RESUMO
Infectious bovine keratoconjunctivitis (IBK) is an ocular disease that affects bovines and has significant economic and health effects worldwide. Gram negative bacteria Moraxella bovis and Moraxella bovoculi are its main etiological agents. Antimicrobial therapy against IBK is often difficult in beef and dairy herds and, although vaccines are commercially available, their efficacy is variable and dependent on local strains. The aim of this study was to analyze for the first time the genomes of Uruguayan clinical isolates of M. bovis and M. bovoculi. The genomes were de novo assembled and annotated; the genetic basis of fimbrial synthesis was analyzed and virulence factors were identified. A 94% coverage in the reference genomes of both species, and more than 80% similarity to the reference genomes were observed. The mechanism of fimbrial phase variation in M. bovis was detected, and the tfpQ orientation of these genes confirmed, in an inversion region of approximately 2.18kb. No phase variation was determined in the fimbrial gene of M. bovoculi. When virulence factors were compared between strains, it was observed that fimbrial genes have 36.2% sequence similarity. In contrast, the TonB-dependent lactoferrin/transferrin receptor exhibited the highest percentage of amino acid similarity (97.7%) between strains, followed by cytotoxins MbxA/MbvA and the ferric uptake regulator. The role of these virulence factors in the pathogenesis of IBK and their potential as vaccine components should be explored.
Assuntos
Doenças dos Bovinos , Genoma Bacteriano , Ceratoconjuntivite Infecciosa , Moraxella bovis , Moraxella , Animais , Moraxella/genética , Moraxella/isolamento & purificação , Bovinos , Moraxella bovis/genética , Ceratoconjuntivite Infecciosa/microbiologia , Doenças dos Bovinos/microbiologia , Infecções por Moraxellaceae/microbiologia , Infecções por Moraxellaceae/veterinária , Uruguai , Fatores de Virulência/genéticaRESUMO
This is the first study of the genetic diversity of Moraxella spp. Isolates were detected in an Eye Hospital in the City of Buenos Aires. Due to the high frequency of Moraxella spp. observed in corneal abscesses, we decided to validate their identification at the species level, determine their drug susceptibility and perform molecular subtyping. Seventeen (17) isolates obtained from corneal abscesses were evaluated. The identification was carried out using a combination of biochemical tests and MALDI-TOF mass spectrometry. Of these isolates, 88.2% were identified as Moraxella lacunata, and 11.8% as Moraxella nonliquefaciens. Molecular subtyping was performed using the pulsed-field gel electrophoresis (PFGE) technique. All isolates were typable and thirteen digestion patterns were identified. Based on the obtained results, the PFGE technique using the SmaI enzyme can be used for epidemiological studies of strains of these species.
Assuntos
Abscesso , Moraxella , Humanos , Moraxella/genética , Eletroforese em Gel de Campo Pulsado , Variação GenéticaRESUMO
Introduction. Infectious keratoconjunctivitis is a significant ocular disease found in confined sheep. Little information about the aetiological agents and their antimicrobial susceptibility is available.Gap Statement. There is limited information on the aetiological agents involved in keratoconjunctivitis outbreaks in sheep.Aim. The present research aimed to determine the bacterial aetiological factors involved in an outbreak of infectious keratoconjunctivitis in confined lambs.Methodology. Ocular swabs were collected from 23 randomly selected lambs, which were classified into three groups according to the severity of the lesion: group I (N=6; no ocular involvement), group II (N=8; less severe injuries) and group III (N=9; more severe injuries). Isolation of aerobic bacteria and antimicrobial susceptibility tests were carried out. Molecular detection of Mollicutes was performed, and positive samples were tested to confirm the presence of the following species: Mycoplasma conjunctivae, Mycoplasma agalactiae and Mycoplasma mycoides subsp. capri.Results. Moraxella sp. and Mollicutes were detected in all groups, but we inferred that Moraxella sp. are only significant in the early stages of the disease. M. conjunctivae was detected in all tested groups, while M. agalactiae was detected in samples of group III only. One strain of Moraxella sp. was resistant to erythromycin and showed intermedite resistance to tetracycline.Conclusion. The presence of these species confirms their importance in the aetiology of this disease, and the low resistance profile observed in the studied farm suggested an increased cure success rate.
Assuntos
Ceratoconjuntivite Infecciosa , Ceratoconjuntivite , Infecções por Mycoplasma , Mycoplasma conjunctivae , Doenças dos Ovinos , Animais , Antibacterianos/farmacologia , Surtos de Doenças/veterinária , Cabras , Ceratoconjuntivite/epidemiologia , Ceratoconjuntivite/microbiologia , Ceratoconjuntivite/veterinária , Ceratoconjuntivite Infecciosa/epidemiologia , Ceratoconjuntivite Infecciosa/microbiologia , Moraxella/genética , Mycoplasma , Infecções por Mycoplasma/microbiologia , Ovinos , Doenças dos Ovinos/microbiologiaRESUMO
Infectious bovine keratoconjunctivitis (IBK) is an ocular disease affecting bovine herds worldwide, and it causes significant economic loss. The etiologic agent of IBK is considered to be Moraxella bovis, but M. ovis and M. bovoculi are frequently recovered of animals presenting clinical signs of IBK. The therapeutic measures available for its control have limited efficacy. Antimicrobial photodynamic therapy (aPDT) using porphyrins as photosensitizing molecules is an alternative method that can be used to reduce microbial growth. We evaluated the antibacterial activity of aPDT using two water-soluble tetra-cationic porphyrins (H2TMeP and ZnTMeP) against 22 clinical isolates and standard strains of Moraxella spp. in vitro and in an ex vivo model. For the in vitro assay, 4.0 µM of porphyrin was incubated with approximately 1.0 × 104 CFU/mL of each Moraxella sp. isolate and exposed to artificial light for 0, 2.5, 5, and 7.5 min. Next, 50 µL of this solution was plated and incubated for 24 h until CFU measurement. For the ex vivo assay, corneas excised from the eyeballs of slaughtered cattle were irrigated with Moraxella spp. culture, followed by the addition of zinc(II) porphyrin ZnTMeP (4.0 µM). The corneal samples were irradiated for 0, 7.5, and 30 min, followed by swab collection, plating, and CFU count. The results demonstrated the in vitro inactivation of the strains and clinical isolates of Moraxella spp. after 2.5 min of irradiation using ZnTMeP, reaching complete inactivation until 7.5 min. In the ex vivo experiment, the use of ZnTMeP resulted in the most significant reduction in bacterial concentration after 30 min of irradiation. These results encourage future in vivo experiments to investigate the role of metalloporphyrin ZnTMeP in the inactivation of Moraxella spp. isolates causing IBK.
Assuntos
Anti-Infecciosos , Doenças dos Bovinos , Ceratoconjuntivite Infecciosa , Ceratoconjuntivite , Infecções por Moraxellaceae , Fotoquimioterapia , Porfirinas , Animais , Antibacterianos/farmacologia , Bovinos , Doenças dos Bovinos/microbiologia , Ceratoconjuntivite Infecciosa/tratamento farmacológico , Ceratoconjuntivite Infecciosa/microbiologia , Moraxella , Infecções por Moraxellaceae/tratamento farmacológico , Infecções por Moraxellaceae/microbiologia , Infecções por Moraxellaceae/veterinária , Porfirinas/farmacologia , OvinosRESUMO
Infectious keratoconjunctivitis (IKC) is the most frequent ocular disease in livestock worldwide and is primarily caused by Moraxella bovis, M. ovis, and/or M. bovoculi. The economic impact of IKC is mainly due to ocular damage, which leads to weight loss, management difficulties, pain and discomfort, and cost of treatments. In horses, limited information is available on the association of Moraxella spp. with keratoconjunctivitis. The present report describes two cases of equine keratoconjunctivitis caused by members of the genus Moraxella. Both animals presented with lacrimation, conjunctivitis, photophobia, mucoid or purulent secretions, blepharitis, and conjunctival hyperemia. The diagnosis of IKC was based on the epidemiological and clinical findings; the etiological agent was identified through bacteriological (culture and biochemistry assays) and molecular testing (PCR and nucleotide sequencing). Our study reports the isolation of Moraxella bovoculi (SBP 88/19) and a putative new species/mutant of Moraxella (SBP 39/19) recovered from ocular secretions in horses. Thus, we suggest the inclusion of Moraxella spp. infection in the differential diagnosis of conjunctivitis in horses in Southern Brazil.
Assuntos
Cavalos/microbiologia , Ceratoconjuntivite Infecciosa , Moraxella , Infecções por Moraxellaceae , Animais , Brasil , Ceratoconjuntivite Infecciosa/diagnóstico , Moraxella/genética , Moraxella/isolamento & purificação , Infecções por Moraxellaceae/diagnóstico , Infecções por Moraxellaceae/veterináriaRESUMO
AIMS: This study aimed to verify the formation of biofilms by Moraxella bovis, Moraxella ovis and Moraxella bovoculi isolates from ruminants. In addition, the lysozyme activity against the isolates of M. bovis, M. ovis and M. bovoculi in free form and in biofilms was determined. METHODS AND RESULTS: In this study, 54 isolates of Moraxella sp. obtained from bovine and ovine clinical samples were evaluated in vitro for capacity of biofilm formation and lysozyme susceptibility in planktonic and sessile cells. In addition, biofilms produced by four Moraxella sp. isolates were visualized under scanning electron microscope (SEM). It was possible to demonstrate, for the first time, the ability to form biofilms by M. ovis and M. bovoculi. The isolates of Moraxella sp. have the capacity to form biofilms in different intensities, varying among weak, moderate and strong. It was verified that the lysozyme shows activity on Moraxella sp. in planktonic form. However, on biofilms there was a reduction in the production, but without impairing its formation, and on consolidated biofilms the lysozyme did not have the capacity to eradicate the preformed biofilms. CONCLUSIONS: This work shows the capacity of biofilm formation by Moraxella sp. of veterinary importance. The lysozyme susceptibility of Moraxella sp. in planktonic form shows that this enzyme has bacteriostatic activity on this micro-organism and it reduced the production of biofilms. SIGNIFICANCE AND IMPACT OF THE STUDY: Based on the results, it is possible to infer that the biofilm formation capacity by Moraxella sp. and the resistance to lysozyme concentrations equal to or greater than the physiological levels of the ruminant tear may be linked not only to the capacity to colonize the conjunctiva, but also to remain in this place even after healing of the lesions, being a reservoir of Moraxella sp. in a herd.
Assuntos
Biofilmes , Moraxella bovis/fisiologia , Moraxella/fisiologia , Muramidase/metabolismo , Animais , Bovinos , Doenças dos Bovinos , Ceratoconjuntivite Infecciosa , Moraxella/isolamento & purificação , Infecções por Moraxellaceae , Ovinos/microbiologiaRESUMO
Herein we report the case of a 10-year-old boy with an autosomal mosaic mutation who developed bacteremia. The causative agent was identified as Moraxella osloensis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and 16S rRNA gene sequencing. In the pediatric population, there have been 13 case reports of infection attributed to M. osloensis and this is the fifth reported case of pediatric bacteremia due to M. osloensis. After Moraxella species infection was confirmed, the patient recovered with appropriate antimicrobial therapy. It is important to consider that M. osloensis can cause serious infections, such as bacteremia, in otherwise healthy children.
Assuntos
Bacteriemia/microbiologia , Moraxella/isolamento & purificação , Infecções por Moraxellaceae/microbiologia , Antibacterianos/uso terapêutico , Bacteriemia/tratamento farmacológico , Criança , Humanos , Masculino , Infecções por Moraxellaceae/tratamento farmacológico , Reação em Cadeia da Polimerase , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Resultado do TratamentoRESUMO
ABSTRACT Herein we report the case of a 10-year-old boy with an autosomal mosaic mutation who developed bacteremia. The causative agent was identified as Moraxella osloensis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and 16S rRNA gene sequencing. In the pediatric population, there have been 13 case reports of infection attributed to M. osloensis and this is the fifth reported case of pediatric bacteremia due to M. osloensis. After Moraxella species infection was confirmed, the patient recovered with appropriate antimicrobial therapy. It is important to consider that M. osloensis can cause serious infections, such as bacteremia, in otherwise healthy children.
Assuntos
Humanos , Masculino , Criança , Bacteriemia/microbiologia , Infecções por Moraxellaceae/microbiologia , Moraxella/isolamento & purificação , Reação em Cadeia da Polimerase , Resultado do Tratamento , Bacteriemia/tratamento farmacológico , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Infecções por Moraxellaceae/tratamento farmacológico , Antibacterianos/uso terapêuticoRESUMO
INTRODUCTION: Infectious bovine keratoconjunctivitis (IBK) is an important ocular disease which affects cattle worldwide. To advance towards IBK effective prevention and treatment strategies, it is important to define the distribution and genetic diversity of potential virulence factors present in M. bovis and M. bovoculi. The objective of this work was to identify and to analyze Moraxella spp. potential virulence factor genes in a collection of clinical isolates. METHODOLOGY: The presence and diversity of virulence factors in a collection of Moraxella spp. strains isolated since 1983 to 2009 in Uruguay was analyzed by PCR using primers for partial amplification of tolC, omp79, plb, fur and mbxA. The selection criterion of these genes was based on the fact that they encode virulence factors which could be present and conserved within strains, an important issue for the development of vaccines. RESULTS: Differences in PCR amplification were observed within tolC (84%), omp79 (80%), plb (76%) and fur (44%) in M. bovis strains, whereas mbxA was amplified in all M. bovis and M. bovoculi strains. Regarding genetic diversity, the tolC nucleotide sequences were the less diverse within all M. bovis and mbxA were the less diverse within all M. bovis and M. bovoculi strains. CONCLUSIONS: PCR amplifications suggest the occurrence of differences between both Moraxella species, related to evaluated genes within Moraxella spp. strains and suggests that both species may have different pathogenic attributes. MbxA and the outer membrane protein TolC might be considered for future studies to develop new vaccines against IBK.
Assuntos
Doenças dos Bovinos/microbiologia , Variação Genética , Ceratoconjuntivite Infecciosa/microbiologia , Moraxella/classificação , Moraxella/genética , Fatores de Virulência/genética , Animais , Bovinos , DNA Bacteriano/genética , Moraxella/isolamento & purificação , Reação em Cadeia da Polimerase , UruguaiRESUMO
The aim of this study was to determine the differences in the antimicrobial susceptibility profiles of Moraxella bovis, M. bovoculi and M. ovis. Thirty-two strains of Moraxella spp. isolated from cattle and sheep with infectious keratoconjunctivitis were tested via broth microdilution method to determine their susceptibility to ampicillin, cefoperazone, ceftiofur, cloxacillin, enrofloxacin, florfenicol, gentamicin, neomycin, oxytetracycline and penicillin. The results demonstrated that Moraxella spp. strains could be considered sensitive for most of the antimicrobials tested in this study, but differences between the antimicrobial susceptibility profiles of these three Moraxella species were found. M. bovis might differ from other species due to the higher MIC and MBC values it presented.