Abstract

Antimicrobial resistance is a major public health crisis. Antimicrobial stewardship programs and guidelines for judicious use of antimicrobials have become commonplace in human and small animal medicine, but the practice of exotic animal medicine lacks such guidance. This master class will review the current situation of the antimicrobial resistance crisis, mechanisms for the spread of resistance, nuances of diagnosing and treating bacterial infections, and review general guidance for antimicrobial stewardship.

Introduction

Antimicrobial resistance is a major public health crisis. Bacteria, fungi, and parasites are all rapidly acquiring the ability to defeat the drugs designed to destroy them. In human health, at least an estimated 2,868,700 people are sickened by an antimicrobial-resistant infection … in the United States … each year. An estimated 35,900 of these people will die. Clostidioides difficile (formerly Clostridium difficile) infections related to antimicrobial use and resistance account for an additional 223,900 cases and 12,800 deaths annually.¹ Put those numbers together and we find that over 3 million people get infections related to overuse of antimicrobials, and nearly 50,000 of them will die in the United States each year.

The days of the miraculous cure with the first dose of antibiotics are gone. “Stop referring to a coming post-antibiotic era,” writes Robert Redfield, director of the United States Centers for Disease Control & Prevention, “it’s already here. You and I are living in a time when some miracle drugs no longer perform miracles and families are being ripped apart by a microscopic enemy. The time for action is now, and we can be part of the solution.”¹

Antimicrobial resistance is an ecological problem. With microbes inhabiting almost all corners of our world and our bodies, exposure to antimicrobials does not stop outside of a living animal. Drugs may leak into the environment at the manufacturing plant.² Antimicrobials administered to patients are eventually excreted in urine or feces, leading to direct environmental exposure or concentration in wastewater treatment facilities. Surface water run-off and effluent from water treatment plants allows the drugs to concentrate in natural bodies of water.³ Reservoirs of antimicrobial resistance genes have been found in environmental bacteria^4,5 and in organisms used as biopesticides⁶.

Environmental microbes may be exposed to antimicrobials, causing a selection pressure to express a variety of antimicrobial resistance genes. While microbes in the environment often naturally produce antimicrobial substances, these do not appear to have a substantial effect on overall expression of resistance genes. It is the additional selection pressure exerted by manufactured antimicrobials that results in higher resistance gene expression.⁷ Antimicrobials, bacteria, and resistance genes move between the compartments of animals (including humans), soil, and water, creating conditions in which resistant bacteria may emerge and proliferate.⁸

There are two ways that microbes acquire resistance genes: mutation and horizontal transfer. Horizontal transfer of resistance genes overwhelmingly occurs via plasmid, a “raft” of genetic material that encodes for a specific trait.⁹ This allows the spread of antimicrobial resistance genes across multiple species of bacteria. The sharing of resistance genes in the environment may flow both ways between human pathogens and non-pathogenic environmental bacteria.⁷

Evidence for the role of point sources (e.g., pharmaceutical plants, wastewater treatment facilities) in development and propagation of antimicrobial resistance in the environment is incomplete. A 2017 systematic review of the topic found that many studies had unclear assessment of bias, limited statistical analysis, and no measurement of effect (e.g., odds ratio, mean difference). The authors noted the difficulty of studying antimicrobial resistance in the environment due to the complex processes, field work, and numerous factors influencing relationships, and provided a framework for further studies of the issue.¹⁰

It is undeniable that overuse of antibiotics plays a major role in the increasing prevalence of antimicrobial resistance. Within a given species, antimicrobial use is considered to be the main driver of antimicrobial resistance.¹¹ The influence of these prescriptions on antimicrobial resistance in other species is less well studied, but some examples stand out.

The use of ceftiofur in chicken hatcheries was linked to increased resistance to ceftriaxone in Salmonella Heidelberg infection in humans (especially those associated with chicken meat). A temporary ban on using ceftiofur in hatcheries resulted in a reduction of resistance, and reintroduction of ceftiofur use resulted in increased resistance.¹² Avoparcin, a glycopeptide antibiotic previously used as a growth promoter in poultry and swine, has been shown to cause high levels of vancomycin-resistant Enterococcus faecium in the feces of treated animals. The resistance was concurrently seen in enterococcal infections in human medicine, where vancomycin is used in treatment of multidrug resistant infections.¹³

Bacteria-Host-Environment Relationship

It is essential to consider the symbiotic relationship between bacteria, the colonized host organism, and the environment as a complex ecosystem. Prokaryotic cells outnumber eukaryotic cells in most animals. Bacteria are essential players in many life functions, including digestion and immune function. To consider bacteria dichotomously as either “good” or “bad” is to vastly oversimply the relationship.¹⁴

Bacteria, like all forms of life, seek to survive and reproduce. In general, host organisms allow bacteria to do this and variably extract some benefit from the relationship. In some cases, the organism finds itself in a niche that causes disease to the host. Often the host environment plays a role in allowing this to occur. This scenario is often seen in exotic pets that have substandard husbandry; the host ecosystem is disrupted, and disease ensues. Unless the underlying problems are corrected, disease may recur after initially successful treatment.

Many factors that can influence the development of microbial disease in a patient. A partial list includes:

• Housing density
• Environmental temperature
• Environmental humidity
• Environmental lighting
• Air quality
• Nutrition
• Opportunities to engage in natural behaviors
• Housing substrate
• Perception of safety

The administration of antimicrobials may have substantial impact on a patient’s microbial ecosystem. Most systemic antimicrobials diffuse throughout the body in varying concentrations, exposing far more than the targeted microbes. The consequences of this may be rapid and life threatening, as seen in antibiotic-associated dysbiosis in hindgut fermenters or C. difficile infections in humans. However, more long-term effects may be seen: recent administration of antimicrobials in dogs increases the likelihood of carrying methicillin-resistant Staphylococcus pseudintermedius;¹⁵ multidrug resistance may be induced in E. coli present in the gastrointestinal tract of dogs^16,17. In addition, idiosyncratic drug reactions, hypersensitivity reactions, accidental overdose, stress from administration, and pain from injections may occur and have a negative impact on health status.

Diagnostic Considerations

Ockham’s razor— “The simplest explanation is the most likely explanation”—becomes a trap in exotic animal medicine. The sneezing rabbit must be infected with Pasteurella multocida. The alopecic guinea pig has follicular ovarian cysts. The dyspneic bird has bacterial pneumonia or air sacculitis. While these diagnoses may be commonly reported in the overall patient population, it is important to remember Hickam’s dictum when considering the individual animal: “[The patient] may have as many diseases as [it] damn well pleases.”

Every effort must be made to localize a disease and determine its nature and underlying causes. In exotic medicine, a thorough review of the husbandry is essential. Do not just ask, “What?” but also, “How?” and “When?” For example:

• What is the temperature on the cool side of the cage? How do you know? What type of thermometer is used? When was it purchased?
• What is the cage cleaning frequency? How is it performed? What cleaners are used and how are they applied?

Diagnostic testing should be applied in a logical manner. In general, CBC, serum chemistry, and radiography are considered baseline diagnostics and should be recommended in most cases to evaluate for evidence of infection versus other diseases prior to the commencement of therapy. Computed tomography (CT) has become increasingly useful as a first-line imaging technique, providing superior diagnostic information compared to traditional radiography, especially in small patients.

Cytology may be used to guide therapy pending results of culture or PCR. A simple Gram stain and Romanowsky stain (e.g., Diff-Quik) yield basic information that can inform drug selection and provide context for interpretation of culture/PCR results. In general, cytology is considered supportive of bacterial infection if bacteria are visualized within phagocytes (e.g., neutrophils, heterophils). Extracellular bacteria may be pathogens, contaminants, or commensals.

In confronting infectious diseases, PCR and microbial culture become especially important tools for guiding treatment. One must remember that the result is only as good as the sample. Submitting a sample from a contaminated surface may produce a spurious result. Inappropriate handling of a sample (e.g., temperature, transport medium) may lead to a false negative result.

In general, swabs may not be the best tools for acquiring culture specimens, as few of the colony forming units (CFUs) within a swab will actually be cultured. Air trapped between the cotton fibers may inhibit anaerobic growth. Necrotic and purulent debris are similarly suboptimal candidate samples for culture, as they may not contain viable organisms. A piece of infected tissue is generally the preferred sample for culture.¹⁸ For example, abscess capsules (or entire abscesses) are considered to be superior to pus in culturing certain organisms.¹⁹ Tissue culture samples may be submitted in a no-additive tube (e.g., white-top) with one or two drops of sterile saline added to prevent desiccation (if needed). If a tissue sample is submitted in a tube containing transport medium and a swab, it should be clearly indicated on the submission form that the sample being submitted is the tissue, not the swab. Not indicating the thing to be cultured runs the risk of the laboratory disregarding the presence of tissue and simply rubbing the accompanying swab on culture media.

Interpretation of culture results requires nuance and knowledge of the possible normal microbial flora at the site cultured. As mentioned above, cytology can provide an especially useful perspective. Isolation of multiple organisms from potentially contaminated sites may simply represent colonization rather than infection. Pure growth of a single organism is more consistent with the need for antimicrobial therapy. The degree of growth should also be considered, as abundant growth is generally considered more supportive of infection. However, not all bacteria exhibit the same rates of growth, and more rapidly growing bacteria may outcompete slowly growing bacteria for resources. This effect may be reduced by refrigeration of the sample after collection and prompt submission to the laboratory.¹⁸

Susceptibility results may be determined using methods including disk diffusion, broth dilution, or Epsilon test (E-test), ideally according to standards set by the Clinical and Laboratory Standards Institute (CLSI). Broth dilution and E-testing are generally preferred, as they provide a quantitative concentration of drug that will inhibit the isolate. This can then be used to guide drug dosing (see Treatment Considerations). Isolates reported as having a minimum inhibitory concentration (MIC) of less than (<) the lowest tested concentration are generally considered highly susceptible, as growth of the isolate is inhibited by even the lowest concentration of drug tested. Isolates that have an MIC above the lowest tested concentration but less than the MIC breakpoint of a resistant organism have likely undergone some of the initial steps towards developing resistance, and continued progress toward full resistance may be more likely for these isolates.¹⁸

Treatment Considerations

“Many veterinarians have the mentality that in exotics species … enrofloxacin is a panacea. The routine use of advanced, broad spectrum antibiotics implies a low level of skill on the part of the clinician. It is our duty to develop appropriate antimicrobial stewardship. Veterinarians should limit antibiotic use to those cases in which their use is warranted.”—Sean Perry and Mark Mitchell²⁰

Treatment with antimicrobials should be reserved only for cases in which there is evidence—not just suspicion—of a bacterial infection. Viral infection, immune-mediated disease, and non-infectious inflammation may all appear grossly similar to bacterial infection. The drug chosen should ideally be based on PCR or culture/sensitivity results. If therapy must be initiated prior to results of these tests, choose a relatively narrow-spectrum drug that will affect the suspected pathogen(s) and a route of administration that will have the least impact on the body’s microflora.²¹

Categorization of antimicrobials into tiers to guide selection have been proposed and designed, often around specific diseases.^22,23,24 Efforts to produce similar guidelines for exotic species have been few. The Zoological Medicine Service at the University of Georgia devised antimicrobial tiers for exotic patients based on retrospective evaluation of their cases (Table 1).²⁵ This should be considered but not as absolute, as infection agent prevalence and susceptibility patterns may differ dramatically based on region and caseload.

Table 1. University of Georgia Zoological Medicine Service antibiotic tiers²⁵

Drug selection should also consider the importance of antimicrobials to human medicine. The World Health Organization (WHO) maintains a list of drugs considered critical for human medicine, divided into the three major tiers—Important, Highly Important, and Critically Important—based on five criteria. The Critically Important category is further subdivided into High Priority and Highest Priority subdivisions (Table 2).²⁶

Table 2. WHO critically important antimicrobials for human medicine²⁶

The World Organization for Animal Health (OIE—Office International des Epizooties) has also categorized antimicrobials into three tiers: Veterinary Important, Veterinary Highly Important, and Veterinary Critically Important (Table 3). These categorizations were made with an emphasis on production animal medicine; companion animals do not appear to have been a major consideration.²⁷

Table 3. OIE categorization of veterinary important antimicrobials for food-producing animals²⁷

The OIE also notes that fluoroquinolones and third and fourth generation cephalosporins, along with colistin, are considered critically important for both human and animal health. Therefore, the OIE has issued the following guidelines for use of these drugs (underling added by the author; two additional guidelines are omitted as the author deems them not applicable to exotic animal practice):²⁷

• Not to be used as preventive treatment applied by feed or water in the absence of clinical signs in the animals to be treated
• Not to be used as a first line treatment unless justified; when used as a second line treatment, it should ideally be based on the results of bacteriological tests.

Some of the key take-home points from the above tables and lists:

• Enrofloxacin is not a first line drug
• Third generation cephalosporins (e.g., ceftazidime) are not first line drugs
• Enrofloxacin is not a first line drug
• Some acceptable first-line drugs include trimethoprim/sulfa combinations, doxycycline, first generation cephalosporins, and basic potentiated penicillins (ampicillin, amoxicillin/clavulanate)
• Enrofloxacin is not a first line drug
• Third generation cephalosporins (e.g., ceftazidime) are not first line drugs
• Enrofloxacin is not a first line drug

Whenever possible, drug dosing should be guided by published pharmacokinetic and pharmacodynamic studies rather than anecdotal use and perceived success. Without pharmacokinetic guidance, the practitioner has no real idea if the chosen therapy will even reach therapeutic concentrations at the site of infection. The most up-to-date editions of common exotic animal formularies should be used, and the cited source of prospective dosing regimens assessed.

Antimicrobials are generally grouped into two classifications based on their effect on bacteria: those that simply inhibit the growth of microbes (i.e., bacteriostatic), and those that actively kill microbes (i.e., bactericidal) (Table 4). This stratification is based on in vitro models and may not reflect the way the drug performs in the patient at the site of infection. Certain bacteriostatic drugs (e.g., macrolides) may reach higher concentrations in white blood cells and at sites of infection, resulting in a bactericidal effect.

Table 4. Antimicrobial drug classes separated into bacteriostatic and bactericidal groups¹⁸

* = accumulation in white blood cells may allow achievement of bactericidal concentrations at site of infection.

Antimicrobials may also be classified based on how their effect relates to the concentration of drug (Table 5). Antimicrobials whose efficacy is best determined by their maximum plasma concentration compared to the MIC are considered concentration dependent. Drugs whose effect is best determined by the amount of time plasma drug concentrations remain above the MIC are considered time dependent.

Table 5. Antimicrobial drug classes separated into concentration dependent and time dependent groups¹⁸

The goal of antimicrobial therapy is to achieve sufficient concentrations of the drug at the site of infection in order to kill the infecting organisms, while concurrently avoiding the patient side-effects of the drug.¹⁸ Culture and susceptibility results providing an MIC for each isolate may be used in the formulation of an antimicrobial therapeutic regimen, especially by comparing the MIC to the expected maximum plasma concentrations of the drug (C_max) or the area under the concentration-time curve (AUC₂₄). Guidelines exist for dosing some antimicrobial agents:^18,28

• Aminoglycosides: the C_max-to-MIC ratio should be between 8 and 10 (C_max:MIC = 8 to 10)
• Fluoroquinolones:
• The ratio of the C_max to the MIC should be between 8 and 10 (C_max:MIC = 8 to 10)
  or
• The ratio of the area under the concentration-time curve to the MIC should be 100 to 125 (AUC₂₄:MIC = 100 to 125)
• Beta-lactams (penicillins, cephalosporins, monobactams, carbapenems)
• The ratio of the C_max and the MIC should be 2 to 4
  and
• The plasma drug concentration should remain above the MIC for as long as possible, ideally at least 50% of the dosing interval

The route of administration may substantially influence the chances for development of resistance. Topical medication, whether applied directly in the form of creams and ointments, or inhaled via nebulization, allows specific delivery of mediation to the intended area with minimal impact on the host microbiome. Care should be used in selecting topical treatment of an area that the patient can lick, and medications that will not cause harm if ingested should be used.

The oral route is commonly employed but is not always the most appropriate. Interspecies differences in gastrointestinal anatomy and physiology result in highly variable bioavailability.²⁸ Orally administered antibiotics result in direct exposure of the gastrointestinal microflora to the chosen drug, which may cause side effects ranging from mild gastroenteritis to death (especially in hindgut fermenter species—rabbits, guinea pigs, chinchillas, hamsters, etc.).²⁹ Furthermore, the patient spitting out a portion of the drug may result in subtherapeutic dosing. Reptile patients being medicated by mouth should be maintained within their preferred optimal temperature range to ensure appropriate gastrointestinal function and metabolism.³⁰

The subcutaneous and intramuscular routes, while less commonly used, can circumvent the first-pass metabolic effect and deliver drug into the circulation. The volume to be administered is often small, and owners may be instructed to perform administration at home. Care should be taken not to administer injectable medications into the caudal half of the body of birds and reptiles as the renal and hepatic portal systems may shunt drug through the kidneys or liver prior to systemic distribution. While the significance of this shunting varies widely by the drug, in general the practice is advised to avoid inadvertent weakening of therapy.³¹

Antimicrobial Stewardship

Antimicrobial stewardship practices have gained popularity in human and small animal medicine but are lacking in exotic animal medicine. The American Veterinary Medical Association (AVMA) has defined antimicrobial stewardship as the actions veterinarians take individually and as a profession to preserve the effectiveness and availability of antimicrobial drugs through conscientious oversight and responsible medical decision-making while safeguarding animal, public, and environmental health.

A number of authors have outlined the core principles of antimicrobial stewardship in veterinary medicine. One group describes the principles as the 5 Rs:³²

• Responsibility
• Reduction
• Replacement
• Refinement
• Review

The AVMA has developed a similar but more detailed list of core principles and the components of each principle (Table 6). A checklist based on these principles is available to aid practitioners in developing and implementing antimicrobial stewardship policies in their practices.^a

Table 6. AVMA’s core principles of antimicrobial stewardship in veterinary medicine³³

The AVMA has also provided a list of principles for judicious use of antimicrobials (bold performed by the author for added emphasis):³⁴

• Antimicrobials should be used in animals only after careful review and consideration of the following points:
• A general or preliminary diagnosis has been made which indicates that antimicrobial therapy is appropriate.
• Culture and antimicrobial susceptibility testing should be performed when possible to guide the selection of antimicrobials.
• Regimens for antimicrobial prevention, control and treatment should be established using accepted scientific and clinical principles, such as microbiological and pharmacological tenets.
• Antimicrobial therapy for uncomplicated viral infections and non-septic inflammatory conditions should be avoided.
• Duration of therapy should be based on scientific and clinical evidence in order to obtain the desired health outcome while minimizing selection for antimicrobial resistance, as allowed by law.
• Antimicrobial therapy should be targeted to ill or at-risk animals when possible, and other management strategies should be used to prevent infection in healthy individuals.
• Accurate records of therapy and outcome should be maintained.
• Environmental contamination with antimicrobials must be avoided whenever possible.

A variety of interventions to improve antimicrobial prescribing practices have been undertaken in human medicine. A systematic review and meta-analysis of published intervention strategies found an overall 15% improvement in prescribing practices regardless of intervention type. A summary of interventions is provided (Table 7). Interventions were associated with a reduction in length of hospitalization by 1.12 days, and no increase in patient morbidity or mortality. However, one restrictive intervention—requiring prior approval for antibiotic use—showed unintended detrimental consequences: negative professional culture due to breakdown in trust, and delay in time to first antibiotic use (when such use was indicated). Further comparison of studies suggested that interventions featuring some element of enablement provided a greater effect, though this was only supported by some studies. Additionally, interventions that included feedback appeared to be more effective.³⁵

Table 7. Definition of behavior change techniques and intervention functions³⁵

Veterinarians in Australia were recently surveyed regarding barriers to and enablers of antimicrobial stewardship. A summary of the major findings is provided (Table 8). The investigators also identified three major reasons for dispensing antimicrobials without a formal consultation in equine and cattle practice:³⁶

1.  Pressure to “keep clients happy” and fear that clients would be driven to competing practices if not provided with the desired antimicrobials.

2.  Some clients felt they were able to diagnose common diseases and were unwilling to pay for a consultation for routine disease management.

3.  Long work hours and lack of time made practitioners inclined to dispense the desired antimicrobial rather than spend time convincing clients of the importance of formal consultation or the lack of need for antimicrobials.

Table 8. Summary of major barriers and enablers for implementing antimicrobial stewardship programs in veterinary practices in Australia³⁶

The investigators noted that requests for antimicrobials without formal consultation was not reported by companion animal practitioners. However, it is the author’s experience that exotic animal practitioners encounter these requests from time to time. Explanations for this behavior have included:

1.  Ownership of a large collection physically limiting the owner’s ability to present all affected patients

2.  Recurrence of clinical signs for which the patient has been presented in the past

3.  Development of clinical signs commonly encountered in the species and which frequently have an infectious component (e.g., dyspnea in rats, sneezing in rabbits, dermatitis in reptiles)

4.  The perceived value of the pet is less than the cost of consultation

5.  The veterinary practice caters almost exclusively towards individual pets, with no accommodations for herd medicine (e.g. no pricing for group/herd consultations, no equipment or predefined workflow for mobile consultations)

The author suggests that exotic animal practitioners accommodate herd medicine in their practices or develop a relationship with a mobile veterinary practice to which these clients may be referred.

Antibiograms—surveys of bacterial isolates and susceptibility patterns—are being more widely used in small animal medicine but are less commonly encountered in exotic pet practice. Published antibiograms vary from surveys of all cultures isolated from a class of patients (e.g., reptiles)^37,38 to summaries of results from specific lesions in one species^39,40. Pathogen prevalence and susceptibility may vary widely not just by species and site of infection but also geographic area and client base. Ideally each practice would develop its own antibiogram in order to better understand the susceptibility patterns being encountered.⁴¹ An antibiogram when combined with point-of-care cytology and Gram stain from a lesion can prove very useful in guiding drug selection pending culture and sensitivity results.

The field of exotic animal medicine currently suffers from a paucity of antimicrobial pharmacokinetic and pharmacodynamic data. Many drug dosing regimens employed in our field are based on anecdotal experience and perceived response. There is little doubt that this leads to inappropriate treatment and increased chances of resistance developing. The importance of continued determination of pharmacokinetic profiles in exotic species cannot be overemphasized.

Client Communication

Clients commonly present sick pets with the expectation of receiving antibiotics. This is especially true with exotic pets, which for years have been regarded by many veterinarians as having an inherent Baytril deficiency. It is essential to convey to pet owners the importance of proper antimicrobial use, as these drugs are not benign. The author frequently references the current worldwide public health crisis of antimicrobial resistance of the development of “superbugs,” and often finds that clients have encountered this information before and understand the need for judicious use of these drugs.

The costs of diagnostics and therapy should be communicated upfront, even if the client says that the cost does not matter. It is especially important not to make assumptions about what the client will or will not pay based on their appearance or the species of pet (the author regularly encounters clients who are willing to fund thorough diagnostic investigations of their exotic pets). The costs should be placed in the context of the patient’s health and prognosis, with the additional perspective of the potential impact on the client’s health if antimicrobial resistance were to emerge in the patient. An estimate is only part of the discussion; a discussion with the owner must be had in order to put the recommendations in context, with a focus on outcome and well-being for the pet.⁴²

Conclusion

Antimicrobial resistance is a towering problem, one that can only be solved through all medical professions collaborating to achieve a high level of expertise in the prescription of antimicrobials. As a clinician, the refusal to just send home two weeks of Baytril is empowering. It requires the practitioner to investigate, think, and develop a level of expertise that most exotic pet owners welcome. They want to know what is wrong with their pet and how to fix it; they don’t want you to shrug your shoulders and dispense medication that might help. We can do better, and there is a way forward.

Endnotes

a. AVMA Veterinary Checklist for Antimicrobial Stewardship: https://www/avma.org/system/files?file=2020-03/Veterinary-Checklist-Antimicrobial-Stewardship.pdf.

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