MINI REVIEW
published: 22 July 2020
doi: 10.3389/fmicb.2020.01416
Exploring the Animal Waste
Resistome: The Spread of
Antimicrobial Resistance Genes
Through the Use of Livestock
Manure
Alice Checcucci, Paolo Trevisi, Diana Luise, Monica Modesto, Sonia Blasioli,
Ilaria Braschi* and Paola Mattarelli
Department of Agricultural and Food Science, University of Bologna, Bologna, Italy
Edited by:
Margherita Sosio,
Naicons Srl, Italy
Reviewed by:
Bongkeun Song,
College of William & Mary,
United States
Michael Benedik,
Texas A&M University, United States
*Correspondence:
Ilaria Braschi
ilaria.braschi@unibo.it
Specialty section:
This article was submitted to
Microbiotechnology,
a section of the journal
Frontiers in Microbiology
Received: 19 February 2020
Accepted: 02 June 2020
Published: 22 July 2020
Citation:
Checcucci A, Trevisi P, Luise D,
Modesto M, Blasioli S, Braschi I and
Mattarelli P (2020) Exploring
the Animal Waste Resistome:
The Spread of Antimicrobial
Resistance Genes Through the Use
of Livestock Manure.
Front. Microbiol. 11:1416.
doi: 10.3389/fmicb.2020.01416
Antibiotic resistance is a public health problem of growing concern. Animal manure
application to soil is considered to be a main cause of the propagation and dissemination
of antibiotic residues, antibiotic-resistant bacteria (ARB), and antibiotic resistance genes
(ARGs) in the soil-water system. In recent decades, studies on the impact of antibioticcontaminated manure on soil microbiomes have increased exponentially, in particular
for taxonomical diversity and ARGs’ diffusion. Antibiotic resistance genes are often
located on mobile genetic elements (MGEs). Horizontal transfer of MGEs toward a
broad range of bacteria (pathogens and human commensals included) has been
identified as the main cause for their persistence and dissemination. Chemical and
bio-sanitizing treatments reduce the antibiotic load and ARB. Nevertheless, effects of
these treatments on the persistence of resistance genes must be carefully considered.
This review analyzed the most recent research on antibiotic and ARG environmental
dissemination conveyed by livestock waste. Strategies to control ARG dissemination
and antibiotic persistence were reviewed with the aim to identify methods for monitoring
DNA transferability and environmental conditions promoting such diffusion.
Keywords: veterinary antibiotics, animal manure, antibiotic resistance genes, crop soils, antimicrobial resistance
INTRODUCTION
In recent decades, the overuse and misuse of antibiotics in human and veterinary medicine has
become a serious public health issue (World Health Organization, 2014; Aidara-Kane et al., 2018).
The increased number of resistant pathogens and commensal bacteria has been associated with the
environmental spread of antibiotics and the propagation of antimicrobial resistant genes (ARGs;
Levy, 1998; Witte, 1998; He et al., 2020). Furthermore, the environmental diffusion of antibiotics
may lead to the change (Han et al., 2018) and loss (Chen et al., 2019) of microbial community
diversity in soil (Kemper, 2008).
Antibiotics are used worldwide in livestock production, thus increasing the risk of antimicrobial
resistance (AMR) spread. When administered for prophylactic treatments, antibiotics can directly
increase selective pressure, thus favoring the generation of antibiotic-resistant bacteria (ARB;
Frontiers in Microbiology | www.frontiersin.org
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Antibiotic Resistance Genes in Animal Manure
site for exogenous gene cassettes, which can be acquired and
converted in functional and expressed genes (Mazel, 2006).
Integrons can move horizontally in bacterial populations by
frequent integration in plasmids or in transposons (RoweMagnus and Mazel, 2002). According to their aminoacidic
sequence, integrases are divided into several classes. Classes
1, 2, and 3 (Inti1, Inti2, Inti3) were the first to be identified
as associated with MGEs, while class 4 (Inti4) was associated
with chromosomal integration (Deng et al., 2015). Among
elements which facilitate DNA transfer, class 1 integron (int1)
is the most frequently identified as responsible for spreading
antibiotic resistance determinants amongst commensals and
pathogens of humans and domesticated animals. Moreover, int1
cassette was found in different environments, such as fresh
water, sediments, and sludge (Collis and Hall, 1995; Hall and
Collis, 1998; Nardelli et al., 2012; Borruso et al., 2016), where it
showed significantly positive correlations with the relative ARG
abundance (Zhao et al., 2019).
Antibiotic residues, once entered into soil through manure
application, can enhance persistence and HGT of ARGs (Binh
et al., 2007; Zhao et al., 2019) through plasmids and integrons
(Gotz and Smalla, 1997; Smalla et al., 2000; Sengeløv et al.,
2003), promoting the spread of ARB in the environment and
affecting the microbial community composition (Chen et al.,
2019). Although manure-derived bacteria cannot always adapt to
new environments, the antimicrobials can favor the enrichment
of specific bacterial taxa in soils (through positive selection) and
suppress others (Ding et al., 2014). In addition, the concentration
of antibiotics in manure, usually at a sub-inhibitory level,
can affect the interactions among strains and impact on gene
expression and regulation (Gillings, 2013; Jechalke et al., 2014;
Brüssow, 2015).
When manure is used as a fertilizer for crop production, both
the increased ARB load and the antibiotic residues contained
within may have negative effects on plant development and
food product quality (Verraes et al., 2013; Mirza et al., 2020;
Muhammad et al., 2020). In addition, antibiotic residues can
persist and accumulate in the environment (Jechalke et al., 2014)
by adsorption on soil solid phases (Du and Liu, 2012).
Pruden et al., 2013; Troiano et al., 2018; Blau et al., 2019).
For these reasons, improved livestock and waste management
strategies (i.e., diets, proximity between animals, waste treatment,
use of additives, and operating conditions) should be adopted to
limit the use of antibiotics in animal husbandry.
Antimicrobial resistant genes can enter and persist in
ecosystem through multiple pathways. They spread across soil
(Binh et al., 2007), crops (Su et al., 2015), and gut microbial
communities of wild and livestock animals and of humans
(Yadav and Kapley, 2019). Antimicrobial resistant genes’ spread
occurs through horizontal gene transfer (HGT) of mobile
genetic elements (MGEs), as phages, plasmids (Fondi and Fani,
2010), transposons, or integron gene cassettes (Figure 1). The
acquisition of AMR by bacteria may be due to spontaneous
mutations (Woodford and Ellington, 2007) or, more frequently,
by gaining specific ARGs from other bacteria through HGT.
High density of microbial cells in the presence of antimicrobial
compounds and nutrients, as observable in manure (Blau
et al., 2018), triggers HGT events among bacteria, thus
conferring selective advantage to the hosts (Thomas and Nielsen,
2005). Mutations are essential for the continuous evolution
of ARGs, producing hundreds of variants which are hardly
identifiable and increasingly dangerous for the environment
(Woodford and Ellington, 2007).
In this review, the effect of antibiotic occurrence in animal
manure on the dissemination of AMR and ARGs in agricultural
fields are discussed in a critical way. The main strategies
to mitigate ARGs’ dissemination and to control antibiotic
persistence are also reported. Methods monitoring changes in
microbial communities and transferability and environmental
diffusion of DNA were addressed as well.
THE DISSEMINATION MECHANISMS OF
ENVIRONMENTAL RESISTOME
The “resistome,” i.e., the total amount of resistance genes
associated with an ecosystem (Finley et al., 2013), is generally
mediated by conjugative plasmids. The resistome confers
resistance of antibiotics and heavy metals to microorganisms,
thus enhancing their survival in hostile environments (Bennett,
2008; Song et al., 2017). IncP-1, a common environmental
plasmid group, is largely known for its efficient conjugative
transferability potential and stable replication in a wide range of
Gram-negative bacteria (Heuer et al., 2012). Conversely, plasmids
IncF (Villa et al., 2010), IncI (Blau et al., 2018), and IncQ
(Rawlings and Tietze, 2001) show a narrower host range. These
plasmids are assumed to be important for the dissemination of
ARG in Escherichia coli and other Enterobacteriaceae (Johnson
and Nolan, 2009; Suzuki et al., 2010; Heuer et al., 2012; Van
Houdt et al., 2013). As evidence, the study of the mechanisms
of diffusion of these plasmids (Teuber, 2001) and compatibility
evolution with broad or narrow host ranges should allow for ARG
diffusion prediction.
Integrons play a key role in the fast spread of resistance
determinants toward antibiotics. They are genetic elements
composed of a gene encoding an integrase and an integration
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ARGs IN THE ENVIRONMENT
The majority of antibiotics are naturally produced by microbes
as a self-protection mechanism against other microorganisms.
ARGs have been always present in the environment. ARGs
encoding resistance for a large set of antibiotics have been
found in 30,000-year-old Beringian permafrost and in bacteria
isolated from prehistoric caves (D’Costa et al., 2011; Berglund,
2015). When present in the environment at a sub-inhibitory
concentration, antibiotics frequently play a role in transcription
regulation and in the exchange of signals among cells (i.e.,
quorum sensing mechanism and conjugation) (Reygaert, 2018).
Antibiotic resistance consists of a large variety of mechanisms,
such as inactivation by specific cleaving enzymes, exclusion
from cells via efflux pumps, interference with protein synthesis,
limitation of drug uptake, and modification of antibiotic target.
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Antibiotic Resistance Genes in Animal Manure
FIGURE 1 | Spread of ARGs and ARB in farm-related environments. ARB, antimicrobial resistant bacteria; ARG, antibiotic resistance gene; AMS, antimicrobial
sensitive; HGT, horizontal gene transfer.
ARGs’ maintenance depends on their considerably low fitness
cost. In fact, once a specific ARG has been acquired by a bacterial
cell, it must evolve to produce more benefits than costs in
order for multiple copies of the same gene to be kept and to
maintain the expression control of genes in MGEs (BengtssonPalme et al., 2017). Furthermore, as already mentioned, nutrient
rich environments can positively influence the ARGs’ spread and
facilitate cell–cell interactions (Manaia et al., 2018) (Figure 1).
Resistance acquired through MGEs and plasmids is responsible
for the last two mechanisms in which the resistance extent
depends on bacterial species and acquired ARGs (Reygaert, 2018;
Kraemer et al., 2019). The antibiotic selective pressure driving
the acquired resistance determines accurate ARGs’ specialization,
thus making the environment a potential reservoir.
Anthropogenic activities affect antibiotic and ARGs’ spread
with somewhat predictable effects (Vikesland et al., 2017).
In livestock farming, the use of antibiotics varies depending
on the farming type and location, having a considerable
effect on ARGs’ concentration. Among the ARGs most
frequently detected in livestock production, those related to
sulfonamide resistance (sul) (Table 1) are particularly diffused
in aquatic systems (Chen et al., 2015; Makowska et al.,
2016). In surface and fresh waters, sul genes were found in
IncQ plasmid group (Sköld, 2001; Berglund, 2015). Similarly,
diaminopyrimidine genes (dfr), which confer resistance to
antimicrobial trimethoprim, have been identified in both
class 1 and class 2 integrons (Deng et al., 2015). Similarly,
quinolone resistance qnr genes have been frequently associated
with different plasmid groups. Both dfr and qnr genes easily
disseminate in the environment, being found in surface
waters (Berglund, 2015), wastewaters, and related irrigated soils
(Dalkmann et al., 2012). Tetracycline resistance genes (tet)
are widely diffused in different pathogenic and environmental
bacteria (Roberts, 2005) and are often detected in sewage
treatment plants, soil, and surface and ground water (CheeSanford et al., 2001; Berglund, 2015). In the same environments,
erm genes, which are the most widespread macrolides resistance
gene, were isolated.
Essentially, ARGs’ diffusion is associated with a stress
response activated by exposure to antibiotics as well as with
the mobilization of several integrative and conjugative elements.
Frontiers in Microbiology | www.frontiersin.org
THE USE OF VETERINARY ANTIBIOTICS
In veterinary medicine, antimicrobials can be used as
therapeutics and/or growth promoters. Antibiotic growth
promoters (AGPs) are antimicrobial substances administered
at a sub-therapeutic dose for a prolonged time with the main
purpose being to improve the feed conversion rate, especially in
young animals, raising the economical profit of farmers. Since
2006, both the European Union and Australia have forbidden
the use of AGPs. Nevertheless, in most other countries the use of
AGPs is still permitted (Guardabassi et al., 2009).
Among breeding farms, poultry and pig livestock have
received the majority of antibiotics for therapeutic or
prophylactic use (Ungemach, 2000; Kim et al., 2011), resulting in
an abundance of ARGs greater than three orders of magnitude
compared to other farming systems, such as fish and cattle
farming. Several studies confirmed swine farms as a hot-spot for
ARB and ARGs (Rosen, 1995; Cromwell, 2002; de Greeff et al.,
2019; Petrin et al., 2019). Recently, the scientific community
investigated prevalence, abundance, and possible mobilization of
ARGs in pig farms and surrounding environments (Hölzel et al.,
2010; Marti Serrano, 2014; Petrin et al., 2019; Van den Meersche
et al., 2019; Wu et al., 2019).
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Antibiotic family
Most used
Animal
Farming
Use
Contrasted bacteria and recognized
main targets
Resistance mechanism
Main ARGs
Macrolides
Tylosin
Cattle
Gastrointestinal and
respiratory infections
Gram-positive bacteria.
Interference with protein synthesis
(sequestration of mRNA
ribosome-binding site)
erm, msr, mef
genes
Erytromycin
Pig
Clarithromycin
Poultry
Sulfamethazine
Cattle
Urinary tract infections
Gram-positive and Gram-negative bacteria.
Main target: Enterobacteriaceae,
Pasteurellaceae
Interference with folic acid synthesis
competing for the enzyme DHPS
sulI, sulII genes
Pig
Respiratory infections
Gram-positive and Gram-negative bacteria
Interference with efflux pump systems
tet genes
Intestinal infections
Gram-positive and Gram-negative bacteria,
including mycobacteria, and anaerobes
Mutations in the genes encoding
quinolone target DNA gyrase and
topoisomerase IV, interference with
efflux pump systems
qnr genes
Pig
Respiratory diseases
Gram-positive and Gram-negative bacteria
Interference with cell wall synthesis and
permeability, inactivation through
β-Lactamase enzyme
bla, amp, pen
genes,
Cattle
Necrotic enteritis
Intestinal infections
Gram-positive, and Gram-negative
bacteria, if aerobic
Inhibition of protein synthesis
(rhibosome interference)
aac, aad, aad aph
genes
Pig
Respiratory disease, foot
rot
Broad spectrum. Main target:
Photobacterium, Salmonella, E. coli
Enzymatic modification of antibiotic
molecules
cat, pp-flo, flo
genes
Horse
Post-weaning scours
Gram-positive and many Gram-negative
bacteria. Main target: Enterobacteriaceae
Interference with folic acid synthesis by
binding the enzyme DHFR
dfr genes
Sulfonamides
Checcucci et al.
Frontiers in Microbiology | www.frontiersin.org
TABLE 1 | The most commonly used antibiotics and the relative ARGs in livestock production (DHPS, dihydropteroate synthase; DHPR, dihydropyridine-resistant).
Main target: Lawsonia intracellularis
Staphylococcus aureus
Poultry
Tetracyclines
Chlortetracycline
Cattle
Systemic and local
infections
Oxytetracyclines
Pig
Gastrointestinal and
respiratory infections
Poultry
Fluoroquinolones (Enrofloxacin,
Danofloxacin, Marbofloxacine)
Pig
β-lactams
Penicillins (Amoxycilline,
Ampicillines) Cephalosporins,
Carbapenems
4
Doxycycline
Quinolones
Cattle
Poultry
Dog
Cat
Streptomycin, Spectinomycin,
Neomycin, Aspramycin,
Gentamycin, Lincomycin
Phenicols
Chloramphenicol
Pig
Poultry
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Thiamphenicols (thiamphenicol,
florfenicol)
Diaminopyrimidines
Trimethoprim
Pig
(Continued)
Antibiotic Resistance Genes in Animal Manure
Aminoglycosides
Pig
Poultry
Valnemulin
Cattle
Tiamulin
Pig
Pleuromutilins
Poultry
Poultry
Lincomycin
Lincosamides
Pig
Bacitracin, Colistin
Polypeptides
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Table 1 summarizes the main antibiotic families and the
most used antimicrobics in livestock animals for therapeutic
use. Nowadays, more than 150 antimicrobial compounds in
livestock production are used. The residues inevitably end up in
the environment because of manure application on agricultural
lands (Baguer et al., 2000). In 2010, more than 63,000 tons
of antimicrobials were consumed by livestock across the globe.
The predicted growth of the world’s population allows for an
estimated increase in antibiotic consumption of up to 105,000
tons by 2030 (Tasho and Cho, 2016). For this reason, specific
action plans have been defined to reduce the use of antibiotics as
therapeutics for livestock in several countries (i.e., the European
One Health Action Plan against Antimicrobial Resistance, 2017;
the National Strategy to Combat Antibiotic-Resistant Bacteria,
proposed by the White House, 2014; the National Action Plan to
Contain Antimicrobial Resistance issued by the Chinese National
Health and Family Planning Commission, 2016–2020).
References: (Schwarz et al., 2001; Petinaki et al., 2008; Guardabassi et al., 2009; Abbas et al., 2011; Van Hoek et al., 2011; Li et al., 2013; Shang et al., 2013; Tasho and Cho, 2016; Deng et al., 2017; Aghapour et al.,
2019; https://www.msdvetmanual.com/pharmacology/antibacterial-agents).
vga, sal, lsa genes
Alteration/protection of the antibiotic
target site
Pasteurellaceae, Brachyspira, Micoplasma
Respiratory and Intestinal
infections
lnu, lin, erm genes
Alteration of the antibiotic target site
Gram positive bacteria, most anaerobic and
some mycolpasma species. Main target:
Staphylococcus aureus
Respiratory and Intestinal
infections
Main Gram positive target: Campylobacter
pmr,pho, mcr, kpn
genes
LPS modification, efflux pump systems
regulation
Gram-positive (Bacitracin) or Gram negative
(Colistin) bacteria. Main Gram negative
target: E. coli Salmonela spp.
Pseudomonas aeruginosa, Klebsiella
pneumoniae, or Acinetobacter.
Intestinal diseases
Main ARGs
Resistance mechanism
Use
Animal
Farming
Most used
Antibiotic family
TABLE 1 | Continued
Antibiotic Resistance Genes in Animal Manure
Contrasted bacteria and recognized
main targets
Checcucci et al.
MANURE TREATMENTS
Besides direct collection into aerobic or anaerobic lagoons,
animal manure can undergo drying and liquid-solid phase
separation. Manure solid phase, as well as whole manure if
shovellable, is traditionally composted to produce biofertilizer.
Currently, anaerobic digestion and biological treatments of
animal manure are often adopted on intensive animal farms
(Van Epps and Blaney, 2016).
Composting can substantially reduce the antibiotic load,
especially during the thermophilic phase (Zhang et al., 2019),
but recalcitrant antibiotics accumulate in compost products
and in amended soil (Bohrer et al., 2019; Zang et al., 2019).
A general ARG abatement (0.7–2.0 log decrease) is obtained
through thermophilic composting of swine, cattle, and poultry
manure, depending on manure type and operational conditions
(He et al., 2020).
Biological treatments of animal manure and wastewater,
which are adopted to reduce the environmental input of nitrates,
slightly decreases the levels of antibiotic residues and pathogenic
bacteria (Van den Meersche et al., 2019). Antimicrobial resistant
gene reduction of 0.1–3.3 log is observed in swine manure after
treatment (He et al., 2020).
Anerobic digestion (AD) is adopted to stabilize manure with
a final production of methane (Fubin et al., 2016, 2017). A 0.3–
52 log decrease of ARGs was observed in digestate from swine
wastewater (He et al., 2020). Interestingly, the higher the content
of volatile solids in manure and the mixing rate, the higher
the ARGs number in the digestate (Turker et al., 2018). The
combined pasteurization and AD of swine manure reduced sole
archaeal communities, whereas simple AD affected bacteria and
archea (Fubin et al., 2020). Manure pretreatment with bacterial
strains is effective in degrading antibiotics (Liu et al., 2019) and
enhancing biogas production, but the overall effect on ARB and
ARGs was not addressed.
Constructed wetlands are vegetated aquatic systems that can
be adopted for the treatment of wastewater and agricultural
drainage water (Lavrnic et al., 2018). Their ability to reduce
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Antibiotic Resistance Genes in Animal Manure
environments. Recently, plasmids from municipal sewage sludge
and recipient bacteria were analyzed for their transferability
by exogenous isolation (Blau et al., 2018; Wolters et al.,
2018). Referring to pig manure samples, four IncQ-like
plasmids were isolated in recipient strains: Pseudomonas putida
UWC1, Acinetobacter sp., Ralstonia eutropha, Agrobacterium
tumefaciens, and E. coli. The plasmid transferability in E. coli
strains was not efficient, underlying a broad but highly specific
host range (Smalla et al., 2000).
Recently, simplified mathematical models have been applied
to predict and quantify ARGs’ spread in livestock animal
gut microbiomes (Andersen et al., 2020) and in agricultural
waste (Baker et al., 2016). In such environments, the variables
involved in the ARGs’ spread are countless and depend on a
wide range of intrinsic and extrinsic factors, such as genetic
mechanisms of ARB replication, HGT dynamics, environmental
and stressor conditions, and microbiota composition. Therefore,
future research should focus on the improvement of predictive
models of ARGs’ dissemination mechanism, exploitable for
targeted operations in livestock waste management.
ARGs in swine wastewater resulted in a 0.18–3 log decrease
(He et al., 2020).
Oxidizing post-treatments, as ozonation or Fenton conditions,
can be used on animal or treated wastewaters to degrade
antibiotics and bacteria thanks to the activity of reactive oxygen
species (Balcıoğlu and Ötker, 2003; Ikehata et al., 2006; Uslu and
Balcıoğlu, 2009). Among advanced oxidation processes, highly
costly ionizing radiations are known for their ability to destroy
microbial DNA. Therefore, affordable combinations of ionizing
radiation and oxidation allows for the degradation of antibiotics
and ARGs in organic matrices, although with a high biological
and environmental risk (Chu et al., 2019, 2020).
DIFFERENT APPROACHES TO
RESISTOME PROFILING STUDY
Even though AMRs introduced in the environment with animal
manure have been largely explored (Dolliver et al., 2008; Selvam
et al., 2012b), contradictory information exists regarding the fate
of ARGs (Selvam et al., 2012a; Wang et al., 2015; Xie et al., 2016).
The growing need for the control of ARGs’ spread prompted the
scientific community to set up and to validate refined molecular
methods for the study of ARGs’ dissemination dynamics among
environmental microbial communities.
Both 16S rRNA amplicon and untargeted sequencing can be
considered exhaustive methods for the exploration of microbial
community structure in manure-fertilized soil and farm waste.
Several studies on resistome diffusion in wastewater treatment
plants (Yadav and Kapley, 2019), sewage sludge composting units
(Su et al., 2015), and urban sewage support the metagenomic
approach (Hendriksen et al., 2019) in monitoring ARGs’ level
during treatments and seasonal changes. A recent work (Han
et al., 2018) showed that the shift in soil bacterial communities
caused by manure application leads to changes in the soil
bacteria resistome.
Recently, studies on the detection of genetic markers
associated with AMR (transposases and class 1 integronintegrase genes) and ARGs have been markedly increasing.
The quantification of ARGs in soils amended with livestock
and swine manure (Brooks et al., 2014; Tao et al., 2014) was
performed with high-throughput qPCR assay (Rocha et al., 2018;
Blau et al., 2019). In a recent study, both intracellular and
extracellular DNA containing ARGs were quantified in sludge
at about 1010 and 1012 copies per gram, respectively (Dong
et al., 2019). Here, the intracellular ARGs were assessed through
conjugation with cell-cell contact, whereas the extracellular ARGs
were assessed through natural transformation. Several works on
different manure types focused on the quantification of targeted
genes intI1 and intI2 for class 1 and 2 integron-integrase genes
and korB gene, specific for IncP-1 plasmids, together with ARGs
(Hu et al., 2016; Blau et al., 2018, 2019).
As already reported, plasmid-mediated ARGs’ diffusion is
frequently used, especially for the role of plasmids in the
rapid bacterial adaptation and fitness improvement (Smalla
et al., 2000). Exogenous plasmid isolation techniques (Bale
et al., 1988) clarified how plasmids diffuse in different
Frontiers in Microbiology | www.frontiersin.org
CONCLUSION
Although a decrease in the use of antibiotics in livestock
production is highly recommended, antibiotics’ overuse remains
an important issue to solve. The uncontrolled spread of ARB
and ARGs in the environment due to soil manuring is of serious
concern. Many studies highlight ARGs’ presence in microbial
communities of livestock manure and manured agricultural
fields, despite the improved livestock and waste management
strategies to contain in-farm ARGs’ spread. In the last thirty
years, knowledge on pathways of ARGs’ diffusion from animal
waste to the environment was enriched by multidisciplinary
research approaches.
In light of the current knowledge, the study of the dynamics
of AMR and ARGs’ spread in manure and environments
surrounding livestock farms should combine molecular
and functional genetics strategies with prediction models
of the diffusion of MGEs (integrons and plasmids) and
metagenomic data.
AUTHOR CONTRIBUTIONS
AC: original draft preparation, figure and table conceptualization,
review, and editing. PT, MM, and SB: review. DL: original draft
preparation and table preparation. IB and PM: original draft
preparation and review. All authors contributed to critically
revising the manuscript and gave final approval for publication.
FUNDING
This research was supported by Programma di Sviluppo Rurale
2014-2020 Regione Lombardia (Project REFLUA: Swine manure
and environment).
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Frontiers in Microbiology | www.frontiersin.org
Conflict of Interest: The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could be construed as a
potential conflict of interest.
Copyright © 2020 Checcucci, Trevisi, Luise, Modesto, Blasioli, Braschi and Mattarelli.
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