The contribution of the animal environments to the worsening of the global antimicrobial resistance framework is related to the use of antimicrobials in subtherapeutic doses and, for long periods, establishing ideal conditions for the circulation of resistance genes, which can be transmitted to pathogens adapted to the human microbiota. The study of the animal environment as conducive to the acceleration of resistance evolution is an emerging and critical area for understanding the development and dissemination of resistance genes among the circulating bacteria. The connection between people, animals, and the environment allows us to consider antimicrobial resistance in an approach within the “One Health” concept, which provides a global strategy for expanding collaboration and interdisciplinary communication. This chapter will highlight the emergence of colistin resistance, a great challenge in antimicrobial resistance field. Also, it will focus on some agents included in the priority list of superbugs of the World Health Organization (WHO) or correlated species already identified in veterinary medicine, such as the critical superbugs; priority level 1, Carbapenem-resistant Acinetobacter baumannii, Carbapenem-resistant Pseudomonas aeruginosa, and ESBL-producing Carbapenemic-resistant Enterobacteriaceae; and the high-priority, level 2, methicillin-resistant Staphylococcus aureus (MRSA).
- one health
- Staphylococcus pseudintermedius
- Acinetobacter baumannii
- mecA gene
- mcr genes
Global antimicrobial resistance indices are the subject of concern once it has been predicted that nearly 10 million annual deaths will be attributable to resistant pathogen infections by 2050 [1, 2]. The World Health Organization (WHO), the US Center for Disease Control and Prevention (CDC), and the European Center for Disease Prevention and Control (ECDC) classified the emergence and the spread of antimicrobial-resistant bacteria as one of the three major threats to public health in the twenty first century .
Importantly, the emergence of resistance is a natural evolutionary response to antimicrobial exposure. Over thousands of years, fungi and bacteria in the natural environment have developed complex mechanisms to prevent their destruction by toxic substances originating from the microbial competition, and these substances have made it possible to synthesize most antibiotics. Therefore, soils should be evaluated as potential reservoirs of antimicrobial-resistant bacteria and should be considered in assessing risk factors that contribute to the global spread of antimicrobial resistance. Moreover, the active collaboration of the human being in the propitiation of this emergency is undeniable due to the increased selection pressure, mainly given by the indiscriminate use of these drugs in human and veterinary medicine .
Antimicrobials not only kill sensitive and select resistant bacteria but also influence the mechanisms of genetic variation such as mutation, recombination, transposition, and gene exchange. Such phenomena can be observed from the soil to the intestinal microbiota of humans or animals exposed to antimicrobial underdosing, as the population of commensal microorganisms includes species that are naturally resistant to some antimicrobials. This selective pressure and subsequent imbalance due to the death of sensitive microorganisms allow bacteria with intrinsic or newly acquired resistance to survive and proliferate .
Despite this general understanding, the multifactorial origin of the current worldwide antimicrobial resistance scenario makes the picture complex and challenging to intervene. Although studies point to the hospital environment as the main reservoir for the resistance genes of bacteria that colonize and infect humans, the community environment indeed contributes to the establishment of a diverse set of resistance genes .
In 2012, Bhullar and colleagues  found multiresistant bacteria from an isolated cave microbiome over 4 million years ago in New Mexico, and some of the microorganisms were resistant to up to 14 commercial antibiotics. In another study, the ability of bacteria to use antibiotics as their sole carbon source was detected, making them a significant reservoir of antimicrobial resistance genes .
In this context, little is known about the contribution of animal production and veterinary hospital care environments in the maintenance of resistance genes and consequent resistance dissemination. The study of the contribution of various animal-related environments in accelerating the evolution of resistance is an emerging and critical area for understanding its development and as a model for the dissemination of resistance genes among the circulating bacteria. The connection between people, animals, and environment allows for the consideration of antimicrobial resistance within the One Health concept.
2. Distinct animal environment and its impact on antimicrobial resistance
2.1 The poultry production environment as a source of emerging colistin resistance
The increase in antibiotic resistance is now a global concern, including in food-producing animals. They can serve as a reservoir of antibiotic-resistant bacteria and antibiotic resistance determinants that may be transferred to humans [8, 9]. The systematic use of antibiotics in food-producing animals has been increasing the selection pressure for antibiotic-resistant bacteria, especially in Enterobacteriales such as
2.1.1 The silent colistin transferable plasmid-mediated resistance dissemination
The chromosomal polymyxin resistance is most associated with the modification of the lipopolysaccharide (LPS) following the addition of 4-amino-4-deoxy-L-arabinose to lipid A. Modifications of Ara4N are regulated by two-component systems: PhoP/PhoQ , PmrA/PmrB, and MgrB regulator. Mutations in genes involved in the production of these systems may result in lower antibiotic fixation . However, in 2015, a Chinese research group reported the emergence of a transferable plasmid-mediated resistance gene (mcr-1) from human, porcine, and poultry samples, shifting colistin resistance from a contained problem to a global issue . After identification of mcr-1, full scientific attention led to the recognition of multiple mcr-1 variants [16, 17, 18] and eight additional mcr genes. Subsequently, the mcr-2 plasmid-mediated colistin resistance gene was detected from poultry, porcine, and bovine
Nevertheless, colistin is often added to feed at low doses and used as a growth promoter in different countries. This practice may be the leading cause of the high rate of colistin-resistant bacteria carrying the mcr genes isolated from food-producing animals compared with humans and accelerate the dissemination of mcr genes from animals to humans [15, 27]. Furthermore, the mcr genes may have originated from food-producing animals. The mcr-1 gene was associated with
2.1.2 Data on colistin resistance in Brazil
In 2015, Brazil overtook China as the world’s second largest poultry producer. Nowadays, about 150 countries from all continents consume Brazilian broiler meat, according to the Brazilian Ministry of Agriculture Livestock and Farming . It is noticeable that scientific and technological advancements have transformed poultry from rural farming to full-fledged industry in the last few decades. However, despite this significant expansion, the Brazilian poultry industry is still highly dependent on antibiotic prescription. Prevalence data on colistin resistance in poultry and broiler are overall scarce in South America, including Brazil, in particular, data regarding the plasmid-mediated resistance to colistin [36, 37, 38]. In 2016, a Brazilian research group developed a retrospective antimicrobial resistance study and screened 4.620 Enterobacteriales strains isolated from human, animal, food, and environmental samples for the presence of the
Furthermore, between 2015 and 2016, Pimenta  also detected a high prevalence of
2.1.3 Colistin resistance genes in soils
Colistin, also known as polymyxin E, is produced by some strains of , a bacterium commonly found in soils associated with plant roots . In some places around the world, the use of poultry litter is an ordinary measure to improve the physical, chemical, and biological properties of soils in agricultural production. However, animal manure, such as poultry litter, a mixture of organic materials including feces, feed, and bedding, is a valuable nutrient-rich soil fertilizer also has been considered an important reservoir of antibiotic residues, antibiotic-resistant bacteria, and antibiotic resistance genes [41, 42]. The enhancement of the concentration and diversity of antibiotic resistance determinants in soils treated with this organic fertilizer is of concern, even considering that untreated soil environments harbor a natural source of both antibiotics and antibiotic resistance genes [43, 44, 45, 46]. The colistin resistance
2.2 Animal production environmental impact on genetic markers mutations: a study of
mecA gene of Staphylococcus aureusisolated from dairy system
In 2011, the report of MRSA strains presenting unusual features in bovine milk samples from the United Kingdom led to the discovery of a novel
2.2.2 A universal primer design experiment
Previous studies [62, 63] reported several phenotypic methicillin-resistant
To validate the newly designed primers, a set of 107 strains was tested for the presence of the
2.3 Companion animals environmental impact on antimicrobial resistance
Companion animals are part of human societies around the world . In veterinary medicine clinical practice, diseases such as pyodermitis, external otitis, urinary tract, and respiratory infections are the most frequent causes for the implementation of antibiotic therapy in dogs and cats. Wide-spectrum antimicrobials also prescribed in human medicine are commonly used in these treatments, such as aminopenicillins with beta-lactamase inhibitors, cephalosporins, fluoroquinolones, macrolides, aminoglycosides, and potentiated sulfonamides . As a result, the extensive and indiscriminate use of such antimicrobials in companion animals, coupled with their proximity to humans, gives canine and feline species importance as sources of antimicrobial resistance spread . In the last decade, the escalation of infectious conditions in the veterinary clinic of pet animals related to hitherto unknown or low prevalence agents such as
Acinetobacter calcoaceticus- Acinetobacter baumanniicomplex (Acb complex): an emerging challenge in companion animal environment
The analyses developed by our research group have identified all three species of clinical relevance of Acb complex, with the prevalence of
Staphylococcus pseudintermedius: an underestimated risk for animal and men
Although dogs are the natural hosts,
18.104.22.168 Is methicillin-resistant
Staphylococcus pseudintermedius(MRSP) the novel MRSA?
Two cases of methicillin-resistant
The relevance of
Staphylococcus pseudintermedius: genetic diversity and clonal distribution
In addition to the challenges of identifying
In Brazil, the prevalence of methicillin-resistant
2.4 β-Lactamase-producing Gram-negative bacteria in a one health approach
Most Enterobacteria pathogens associated with human enteric illness originate from animals and can be transmitted directly to humans or indirectly through animal origin food, contaminated water, or a common reservoir . Currently, β-lactamase-producing strains have been recovered from urban environments, companion/production animals, and animal source foods, which indicate a possible route of dissemination in different ecosystems.
To better understand these links and to identify control measures to reduce the bacterial resistant infections in humans and animals, a One Health approach is needed [121, 122]. The application of a global concept of cross-linking data will improve the prevention, prediction, and control of zoonotic diseases [123, 124].
Undoubtedly, the mobilization of resistance genes through plasmids, transposons, and integrons is intimately linked with widespread of β-lactamases, facilitating the exchange of genetic elements among various bacteria species that can later colonize different hosts and ecosystems and can be spread by different routes .
The detection of ESBLs in bacterial isolates of animal origin, such as
ESBL or plasmidial AmpC-β-lactamase producers are also frequently resistant to aminoglycosides and fluoroquinolones. The rate of resistance to these antibiotics among
2.4.1 β-Lactamases resistance in Gram-negative bacteria
The most common mechanism of resistance to beta-lactam antibiotics in Gram-negative bacteria is the production of hydrolytic enzymes of antimicrobial agents, including extended-spectrum beta-lactamases (ESBLs) . Two systems of classifying this array of enzymes are in use: the Bush-Jacoby-Medeiros activity-based system  and the Ambler system  based on nucleotide and amino acid sequence information . The resistance to beta-lactamase inhibitors characterizes the group I (Ambler class C) beta-lactamases (also known as AmpC enzymes). AmpC is mostly found on chromosomes, and its production is inducible. Group 2 (Ambler Class A) beta-lactamases could easily be transmitted into different bacterial cells once plasmids carry them. This group comprises the largest number of characterized enzymes divided into subgroup 2b hydrolyzing penicillins and cephalosporins and its variation 2be (known as “ESBL”). ESBL present a broad spectrum of various antimicrobials as ceftazidime, cefotaxime, and aztreonam. Clavulanic acid exerts potent inhibition towards them. Group 3 (Ambler Class B) enzymes are metalloenzymes capable of destroying carbapenems. Finally, group 4 beta-lactamases contain those unusual penicillinases not inhibited by clavulanic acid, and four of these enzymes exhibit high rates of hydrolysis with carbenicillin or cloxacillin .
The spread of extended-spectrum β-lactamase-producing Gram-negative bacteria has dramatically increased worldwide regarding as one of the most important public health threats. Therefore, their appropriate classification and epidemiological data on the main enzymes disseminated in humans, animals, and the environment are of utmost importance.
2.4.2 An historical approach
The first plasmid-mediated beta-lactamase in Gram-negative bacteria was reported in Greece in the 1960s. At the end of the 1970s, most
2.4.3 The CTX-M-type β-lactamase resistance dissemination
The CTX-M-type β-lactamases can be further differentiated into at least six sub-lineages or groups, namely, CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, CTX-M-25, and KLUC . The impressive worldwide spread of CTX-M-producing Gram-negative bacteria turned them to be considered the primary ESBL producers associated with community-acquired infections. The CTX-M family is described as predominant in South America, as well as in Spain and Eastern Europe . Therefore, according to the increasing number of reports describing these enzymes in Brazil, it appears that CTX-M variants are also prevalent in the country compared to TEM and SHV enzymes, prevalent in North America and Western Europe, respectively . In Brazil, CTX-M has been reported in several states; CTX-M-2, CTX-M-8, and CTX-M-9 subtypes are the most prevalent in human samples. In animal production species such as poultry, swine, cattle, and horses, the prevalent enzymes are CTX-M-2, CTX-M-8, and CTX-M-15 . Unfortunately, there are no nationwide surveillance programs on bacterial resistance and its mechanisms, making it difficult to estimate the proportion of ESBL producers .
2.4.4 The AmpC-type β-lactamases
Another enzyme group of the β-lactamases type is AmpC. They are relevant enzymes produced constitutively or induced by chromosomal or plasmidial genes expressed by members of Enterobacterales and other Gram-negative bacteria. This class of β-lactamases belongs to the functional groups 1 and C of the Bush and Ambler’s classification, respectively . They are often overlooked because they are not within groups 2b or 2b, as CTX-M, TEM, and SHV. AmpC producers hydrolyze almost all β-lactam antibiotics, including cephalosporins, cephamycins, and penicillins, solely or associated with Β -lactamase inhibitors, limiting therapeutic options to treat infections caused by these resistant bacteria.
Of major concern is the hyperproduction of this enzyme in
2.4.5 The carbapenemases
The carbapenem resistance is related to the production of β-lactamases with versatile hydrolytic capacities. Currently, the most important type of class A carbapenemases are KPC enzymes, whereas VIM, IMP, and (particularly) NDM in class B and OXA-48 (and related) in class D are the more relevant enzymes. Most carbapenemases are plasmid-mediated (with genes frequently located in integrons), favoring its dissemination . Since carbapenemase-producing Gram-negative bacteria generally also contain gene coding for other beta-lactam resistance mechanisms, it is not uncommon for organisms to exhibit complex beta-lactam resistance phenotypes. Besides, these organisms often contain other genes that confer resistance to quinolones, aminoglycosides, tetracyclines, sulfonamides, and other families of antimicrobial agents that cause multidrug resistance (AMR) or even pan-resistance. The emergence of new variants and the prevalence of β-lactamases in isolates of community, environmental, and animal origin has demonstrated the complexity of establishing the origin of resistance.
2.4.6 Challenges in detecting the prevalence of β-lactamases
The incidence of large-scale beta-lactamase-producing organisms’ spectrum is difficult to determine. There are significant differences between the detection and interpretation methods used by countries and health institutions throughout the study . Considerable phenotypic confirmatory tests for ESBL (2be and 2b) producers have been described in the literature, and all methods utilize the characteristics of ESBL production inhibition by clavulanic acid.
The Clinical and Laboratory Standard Institute (CLSI) recommended test consists of an initial screening by disk diffusion or by the broth dilution method with ceftazidime, ceftriaxone, cefotaxime, cefpodoxime, and aztreonam followed by a phenotypic confirmatory test with cefotaxime and ceftazidime in the presence and absence of clavulanate . The European Antimicrobial Susceptibility Testing Committee (EUCAST)  also recommends these tests, but both documents preconize different disk concentrations, and there are also differences in susceptibility zone sizes for consideration of resistance patterns. These factors lead to difficult interlaboratory standardization and consequently to the correct definition of local, regional, and national epidemiological data.
Specifically, regarding AmpC, the Clinical and Laboratory Standards Institute (CLSI) offers no standard test to detect AmpC producer isolates. There are few antimicrobial agents safely effective against these isolates, and many of them are not available or even not approved for animal use. Although different detection methods are available, the lack of international standardization limits the reporting of AmpC by clinical laboratories, which may underestimate this important mechanism of antimicrobial resistance .
Different environments related to animal production and clinical care can act as a source of the emergence of resistance genes. Studies developed over two decades show that there are relevant peculiarities that must be considered in the detection and understanding of emerging resistance in animal environments to achieve a systemic and practical approach to control antimicrobial resistance worldwide. This chapter discussed some current challenges, the importance of the poultry production environment in the significant emergence of colistin resistance, the development of a universal primer that made it possible to detect a variant of the
We are grateful to the Foundation for Research Support of the State of Rio de Janeiro (FAPERJ), National Council for Scientific and Technological Development (CNPq), and Coordination for the Improvement of Higher Education Personnel (CAPES) for grants that supported this work. We express our sincere thanks to Dr. Catherine Logue, Dr. Lisa Nolan, and Dr. Nicolle Barbieri from Poultry Disease Research Center, University of Georgia, United States of America, for providing us with technical conditions for the development of part of the reported work. We also thank Dr. Helena Maria Neto Ferreira from the Department of Pharmacy, University of Porto, Portugal, for helping us perform the initial beta-lactamases experiments. Our deep gratitude goes to all the former and present staff of the Veterinary Bacteriology Laboratory at Federal Rural University of Rio de Janeiro.