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1.
J Appl Microbiol ; 135(4)2024 Apr 01.
Article in English | MEDLINE | ID: mdl-38503568

ABSTRACT

AIMS: The purpose was to evaluate the antimicrobial activity of highly soluble polypyrrole (Hs-PPy), alone or combined with oxacillin, as well as its antibiofilm potential against methicillin-resistant Staphylococcus aureus strains. Furthermore, the in silico inhibitory mechanism in efflux pumps was also investigated. METHODS AND RESULTS: Ten clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and two reference strains were used. Antimicrobial activity was determined by broth microdilution, and the combination effect with oxacillin was evaluated by the checkerboard assay. The biofilm formation capacity of MRSA and the interference of Hs-PPy were evaluated. The inhibitory action of Hs-PPy on the efflux pump was evaluated in silico through molecular docking. Hs-PPy showed activity against the isolates, with inhibitory action between 62.5 and 125 µg ml-1 and bactericidal action at 62.5 µg ml-1, as well as synergism in association with oxacillin. The isolates ranged from moderate to strong biofilm producers, and Hs-PPy interfered with the formation of this structure, but not with mature biofilm. There was no in silico interaction with the efflux protein EmrD, the closest homolog to NorA. CONCLUSIONS: Hs-PPy interferes with biofilm formation by MRSA, has synergistic potential, and is an efflux pump inhibitor.


Subject(s)
Anti-Infective Agents , Methicillin-Resistant Staphylococcus aureus , Anti-Bacterial Agents/pharmacology , Polymers/pharmacology , Pyrroles/pharmacology , Molecular Docking Simulation , Oxacillin/pharmacology , Anti-Infective Agents/pharmacology , Biofilms , Microbial Sensitivity Tests
2.
Braz J Microbiol ; 55(1): 1011-1015, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38012513

ABSTRACT

Mycoplasma hyopneumoniae (M. hyopneumoniae) is one of the smallest free-living bacteria found in nature; it has an extremely small genome and lacks a cell wall. It is the main etiological agent of porcine enzootic pneumonia (EP), a chronic respiratory disease with worldwide distribution that causes significant losses in swine production. Due to the great economic impact caused by EP, new strategies for treating and controlling this agent are researched. The objective of this study was to verify the anti-M. hyopneumoniae activity of compounds derived from Garcinia brasiliensis and the synergism with the main antimicrobials used in the treatment of EP; this is the first study assessing the synergism between bioactive molecules and antimicrobial compounds in vitro against isolates of M. hyopneumoniae. The minimum inhibitory concentrations (MICs) of the antimicrobials tiamulin, valnemulin, and enrofloxacin, as well as the bioactive compounds guttiferone-A (Gut-A), 7-epiculsone (7-Epic), copper 7-epiculsone (7-Epic-Cu), and benzophenone, were determined. Subsequently, the interactions of antibiotics with the compounds were evaluated using the checkerboard method. Three field M. hyopneumoniae isolates were used, and the J strain was used as a control. The MIC values of the antimicrobials compared to the field isolates were equal to and lower than those of the reference strain J. Among the compounds used, 7-Epic-Cu showed the lowest MIC value. Synergistic association was observed for Gut-A with tiamulin and valnemulin, whereas 7-Epic and 7-Epic-Cu showed synergistic action with enrofloxacin. No synergistic effect was observed for benzophenone. Despite being an initial study, the results suggest that these combinations hold promise for the treatment of infections caused by M. hyopneumoniae.


Subject(s)
Anti-Infective Agents , Mycoplasma hyopneumoniae , Swine , Animals , Enrofloxacin/pharmacology , Copper/pharmacology , Anti-Infective Agents/pharmacology , Benzophenones/pharmacology , Diterpenes
4.
J Proteomics ; 292: 105058, 2024 02 10.
Article in English | MEDLINE | ID: mdl-38065354

ABSTRACT

Bacteria typically produce membrane vesicles (MVs) at varying levels depending on the surrounding environments. Gram-negative bacterial outer membrane vesicles (OMVs) have been extensively studied for over 30 years, but MVs from Gram-positive bacteria only recently have been a focus of research. In the present study, we isolated MVs from Mycobacterium avium subsp. paratuberculosis (MAP) and analyzed their protein composition using LC-MS/MS. A total of 316 overlapping proteins from two independent preparations were identified in our study, and topology prediction showed these cargo proteins have different subcellular localization patterns. When MVs were administered to bovine-derived macrophages, significant up-regulation of pro-inflammatory cytokines was observed via qRT-PCR. Proteome functional annotation revealed that many of these proteins are involved in the cellular protein metabolic process, tRNA aminoacylation, and ATP synthesis. Secretory proteins with high antigenicity and adhesion capability were mapped for B-cell and T-cell epitopes. Antigenic, Immunogenic and IFN-γ inducing B-cell, MHC-I, and MHC-II epitopes were stitched together through linkers to form multi-epitope vaccine (MEV) construct against MAP. Strong binding energy was observed during the docking of the 3D structure of the MEV with the bovine TLR2, suggesting that the putative MEV may be a promising vaccine candidate against MAP. However, in vitro and in vivo analysis is required to prove the immunogenic concept of the MEV which we will follow in our future studies. SIGNIFICANCE: Johne's disease is a chronic infection caused by Mycobacterium avium subsp. paratuberculosis that has a potential link to Crohn's disease in humans. The disease is characterized by persistent diarrhea and enteritis, resulting in significant economic losses due to reduced milk yield and premature culling of infected animals. The dairy industry in the United States alone experiences losses of approximately USD 250 million due to Johne's disease. The current vaccine against Johne's disease is limited by several factors, including variable efficacy, limited duration of protection, interference with diagnostic tests, inability to prevent infection, and logistical and cost-related challenges. Nevertheless, a multiepitope vaccine design approach targeting M. avium subsp. paratuberculosis has the potential to overcome these challenges and offer improved protection against Johne's disease.


Subject(s)
Mycobacterium avium subsp. paratuberculosis , Paratuberculosis , Vaccines , Humans , Animals , Cattle , Paratuberculosis/diagnosis , Paratuberculosis/microbiology , Mycobacterium avium subsp. paratuberculosis/genetics , Membrane Proteins , Epitopes , Chromatography, Liquid , Proteomics , Tandem Mass Spectrometry
5.
Front Microbiol ; 14: 1280588, 2023.
Article in English | MEDLINE | ID: mdl-38075868

ABSTRACT

Mycoplasma hyopneumoniae (M. hyopneumoniae) is considered the primary causative agent of porcine enzootic pneumonia (EP), a chronic contagious respiratory disease that causes economic losses. Obtaining new pathogenic isolates and studying the genome and virulence factors are necessary. This study performed a complete sequencing analysis of two Brazilian strains, UFV01 and UFV02, aiming to characterize the isolates in terms of the virulence factors and sequence type. The complete genome analysis revealed the main virulence genes (mhp385, mhp271, MHP_RS03455, p102, p97, p216, MHP_RS00555, mhp107) and ST-123, the presence of three toxin-related genes (tlyC, PLDc_2 and hcnC), and some genetic groups specific to these two isolates. Subsequently, the pathogenicity of the isolates was evaluated via an experimental infection conducted in a swine model. The study was divided into three groups, namely a negative control group (n = 4) and two test groups (n = 8), totaling 20 animals. They were challenged at 35 days of age with 107 CCU (Color Changing Units) M. hyopneumoniae via the intratracheal route. The UFV01 group showed earlier and higher seroconversion (IgG) (100%), while only 50% of the UFV02 group seroconverted. The same trend was observed when analyzing the presence of IgA in the bronchoalveolar lavage fluid (BALF) at 35 days post-infection (dpi). The UFV01 group had a mean macroscopic lesion score of 11.75% at 35 dpi, while UFV02 had 3.125%. Microscopic lesions were more severe in the UFV01 group. Based on laryngeal swab samples evaluated by qPCR, and the detection began at 14 days. The UFV01 group showed 75% positivity at 14 dpi. The UFV02 group also started excreting at 14 dpi, with a positivity rate of 37.5%. The results indicate that the UFV01 isolate exhibits higher virulence than UFV02. These findings may aid in developing new vaccines and diagnostic kits and establishing experimental models for testing.

6.
Curr Microbiol ; 81(1): 40, 2023 Dec 16.
Article in English | MEDLINE | ID: mdl-38103072

ABSTRACT

Nosocomial infections caused by multidrug-resistant enterobacteria have become a major challenge in global public health. Previous studies have indicated that use of antibiotics in livestock production chains is linked to the rising threat of antibiotic resistance in humans. In this study, we aimed to evaluate the distribution of genes encoding resistance to tetracycline, ß-lactams, and colistin in multidrug-resistant enterobacteria isolated from feces of weaned pigs. Ninety-four enterobacteria isolates were submitted to antibiotic susceptibility test by minimum inhibitory concentration (MIC). In addition, we performed conjugation experiments to verify if plasmid-bearing isolates containing the mcr-1 gene could transfer their resistance determinant to a colistin-sensitive recipient strain. Our results demonstrated a positive association between the detection of antibiotic resistance genes in enterobacteria and the phenotypic resistance profiles of the bacterial isolates. At least one of the extended-spectrum ß-lactamases (ESBL) genes (blaCTX-M, blaTEM, or bla SHV) and tetA was found among most bacterial genera analyzed. In addition, results revealed that the mcr-1 gene can be transferred from E. coli UFV-627 isolate to an F- recipient (Escherichia coli K12) by conjugation. Our findings support the hypothesis that swine represents an important reservoir of antibiotic resistance genes and suggest that horizontal transfer mechanisms (e.g., conjugation) may mediate the spread of these genes in the swine gastrointestinal tract.


Subject(s)
Escherichia coli Infections , Escherichia coli Proteins , Humans , Animals , Swine , Escherichia coli/genetics , Anti-Bacterial Agents/pharmacology , Colistin/pharmacology , Escherichia coli Proteins/genetics , Escherichia coli Infections/microbiology , Enterobacteriaceae/genetics , Drug Resistance, Bacterial , beta-Lactamases/genetics , Plasmids/genetics , Feces/microbiology
7.
Viruses ; 15(4)2023 04 20.
Article in English | MEDLINE | ID: mdl-37112990

ABSTRACT

African swine fever (ASF) is a transboundary infectious disease that can infect wild and domestic swine and requires enhanced surveillance between countries. In Mozambique, ASF has been reported across the country, spreading between provinces, mainly through the movement of pigs and their by-products. Subsequently, pigs from bordering countries were at risk of exposure. This study evaluated the spatiotemporal distribution and temporal trends of ASF in swine in Mozambique between 2000 and 2020. During this period, 28,624 cases of ASF were reported across three regions of the country. In total, the northern, central, and southern regions presented 64.9, 17.8, and 17.3% of the total cases, respectively. When analyzing the incidence risk (IR) of ASF per 100,000 pigs, the Cabo Delgado province had the highest IR (17,301.1), followed by the Maputo province (8868.6). In the space-time analysis, three clusters were formed in each region: (i) Cluster A involved the provinces of Cabo Delgado and Nampula (north), (ii) Cluster B involved the province of Maputo and the city of Maputo (south), and (iii) Cluster C consisted of the provinces of Manica and Sofala (central) in 2006. However, when analyzing the temporal trend in the provinces, most were found to be decreasing, except for Sofala, Inhambane, and Maputo, which had a stationary trend. To the best of our knowledge, this is the first study to evaluate the spatial distribution of ASF in Mozambique. These findings will contribute to increasing official ASF control programs by identifying high-risk areas and raising awareness of the importance of controlling the borders between provinces and countries to prevent their spread to other regions of the world.


Subject(s)
African Swine Fever Virus , African Swine Fever , Swine Diseases , Swine , Animals , African Swine Fever/epidemiology , African Swine Fever/prevention & control , Mozambique/epidemiology , Spatio-Temporal Analysis , Sus scrofa , Disease Outbreaks , Swine Diseases/epidemiology
9.
Front Microbiol ; 13: 918706, 2022.
Article in English | MEDLINE | ID: mdl-36090116

ABSTRACT

Mastitis, mainly caused by bacterial intramammary infections, is the main problem in the breeding of dairy animals. The inflammations of the mammary gland is separated by types of mastitis, being subclinical, clinical, and the most severe, gangrenous mastitis. Here, we used 16S rRNA amplicon sequencing to characterize the bacterial microbiota of goat milk in the different types of goat mastitis caused by bacteria. We used 72 goat milk samples from a region of the state of Minas Gerais in Brazil, of which 12 were from clinically healthy animals, 42 from animals diagnosed with subclinical mastitis, 16 from animals with clinical mastitis, and 2 from animals with gangrenous mastitis. The group related to gangrenous mastitis was the most divergent in terms of alpha and beta diversity. The most abundant genus among samples of the groups was Staphylococcus spp., and we found a high abundance of Mycoplasma sp. in the milk of animals diagnosed with clinical mastitis. The most statistically relevant microorganisms among the groups were Prevotella sp., Ruminococcaceae, Prevotella ruminicola sp., and Providencia sp. We highlight a new association of bacterial agents in gangrenous mastitis among Escherichia sp./Shigella sp. and Enterococcus sp. and provide the second report of the genus Alkalibacterium sp., in milk samples. Only the taxa Staphylococcus sp., Bacteroides sp., Enterococcus, and Brevidabacterium sp., were present in all groups. The superpathway of L-tryptophan biosynthesis metabolites and the sucrose degradation III (sucrose invertase) pathway were the most prominent ones among the groups. In this study, we demonstrate how a rich microbiota of goat milk from healthy animals can be altered during the aggravation of different types of mastitis, in addition to demonstrating new bacterial genera in milk not previously detected in other studies as well as new associations between agents.

10.
Braz J Microbiol ; 53(2): 1081-1084, 2022 Jun.
Article in English | MEDLINE | ID: mdl-35286665

ABSTRACT

Respiratory diseases constitute a major health challenge for the worldwide pork industry. Porcine enzootic pneumonia (PES) is caused by Mycoplasma hyopneumoniae (Mhyo). Mycoplasmas have the ability to produce extracellular vesicles (EVs), which can be useful for pathogenicity studies and as delivery systems for vaccines. The aim of this study was to demonstrate and compare, under laboratory conditions, EVs produced by Mhyo strain J and wild isolate in stressed and non-stressed in vitro conditions. Using differential centrifugation, density gradient ultracentrifugation, and transmission electron microscopy, the ability of Mhyo strains to produce EVs was demonstrated under favorable and unfavorable conditions.


Subject(s)
Extracellular Vesicles , Mycoplasma hyopneumoniae , Pneumonia of Swine, Mycoplasmal , Swine Diseases , Animals , Pneumonia of Swine, Mycoplasmal/microbiology , Swine , Swine Diseases/microbiology , Virulence
11.
Sci Rep ; 11(1): 17252, 2021 08 26.
Article in English | MEDLINE | ID: mdl-34446803

ABSTRACT

Staphylococcus aureus is one of the main bacterial agents responsible for cases of mastitis in ruminants, playing an important role in the persistence and chronicity of diseases treated with antimicrobials. Using the multilocus sequence typing technique, network approaches and study of the population diversity of microorganisms, we performed analyzes of S. aureus (ES-GPM) isolated from goats with persistent mastitis (GPM). The most strains of ES-GPM were categorically different phylogenetically from the others and could be divided into two lineages: one with a majority belonging to ES-GPM and the other to varied strains. These two lineages were separated by 27 nuclear polymorphisms. The 43 strains comprised 22 clonal complexes (CCs), of which the ES-GPM strains were present in CC133, CC5 and a new complex formed by the sequence type 4966. The genetic diversity of some alleles showed be greater diversity and polymorphism than others, such as of the aroE and yqiL genes less than glpF gene. In addition, the sequences ES-GPM to the arc gene and glpF alleles showed the greatest number of mutations for ES-GPM in relation to non-ES-GPM. Therefore, this study identified genetic polymorphisms characteristic of S. aureus isolated from milk of goats diagnosed with persistent mastitis after the failed treatment with the antibiotic enrofloxacin. This study may help in the future to identify and discriminate this agent in cases of mastitis, and with that, the most appropriate antibiotic treatment can be performed in advance of the appearance of persistent mastitis caused by the agent, reducing the chances of premature culling and animal suffering.


Subject(s)
Enrofloxacin/pharmacology , Genetic Variation , Goat Diseases/drug therapy , Mastitis/drug therapy , Multilocus Sequence Typing/methods , Staphylococcal Infections/drug therapy , Staphylococcus aureus/genetics , Animals , Anti-Bacterial Agents/pharmacology , Brazil , Drug Resistance, Bacterial/genetics , Female , Geography , Goat Diseases/diagnosis , Goat Diseases/microbiology , Goats , Mastitis/diagnosis , Mastitis/microbiology , Microbial Sensitivity Tests/methods , Milk/microbiology , Phylogeny , Staphylococcal Infections/diagnosis , Staphylococcal Infections/microbiology , Staphylococcus aureus/classification , Staphylococcus aureus/physiology
12.
Front Public Health ; 9: 644748, 2021.
Article in English | MEDLINE | ID: mdl-34150698

ABSTRACT

Working the One health strategy in developing countries is a challenge, due to structural weaknesses or deprivation of financial, human, and material resources. Brazil has policies and programs that would allow continuous and systematic monitoring of human, animal, and environmental health, recommending strategies for control and prevention. For animals, there are components of the Epidemiological Surveillance of zoonosis and Animal Health Programs. To guarantee food safety, there are Health Surveillance services and support of the Agropecuary Defense in the inspection of these products, productive environments, and their inputs. Environmental Surveillance Services monitor water and air quality, which may influence health. For human health, these and other services related to Health Surveillance, such as Worker Health and Epidemiological Surveillance, which has a training program responsible for forming professionals groups to respond effectively to emergencies in public health are available. Therefore, Brazil has instruments that may allow integrated planning and intervention based on the One Health initiative. However, the consolidation of this faces several challenges, such as insufficient resources, professional alienation, and lack of the recognition of the importance of animal and environmental health for the maintenance of human and planetary well-being. This culminates in disarticulation, lack of communication, and integration between organizations. Thus, efforts to share attributions and responsibilities must be consolidated, overcoming the verticality of the actions, promoting efficiency and effectiveness. Finally, this perspective aims to describe the government instruments that constitute potential national efforts and the challenges for the consolidation of the One Health initiative in Brazil.


Subject(s)
One Health , Animals , Brazil/epidemiology , Disarticulation , Humans , Organizations , Public Policy
13.
Prev Vet Med ; 193: 105417, 2021 Aug.
Article in English | MEDLINE | ID: mdl-34192652

ABSTRACT

The buffalo herds in Brazil have been an alternative for increasing the economy in different biomes. For this reason, knowledge of the spatial distribution of diseases of mandatory notification in buffalo herds, such as brucellosis and tuberculosis, is essential to guarantee the quality of exported animal products, as well as assist in strategies of national control and eradication programs. In the present study, we aimed to evaluate the spatiotemporal distribution and temporal trends of brucellosis and tuberculosis in buffalo in Brazilian states between 2012-2019. During this period, 474 cases of brucellosis and 604 cases of tuberculosis were observed in buffalo in Brazil, with no significant differences between the total number of cases and incidence risk. The spatial distribution for the states was mostly heterogeneous, showing similarities of occurrences for both diseases in the south, north, and the states of Minas Gerais and Pernambuco. In the eight years evaluated, tuberculosis showed cyclical variation every 1-2 years; however, for brucellosis, there was a cyclical trend only between 2012-2015, with a significant decrease until 2018. Among Brazilian states, Pará had greater disease case numbers, with 34 % for brucellosis and 40.6 % for tuberculosis. Temporal trend analysis showed an increase for Pernambuco (annual percentage change [APC]: 21.0 [CI = 20.3; 21.8]), Paraná (APC: 27.1 [CI = 5.6; 53.0]), and Santa Catarina (APC: 10.4 [CI = 0.8; 21.0]) for brucellosis, and for tuberculosis, only for Santa Catarina state (APC: 24.1 [CI = 15.5; 33.3]). Spatiotemporally, there were four high-risk brucellosis clusters with a primary cluster, cluster A (relative risk [RRs] = 53.42, P < 0.001), involving the state of Amazonas between 2014-2015. For tuberculosis, there were three states of high risk, with a primary cluster, cluster E (RRs = 28.18, P < 0.001), involving the states of Pará, Roraima, Amazonas, Rondônia, and Acre in 2014. In conclusion, brucellosis and tuberculosis in buffalo in Brazil are heterogeneously distributed, with well-defined regions of high historical risk of infection. Among these regions, the states of Amazonas, Pará, Amapá, and Minas Gerais stand out due to the higher risk of infection for both brucellosis and tuberculosis, alerting Brazilian authorities to the need for disease control actions.


Subject(s)
Brucellosis , Buffaloes/microbiology , Tuberculosis, Bovine , Animals , Brazil/epidemiology , Brucellosis/epidemiology , Brucellosis/veterinary , Cattle , Incidence , Spatio-Temporal Analysis , Tuberculosis, Bovine/epidemiology
14.
Theriogenology ; 161: 228-236, 2021 Feb.
Article in English | MEDLINE | ID: mdl-33340756

ABSTRACT

Mycobacterium avium subsp. paratuberculosis (MAP) is responsible for Paratuberculosis mainly affecting domestic ruminants. The interaction between MAP and sperm and/or germ cells has not yet been established, however the adherence between MAP and the host cell surface is associated to the 85 complex proteins that bind to the host cell's fibronectin. Therefore, this study aimed to evaluate the binding of MAP to bovine sperm and to verify changes in these cells by the presence of MAP before and after sperm cryopreservation. Polyclonal antibodies to MAP 85 complex proteins were produced and utilized in the analyzes. Two Nelore bulls were used for semen collection and MAP dilutions (103-108 CFU/mL) were inoculated in the samples; sperm motility and vigor were evaluated using light microscopy at different times before and after cryopreservation and in the presence and absence of the antibodies 85A and 85B. Interaction of MAP and sperm, interaction of MAP and sperm in the presence of Ab 85A and in the presence of Ab 85B were analyzed by scanning electron microscopy. The viability of MAP after sperm cryopreservation were evaluated by plating the samples after thawing. It was observed that sperm in the presence of MAP shows a decrease in motility and vigor, and that the higher the MAP concentration, the lower the sperm performance. It was possible to determine the viability of MAP after cryopreservation in samples of higher concentrations, which demonstrates the potential of transmission of this pathogen through artificial insemination. The interaction of MAP with bovine sperm occurs mainly in the midpiece and may be linked to the proteins 85A and 85B present in the MAP membrane.


Subject(s)
Cattle Diseases , Mycobacterium avium subsp. paratuberculosis , Paratuberculosis , Animals , Cattle , Male , Sperm Motility , Spermatozoa
15.
J Dairy Res ; 87(3): 309-314, 2020 Aug.
Article in English | MEDLINE | ID: mdl-32958093

ABSTRACT

Our objectives were to identify genes of the multi-drug efflux system and to evaluate the antimicrobial activities of polypyrrole nanoparticles (PPy-NPs) and aqueous extract of Moringa oleifera against Staphylococcus spp. isolated from dairy farms in Northeast Brazil. Initially, 162 Staphylococcus spp. isolates were subjected to in vitro antimicrobial sensitivity tests. Of these, 35 presented antimicrobial multi-drug resistance phenotypes. These 35 isolates were then referred for the detection of norA, norB, norC, msrA, mgrA, tet-38, and lmrS genes, all of which feature in multi-drug efflux systems. In the isolates carrying the genes, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of PPy-NPs and Moringa oleifera aqueous extract were determined. In the molecular analysis of the 35 isolates norA, norC, tet-38, and msrA genes were detected and for the other genes norB, lmrS and mgrA there was no amplification. Antimicrobial activity was verified of PPy-NPs and aqueous extract of Moringa oleifera in Staphylococcus spp. carrying multi-drug efflux system genes. We concluded that there are multi-drug efflux system genes present in the Staphylococcus spp. from the agricultural environment in Northeast Brazil, and that aqueous extract of Moringa oleifera and PPy-NPs show bactericidal activity against these isolates.


Subject(s)
Anti-Bacterial Agents/pharmacology , Moringa oleifera/chemistry , Nanoparticles/chemistry , Plant Extracts/pharmacology , Polymers/chemistry , Pyrroles/chemistry , Staphylococcus/drug effects , Animals , Anti-Bacterial Agents/chemistry , Cattle , Dairying , Drug Resistance, Multiple, Bacterial , Gene Expression Regulation, Bacterial/drug effects , Plant Extracts/chemistry , Staphylococcus/genetics
16.
BMC Microbiol ; 20(1): 127, 2020 05 24.
Article in English | MEDLINE | ID: mdl-32448145

ABSTRACT

BACKGROUND: Staphylococcus aureus is one of the main causative agents of mastitis in small ruminants. Antimicrobial use is the major treatment, but there are many flaws linked to resistance, tolerance or persistence. This study aimed to verify changes in resistance, virulence and clonal profiles of S. aureus isolated from persistent mastitis goat milk before and after enrofloxacin treatment. RESULTS: MIC increased to at least one antimicrobial in S. aureus isolates after enrofloxacin treatment compared to before. The most detected resistance genes before and after treatment were tetK, tetM, and blaZ, with more resistance genes detected after enrofloxacin treatment (p < 0.05). Occasional variations in efflux system gene detection were observed before and after treatment. Nine virulence genes (hla, fnbA, fnbB, eta, etb, sea, sec, seh, and sej) were detected at both times, and between these, the hla and eta genes were detected more in isolates after treatment. All isolates of S. aureus belonged to the same sequence type (ST) 133, except for two S. aureus isolates prior to enrofloxacin treatment which were classified as ST5 and the other as a new one, ST4966. Isolates of S. aureus 4, 8, and 100 from before and after treatment had identical pulse types, while others obtained from other animals before and after treatment were classified into distinct pulse types. CONCLUSION: There were occasional changes in the studied profiles of S. aureus isolated before and after treatment of animals with enrofloxacin, which may have contributed to the permanence of bacteria in the mammary gland, even when using traditional treatment, resulting in persistent mastitis.


Subject(s)
Anti-Bacterial Agents/therapeutic use , Enrofloxacin/therapeutic use , Goat Diseases/drug therapy , Mastitis/veterinary , Staphylococcal Infections/drug therapy , Staphylococcus aureus/classification , Animals , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Drug Resistance, Bacterial , Enrofloxacin/pharmacology , Female , Gene Expression Regulation, Bacterial/drug effects , Goat Diseases/microbiology , Goats , Mastitis/drug therapy , Mastitis/microbiology , Microbial Sensitivity Tests , Multilocus Sequence Typing , Phylogeny , Staphylococcal Infections/veterinary , Staphylococcus aureus/drug effects , Staphylococcus aureus/isolation & purification , Staphylococcus aureus/pathogenicity , Virulence , Virulence Factors/genetics
17.
Sci Rep ; 10(1): 4421, 2020 03 10.
Article in English | MEDLINE | ID: mdl-32157153

ABSTRACT

Antibiotic resistance has become a major concern for human and animal health. As fluoroquinolones have been extensively used in human and veterinary medicine, there has also been the rapid emergence and spread of antimicrobial resistance around the world. Here, we analysed the microbiome of goat milk using samples from healthy goats and those diagnosed with persistent mastitis and treated using the antibiotic enrofloxacin with 16S rRNA amplicon sequencing. We selected a group of 11 goats and 22 samples of milk that did not respond clinically to enrofloxacin treatment. Milk samples were evaluated before and after treatment to verify changes of the microbiota; the three first lactating goats were selected from the healthy control group. The milk samples from the healthy control animals presented a larger abundance of different species of bacteria of the Staphylococcus genus, but a smaller number of different genera, which indicated a more specific niche of resident bacteria. The Firmicutes phylum was predominantly different between the studied groups. Samples from before-treatment animals had a higher number of new species than those from the control group, and after being treated again. These microbiota received new bacteria, increasing the differences in bacteria even more in relation to the control group. Genotypes such as Trueperella and Mannheimia, between other genera, had a high abundance in the samples from animals with persistent mastitis. The dysbiosis in this study, with marked evidence of a complex microbiota in activity in cases of the failure of antimicrobial treatment for persistent chronic mastitis, demonstrates a need to improve the accuracy of pathogen identification and increases concern regarding antibiotic treatments in milk production herds.


Subject(s)
Anti-Bacterial Agents/administration & dosage , Bacteria/classification , Enrofloxacin/administration & dosage , Goat Diseases/drug therapy , Mastitis/veterinary , Milk/microbiology , Animals , Anti-Bacterial Agents/pharmacology , Bacteria/drug effects , Bacteria/genetics , Bacteria/isolation & purification , Case-Control Studies , DNA, Bacterial/drug effects , DNA, Ribosomal/genetics , Enrofloxacin/pharmacology , Female , Goats , Mastitis/drug therapy , Microbiota/drug effects , Milk/drug effects , Phylogeny , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA/veterinary
18.
Braz J Microbiol ; 51(1): 377-384, 2020 Mar.
Article in English | MEDLINE | ID: mdl-31797326

ABSTRACT

Mycoplasma hyopneumoniae is the etiologic agent of porcine enzootic pneumonia, responsible for major production losses worldwide. The bacteria have a limited metabolism and need to obtain molecules from the growth environment, which causes multiple difficulties for in vitro culture. These limitations have a negative influence on the ability to carry out research for the development of the rational use of antimicrobials and vaccines. The objective of this investigation was to evaluate the genetic profile and in vitro susceptibility of field isolates of M. hyopneumoniae to different antimicrobials. All 16 isolates obtained from the samples presented 100% of identity in the partial sequence of 16S rRNA gene when compared to M. hyopneumoniae. A dendrogram was created using the PCR results of the genes related to pathogenicity, and the isolates were distributed into four clusters, suggesting genetic variability among four different isolates circulating on the same farm. The minimum inhibitory concentration of the isolates was higher for the antimicrobials tylosin (< 0.001-16 mg/L) and spiramycin (< 0.001-16 mg/L) than for enrofloxacin (< 0.001-0.125 mg/L) and tiamulin (< 0.001-0.125 mg/L). Our results demonstrate the genetic variability among M. hyopneumoniae isolates from pigs of the same farm, with differences in their susceptibility to antimicrobial agents.


Subject(s)
Anti-Bacterial Agents/pharmacology , Mycoplasma Infections/veterinary , Mycoplasma hyopneumoniae , Swine/microbiology , Animals , Brazil , Genes, Bacterial , Genetic Profile , Genetic Variation , Microbial Sensitivity Tests , Mycoplasma Infections/drug therapy , Mycoplasma Infections/microbiology , Mycoplasma hyopneumoniae/drug effects , Mycoplasma hyopneumoniae/genetics , Mycoplasma hyopneumoniae/isolation & purification , Mycoplasma hyopneumoniae/pathogenicity , Pneumonia of Swine, Mycoplasmal/drug therapy , Pneumonia of Swine, Mycoplasmal/microbiology , RNA, Ribosomal, 16S , Swine Diseases/microbiology , Virulence/genetics
20.
BMC Microbiol ; 19(1): 234, 2019 10 28.
Article in English | MEDLINE | ID: mdl-31660853

ABSTRACT

BACKGROUND: Porcine enzootic pneumonia is a worldwide problem in swine production. The infected host demonstrates a respiratory disease whose etiologic agent is Mycoplasma hyopneumoniae (Mhp). A total of 266 lung samples with Mycoplasma-like lesions were collected from two slaughterhouses. We analyzed the genetic profile of Mhp field samples using 16 genes that encode proteins involved in the mechanisms of bacterial pathogenesis and/or the immune responses of the host. Bioinformatic analyses were performed to classify the Mhp field samples based on their similarity according to the presence of the studied genes. RESULTS: Our results showed variations in the frequency of the 16 studied genes among different Mhp field samples. It was also noted that samples from the same farm were genetically different from each other and samples from different regions could be genetically similar, which is evidence of the presence of different genetic profiles among the Mhp field strains that circulate in Brazilian swine herds. CONCLUSION: This work demonstrated the genetic diversity of several Mhp field strains based on 16 selected genes related to virulence and/or immune response in Brazil. Our findings demonstrate the difference between Mhp field strains could influence the virulence, and we hypothesize that the most frequent genes in Mhp field strains could possibly be used as vaccine candidates. Based on our results, we suspect that Mhp genetic variability may be associated with the frequency of genes among the field strains and we have demonstrated that some Mhp field samples could not have many important genes described in the literature.


Subject(s)
Bacterial Proteins/genetics , Genetic Variation , Mycoplasma hyopneumoniae/genetics , Pneumonia of Swine, Mycoplasmal/microbiology , Abattoirs , Animals , Antigens, Bacterial/genetics , Brazil , Evolution, Molecular , Mycoplasma hyopneumoniae/immunology , Mycoplasma hyopneumoniae/pathogenicity , Sequence Analysis, DNA/methods , Swine , Virulence Factors/genetics
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