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Respiratory microbiota of healthy broilers can act as reservoirs for multidrug-resistant Escherichia coli
Abstract
This study aimed at evaluate the presence and to study characteristics of Escherichia coli in the respiratory system microbiota of healthy broilers. Trachea, air sacs, and lungs of 20 broilers were analyzed at 21 days of age, reared in experimental conditions, without receiving antimicrobials. E. coli strains were isolated and identified using conventional bacteriology through morphological and biochemical characterization. The production of bacteriocin-like substances, the presence of virulence-associated genes (VAGs) of APEC (Avian Pathogenic Escherichia coli) predictors, and the antimicrobial susceptibility were evaluated. E. coli was found in 85% of the animals (17/20), in the trachea, air sacs or lungs; and it was not found in 15% of the animals (3/20). A total of 34 isolates were recovered, 13 from the air sacs, 13 from the lungs, and 8 from the trachea, which showed no production of bacteriocin-like substances nor virulence genes associated with APEC. Most isolates, 59% (20/34), showed resistance to at least one of the tested antimicrobials, and six multiresistant strains were identified. The results demonstrated that strains of E. coli were commensal of the respiratory microbiota, and that they did not present pathogenicity to the host, since there were no clinical signs of disease, macroscopic lesions in the organs of the evaluated broilers, production of bacteriocin-like substances, nor virulence-associated genes considered as predictors of APEC in bacteria. These strains of E. coli were mostly susceptible to antimicrobials. However, the occurrence of multidrug-resistant strains suggests that these animals can act as reservoirs of resistant to antimicrobials E. coli.
... Effective probiotics, live microorganisms that confer a health benefit to the host, must present high survival and multiplication rates in the host's environment, high capacity to adhere to the epithelium, high stability under feed processing and distribution conditions, capacity to reduce pathogen numbers and to modulate the immune response (Khan and Naz 2013;Silva et al. 2016). In addition, one of the most important advantages of probiotics is that, unlike antibiotics, they do not leave any residues in the meat, which may damage consumers' health (Khan and Naz 2013;Landers et al. 2012;Soares et al. 2021). Probiotics do not replace antimicrobial agents, but may be used as growth promoters, saving antibiotics to be used when they really are needed (Macari and Furlan 2005). ...
... Evaluating the respiratory bacterial microbiota of clinically healthy broilers, was found higher frequencies of E. coli, coagulase-negative Staphylococcus and Streptococcus spp. In the present study, E. coli was also predominant, followed by Staphylococcus spp., Streptococcus sp., Klebsiella spp., Bacillus spp., and P. pseudoalcaligenes (Soares et al. 2021). ...
... Several substances with antimicrobial activity, such as bacteriocins, short-chain fatty acids, and hydrogen peroxide are included in the composition of probiotics with the aim the fosA3 gene. In another work was observed that 20% of the E. coli from the respiratory tract of clinically healthy broilers presented MDR profile (Soares et al. 2021). 20% of E. coli environmental samples collected in a broiler house presented MDR (Carvalho et al. 2015). ...
Desirable characteristics of Staphylococcus sp., Streptococcus sp., Bacillus sp., Klebsiella sp., Escherichia coli, and Pseudomonas pseudoalcaligenes isolated from the trachea of healthy turkeys were evaluated as probiotic candidates in the search for new alternatives to solve antimicrobial resistance issues in poultry. In current study phenotypic and genotypic capacity to produce bacteriocin-like substances, efficacy to inhibit the growth of avian pathogens, susceptibility to antimicrobials of bacteria isolated from the respiratory microbiota of healthy turkeys, and the presence of virulence-associated genes (VAGs) predictors of Avian Pathogenic Escherichia coli (APEC) were evaluated. Nine E. coli and one Klebsiella sp. strains produced bacteriocin-like substances, and all harbored the cvaA gene. Some strains also showed antagonistic activity against APEC. Multidrug-resistant profile was found in 54% of the strains. Six strains of bacteriocin-like substances producing E. coli also harbored 3–5 VAGs. The study showed that two bacterial genuses (Klebsiella sp. and E. coli) present desirable probiotic characteristics. Our results identified strains with potential for poultry’s respiratory probiotic.
... Although the influence of the microbiota on this disease has been documented in the gastrointestinal tract [130,132], a few studies have also confirmed the presence of Escherichia coli in the respiratory microbiota of poultry [33,133,134]. However, the isolated Escherichia coli strains from these studies were confirmed to be nonpathogenic to the host [133,134]. ...
... Although the influence of the microbiota on this disease has been documented in the gastrointestinal tract [130,132], a few studies have also confirmed the presence of Escherichia coli in the respiratory microbiota of poultry [33,133,134]. However, the isolated Escherichia coli strains from these studies were confirmed to be nonpathogenic to the host [133,134]. This observation is not surprising as the severity of APEC infection is dependent on strain pathogenicity and gene virulence [135,136]. ...
An accurate understanding of the ecology and complexity of the poultry respiratory microbiota is of utmost importance for elucidating the roles of commensal or pathogenic microorganisms in the respiratory tract, as well as their associations with health or disease outcomes in poultry. This comprehensive review delves into the intricate aspects of the poultry respiratory microbiota, focusing on its colonization patterns, composition, and impact on poultry health. Firstly, an updated overview of the current knowledge concerning the composition of the microbiota in the respiratory tract of poultry is provided, as well as the factors that influence the dynamics of community structure and diversity. Additionally, the significant role that the poultry respiratory microbiota plays in economically relevant respiratory pathobiologies that affect poultry is explored. In addition, the challenges encountered when studying the poultry respiratory microbiota are addressed, including the dynamic nature of microbial communities, site-specific variations, the need for standardized protocols, the appropriate sequencing technologies, and the limitations associated with sampling methodology. Furthermore, emerging evidence that suggests bidirectional communication between the gut and respiratory microbiota in poultry is described, where disturbances in one microbiota can impact the other. Understanding this intricate cross talk holds the potential to provide valuable insights for enhancing poultry health and disease control. It becomes evident that gaining a comprehensive understanding of the multifaceted roles of the poultry respiratory microbiota, as presented in this review, is crucial for optimizing poultry health management and improving overall outcomes in poultry production.
... Despite ongoing research, effective and broadly protective vaccines are still under development, leaving antimicrobial treatments, such as florfenicol, apramycin, and danofloxacin, as the primary control measures. However, the widespread use of antibiotics has raised concerns about antimicrobial resistance and heightened consumer demand for antibiotic-free poultry products, necessitating alternative control strategies [10,[12][13][14][15][16][17]. ...
Introduction: Ostrich farming is increasingly recognized for its economic potential but poses significant health challenges due to the risk of pathogen transmission, including multidrug-resistant Escherichia coli. Case study: This study reports a case of a four-month-old female ostrich from western Romania presenting with severe respiratory and digestive infections, progressing to septicemia and death. A post-mortem examination revealed extensive mucus in the trachea, pulmonary congestion, hemorrhagic enteritis, and approximately 1250 g of metal objects in the ventriculus. Pure cultures of E. coli were isolated from the lungs and bone marrow and identified via MALDI-TOF MS. The strain exhibited multidrug resistance to several antibiotics, including enrofloxacin, doxycycline, and amoxicillin, highlighting the critical issue of antimicrobial resistance in veterinary medicine. Discussions: This case underscores the need for enhanced management practices in ostrich farming to mitigate environmental and pathogenic risks, as well as the urgency of developing alternative strategies for controlling resistant bacterial infections in avian species. Conclusions: This case highlights the need for alternative treatments and stricter antimicrobial stewardship to combat multidrug-resistant E. coli in ostriches.
... More specifically in Brazil, studies showed that less than 25% of isolates from chicken treated with antimicrobials were resistant to gentamicin and neomycin [42,43]. In addition, florfenicol (amphenicol class) also showed a low resistance result of 26.2% in the antimicrobial susceptibility test, a rate similar to other reports [8,11]. ...
Colibacillosis is a chicken disease caused by avian pathogenic Escherichia coli (APEC). Pathogenicity in birds is determined by the occurrence of bacterial genes encoding virulence factors in APEC strains. Furthermore, APEC and other bacterial infections in commercial poultry farms have been treated with intensive use of antimicrobials for decades. Currently, many APEC strains are no longer susceptible to frequently used antibiotics due to increasing antimicrobial resistance (AMR) associated with the acquisition and mutation of other specific bacterial genes. The present study aimed to isolate and detect APEC isolates in broiler farms from different poultry-producing regions of Brazil and to determine their AMR profile. A total of 126 E. coli isolates were obtained from necropsied chickens with colibacillosis. All of these E. coli isolates were analyzed with one species-specific qPCR (targeting uspA gene) and five virulence factors genes qPCRs (targeting iroN, hlyF, iutA, iss, and ompT). AMR was determined by disk diffusion method using ten drugs frequently used to treat colibacillosis in Brazilian poultry farms. The results demonstrated that 109 (86.5%) isolates were classified as APEC. AMR was commonly observed in APEC and AFEC isolates, highlighting resistance for amoxicillin (85; 67.4%) and ceftiofur (72; 57.1%). A total of 41 (32.5%) E. coli isolates presented a multidrug resistance (MDR) profile. These results can contribute to implementing more effective colibacillosis prevention and control programs on Brazilian poultry farms.
... infectious diseases in poultry because multidrug-resistant E. coli can live in chicken respiratory microbiota, which forms competitive exclusion of normal bacteria [3]. As a result, immunization against salmonellosis and colibacillosis is still required in poultry farms. ...
Attenuated Salmonella Typhimurium is a promising antigen delivery system for live vaccines such as polysaccharides. The length of polysaccharides is a well-known key factor in modulating the immune response induced by glycoconjugates. However, the relationship between the length of Lipopolysaccharide (LPS) O-antigen (OAg) and the immunogenicity of S. Typhimurium remains unclear. In this study, we assessed the effect of OAg length determined by wzzST on Salmonella colonization, cell membrane permeability, antimicrobial activity, and immunogenicity by comparing the S. Typhimurium wild-type ATCC14028 strain to those with various OAg lengths of the ΔwzzST mutant and ΔwzzST::wzzECO2. The analysis of the OAg length distribution revealed that, except for the very long OAg, the short OAg length of 2-7 repeat units (RUs) was obtained from the ΔwzzST mutant, the intermediate OAg length of 13-21 RUs was gained from ΔwzzST::wzzECO2, and the long OAg length of over 20 RUs was gained from the wild-type. In addition, we found that the OAg length affected Salmonella colonization, cell permeability, and antibiotic resistance. Immunization of mice revealed that shortening the OAg length by altering wzzST had an effect on serum bactericidal ability, complement deposition, and humoral immune response. S. Typhimurium mutant strain ΔwzzST::wzzECO2 possessed good immunogenicity and was the optimum option for delivering E. coli O2 O-polysaccharides. Furthermore, the attenuated strain ATCC14028 ΔasdΔcrpΔcyaΔrfbPΔwzzST::wzzECO2-delivered E. coli O2 OAg gene cluster outperforms the ATCC14028 ΔasdΔcrpΔcyaΔrfbP in terms of IgG eliciting, cytokine expression, and immune protection in chickens. This study sheds light on the role of OAg length in Salmonella characteristics, which may have a potential application in optimizing the efficacy of delivered polysaccharide vaccines.
... Los microorganismos patógenos se caracterizan por la capacidad de producir daño leve o grave mediado estrictamente por las condiciones del hospedero que lo albergue (inmunosuprimido o inmunocompetente) (Martín et al., 2018). Los factores de virulencia emergen una vez el patógeno supera los mecanismos fisiológicos de respuesta inmune del organismo a infectar y se definen como una medida cuantitativa de la gravedad y número de microorganismos necesarios para causar una enfermedad, es decir, es el grado de patogenicidad (Bag et al., 2021;Soares et al., 2021). Debido a la eficacia de los mecanismos se considera que una bacteria puede ser poco o muy virulenta, estos se identifican en dos grupos de fase temprana y de fase tardía (Nüesch et al., 2021). ...
The virulence factors of pathogens are expressed once they overcome the physiological mechanisms of immune response by the organism, Escherichia coli (E. coli) is a bacterium that is considered a public health problem worldwide due to the high prevalence of resistance and pathogenicity mechanisms expressed , it is mainly related to intestinal infections and is transmitted in the food chain; the genetic determinants encoding virulence factors are transferred between different species or the same, through the mechanism of horizontal gene transfer. The present review aims to describe virulence factors and genes encoding them in E. coli strains isolated from production animals and food products. The databases Medline, Lilacs, ScienceDirect, Scopus, SciELO, and Dialnet were searched using a combination of validated keywords in English (gene, virulence, virulence factor , infection, horizontal gene transfer, mutation, and production animals). Different virulence mechanisms were identified in different production environments, which vary according to the animal and bacterial species. The genes with the highest circulation were Stx1, Stx2 coding for toxins, Saa for adhesins, ehxA for enterohe-molysin, eaeA for intimin, and IpfA for fimbriae. Microbiological surveillance and control in the food and animal production area are of great importance to avoid possible disease outbreaks in susceptible populations, due to the effect of virulence factors.
... Los microorganismos patógenos se caracterizan por la capacidad de producir daño leve o grave mediado estrictamente por las condiciones del hospedero que lo albergue (inmunosuprimido o inmunocompetente) (Martín et al., 2018). Los factores de virulencia emergen una vez el patógeno supera los mecanismos fisiológicos de respuesta inmune del organismo a infectar y se definen como una medida cuantitativa de la gravedad y número de microorganismos necesarios para causar una enfermedad, es decir, es el grado de patogenicidad (Bag et al., 2021;Soares et al., 2021). Debido a la eficacia de los mecanismos se considera que una bacteria puede ser poco o muy virulenta, estos se identifican en dos grupos de fase temprana y de fase tardía (Nüesch et al., 2021). ...
Los factores de virulencia de patógenos se expresan una vez superan los mecanismos fisiológicos de respuesta inmune por parte del organismo, Escherichia coli (E. coli) es una bacteria que se considera un problema en salud pública en todo el mundo debido a la alta prevalencia de mecanismos de resistencia y patogenicidad que expresa, se relaciona principalmente con infecciones intestinales y es transmitida en la cadena alimenticia; los determinantes genéticos que codifican factores de virulencia se trasladan entre diferentes especies o la misma, mediante el mecanismo de transferencia horizontal de genes. El objetivo de la presente revisión es describir factores de virulencia y genes que los codifican en cepas de E. coli aisladas de animales de producción y de productos alimenticios. En las bases de datos Medline, Lilacs, ScienceDirect, Scopus, SciELO y Dialnet, se realizó búsqueda utilizando una combinación de palabras claves validadas en inglés (gen, virulence, virulence factor, infection, horizontal gene transfer, mutation and production animals). En diferentes ambientes de producción se identificó la presencia de diferentes mecanismos de virulencia que varían según la especie animal y bacteriana, los genes con mayor circulación con Stx1, Stx2 codificantes de toxinas, Saa de adhesinas, ehxA de enterohemolisina, eaeA de intimina, IpfA de fimbrias. La vigilancia y control microbiológico en el área alimenticia y de producción animal es de gran importancia para evitar posibles brotes de enfermedades en población susceptible, por efecto de los factores de virulencia.
The respiratory tracts of turkeys play important roles in the overall health and performance of the birds. Understanding the bacterial communities present in the respiratory tracts of turkeys can be helpful to better understand the interactions between commensal or symbiotic microorganisms and other pathogenic bacteria or viral infections. The aim of this study was the characterization of the bacterial communities of upper respiratory tracks in commercial turkeys using NGS sequencing by the amplification of 16S rRNA gene with primers designed for hypervariable regions V3 and V4 (MiSeq, Illumina). From 10 phyla identified in upper respiratory tract in turkeys, the most dominated phyla were Firmicutes and Proteobacteria. Differences in composition of bacterial diversity were found at the family and genus level. At the genus level, the turkey sequences present in respiratory tract represent 144 established bacteria. Several respiratory pathogens that contribute to the development of infections in the respiratory system of birds were identified, including the presence of Ornithobacterium and Mycoplasma OTUs. These results obtained in this study supply information about bacterial composition and diversity of the turkey upper respiratory tract. Knowledge about bacteria present in the respiratory tract and the roles they can play in infections can be useful in controlling, diagnosing and treating commercial turkey flocks.
Diarrheagenic (DEC) and avian pathogenic Escherichia coli (APEC) are associated with intestinal and extra-intestinal infections (ExPEC), respectively. We aimed to analyze the antimicrobial susceptibility, gene encoding virulence factors associated to DEC and APEC, and phylogenetic classification in E. coli isolated from 320 samples of feed and ingredients. Antimicrobial susceptibility was performed using the disk diffusion method and Multiple Antibiotic Resistance (MAR) Index and Multi-Drug Resistance (MDR) were calculated. Phylogenetic classification was performed on samples harboring DEC and/or APEC virulence-associated genes. A total of 110 E. coli strains were isolated in 15% (49/320) of the evaluated inputs (n=13 vegetable meal; n=33 animal meal, n=3 feed). In general, the isolates showed the highest rates of antimicrobial resistance to sulfonamide and cefazolin and 18% (20/110) were multi-drug resistant. MAR index of feed samples was the highest (0.467). Six and five strains had APEC and DEC virulence-associated genes, respectively, and belonging to phylogenetic groups A and B1. These findings point to the need for strict microbiological control during the production process of these foods.
The poultry industry is faced with numerous challenges associated with infectious diseases and suboptimal performance of flocks. As microbiome research continues to grow, it is becoming clear that poultry health and production performance are partly influenced by nonpathogenic symbionts that occupy different habitats within the bird. This study has defined the baseline composition and overlaps between respiratory and gut bacteria in healthy, optimally performing chicken layers across all stages of the commercial farm sequence. Consequently, the study has set the groundwork for the development of interventions that seek to enhance production performance and to prevent and control infectious diseases through the modulation of gut and respiratory bacteria.
Pathogenic Escherichia coli found in humans and poultry carcasses harbor similar virulence and resistance genes. The present study aimed to analyze the distribution of extraintestinal pathogenic E. coli (ExPEC) virulence factors (VF), blaCTX−M groups, fosA3, and mcr-1 genes in E. coli isolated from commercialized chicken carcasses in southern Brazil and to evaluate their pathogenic risk. A total of 409 E. coli strains were isolated and characterized for genes encoding virulence factors described in ExPEC. Results of antimicrobial susceptibility testing confirmed that the strains were resistant to β-lactams, fosfomycin, colistin, and others resistance groups. The highest prevalence of VFs was observed in isolates belonging to the CTX-M groups, especially the CTX-M-2 group, when compared to those in other susceptible strains or strains with different mechanisms of resistance. Furthermore, ESBL strains were found to be 1.40 times more likely to contain three to five ExPEC virulence genes than non-ESBL strains. Our findings revealed the successful conjugation between ESBL-producing E. coli isolated from chicken carcass and the E. coli recipient strain J53, which suggested that genetic determinants encoding CTX-M enzymes may have originated from animals and could be transmitted to humans via food chain. In summary, chicken meat is a potential reservoir of MDR E. coli strains harboring resistance and virulence genes that could pose serious risks to human public health.
The study was undertaken for isolation, identification and in vitro antibiotic sensitivity assay of bacteria present in the respiratory tract of apparently healthy Japanese quails. A total of 50 samples comprised of tracheal swabs (n = 18), tracheal washings (n = 8), air sacs (n = 8), lungs (n = 8) and exudates of infraorbital sinuses (n = 8) were aseptically collected from 26 apparently healthy Japanese quails. The samples were inoculated onto a variety of media for isolation of bacteria. Identification of bacteria was performed by colony morphology, Gram’s staining and biochemical tests. In total, 25 (50%), 9 (18%), 22 (44%), 20 (40%) and 24 (48%) isolates were identified as Escherichia coli , Salmonella spp., Pasteurella spp., Bacillus spp. and Staphylococcus spp. respectively. Antibiotic sensitivity tests of one randomly selected isolate from each genus of bacteria were performed against 10 common antibiotics. E. coli showed resistance to four antibiotics (amoxicillin, gentamycin, nalidixic acid and tetracycline); Salmonella spp. showed resistance to seven antibiotics (amoxicillin, ampicillin, erythromycin, gentamycin, kanamycin, nalidixic acid and tetracycline); Pasteurella spp. to three antibiotics (erythromycin, sulphamethoxazole and tetracycline); Bacillus spp. to chloramphenicol and nalidixic acid and Staphylococcus spp. to amoxicillin and ampicillin. All the isolates were susceptible to ciprofloxacin. Data of this study indicate that multidrug resistant bacteria are present in the respiratory tract of clinically healthy quails. This is the first report of isolation, identification and antibiogram profile of bacteria present in the respiratory tract of quails in Bangladesh. DOI: http://dx.doi.org/10.3329/mh.v1i2.14088 Microbes and Health, 2012 1(2): 46-49
Microbiological contamination in commercial poultry production has caused concerns for human health because of both the presence of pathogenic microorganisms and the increase in antimicrobial resistance in bacterial strains that can cause treatment failure of human infections. The aim of our study was to analyze the profile of antimicrobial resistance and virulence factors of
E. coli
isolates from chicken carcasses obtained from different farming systems (conventional and free-range poultry). A total of 156
E. coli
strains were isolated and characterized for genes encoding virulence factors described in extraintestinal pathogenic
E. coli
(ExPEC). Antimicrobial susceptibility testing was performed for 15 antimicrobials, and strains were confirmed as extended spectrum of
β
-lactamases- (ESBLs-) producing
E. coli
by phenotypic and genotypic tests. The results indicated that strains from free-range poultry have fewer virulence factors than strains from conventional poultry. Strains from conventionally raised chickens had a higher frequency of antimicrobial resistance for all antibiotics tested and also exhibited genes encoding ESBL and AmpC, unlike free-range poultry isolates, which did not. Group 2 CTX-M and CIT were the most prevalent ESBL and AmpC genes, respectively. The farming systems of poultries can be related with the frequency of virulence factors and resistance to antimicrobials in bacteria.
Escherichia coli is a common inhabitant of the gastrointestinal tracts of humans and animals. Commensal E. coli, designated as nonpathogenic, are considered to be the majority. This microorganism can be divided into two large groups: pathogenic and commensal E. coli. The pathogenic group is characterized by two more pathotypes: diarrheagenic E. coli (DEC) (with eight subpathotypes) and extraintestinal pathogenic E. coli (ExPEC) (with six subpathotypes). ExPEC leads to disease in humans and animals. Avian colibacillosis is the main disease caused by ExPEC and has led to millions of dollars in losses to the worldwide poultry industry. Another problem associated with ExPEC strains is the high rates of antibiotic resistance. The indiscriminate use of antibiotics in animal production may have contributed to the resistant strains of ExPEC. E. coli strains of animal origin have been reported in humans. Herein we discuss the major diseases associated with E. coli and the problems of resistance to antibiotics in humans and the poultry industry, as well as alternative methods to control colibacillosis in poultry.
Avian pathogenic Escherichia coli (APEC) causes extraintestinal infections in birds, leading to an increase in the cost of poultry production. The ColV plasmid-linked genes iroN, ompT, hlyF, iss, and iutA have previously been suggested to be predictors of the virulence of APEC. In this research, we analyzed the frequencies of these genes in a Brazilian collection of E. coli isolated from birds with colibacillosis (APEC) and from apparently healthy birds (avian fecal [Afecal]), as well as from the litter of poultry houses of apparently healthy flocks (avian litter [Alitter]). All the isolates that harbored ompT also harbored hlyF, so they were considered as one trait for statistical analysis. The relationship between in vivo virulence in 1-day-old chicks, expressed as a pathogenicity score, and the number of genes in each isolate showed that isolates with less than two of the four genes were rarely pathogenic, while most pathogenic isolates contained two or more genes. Nevertheless, about half of the nonpathogenic isolates also harbored two or more genes, in agreement with previous observations that commensal E. coli isolates from the birds' microbiota can serve as a reservoir of virulence genes. Thus, the pentaplex polymerase chain reaction can be used to indicate that a strain carrying none or only one gene would be nonpathogenic, but it cannot be used to indicate that a strain with two to four genes would be an APEC. Isolates allocated to phylogenetic group B2, which is frequently associated with extraintestinal infections, had the highest pathogenicity scores, while isolates allocated to group B1 had the lowest.
O estudo teve como objetivo avaliar a suscetibilidade antimicrobiana de 109 amostras de Escherichia coli (E. coli) de origem ambiental frente a antibióticos e caracterizar esses isolados quanto ao grau de patogenicidade in vivo, verificando-se uma possível relação entre esta variável e a suscetibilidade aos princípios ativos testados. Os isolados foram submetidos ao teste de disco-difusão para 14 antibióticos. Entre 16.5% a 90% das amostras foram sensíveis, 1-28.5% apresentaram grau de suscetibilidade intermediário e entre 9-78% das E. coli analisadas foram resistentes. Os maiores percentuais de resistência foram encontrados para a classe das quinolonas e das tetraciclinas (>75%), e de sensibilidade para a classe dos anfenicóis (68.8%). Por meio da inoculação em pintinhos de um dia de idade, os isolados foram classificados como sendo de patogenicidade alta (2.7%), intermediária (10.1%), baixa (42.2%) e apatogênicos (45%). Foi observada uma ampla variação no perfil de suscetibilidade das amostras frente aos antimicrobianos. Verificou-se também que a maioria apresentou potencial patogênico (55%), sendo, portanto, consideradas APEC (E. coli patogênica para aves). Não foi observada relação entre a patogenicidade e a suscetibilidade aos antimicrobianos (P≤0.05).
Abstract This study characterized 52 Escherichia coli isolates from distinct diseased organs of 29 broiler chickens with clinical symptoms of colibacillosis in the Southern Brazilian state of Rio Grande do Sul. Thirty-eight isolates were highly virulent and 14 were virtually avirulent in 1-day-old chicks, yet all isolates harbored virulence factors characteristic of avian pathogenic E. coli (APEC), including those related to adhesion, iron acquisition, and serum resistance. E. coli reference collection phylogenetic typing showed that isolates belonged mostly to group D (39%), followed by group A (29%), group B1 (17%), and group B2 (15%). Phylogenetic analyses using the Amplified Ribosomal DNA Restriction Analysis and pulse-field gel electrophoresis methods were used to discriminate among isolates displaying the same serotype, revealing that five birds were infected with two distinct APEC strains. Among the 52 avian isolates, 2 were members of the pandemic E. coli O25:H4-B2-ST131 clone.
Recent advances in the technology available for culture-independent methods for identification and enumeration of environmental bacteria have invigorated interest in the study of the role of chicken intestinal microbiota in health and productivity. Chickens harbour unique and diverse bacterial communities that include human and animal pathogens. Increasing public concern about the use of antibiotics in the poultry industry has influenced the ways in which poultry producers are working towards improving birds' intestinal health. Effective means of antibiotic-independent pathogen control through competitive exclusion and promotion of good protective microbiota are being actively investigated. With the realisation that just about any change in environment influences the highly responsive microbial communities and with the abandonment of the notion that we can isolate and investigate a single species of interest outside of the community, came a flood of studies that have attempted to profile the intestinal microbiota of chickens under numerous conditions. This review aims to address the main issues in investigating chicken microbiota and to summarise the data acquired to date.
We characterized 144 Escherichia coli isolates from severe cellulitis lesions in broiler chickens from South Brazil. Analysis of susceptibility to 15 antimicrobials revealed frequencies of resistance of less than 30% for most antimicrobials except tetracycline (70%) and sulphonamides (60%). The genotyping of 34 virulence-associated genes revealed that all the isolates harbored virulence factors related to adhesion, iron acquisition and serum resistance, which are characteristic of the avian pathogenic E. coli (APEC) pathotype. ColV plasmid-associated genes (cvi/cva, iroN, iss, iucD, sitD, traT, tsh) were especially frequent among the isolates (from 66.6% to 89.6%). According to the Clermont method of ECOR phylogenetic typing, isolates belonged to group D (47.2%), to group A (27.8%), to group B2 (17.4%) and to group B1 (7.6%); the group B2 isolates contained the highest number of virulence-associated genes. Clonal relationship analysis using the ARDRA method revealed a similarity level of 57% or higher among isolates, but no endemic clone. The virulence of the isolates was confirmed in vivo in one-day-old chicks. Most isolates (72.9%) killed all infected chicks within 7 days, and 65 isolates (38.1%) killed most of them within 24 hours. In order to analyze differences in virulence among the APEC isolates, we created a pathogenicity score by combining the times of death with the clinical symptoms noted. By looking for significant associations between the presence of virulence-associated genes and the pathogenicity score, we found that the presence of genes for invasins ibeA and gimB and for group II capsule KpsMTII increased virulence, while the presence of pic decreased virulence. The fact that ibeA, gimB and KpsMTII are characteristic of neonatal meningitis E. coli (NMEC) suggests that genes of NMEC in APEC increase virulence of strains.
We examined 216 Escherichia coli (E. coli) isolated from chickens and environmental specimens from hatcheries between 2005 and 2006 in order to evaluate the epidemiological prevalence of avian pathogenic E. coli (APEC) in Korea tentatively by multiplex PCR. The multiplex PCR which was used as tentative criteria of APEC targets 8 virulence-associated genes; enteroaggregative toxin (astA), increased serum survival protein (iss), iron-repressible protein (irp2), P fimbriae (papC), aerobactin (iucD), temperature-sensitive hemagglutinin (tsh), vacuolating autotransporter toxin (vat), and colicin V plasmid operon (cva/cvi) genes. The number of detected genes could be used as a reliable index of their virulence. It was demonstrated that E. coli strains already typed as APEC always harbor 5 to 8 genes, but non-APEC strains harbor less than 4 genes. Assuming the criteria of APEC is a possession of more than 5 virulence-associated genes, we discriminated 24 APEC strains among the 216 E. coli strains. Contamination rates of APEC in the field were 31.3% in layers, 14.0% in broilers, 2.7% in broiler breeders, and 0.0% in environmental specimens from hatcheries. The combinational tendency of APEC examined is a fundamental possession of astA, iss and iucD genes and addition of cva/cvi, tsh, vat, and irp2 genes which have a critical importance for virulent traits of APEC. Compared with intravenous chicken challenge or embryo lethality assay, multiplex PCR method could be useful to discriminate APEC rapidly for convenient diagnosis.
This review article presents fundamental mechanisms of the local mucosal immunity in selected regions of the respiratory tract in healthy birds and in some pathological conditions. The respiratory system, whose mucosa come into direct contact with microorganisms contaminating inhaled air, has some associated structures, such as Harderian gland (HG), conjunctive-associated lymphoid tissue (CALT) and paranasal glands (PG), whose participation in local mechanisms of the mucosal immunity has been corroborated by numerous scientific studies. The nasal mucosa, with structured clusters of lymphoid tissue (NALT - nasal-associated lymphoid tissue) is the first to come into contact with microorganisms which contaminate inhaled air. Lymphoid nodules, made up of B cells with frequently developed germinal centres (GC), surrounded by a coat of CD4+ cells, are the major NALT structures in chickens, whereas CD8+ cells are situated in the epithelium and in the lamina propria of the nasal cavity mucosa. Studies into respiratory system infections (e.g. Mycoplasma gallisepticum) have shown the reactivity of the tracheal mucosa to infection, despite a lack of essential lymphoid tissue. Bronchus-associated lymphoid tissue (BALT) takes part in bronchial immune processes and its structure, topography and ability to perform defensive function in birds is largely age-dependent. Mature BALT is covered by a delicate layer of epithelial cells, called follicle-associated epithelium (FAE). Germinal centres (GC), surrounded by CD4+ cells are developed in most mature BALT nodules, while CD8+ lymphocytes are dispersed among lymphoid nodules and in the epithelium, and they are rarely present in GC. Macrophages make up the first line of defence mechanisms through which the host rapidly responds to microorganisms and their products in the respiratory mucosal system. Another very important element are polymorphonuclear cells, with heterophils being the most important of them. Phagocytic cells obtained from lung lavages in birds are referred to as FARM (free avian respiratory macrophages). Their number in chickens and turkeys is estimated to be 20 times lower than that in mice and rats, which indicates a deficit in the first-line of defence in the birds' respiratory system. There are numerous B cells and antibody secreting cells (ASC) present throughout the respiratory system in birds. Their role comes down to perform antigen-specific protection by producing antibodies (IgM, IgY or IgA class) as a result of contact with pathogenic factors.
To identify traits that predict avian pathogenic Escherichia coli (APEC) virulence, 124 avian E. coli isolates of known pathogenicity and serogroup were subjected to virulence genotyping and phylogenetic typing. The results
were analyzed by multiple-correspondence analysis. From this analysis, five genes carried by plasmids were identified as being
the most significantly associated with highly pathogenic APEC strains: iutA, hlyF, iss, iroN, and ompT. A multiplex PCR panel targeting these five genes was used to screen a collection of 994 avian E. coli isolates. APEC isolates were clearly distinguished from the avian fecal E. coli isolates by their possession of these genes, suggesting that this pentaplex panel has diagnostic applications and underscoring
the close association between avian E. coli virulence and the possession of ColV plasmids. Also, the sharp demarcation between APEC isolates and avian fecal E. coli isolates in their plasmid-associated virulence gene content suggests that APEC isolates are well equipped for a pathogenic
lifestyle, which is contrary to the widely held belief that most APEC isolates are opportunistic pathogens. Regardless, APEC
isolates remain an important problem for poultry producers and a potential concern for public health professionals, as growing
evidence suggests a possible role for APEC in human disease. Thus, the pentaplex panel described here may be useful in detecting
APEC-like strains occurring in poultry production, along the food chain, and in human disease. This panel may be helpful toward
clarifying potential roles of APEC in human disease, ascertaining the source of APEC in animal outbreaks, and identifying
effective targets of avian colibacillosis control.
The production of antimicrobial compounds known as colicins has been shown to be an important mediator of competitive interactions among Escherichia coli genotypes. There is some understanding of the forces responsible for determining the frequency of colicin production in E. coli populations; however, this understanding cannot explain all of the observed variation. A survey of colicin production in E. coli isolated from native Australian mammals revealed that the frequency of colicin production in strains isolated from carnivores was significantly lower than the frequency of production in strains recovered from herbivores or omnivores. The intestine of Australian carnivores is tube-like and gut turnover rates are rapid compared with the turnover rates of the intestinal tracts of herbivores and omnivores, all of which possess a hindgut fermentation chamber. A mathematical model was developed in order to determine if variation in gut turnover rates could determine if a host was more likely to harbour a colicin-producing strain or a non-producer. The model predicted that a colicin producer was more likely to dominate in the gut of a host with lower gut turnover rates, and a non-producer to dominate in hosts with rapid gut turnover rates.
Colibacillosis refers to any localized or systemic infection caused entirely or partly by avian pathogenic Escherichia coli. Syndromes of disease include colisepticemia, hemorrhagic septicemia, coligranuloma, airsacculitis, swollen‐head syndrome, venereal colibacillosis, coliform cellulitis, peritonitis, salpingitis, orchitis, osteomyelitis/synovitis, panophthalmitis, omphalitis/yolk sac infection, and enteritis. This chapter offers detailed coverage of the history, etiology, pathobiology, epidemiology, diagnosis, and intervention strategies of campylobacteriosis. The various forms of colibacillosis are considered to be the most common infectious bacterial disease of broiler chickens and turkeys. Diagnosis is based on isolation and identification of E. coli from lesions typical of colibacillosis. Isolation of the organism from visceral organs of birds undergoing decomposition must be interpreted cautiously as E. coli rapidly spreads from the intestinal tract into surrounding tissues of dead birds. Interventions for colibacillosis include hatchery and flock management practices, with emphasis on air quality, temperature, litter and housing environment, and sanitation.
Explaining the coexistence of competing species is a major challenge in community ecology. In bacterial systems, competition
is often driven by the production of bacteriocins; narrow spectrum proteinaceous toxins that serve to kill closely related
species providing the producer better access to limited resources. Bacteriocin producers have been shown to competitively
exclude sensitive, nonproducing strains. However, the interaction dynamics between bacteriocin producers, each lethal to its
competitor, are largely unknown. Several recent studies have revealed some of the complexity of these interactions, employing
a suite of in vitro, in vivo, and in silico bacterial model systems. This chapter describes the current state of knowledge
regarding the population and community ecology of this potent family of toxins.
Many different definitions for multidrug-resistant (MDR), extensively drug-resistant (XDR) and pandrug-resistant (PDR) bacteria are being used in the medical literature to characterize the different patterns of resistance found in healthcare-associated, antimicrobial-resistant bacteria. A group of international experts came together through a joint initiative by the European Centre for Disease Prevention and Control (ECDC) and the Centers for Disease Control and Prevention (CDC), to create a standardized international terminology with which to describe acquired resistance profiles in Staphylococcus aureus, Enterococcus spp., Enterobacteriaceae (other than Salmonella and Shigella), Pseudomonas aeruginosa and Acinetobacter spp., all bacteria often responsible for healthcare-associated infections and prone to multidrug resistance. Epidemiologically significant antimicrobial categories were constructed for each bacterium. Lists of antimicrobial categories proposed for antimicrobial susceptibility testing were created using documents and breakpoints from the Clinical Laboratory Standards Institute (CLSI), the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the United States Food and Drug Administration (FDA). MDR was defined as acquired non-susceptibility to at least one agent in three or more antimicrobial categories, XDR was defined as non-susceptibility to at least one agent in all but two or fewer antimicrobial categories (i.e. bacterial isolates remain susceptible to only one or two categories) and PDR was defined as non-susceptibility to all agents in all antimicrobial categories. To ensure correct application of these definitions, bacterial isolates should be tested against all or nearly all of the antimicrobial agents within the antimicrobial categories and selective reporting and suppression of results should be avoided.
Several human fecal isolates of Bacteroides have been found to produce bacteriocins. The bacteriocin-producing strain T1-1 was studied in the most detail. Strain T1-1 belongs to the 0061-1 deoxyribonucleic acid (DNA) homology group of Bacteroides. This homology group phenotypically resembles Bacteroides thetaiotaomicron but has little DNA homology with it. The bacteriocin-producing strains T1-12 and T1-48 belong to the 3452-A DNA homology group. This group has DNA homology with B. thetaiotaomicron and Bacteroides ovatus. The bacteriocin-producing strain T1-42 remains unidentified in that it does not belong to any recognized DNA homology group of the saccharolytic intestinal bacteroides. The extracellular bacteriocin produced by strain T1-1 was specifically bactericidal for other bacteria within the genus Bacteroides. The highest bacteriocin titers (32 to 64) were produced in complex media, with only trace amounts being produced in a defined medium. The bacteriocin appeared to have a high molecular weight (>/=300,000) and was unusual because it was stable from pH 1 to 12 and only a 50% reduction in activity resulted after 15 min at 121 degrees C in an autoclave. It was inactivated by trypsin and Pronase. Strain T1-1 was isolated from all three fecal samples obtained over a 25-week period from an individual who was part of a National Aeronautics and Space Administration mock Skylab flight. Strains T1-12, T1-48, and T1-42 were isolated only from the first fecal sample. Each of these strains was immune to the bacteriocins produced by the others. These strains were found to coexist in the colon with a larger population of non-bacteriocin-producing, bacteriocin-susceptible strains of Bacteroides.
The effects of three different types of breeding such as isolator, floor, and cage breedings on the bacterial flora of the respiratory tracts (nasal cavity, tongue, pharygolarynx, trachea and air sac) in chickens were determined. Total viable bacterial numbers on the nasal mucus of chickens in the isolator breeding as control group (Group A) aged of 14 days were 10(4.6)/g of autopsy specimen (wet weight), 10(5.7)/g of sample in the cage breeding (Group B) aged of 28 days, and 10(7.0)/g of sample in the floor breeding (Group C) aged of 28 days. Staphylococci and micrococci were predominant bacteria in the nasal cavities of all groups. Total viable numbers of tongue and pharygolarynx were from 10(5.4) to 10(6.5)/g of autopsy specimen. Lactobacilli were the predominant bacteria in pharyngolarynx of chickens. The incidence of staphylococci and micrococci in trachea was lower than those in the another regions. Staphylococci and micrococci dominated in the air sacs of two groups (B and C), but the number and incidence of lactobacilli in the air sacs of chickens were lower than those in the another respiratory tracts. The only clostridia isolation in the air sacs of Group A was observed. A total of 75 strains of Lactobacillus species was isoalted from all respiratory organs and intestine of chickens. These strains were divided into 19 groups. Lactobacillus salivearius subsp. salivarius was the predominant lactobacilli isolated from tongue and pharyngolarynx. Most of isolates from the chicken intestines were mainly identified as the L. acidophilus group and L. reuteri.(ABSTRACT TRUNCATED AT 250 WORDS)
Escherichia coli isolates taken from environments considered to have low and high enteric disease potential for humans were screened against 12 antibiotics to determine the prevalence of multiple antibiotic resistance among the isolates of these environments. It was determined that multiple-antibiotic-resistant E. coli organisms exist in large numbers within the major reservoirs of enteric diseases for humans while existing in comparatively low numbers elsewhere. These differences provide a method for distinguishing high-risk contamination of foods by indexing the frequency with which multiple-antibiotic-resistant E. coli organisms occur among isolates taken from a sample.
The bacterial flora of the trachea of 17 to 38 day old chickens was quantitatively and qualitatively studied in two groups of animals from two different sources: an experimental disease-free flock and a commercial flock. In both groups of chickens, the aerobic flora consisted predominantly of Escherichia coli (E. coli), Streptococcaceae, Micrococcaceae and members of the genus Lactobacillus. Some differences between the two groups of animals were observed at the species level. Strictly anaerobic bacteria did not appear until the third week. No bacterial group seemed to predominate during the sampling period. The constant presence of Lactobacilli and E. coli in the trachea suggests that these organisms could play a prominent role in the bacterial ecosystem of the trachea.
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