Antimicrobial Resistance

Specific resistance to ionophores in coccidia species has been well documented. In poultry, restoration of sensitivity following the use of vaccines comprising drug‐sensitive strains of Eimeria has been demonstrated for the monensin and salinomycin. Therefore, a yearly rotation program of ionophores alternated in successive flocks with vaccination is recommended (Noack et al., 2019). Unfortunately, in most countries, anticoccidial drugs are purchased and used without veterinary prescription or supervision.

Because of the complexity and high degree of specificity of ionophore resistance, initially it appeared that ionophores did not contribute to the development of antimicrobial resistance to important human drugs, and did not affect fecal shedding of potential pathogens (e.g., E. coli O157:H7), and there was no need to eliminate them from use in animal feeds (Lefebvre et al., 2006; Russell and Houlihan, 2003). However, the risk of cross‐resistance to Critically Important Antimicrobials (CIA) for human medicine has not been fully investigated (Wong, 2019).

Resistance to narasin exists on mobile plasmids in Enterococcus faecium and the resistance mechanism is associated with a two‐gene operon encoding an ABC‐type transporter. The genes encoding the narasin resistance mechanism (NarAB) confer reduced susceptibility to narasin, salinomycin, and maduramicin, but not to monensin. Importantly, NarAB does not affect the susceptibility of E. faecium to antimicrobials that are used in human medicine. However, conjugation in the presence of certain ionophores increases the number of vancomycin‐resistant E. faecium (VRE), suggesting that narasin and other ionophores contribute to the persistence of VRE in poultry populations (Naemi et al., 2020). The European Food Safety Authority Panel on Additives and Products or Substances used in Animal Feed found that the use of monensin as a coccidiostat in chickens did not affect the colonization or shedding of Salmonella in the gastrointestinal tract and that there is no evidence to suggest that exposure of Gram‐positive bacteria to monensin results in the development of cross‐resistance to other antimicrobials used in human and veterinary medicine (European Food Safety Authority, 2019).

Pharmacokinetic Properties

Monensin is rapidly absorbed following oral administration. Ruminants appear to absorb only about 50% of a dose, while monogastric species appear to absorb almost the entire administered dose. Oral bioavailability is 30% in broiler chickens. Ionophores do not accumulate in large amounts in tissues, even when toxic doses are administered (Donoho, 1984; Puschner et al., 2016). Ionophores are rapidly and extensively metabolized by the liver into numerous metabolites, which are excreted in the bile and eliminated in the feces. Horses are not able to eliminate monensin from the blood as rapidly as cattle, which may explain why horses are the species most sensitive to monensin toxicity.

Toxicity and Adverse Effects

The relative toxicities of the ionophores from lowest to highest are salinomycin < lasalocid < narasin < monensin < maduramicin. Ionophore toxicity causes cellular electrolyte imbalances, elevating extracellular potassium and intracellular calcium, resulting in severe cellular damage and death. The dose necessary to cause toxicity is variable among species, with equine species being the most sensitive and turkeys being more sensitive than chickens (Table 18.1). Skeletal and cardiac muscle cells are generally the most severely affected but the specific tissues affected and resulting clinical signs vary from species to species. Skeletal muscle is primarily affected in dogs, ostriches, sheep, and turkeys. Cardiac muscles are affected in cattle, and both myocardium and skeletal muscles are damaged in horses and goats. Age‐related differences in ionophore sensitivity occur in poultry, with adult birds more sensitive to the toxic effects of ionophores than young birds. In dogs, puppies are more sensitive to the toxic effects of narasin than adult dogs. In cattle, calves 5–8 months of age are much more susceptible to the toxic effects of maduramicin exposure than calves aged 9–16 months. In cattle, clinical signs of ionophore toxicity include feed refusal and anorexia within 24 hours of exposure, diarrhea, dullness, weakness, ataxia, dyspnea, prostration, and death (Oruc et al., 2011; Puschner et al., 2016). Animals that recover from acute ionophore toxicosis may also suffer unexpected acute cardiac failure from damage to the myocardium, especially if exercised or stressed (Gy et al., 2020).

Ionophore toxicity occurs from dose errors in mixing with feed, accidental ingestion of treated feed by sensitive species, ingestion by ruminants of poultry litter from ionophore‐treated flocks, concurrent administration with a medication that potentiates toxicosis, or accidental feed mill contamination of presumably untreated feed. Ionophores are highly lipophilic and cross‐contaminated feed is a frequent cause of violative residues in eggs (Olejnik et al., 2014; Vandenberge et al., 2012). Heat stress and water deprivation exacerbate toxicity in chickens when lasalocid is administered at 1–2 times the recommended dose. Cattle and sheep have manifested signs of ionophore toxicity following ingestion of poultry litter from chicken flocks treated with maduramicin. Ionophore toxicosis is potentiated by medications that interfere with hepatic metabolism. Tiamulin administered concurrently with monensin causes severe ionophore toxicity in chickens and pigs.

Clinical Use

Chemistry

Nitroimidazoles are heterocyclic compounds based on a five‐membered nucleus similar to that of the nitrofurans (Figure 18.1).

Mechanism of Action

After entry into the cell, nitroimidazoles undergo reduction of the nitro group to produce a variety of unstable intermediates, including antibacterial products. Reduction occurs under anaerobic conditions but, unlike that of the nitrofurans, is not enzymatically controlled. The reduction system of aerobic bacteria is insufficiently low for reduction to occur, but there is the suggestion that these agents or their metabolites, produced by anaerobic bacteria, may have some activity against aerobic bacteria under anaerobic conditions. Nitroimidazoles cause extensive breakage of DNA strands and inhibition of the DNA repair enzyme DNAase.

Antimicrobial Activity

The antimicrobial activity of the clinically useful nitroimidazoles is similar. They are bactericidal to most Gram‐negative and many Gram‐positive anaerobic bacteria. They are highly active against Brachyspira (Serpulina) hyodysenteriae and a variety of protozoa (Tritrichomonas foetus, Giardia lamblia, Histomonas meleagridis). Campylobacter spp. are moderately susceptible. Heliocobacter pylori are commonly susceptible, but the susceptibility of animal‐origin Helicobacter species has not been sufficiently investigated to substantiate susceptibility.

Resistance

Resistance is rare among usually susceptible bacteria. Resistance involves reduced intracellular drug activation. Cross‐resistance between nitroimidazoles is complete. The plasmid pCD‐METRO confers metronidazole resistance in Clostridioides (Clostridium) difficile (Smits et al., 2022). Equine and canine isolates of Clostridioides difficile and Clostridium perfringens resistant to metronidazole have been described, so susceptibility testing is warranted in patients with clostridial diarrhea. Bacteroides fragilis resistant to metronidazole therapy has been reported in a horse with pleuropneumonia (Dechan, 1997).

Pharmacokinetic Properties

Metronidazole is rapidly and well absorbed after oral administration in horses and dogs, with an oral bioavailability of 75–85% in adult horses, 100% in neonatal foals and 59–100% in dogs. It may be administered per rectum to horses and is rapidly absorbed; however, bioavailability is only 30%. Metronidazole is lipophilic and widely distributed in tissues. It penetrates bone, abscesses, and the central nervous system. The volume of distribution is 0.7–1.7 l/kg in mares, 0.8 in foals, and 0.95 l/kg in dogs. It crosses the placenta and is distributed into milk in concentrations similar to those in plasma. Metronidazole is primarily metabolized in the liver by oxidation and conjugation. Both metabolites and unchanged drug are eliminated in urine and feces. The elimination half‐life is 3–4 hours in horses, 10 hours in foals, and eight hours in dogs.

Drug Interactions

No interference with the susceptibility of anaerobic bacteria has been reported in vitro when metronidazole is combined with a variety of other anaerobe‐active drugs, such as clindamycin, erythromycin, penicillin G, amoxicillin‐clavulanic acid, cefoxitin, and rifampin. Combined with a beta‐lactam and gentamicin or enrofloxacin, metronidazole is commonly used for therapy of bacterial pleuropneumonia in horses (Reuss and Giguère, 2015). The hepatic metabolism of metronidazole may be decreased when administered concurrently with cimetidine, possibly resulting in delayed elimination and increased serum concentrations of metronidazole. Phenobarbital may induce microsomal liver enzymes, increasing the metabolism of metronidazole and decreasing serum concentrations.

Toxicity and Adverse Effects

Nitroimidazoles have been shown to be carcinogenic in some laboratory animals and mutagenic in some in vitro assays. These drugs are banned for use in food animals in the United States, Canada, and the European Union, but metronidazole is still used in veterinary medicine in some countries and directly used in people, without reports of cancer‐associated morbidity. Oral use in horses is associated with anorexia, while vomiting, nausea, and nervousness are sometimes noted in dogs and cats. Human formulations are difficult to administer to cats and they tend to salivate profusely with oral administration.

Adverse effects of metronidazole in the dog and cat have been reported and include vomiting, hepatotoxicity, neutropenia, and neurological signs such as seizures, head tilt, falling, paresis, ataxia, vertical nystagmus, tremors, and rigidity (Caylor and Cassimatis, 2001; Olson et al., 2005; Tauro et al., 2018). Neurological toxicity from metronidazole has been reported in dogs receiving 60 mg/kg/day, but there are reports of toxicity at lower dosages. The mechanism of the neurotoxic effects of metronidazole has not been identified. Initially, the recommended therapy for metronidazole toxicosis was discontinuation of the drug and supportive therapy. With supportive therapy, the reported recovery time of dogs with neurological manifestations of metronidazole toxicosis is 1–2 weeks. The recovery time can be significantly shortened by the administration of diazepam, with an initial IV bolus of 0.5 mg/kg and then PO q8 hours for three days (Evans et al., 2003). Recovery time is markedly shorter for diazepam‐treated dogs (38.8 hours) compared to untreated dogs (11 days). While the mechanism of this effect is unknown, it is likely that diazepam competitively reverses the binding of metronidazole to the benzodiazepine site on the GABA receptor.

Metronidazole has significant effects on the intestinal microbiota of horses (Gomez et al., 2023). A study that utilized a herd of horses with cecal cannulas to investigate the effects of metronidazole on the equine cecal content and fecal microbiome and metabolome had to be abruptly ended due to adverse gastrointestinal effects. All horses in the study were PCR positive for Salmonella in their cecal content, but not in fecal samples, after metronidazole administration, and one horse died of typhlocolitis (Arnold et al., 2020). Daily fecal microbiota transplantation failed to prevent metronidazole‐induced dysbiosis of equine gut microbiota (Kinoshita et al., 2022). Oral administration of metronidazole decreased the number of aerobic bacteria and altered indigenous flora in the small bowel of cats (Johnston et al., 2000). The alteration in bacterial flora appeared to have an impact on nutrients, because serum albumin and cobalamin concentrations increased during administration and returned to preadministration concentrations after therapy was discontinued. Metronidazole also significantly changed the canine fecal microbiome, with effects that lasted four weeks (Pilla et al., 2020).

Administration and Dosage

Since antimicrobial effect is concentration dependent, twice‐daily therapy is now recommended over three times daily therapy. In most countries, there are no veterinary formulations of metronidazole, so human formulations are used. Metronidazole USP induces salivation and inappetence when administered orally to some cats. Products containing metronidazole benzoate are commercially available in some countries and the drug is available for compounding in the United States and Canada. Metronidazole benzoate is very well tolerated by cats. The recommended dose for treatment of Giardia in dogs and cats is 25 mg/kg every 12 hours for 5–7 days. High doses (25–50 mg/kg q12 hours) are sometimes used in the treatment of serious anaerobic infections (peritonitis, meningitis), but there is an increased risk of neurotoxicity. Doses of 10–25 mg/kg PO q12 hours are used in horses; withholding feed for two hours after administration may improve oral bioavailability. Metronidazole can be administered per rectum to horses at 20 mg/kg (Stein et al., 2018).

Clinical Applications

Metronidazole is used to treat anaerobic infections, especially pleuropneumonia and lung abscesses caused by penicillin‐resistant Bacteroides fragilis, and clostridial enterocolitis in horses (Uchida‐Fujii et al., 2023). It is typically administered orally along with a parenteral beta‐lactam and aminoglycoside or enrofloxacin to achieve Gram‐positive, Gram‐negative, and anaerobic coverage. Although rectal absorption is inferior to oral absorption, it is a viable option for treatment when oral administration is not feasible.

Metronidazole is the most prescribed antimicrobial for treatment of acute diarrhea in dogs due to suspected Giardia or Clostridium perfringens infections. While C. perfringens was thought to cause acute hemorrhagic diarrhea syndrome in dogs, in a clinical trial there was no benefit of antimicrobial treatment.

Because of concerns over the use of antimicrobials, which generate dysbiosis, in dogs with an already dysbiotic microbiome, alternative approaches with probiotics and synbiotics should be utilized (Pilla et al., 2020). In randomized controlled clinical trials, dietary management with an easily digestible diet with or without psyllium or probiotics were superior management strategies compared to metronidazole administration (Rudinsky et al., 2022; Shmalberg et al., 2019). Therefore, it is not consistent with antimicrobial stewardship to use metronidazole empirically in dogs with acute diarrhea.

In small animals, metronidazole is also used in the therapy of anaerobic infections, including bacterial stomatitis, osteomyelitis, hepatitis, pneumonia and lung abscessation, and peritonitis. It is also used in the treatment of giardiasis and other protozoal infections (Trichomonas, Balantidium coli). Metronidazole appears efficacious for the treatment of Giardia in dogs, but fenbendazole appears equally efficacious with fewer adverse effects (Ciuca et al., 2021).

Chemistry