Multocida Isolates Research Articles

Comparative evaluation of PCR and loop-mediated isothermal amplification (LAMP) assays for detecting Pasteurella multocida in poultry

ABSTRACT Aims To develop a colourimetric loop-mediated isothermal amplification (LAMP) assay for the detection of Pasteurella multocida in clinical poultry samples and compare the performance of this assay with PCR. A secondary aim was to evaluate a simple DNA extraction method that could enable LAMP-based testing in the field without the need for specialised laboratory equipment. Methods Primer sets for both LAMP and PCR were designed to amplify the KMT1 gene of P. multocida. DNA was extracted from 12 P. multocida isolates using a commercial extraction kit, and subjected to analysis using both LAMP and PCR. The analytical specificity of the LAMP assay was evaluated by testing it against a panel of 12 unrelated bacterial species, and the analytical sensitivity (limit of detection) was determined through testing of serial dilutions of the target DNA and compared to that of PCR. Subsequently, cloacal swabs (n = 40) from a commercial turkey flock were subjected to analysis using both LAMP and PCR assays, using a rapid DNA extraction method and a commercial extraction kit. Clinical sensitivity and specificity of the LAMP assay were calculated in comparison to PCR. Results A single DNA fragment of the expected size (∼ 200 base pairs), was amplified by PCR from 12 P. multocida isolates, which were also all positive by the LAMP assay. The identity of all PCR amplicons was confirmed by sequencing. Both PCR and LAMP showed similar analytical sensitivity, with a LOD of 20 pg of target DNA. As neither PCR nor LAMP assays produced positive results with 12 non-related bacterial species, the analytical specificity was assessed as 100%. However, LAMP demonstrated lower clinical specificity (94.74%) compared to PCR (100%) when 40 clinical samples were tested. None of the DNA samples extracted using the simplified DNA extraction method were amplified by either LAMP or PCR. Conclusion The LAMP assay developed in this study exhibits comparable performance to PCR in detecting P. multocida. Clinical relevance The use of a rapid and portable DNA extraction method, in conjunction with LAMP assays, could create opportunities for point-of-care testing for fowl cholera in field settings.

Genetic basis of macrolide resistance in porcine Pasteurella multocida isolates from the German national resistance monitoring program GERM-Vet 2008-2021.

To analyse porcine Pasteurella multocida isolates obtained from the national resistance monitoring program GERM-Vet 2008-2021 in Germany for phenotypic and genotypic macrolide resistance. The antimicrobial susceptibility of the isolates was determined by broth microdilution according to CLSI standards. Closed genomes were obtained by WGS via Illumina MiSeq and MinION platforms followed by a hybrid assembly. Of 1114 porcine P. multocida isolates, only four isolates (0.36%), one each from 2008, 2010, 2019 and 2021, exhibited resistance to at least one macrolide tested. The isolate from 2010 was only resistant to erythromycin and WGS analysis neither revealed a macrolide resistance gene nor a macrolide resistance-mediating mutation. The isolates from 2008 and 2019 were resistant to erythromycin, tilmicosin, tildipirosin, tulathromycin and gamithromycin and showed either only the A2058G mutation in all six 23S rRNA operons or the chromosomally located macrolide resistance genes msr(E) and mph(E), respectively. The isolate from 2021 was resistant to erythromycin, tulathromycin, gamithromycin and tylosin and carried a novel integrative and conjugative element of 64 966 bp, designated Tn7730, in its chromosomal DNA. It harboured the macrolide resistance genes mef(C), mph(G) and estT, the lincosamide resistance gene lnu(H), and the tetracycline resistance gene tet(Y), the last two were detected for the first time in P. multocida. Macrolide resistance in German porcine P. multocida can be due to resistance-mediating mutations or resistance genes. The presence of the novel Tn7730 carrying three different macrolide resistance genes is alarming and should be monitored.

Clinical outcome of cats with suppurative otitis media and intact tympanum submitted to myringotomy: retrospective findings from 26 cases.

The efficacy of medical treatment associated with single myringotomy in cats with suppurative otitis media (OM) and intact tympanum was retrospectively evaluated. Patients' records were retrieved from cats with suppurative OM. Cats were included in this retrospective study based on fluid content within the tympanic cavity on imaging, intact tympanum on otoendoscopic examination and neutrophils on ear bulla exudate cytological examination. A total of 26 cats with middle ear suppurative otitis were included. Just over half of the cats (54%) presented bilateral involvement. Clinical signs included head tilt (13/26), otalgia (9/26), Horner's syndrome (7/26), external ear discharge (5/26), and nystagmus and facial paralysis (1/26). Cocci were identified on cytological examination in 18/40 samples and rods in 2/40. Bacterial culture results were positive in 15/40 samples, with Pseudomonas species (4/15), Pasteurella multocida (3/15), Staphylococcus felis (3/15), Staphylococcus schleiferi (2/15), Staphylococcus canis (2/15), Escherichia coli (2/15), Staphylococcus pseudintermedius (1/15) and Serratia marcesens (1/15) isolation. After myringotomy and gentle flushing of middle ear bullae (0.5-2 ml saline per ear), all cats were treated with oral corticosteroids and a 1-month course of systemic antibiotics according to sensitivity testing. In total, 19 (73%) cats were clinically healed 60-240 days after treatment. One cat was euthanased because of failure to eradicate a Pseudomonas species infection. Another cat died a few hours after the procedure. The five remaining cats were cured after ventral bulla osteotomy. OM without an aural polyp is a frequent diagnosis in cats, but data on medical treatment are scarce. This simple therapeutic technique, single myringotomy procedure and mild ear flushing, offers a practical, safe and efficient first-line treatment of suppurative OM with an intact tympanum in cats with or without neurological signs.

Antimicrobial in vitro effects of eight essential oils on Pasteurella multocida and Mannheimia haemolytica from nasopharyngeal swab samples of fattening calves

Essential oils are secondary metabolites of aromatic plants and are used in phytotherapy to treat various diseases. In the present study, eight selected essential oils - ajwain oil (Trachyspermum ammi L.), fennel oil (Foeniculum vulgare Mill. subsp. vulgare var. vulgare), thyme oil chemotype (ct.) thymol (Thymus vulgaris L.), tea tree oil (Melaleuca alternifolia Cheel.), oregano oil (Origanum vulgare L.), mountain savory oil (Satureja montana L.), lemongrass oil (Cymbopogon citratus (DC.) Stapf.) and eucalyptus oil (Eucalyptus globulus Labill.) -were examined for their antibacterial effect against Pasteurella (P.) multocida and Mannheimia (M.) haemolytica isolates from deep nasopharyngeal swab samples of fattening calves using agar diffusion and microdilution. All eight essential oils were effective against the tested isolates. Lemongrass oil proved to be the most potent of all eight essential oils, while fennel oil was only weakly effective. Different antimicrobial effects were observed between the two research methods. The effectiveness of ajwain, thyme, oregano and mountain savory oils was comparable in agar diffusion. However, this could not be reproduced using the microdilution method. P. multocida was found to be more sensitive to all essential oils tested than M. haemolytica. This study shows that the tested essential oils have antimicrobial in-vitro effects on P. multocida and M. haemolytica isolates and that the examination method is associated with the test result.

Complete genome sequence of a Pasteurella multocida isolate from a pig in Australia.

Here, we announce the complete genome sequence of a Pasteurella multocida isolate (PM1463) obtained from a diseased pig as a part of routine diagnostic investigations by the field veterinarian in Queensland, Australia. The assembly consists of a 2,321,605-bp chromosome and a 4,769-bp plasmid. The assembly has an average GC content of 40.33%.

Pathogenomic analysis and characterization of Pasteurella multocida strains recovered from human infections.

Pasteurella multocida is an upper respiratory tract commensal in several mammal and bird species but can also cause severe disease in humans and in production animals such as poultry, cattle, and pigs. In this study, we performed whole-genome sequencing of P. multocida isolates recovered from a range of human infections, from the mouths of cats, and from wounds on dogs. Together with publicly available P. multocida genome sequences, we performed phylogenetic and comparative genomic analyses. While isolates from cats and dogs were spread across the phylogenetic tree, human infections were caused almost exclusively by subsp. septica strains. Most of the human isolates were capsule type A and LPS type L1 and L3; however, some strains lacked a capsule biosynthesis locus, and some strains contained a novel LPS outer-core locus, distinct from the eight LPS loci that can currently be identified using an LPS multiplex PCR. In addition, the P. multocida strains isolated from human infections contained novel mobile genetic elements. We compiled a curated database of known P. multocida virulence factor and antibiotic resistance genes (PastyVRDB) allowing for detailed characterization of isolates. The majority of human P. multocida isolates encoded a reduced range of iron receptors and contained only one filamentous hemagglutinin gene. Finally, gene-trait analysis identified a putative L-fucose uptake and utilization pathway that was over-represented in subsp. septica strains and may represent a novel host predilection mechanism in this subspecies. Together, these analyses have identified pathogenic mechanisms likely important for P. multocida zoonotic infections.IMPORTANCEPasteurella multocida can cause serious infections in humans, including skin and wound infections, pneumonia, peritonitis, meningitis, and bacteraemia. Cats and dogs are known vectors of human pasteurellosis, transmitting P. multocida via bite wounds or contact with animal saliva. The mechanisms that underpin P. multocida human predilection and pathogenesis are poorly understood. With increasing identification of antibiotic-resistant P. multocida strains, understanding these mechanisms is vital for developing novel treatments and control strategies to combat P. multocida human infection. Here, we show that a narrow range of P. multocida strains cause disease in humans, while cats and dogs, common vectors for zoonotic infections, can harbor a wide range of P. multocida strains. We also present a curated P. multocida-specific database, allowing quick and detailed characterization of newly sequenced P. multocida isolates.

Properties of Pasteurella multocida isolated from animals during the seasonal migration of saigas.

The paper describes data from the study of cultural, morphological, and biochemical properties and the pathogenicity and virulence of epizootic isolates of Pasteurella multocida obtained from cattle and saigas. The study aimed to investigate the properties of P. multocida isolated from saigas and cattle during their seasonal migration, with a focus on its role in the epizootic process and potential transmission to farm animals. The research was conducted in a laboratory setting at the West Kazakhstan Agrarian-Technical University. White mice, saigas, and cattle were examined, and pathological and bacteriological analyses were performed on tissues and secretions. Pathogenicity, virulence, and toxigenicity of the isolated Pasteurella cultures were determined through biological tests on white mice. The morphological, cultural, and biochemical properties of the isolates were studied using standard microbiological methods. The study found that P. multocida isolates from both saigas and cattle exhibited high pathogenicity and virulence when tested on white mice. The isolates from sick and dead animals displayed 65.3 and 83.3% pathogenicity, respectively. The isolates were toxic to white mice, with filtrate dilutions showing 100% toxigenicity. Comparative analysis showed morphological and cultural similarities between Pasteurella isolates from saigas and cattle, confirming their identity. This research demonstrates that P. multocida, isolated from both saigas and cattle, contributes to the epizootic process and poses a threat to farm animals. Saigas, in particular, play a role in disease transmission during seasonal migrations. Understanding the ecological interactions between wild and farm animals is crucial for implementing preventive measures to control the spread of infectious diseases, emphasizing the need for comprehensive monitoring and intervention strategies.

Susceptibility of Pasteurella multocida isolated from cattle in Egypt to antibiotics, silver, chitosan and curcumin nanoparticles.

Pasteurella multocida is a Gram-negative bacterium causing economically significant diseases in cattle. This study aimed to determine P. multocida susceptibility to different antibiotics and antibiotic alternatives. In this study, 246 samples (180 nasal swabs and 66 lung tissue specimens) were collected from cattle showing respiratory manifestations in Egypt. Suspected P. multocida colonies following culture were subjected to polymerase chain reaction (PCR) for molecular confirmation of the isolates. A multiplex PCR was employed to identify P. multocida capsular groups. Susceptibility of the isolated P. multocida to different antibiotics and nanoparticles as antibiotic alternatives including silver (AgNPs), chitosan (CNPs) and curcumin (CurNPs) were tested using broth microdilution method. Thirty-two P. multocida isolates were obtained, kmt1 gene was detected in these isolates, and molecular capsular types classification revealed that all isolates were belonged to the capsular type A. Based on broth microdilution method findings, 20 (62.50%) isolates were considered as multi-drug resistant (MDR); the isolates were most sensitive to danofloxacin and kanamycin, whereas they were most resistant to doxycycline and tilmicosin. Antibiotic alternatives showed high anti-microbial activity against tested isolates with minimum inhibitory concentrations ranging from 1.56 - 6.25 μg mL-1, 156 - 625 μg mL-1, and 128 - 512 μg mL-1 for AgNPs, CNPs and CurNPs, respectively. Our finding demonstrated that MDR P. multocida was evident in cattle in Egypt. Although antibiotic alternatives showed promising in vitro anti-microbial effects against MDR isolates, additional studies are required to be actually applicable in veterinary practices.

Evaluasi Efektivitas Antibiotik Komersil Terhadap Agen Penyebab Gejala Snot pada Ayam Broiler di Kabupaten Kupang

Snot is a symptom of upper respiratory system infections in poultry, characterized by exudate production from the nasal cavity, swelling of infraorbital sinuses, snoring, sneezing, and dyspnoea. The aetiology of snot that have been isolated are Avibaterium paragallinarum, Pasteurella multocida, Ornithobacterium rhinotracheale, and Mycoplasma sp. Snot symptom can be eradicated with antibiotic treatment; however, antibiotic resistance makes antibiotic treatment ineffective. This study is aimed to evaluate the effectiveness of some commercial antibiotics against bacteria isolated from broiler chicken with snot symptom in Kupang Regency. Amoxycillin, ampicillin, tetracycline, cefoxitin, and ciprofloxacin were tested using the Kirby Bauer method with McFarland Turbidity Standard against Avibacterium paragallinarum, Ornithobacterium rhinotracheale, Pasteurella multocida, and Mycoplasma sp isolates. Inhibition zones were measured and compared to the standard of the Clinical Laboratory Standard Institute (CLSI) to determine the sensitivity or resistance percentage. The result showed that Avibacterium paragallinarum, Ornithobacterium rhinotracheale, Pasteurella multocida, and Mycoplasma sp were highly resistant to amoxycillin and ampicillin, yet most sensitif to ciprofloxacin. This suggests commercial antibiotics that are productive to eradicate snot symptom and implies some antibiotics that are ineffective to overcome snot symptom and hence should not be used in the field.

Genomic analysis reveals the population structure and antimicrobial resistance of avian Pasteurella multocida in China.

To investigate the population structure and antimicrobial resistance (AMR) of avian Pasteurella multocida in China. Utilizing WGS analysis, we explored the phylogeny using a dataset of 546 genomes, comprising avian P. multocida isolates from China (n = 121), the USA (n = 165), Australia(n = 153), Bangladesh (n = 3) and isolates of other hosts from China (n = 104). We examined the integrative and conjugative element (ICE) structures and the distribution of their components carrying resistance genes, and reconstructed the evolutionary history of A:L1:ST129 (n = 110). The population structure of avian P. multocida in China was dominated by the A:L1:ST129 clone with limited genetic diversity. A:L1:ST129 isolates possessed a broader spectrum of resistance genes at comparatively higher frequencies than those from other hosts and countries. The novel putative ICEs harboured complex resistant clusters that were prevalent in A:L1:ST129. Bayesian analysis predicted that the A:L1:ST129 clone emerged around 1923, and evolved slowly. A:L1:ST129 appears to possess a host predilection towards avian species in China, posing a potential health threat to other animals. The complex AMR determinants coupled with high frequencies may strengthen the population dominance of A:L1:ST129. The extensive antimicrobial utilization in poultry farming and the mixed rearing practices could have accelerated AMR accumulation in A:L1:ST129. ICEs, together with their resistant clusters, significantly contribute to resistance gene transfer and facilitate the adaptation of A:L1:ST129 to ecological niches. Despite the genetic stability and slow evolution rate, A:L1:ST129 deserves continued monitoring due to its propensity to retain resistance genes, warranting global attention to preclude substantial economic losses.

Brazilian Clinical Strains of Actinobacillus pleuropneumoniae and Pasteurella multocida: Capsular Diversity, Antimicrobial Susceptibility (In Vitro) and Proof of Concept for Prevention of Natural Colonization by Multi-Doses Protocol of Tildipirosin.

One hundred Actinobacillus pleuropneumoniae (App) and sixty Pasteurella multocida subsp. multocida serogroup A (PmA) isolates were recovered from porcine pneumonic lungs collected from eight central or southern states of Brazil between 2014 and 2018 (App) or between 2017 and 2021 (PmA). A. pleuropneumoniae clinical isolates were typed by multiplex PCR and the most prevalent serovars were 8, 7 and 5 (43, 25% and 18%, respectively). In addition, three virulence genes were assessed in P. multocida isolates, all being positive to capA (PmA) and kmt1 genes, all negative to capD and toxA, and most of them (85%) negative to pfhA gene. The susceptibility of both pathogens to tildipirosin was investigated using a broth microdilution assay. The percentage of isolates susceptible to tildipirosin was 95% for App and 73.3% for PmA. The MIC50 values were 0.25 and 1 μg/mL and the MIC90 values were 4 and >64 μg/mL for App and PmA, respectively. Finally, a multiple-dose protocol of tildipirosin was tested in suckling piglets on a farm endemic for both pathogens. Tildipirosin was able to prevent the natural colonization of the tonsils by App and PmA and significantly (p < 0.0001) reduced the burden of Glaesserella parasuis in this tissue. In summary, our results demonstrate that: (i) tildipirosin can be included in the list of antibiotics to control outbreaks of lung disease caused by App regardless of the capsular type, and (ii) in the case of clinical strains of App and PmA that are sensitive to tildipirosin based on susceptibility testing, the use of this antibiotic in eradication programs for A. pleuropneumoniae and P. multocida can be strongly recommended.

Complete genomes of Pasteurella multocida isolates from porcine, bovine, and cervine samples found in New Zealand farms.

Here, we present complete genome assemblies of Pasteurella multocida strains isolated from porcine, bovine, and cervine farms as part of bacteriology incursion investigations to identify pathogens that might present a sanitary risk to New Zealand.

Susceptibility trends of swine respiratory pathogens from 2019 to 2022 to antimicrobials commonly used in Spain

BackgroundAntimicrobial resistance is one of the most important health challenges in humans and animals. Antibiotic susceptibility determination is used to select the most suitable drug to treat animals according to its success probability following the European legislation in force for these drugs. We have studied the antibiotic susceptibility pattern (ASP) of Actinobacillus pleuropneumoniae (APP) and Pasteurella multocida (PM) isolates, collected during the period 2019–2022 in Spain. ASP was measured by determining minimum inhibitory concentration using standardized laboratory methods and its temporal trend was determined by logistic regression analysis of non-susceptible/susceptible isolates using clinical breakpoints.ResultsIt was not observed any significant temporal trends for susceptibility of Actinobacillus pleuropneumoniae to ceftiofur, florfenicol, sulfamethoxazole/trimethoprim, tulathromycin and tildipirosin during the study period (p > 0.05). Contrarily, a significant temporal trend (p < 0.05) was observed for quinolones (enrofloxacin and marbofloxacin), tetracyclines (doxycycline and oxyteracycline), amoxicillin, tiamulin and tilmicosin. On the other hand, it was not observed any significant temporal trends for susceptibility of Pasteurella multocida to quinolones (enrofloxacin and marbofloxacin), amoxicillin, ceftiofur, florfenicol and macrolides (tildipirosin, tulathromycin and tilmicosin) during the study period (p > 0.05). Contrarily, a significant temporal trend (p < 0.05) was observed for tetracyclines (oxyteracycline), tiamulin and sulfamethoxazole/trimethoprim.ConclusionsIn general terms, pig pathogens (APP and PM) involved in respiratory diseases analysed herein appeared to remain susceptible or tended to increase susceptibility to antimicrobials over the study period (2019–2022), but our data clearly showed a different pattern in the evolution of antimicrobial susceptibility for each combination of drug and microorganism. Our results highlight that the evolution of antimicrobial susceptibility must be studied in a case-by-case situation where generalization for drug families and bacteria is not possible even for bacteria located in the same ecological niche.

Pathognomonic features of Pasteurella multocida isolates among various avian species in Sharkia Governorate, Egypt

The present study aimed to isolate Pasteurella multocida (P. multocida) from pulmonary cases in several avian species and then investigate the histopathological features, antimicrobial resistance determinants, virulence characteristics, and risk factors analysis of the isolates in each species in correlation with epidemiological mapping of pasteurellosis in Sharkia Governorate, Egypt. The obtained data revealed a total occurrence of 9.4% (30/317) of P. multocida among the examined birds (chickens, ducks, quails, and turkeys). The incidence rate was influenced by avian species, climate, breed, age, clinical signs, and sample type. Antimicrobial susceptibility testing revealed that all isolates were sensitive to florfenicol and enrofloxacin, while 86.6 and 73.3% of the isolates displayed resistance to amoxicillin-clavulanic acid and erythromycin, respectively. All of the P. multocida isolates showed a multiple-drug resistant pattern with an average index of 0.43. Molecular characterization revealed that the oma87, sodA, and ptfA virulence genes were detected in the all examined P. multocida isolates. The ermX (erythromycin), blaROB-1 (β-lactam), and mcr-1(colistin) resistance genes were present in 60, 46.6, and 40% of the isolates, respectively. Ducks and quails were the most virulent and harbored species of antimicrobial-resistant genes. These results were in parallel with postmortem and histopathological examinations which detected more severe interstitial pneumonia lesions in the trachea and lung, congestion, and cellular infiltration especially in ducks. Epidemiological mapping revealed that the Fakous district was the most susceptible to pasteurellosis infection. Thus, farmers are recommended to monitor their flocks for signs of respiratory disease, seek veterinary care promptly if any birds are sick, and avoid the random usage of antibiotics. In conclusion, this study presents a comprehensive picture of the risk factors in correlation to the pathognomonic characteristics of P. multocida infection in poultry sectors to help in developing more effective strategies for prevention and control.

MOLECULAR CHARACTERIZATION, ANTIBIOTIC SUSCEPTIBILITY AND BIOFILM FORMING ABILITY OF PASTEURELLA MULTOCIDA ISOLATED FROM THE RESPIRATORY MICROBIOTA OF HEALTHY BOVINES

Livestock plays an important role in the economy of the country because it provides food producing animals. Pasteurella multocida which is an opportunistic pathogen of upper respiratory tract of animals and it has been known to cause many harmful diseases may survive in animal body by forming biofilm in different organs. So, the presence of this bacterium in animals may lead to transfer and threaten the human health by food chain. In the last few years, one of the major problems that alarms and evokes a major horrific threat to human being is the bacterial resistance to antimicrobial agents. Due to deficiency of antimicrobial agents in pipeline and emergence of new mechanisms of intra antimicrobial resistance, many surveillance programs have been set up for its containment. Many factors are responsible for this grave situation like high use of antibiotics in animals. So, the current study concluded that all of the 76 isolates of Pasteurella multocida out of 100 samples were confirmed through PCR and found sensitive to florfenicol (100%), gentamycin (92.11%), chloramphenicol (84.21%) and kanamycin (72.37%) respectively. For tetracycline and neomycin sensitivity were (69.73%) and (64.47%) respectively against the bacteria. Whereas tilmicosin and spectinomycin sensitivity were recorded as (30.27%) and (23.69%). Streptomycin (19.73%), sulfadiazine (7.89%), erythromycin (3.94%) and clindamycin (0.0 %) were least sensitive. It was also concluded that when all pure isolates of Pasteurella multocida were tested for Biofilm assay out of 76 isolates categorized as: non biofilm producer (7.89%), weak biofilm producer (34.21%), moderate biofilm producer (46.05%) and strong biofilm producer (11.84%). Hence, it was concluded that Pasteurella multocida isolated from cattle and buffaloes were found to form biofilm in in-vitro condition.

Genotypic Comparison of Pasteurella multocida from Healthy Animals at Entry to the Feedlots with That and from Bovine Respiratory Disease-Affected Animals during the Fattening Period.

The aim of this study was to investigate the possible genotypic differences between commensal Pasteurella multocida isolates from apparently healthy animals (AHA) at the time of entry to feedlots and those from BRD-affected animals (BRD-AA). A total of 20 batches of beef calves in seven feedlots were followed-up during the fattening period. P. multocida was isolated from 28.1% of AHA and 22.9% of BRD-AA. All isolates belonged to the A: L3 genotype. Most isolates from clinical cases (81.0%) grouped into a PFGE cluster were significantly associated with BRD cases (OR, 24.9; 95% CI, 6.4-96.2). The whole genomes of 14 isolates representative of the pulsotypes most frequently detected in BRD-AA and AHA were sequenced and compared with 53 bovine genomes belonging to the identified ST13, ST79, and ST80 genotypes for a global comparison. No differences were found in the virulence-associated gene content between sequence types (STs) globally or between BRD-AA and AHA isolates in this study. Significantly, ST79 isolates harbored ARGs, conferring resistance to different antimicrobials, including macrolides and tetracyclines, which are commonly used for the treatment of BRD. Two Spanish ST79 isolates carried an ICE highly similar to ICE Tn7407, which was recently detected in Germany, suggesting that ST79 P. multocida isolates in Europe and North America may be associated with different ICEs.

Agreement of antimicrobial susceptibility testing of Pasteurella multocida and Mannheimia haemolytica isolates from preweaned dairy calves with bovine respiratory disease.

Evaluate agreement among the antimicrobial susceptibility profiles of Mannheimia haemolytica or Pasteurella multocida obtained by transtracheal wash, nasal swab, nasopharyngeal swab, and bronchoalveolar lavage. 100 Holstein and Holstein-cross bull calves with bovine respiratory disease. Calves > 30 days old with naturally occurring bovine respiratory disease were sampled sequentially by nasal swab, nasopharyngeal swab, transtracheal wash, and then bronchoalveolar lavage. Samples were cultured, and for each antimicrobial, the MIC of 50% and 90% of isolates was calculated, and isolates were categorized as susceptible or not. Categorical discrepancies were recorded. Percent positive agreement and kappa values were calculated between isolates for each of the sampling methods. Antimicrobial susceptibility varied by pathogen and resistance to enrofloxacin, florfenicol, tilmicosin, and spectinomycin was detected. Minor discrepancies were seen in up to 29% of classifications, with enrofloxacin, penicillin, and florfenicol more frequently represented than other drugs. Very major and major discrepancies were seen when comparing florfenicol (1.9%) and tulathromycin (3.8 to 4.9%) across sampling methods. Some variability was seen in agreement for enrofloxacin for several comparisons (8.3 to 18.4%). Susceptibility testing of isolates from 1 location of the respiratory tract can reliably represent susceptibility in other locations. Nevertheless, the potential for imperfect agreement between sampling methods does exist. The level of restraint available, the skill level of the person performing the sampling, the age and size of the animal, disease status, and treatment history all must be factored into which test is most appropriate for a given situation.

Susceptibility of Actinobacillus pleuropneumoniae, Pasteurella multocida and Streptococcus suis Isolated from Pigs in Hungary between 2018 and 2021.

Porcine respiratory disease complex (PRDC) has been a major animal health, welfare, and economic problem in Hungary; therefore, great emphasis should be put on both the prevention and control of this complex disease. As antibacterial agents are effective tools for control, antibiotic susceptibility testing is indispensable for the proper implementation of antibacterial therapy and to prevent the spread of resistance. The best method for this is to determine the minimum inhibitory concentration (MIC) by the broth microdilution method. In our study, we measured the MIC values of 164 Actinobacillus pleuropneumoniae, 65 Pasteurella multocida, and 118 Streptococcus suis isolates isolated from clinical cases against the following antibacterial agents: amoxicillin, ceftiofur, cefquinome, oxytetracycline, doxycycline, tylosin, tilmicosin, tylvalosin, tulathromycin, lincomycin, tiamulin, florfenicol, colistin, enrofloxacin, and sulfamethoxazole-trimethoprim. Outstanding efficacy against A. pleuropneumoniae isolates was observed with ceftiofur (100%) and tulathromycin (100%), while high levels of resistance were observed against cefquinome (92.7%) and sulfamethoxazole-trimethoprim (90.8%). Ceftiofur (98.4%), enrofloxacin (100%), florfenicol (100%), and tulathromycin (100%) were found to be highly effective against P. multocida isolates, while 100% resistance was detected against the sulfamethoxazole-trimethoprim combination. For the S. suis isolates, only ceftiofur (100%) was not found to be resistant, while the highest rate of resistance was observed against the sulfamethoxazole-trimethoprim combination (94.3%). An increasing number of studies report multi-resistant strains of all three pathogens, making their monitoring a high priority for animal and public health.

Genomic profiling of Pasteurella multocida isolated from feedlot cases of bovine respiratory disease

Pasteurella multocida causes a range of diseases in many host species throughout the world, including bovine respiratory disease (BRD) which is predominantly seen in feedlot cattle. This study assessed genetic diversity among 139 P. multocida isolates obtained from post-mortem lung swabs of BRD-affected feedlot cattle in four Australian states: New South Wales, Queensland, South Australia, and Victoria during 2014–2019. Whole-genome sequencing (WGS) was used to determine capsular serogroup, lipopolysaccharide genotypes, multi-locus sequence types and phylogenetic relationships. Two capsular types (A and D), with most isolates (132/139; 95%) belonging to type A; and three lipopolysaccharide (LPS) genotypes were identified (L1 [6/139; 4.3%], L3 [124/139; 89.2%] and L6 [9/139; 6.4%)]). Multi-locus sequence types (STs) ST9, ST13, ST17, ST20, ST36, ST50, ST58, ST79, ST124, ST125, ST132, ST167, ST185, ST327, ST394, and three novel STs [ST396, ST397, and ST398] were identified, with ST394 (59/139; 42.4%) and ST79 (44/139; 32%) the most prevalent in all four states. Isolates displaying phenotypic resistance to single, dual or multiple antibiotics (macrolide, tetracycline and aminopenicillins) were predominantly ST394 (23/139; 17%). Laterally mobile elements identified in the resistant ST394 isolates included small plasmids, encoding macrolide and/or tetracycline resistance, distributed in all states; and chromosomally located integrative conjugative elements (ICEs) (4 ST394 and 1 ST125) from the same Queensland feedlot. This study highlights the genomic diversity, epidemiological relationships and AMR associations in bovine P. multocida isolates from Australia and provides insight into the unique ST prevalence compared to other major beef-producing countries.

In vitro and ex vivo antibacterial activity of levofloxacin against Pasteurella multocida and Escherichia coli isolated from rabbits (Oryctolagus cuniculus) - A preliminary study.

Levofloxacin veterinary formulations are available in Argentina, China and India for the use in dogs, cattle, pig and sheep, but not currently in the rabbit. Only the extra-label use in rabbits is possible. Levofloxacin is not labelled for veterinary use in the EU or the USA. The activity of levofloxacin against rabbit pathogens Pasteurella multocida (P. multocida) and Escherichia coli (E. coli) was evaluated. Minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were determined in broth and serum for 10 P. multocida isolates and 5 E. coli isolates from rabbits. One isolate of each bacterial species was used for the time-killing curve study in vitro and ex vivo. In vitro AUC24 /MIC ratios were used for building the inhibitory pharmacodynamic Imax model. The P. multocida MIC were 0.008-0.5 μg/mL, MBC - 0.015-0.5 μg/mL. Escherichia coli MIC was 0.008-0.03 μg/mL and MBC - 0.03-0.25 μg/mL. Bacterial counts were reduced to the limit of detection after 24 h with levofloxacin concentrations of 2 MIC and higher. All serum samples from rabbits treated with levofloxacin eliminated the bacteria within 24 h. AUC24 /MIC ratios for bacteriostatic, bactericidal and bacterial elimination effects for P. multocida and E. coli isolates were 21, 29 and 75 h and 27, 32 and 60 h, respectively. Proposed daily doses against P. multocida (MIC = 0.015 μg/mL) and E. coli (MIC = 0.03 μg/mL) isolates were calculated as ≤0.91 and ≤1.43 mg/kg, respectively. Fluoroquinolones are categorized by WHO as 'highest priority critically important antimicrobials'. Considering the increasing importance of antimicrobial stewardship, antimicrobials from a lower importance category that are active against the isolate of interest should be used in preference to fluoroquinolones. Fluoroquinolone use in veterinary medicine should be based on antimicrobial susceptibility testing in order to mitigate the risk to public health and prevent the spread of bacterial resistance.