model antibiotic- Florfenicol (FF) using electrochemical oxidation (EO) with Ti-RuO2/IrO2 anode. The anode material was characterized using SEM-EDS studies expressing stable structure and optimal interaction of the neighboring metal oxides with each other. The EDS results showed the presence of Ru, Ir, Ti, O and C elements with 6.44%, 2.57%, 9.61%, 52.74% and 28.64% atomic weight percentages, respectively. Optimization studies revealed pH 5, 30 mA cm-2 current density and 0.05 M Na2SO4 for 5 mg L-1 FF achieved 90% TOC removal within 360 min treatment time. The degradation followed pseudo-first order kinetics. LC-Q-TOF-MS studies revealed six predominant byproducts illustrating hydroxylation, deflourination, and dechlorination to be the major degradation mechanisms during the electrochemical oxidation of FF. Ion chromatography studies revealed an increase in Cl-, F- and NO3- ions as treatment time progressed with Cl- decreasing after the initial phase of the treatment. Toxicity studies using Zebrafish (Danio rerio) embryo showed the treated sample to be toxic inducing developmental disorders such as pericardial edema, yolk sac edema, spinal curvature and tail malformation at 96 h post fertilization (hpf). Compared to control, delayed hatching and coagulation were observed in treated embryos. Overall, this study sets the stage for understanding the effect of mixed metal oxide (MMO) anodes on the degradation of veterinary antibiotic-polluted water and wastewater sources using electrochemical oxidation.

Protein succinylation modification is a common post-translational modification (PTM) that plays an important role in bacterial metabolic regulation. In this study, quantitative analysis was conducted on the succinylated proteome of wild-type and florfenicol-resistant Vibrio alginolyticus to investigate the mechanism of succinylation regulating antibiotic resistance. Bioinformatic analysis showed that the differentially succinylated proteins were mainly enriched in energy metabolism, and it was found that the succinylation level of phosphoenolpyruvate carboxyl kinase (PEPCK) was highly expressed in the florfenicol-resistant strain. Site-directed mutagenesis was used to mutate the lysine (K) at the succinylation site of PEPCK to glutamic acid (E) and arginine (R), respectively, to investigate the function of lysine succinylation of PEPCK in the florfenicol resistance of V. alginolyticus. The detection of site-directed mutagenesis strain viability under florfenicol revealed that the survival rate of the E mutant was significantly higher than that of the R mutant and wild type, indicating that succinylation modification of PEPCK protein may affect the resistance of V. alginolyticus to florfenicol. This study indicates the important role of PEPCK during V. alginolyticus antibiotic-resistance evolution and provides a theoretical basis for the prevention and control of vibriosis and the development of new antibiotics.

Florfenicol resistance genes (FRGs) are widely present in livestock farms. The aim of this study was to evaluate the removal efficiencies of FRGs as well as the relationships between FRGs, mobile genetic elements (MGEs) and bacterial communities during the natural drying (ND) and anaerobic digestion (AD) processes of manure treatment in swine farms by combining bacterial isolation, quantitative PCR and metagenomic approaches. Solid manure showed a higher abundance of FRGs than fresh manure and was the main contamination source of fexA and fexB in ND farms, whilst biogas slurry displayed a lower abundance of FRGs than the wastewater in AD farms. Moreover, fresh manure and wastewater showed a high abundance of optrA, and wastewater was the main contamination source of cfr in both ND and AD farms. Both optrA/fexA-positive enterococci and cfr/fexA-positive staphylococci were mainly isolated along the farms\' treatment processes. The cfr-positive staphylococci were highly prevalent in wastewater (57.14 % - 100 %) and may be associated with nasal-derived cfr-positive porcine staphylococci. An increased abundance of Enterococcus, Jeotgalibaca and Vagococcus in the bacterial community structures may account for the high optrA abundance in wastewater and Jeotgalibaca may be another potential host of optrA. Furthermore, the abundance of FRG-related MGEs increased by 22.63 % after the ND process and decreased by 66.96 % in AD farms. A significant correlation was observed between cfr and ISEnfa4, whereas no significance was found between optrA and IS1216E, although IS1216E is the predominant insertion sequence involved in the transfer of optrA. In conclusion, manure and wastewater represented independent pollution sources of FRGs in swine farms. Associated MGEs might play a key role in the transfer and persistence of FRGs. The AD process was more efficient in the removal of FRGs than the ND method, nevertheless a longer storage of slurry may be required for a complete removal.

Florfenicol, a widely used veterinary antibiotic, has now been frequently detected in various water environments and human urines, with high concentrations. Accordingly, the ecological risks and health hazards of florfenicol are attracting increasing attention. In recent years, antibiotic exposure has been implicated in the disruption of animal glucose metabolism. However, the specific effects of florfenicol on the glucose metabolism system and the underlying mechanisms are largely unknown. Herein, zebrafish as an animal model were exposed to environmentally relevant concentrations of florfenicol for 28 days. Using biochemical and molecular analyses, we found that exposure to florfenicol disturbed glucose homeostasis, as evidenced by the abnormal levels of blood glucose and hepatic/muscular glycogen, and the altered expression of genes involved in glycogenolysis, gluconeogenesis, glycogenesis, and glycolysis. Considering the efficient antibacterial activity of florfenicol and the crucial role of intestinal flora in host glucose metabolism, we then analyzed changes in the gut microbiome and its key metabolite short-chain fatty acids (SCFAs). Results indicated that exposure to florfenicol caused gut microbiota dysbiosis, inhibited the production of intestinal SCFAs, and ultimately affected the downstream signaling pathways of SCFA involved in glucose metabolism. Moreover, non-targeted metabolomics revealed that arachidonic acid and linoleic acid metabolic pathways may be associated with insulin sensitivity changes in florfenicol-exposed livers. Overall, this study highlighted a crucial aspect of the environmental risks of florfenicol to both non-target organisms and humans, and presented novel insights into the mechanistic elucidation of metabolic toxicity of antibiotics.