Intestinal health and antimicrobial resistance (AMR) in pigs

This article exposes the need of a good “intestinal health” for a successful enterprise, and explores the issue of antimicrobial resistance (AMR) in pigs. The  latest concepts in AMR are conveyed in an accessible and interesting way.  This great review is a good starting point for further reading. 

Maintaining optimum structure and functions of the gastrointestinal tract (GIT) are necessary to achieve sustainable, economically profitable swine production. The GIT is responsible for the regulation of physiological homeostasis that gives the pig the ability of confronting infectious (e.g. enteric pathogens) and non-infectious (e.g. stressors) challenges.

What does the term “intestinal health” mean?

“Intestinal health” is commonly used term that attracts much attention. Although there is not a clear definition for it, Pluske et al. (2018) sates that:

Due to the deep changes in structure and function of the GIT associated with  the post-weaning transition, “intestinal health” of piglets is of major relevance in swine production.

The addition of antimicrobial compounds (comprising also antibiotics), in diet and/or water, kill microorganisms or inhibit their growth. This helps the piglets during this period of change, reducing the impact of post-weaning stress.

However, prohibitions/reductions in the use of certain antimicrobials are being imposed around the world. This has created the need to find alternative feeding strategies, and/or ways to include feed additives (e.g. Pew Charitable Trusts, 2017), putting a major emphasis on the concept of “healthy intestine”.

A deterioration of intestinal health in the pig during the post-weaning period, with the added presence of enteric pathogens, may result in a general impairment of health status of the animal.  Although some direct relationships can be established between performance/efficiency of pigs and a “healthy” GIT, many times such connections are more subtle and less obvious. This is especially relevant when enteric diseases appear without symptoms (subclinical diseases) (Pluske et al., 2018).

One of the key points to comprehend the idea of “intestinal health”, is the concept of microbiota. This is the ecological community of commensal, symbiotic, and pathogenic organisms inhabiting the GIT.

It should be mentioned that, as a synonym, microbiome describes the collective genomes of microorganism that live in the GIT, and the microorganisms themselves. Although the term “microbiome” technically refers to the population of bacteria, yeasts, fungi, viruses and protozoa in an ecosystem, it is usually applied only to the bacterial composition. However, the appropriate terminology to refer to the bacterial population is microbiome or bacterial microbiota. 

The scanning of bacterial microbiome in pigs and its complex interactions with the host and the external environment (e.g. through the diet and or through the surrounding environment) is a relatively new phenomenon. It is greatly driven by the growing availability and lower cost of new technologies of genomic sequencing.

Research on GIT’s microbiota of the pig has acquired a new and more urgent perspective due to the already mentioned bans/reductions in the use of certain antimicrobials in swine production. It is necessary to find ways to maintain a healthy GIT microbiota, when facing diverse stressing factors and infectious diseases (Brüssow, 2017).

However, some antimicrobial compounds are still permitted in some countries. This has created a parallel area of interest (and concern) facing the ability of numerous members of the GIT microbiota community to develop resistance to some antimicrobial compounds. Of particularly importance is the resistance against the CIAs or Critically Important Antibiotics, also used in human medicine.

What is antimicrobial resistance (AMR)? What does it entail?

AMR has been described as one of the greatest long-term threats to human health (Viens y Littmann, 2015), and to animal health and production.

In September 2016, and for the fourth time in its history, the United Nations meet to debate the issue of AMR, considered to affect worldwide health. They committed to take a “comprehensive and coordinated approach to address the causes of AMR in multiple sectors, especially in human and animal health, as well as in agriculture” (OMS, 2016).

The majority of AMR of (human) medical relevance are associated to the use of antimicrobials in the human population. However, AMR in zoonotic pathogens transmitted through food (e.g. Salmonella spp. and Campylobacter spp.), and in commensal bacteria of livestock, such as a E. coliI and Enterococcus spp., are currently under increasing surveillance around the world.

Why does this phenomenon occur?

It is, mainly, because many times, the same drug classes are used for disease treatment in humans and, at the same time, to control bacterial infections in livestock, especially pigs, chickens, and cattle.

However, it is also related to:

➢ The historical utilisation of some antimicrobial types (of common use in humans and animals) at subtherapeutic levels with the objective of promoting growth (antimicrobial growth promoters, or AGPs)

➢ The prophylactic (or metaphylactic) use of antimicrobials in feed and water, to prevent infections in cattle, pigs, and poultry.

Furthermore, the development of resistance to multiple antimicrobials, as observed in enterotoxigenic E. coli, could be the result of the unapproved use CIAs such as wide spectrum cephalosporins (Abraham et al., 2017. 2018).

Most resistances to CIAs are coded in mobile genetical elements, such as plasmids and integrative conjugate elements. This could determine the transference of such genes to occur easily, between bacterial species and between humans and animals.

Additionally, critically important AMR could link with a less relevant AMRs in a process known as co-selection (Mukerji et al., 2017). This phenomenon of co-selection is particularly important for livestock industries, where the transmission of genes of resistance to CIAs could combine with the resistance to first line antibiotics, and even to standard additives, such as zinc and copper, when used at supraphysiological levels.

➢ In this way, sometimes, the use of standard additives and first-line therapies could promote the proliferation of bacteria resistant to CIAs (Mukerji et al., 2017).

In this context, as already indicated by several authors (Wegener, 2003; Marshall and Levy, 2011; Mukerji et al., 2017), pigs constitute a potential reservoir of AMR. This AMR can be transferred to bacteria that are pathogenic to humans through:

• contaminated food
• direct contact with pigs
• and/or spreading of faeces in the environment.

What do we know about AMRs in pigs?

A recent study conducted in Denmark (Birkegård et al., 2017) quantified the relationship between lifelong exposure to antimicrobials and the expression of seven genes of AMR in Danish pig farms.

• The levels of AMR gene expression were quantified by means of qPCR in faecal samples obtained in 681 groups of pigs at slaughter.
• The results demonstrated a complex effect of lifelong exposure to antimicrobials on the expression level of AMR, unique for each individual gen.
• Various classes of antimicrobials showed negative and positive correlations with AMR genes.

In conclusion, it was discovered that the associations between factors were more complex than previously described. They depended on the specific AMR genes, as well as on the type of antimicrobial under study. Furthermore, the results indicated that the exposure to antimicrobials was not the only important factor determining the levels of expression of AMR genes.

Clearly, and in agreement with the literature, numerous influences, other than the exposure to antimicrobials, contribute to the development of AMR in pigs. Those factors include the incomplete understanding of selective pressure of resistance in livestock, the rate of transmission of AMR between animals and humans, as well as management practices helping or hindering the occurrence and persistence of AMRs in farms. Such factors have made difficult the evaluation and prediction of the risk of developing AMRs secondary to the use of antibiotics (Rosengren et al., 2010).

Figure 1. Association between the exposure to antimicrobials and AMR genes. The association map shows the effects of variables of antimicrobial exposure and other factors on the levels of expression of AMR genes. The figure summarises the effect of antimicrobial class and other variables included in the final regression analyses. Dotted blue line represent negative correlation, whilst the continuous red line represents positive correlation (Adapted from Birkegård, A.C., Halasa, T., Græsbøll, K., Clasen, J., Folkesson, A. and Toft, N. (2017). Association be tween selected antimicrobial resistance genes and antimicrobial exposure in Danish pig farms. Scientific Reports 7: 9683 (2017) doi:10.1038/ s4 1 598-017-10092-9)

Investgation on extended spectrum cephalosporins (ESC)

Part of the work carried on at the Murdoch University on AMRs addresses the use and occurrence of resistance against ESC in pig production in Australia.

Currently, there is an important incidence of AMRs against CIAs in bacteria known to cause severe disease in humans around the world. Among these antimicrobials, the WHO (2017) includes:

• extended spectrum cephalosporins (ESC)
• fluoroquinolones (FQ)
• carbapenems
• colistin

Ceftiofur and cefquinome are third generation ESCs (extended spectrum cephalosporins) registered all over the world for the treatment of respiratory infection in livestock animals, including pigs.

» It has already been possible to show that the excessive or inadequate use of ESCs has resulted in the appearance of a reservoir of ESC-resistant Enterobacteriaceae, including E. coli and Salmonella enterica, in livestock worldwide.

Figure 2. Genomic map of the plasmid pCTXM1-MU2 transported by strains of E. coli resistant to ESC. Such strains belonged to multiple lineages of E. coli in an Australian pig farm, after the voluntary elimination of ceftiofur. The positions of gene classes are indicated by colour; A- Expanded view of the genomic island consisting of aadA5 and dfrA17 in a transposase Tn3; B- Amplified view of the region ISEcp1-CTX-M-1. Adapted from Abraham, S., Kirkwood, R., Laird, T., Saputra, S., Mitchell, T., Singh, M., Linn, B., Abraham, R.J., Pang, S., Gordon, D.M., Trott, D.J. and O’Dea, M. (2018). Dissemination and persistence of extended-spectrum cephalosporin resistance encoding IncI1-blaCTXM-1 plasmid among Escherichia coli in pigs. The ISME Journal, 12: 2352-2362.

The first report on resistance to ESCs observed in E. coli and Salmonella serovars isolated from livestock was related to the use of ESCs in those animals. Consequently, there has been an increasing alarm in the public health sector due to the direct transfer of resistant bacteria, or the potential plasmid-mediated transfer of resistance, to humans (Allen & Poppe, 2002).

A more recent study at the Murdoch University (Abraham et al., 2018) investigated the ecology, epidemiology, and characteristics of the plasmid of E. coli resistent to ESC. The plasmid was present in healthy pigs during a 4-year period (2013-2016) after antibiotic withdrawal. During that period, high rates of transfer of E. coli ESC resistent were observed (Fig. 3).

Figure 3. Changes in the rate of transfer of ESC resistance gene during the 4 years following antibiotic withdrawal from the farm.

The study shows that ESC resistance can persist in the environment during a prolonged period, after eliminating direct selection pressure (antibiotic withdrawal). This resulted in the occurrence of ESC resistance in commensal E. coli as well as in atypical isolates of enteropathogenic E. coli, of potential relevance for human and animal health.

Conclusions

“Intestinal health” is of great relevance to pig research, especially regarding the exhaustive examinations of feeding strategies and techniques to help establishing an appropriate microbiome in the GIT. Ultimately, research looks at optimising productivity and efficiency of pigs in absence of certain antimicrobials.

The utilisation of antimicrobials in pig production used to be a tool for improving efficiency and controlling infections. However, its use has become very much compromised due to the increasing prevalence of AMR. This has resulted in prohibitions/reductions of their use.

However, some antimicrobials are recognised as a necessary intervention for veterinary treatments and the welfare of the animals. Hence, it is necessary to preserve its appropriate antimicrobial use.

This changing situation on the use of antimicrobials has become even more crucial since the resistance to CIAs began to appear in livestock systems all around the world. This underlines the need of continuous surveillance and genomic characterisation of CIAs resistant bacteria. The ultimate objective is to control the propagation of AMR in pigs and reduce the probability of an impact on human health.

In this sense, future research could investigate the overgrowth potential of resistant bacteria that may affect the standard microbiota. Similar to the development of dysbiosis after the administration of a short course of therapeutic antibiotics, resistant bacteria can overpopulate the GIT when prophylactic treatments are implemented. This will result in a reduction of “intestinal health” and a drop in production.

Therefore, nutritionists and feed manufacturers should not only know the current norms and regulations on the use of antimicrobials, and how they affect the formulation/supply of feed. They also should take part in projects and discussions, with the objective of confirming the efficacy of the implemented nutritional solutions. This will help them to better comprehend the mechanisms and interactions on which to build measures to improve animal health and immune status.

Furthermore, pork producers should have a better comprehension of socioeconomical factors. They are the ones making the decisions on the use of nutritionally optimized feeds (i.e. containing antimicrobials), as well as on whether to implement or not the professional advice given by the experts.

This article was published in nutriNews Spain, with the title “Salud intestinal y resistencia antimicrobiana”