Chloramphenicol and Derivatives, Macrolides, Lincosamides, and Miscellaneous Antimicrobials

36
Chloramphenicol and Derivatives, Macrolides, Lincosamides, and Miscellaneous Antimicrobials

Mark G. Papich

Drugs listed in this chapter do not warrant a separate chapter and are included together, as they comprise narrow uses in veterinary medicine. They have some features in common – for example they inhibit protein synthesis in bacteria (with macrolides, lincosamides, and chloramphenicol acting at a similar site), and have some similar pharmacokinetic features.

Some of these drugs are not as common or available as in previous years. Some older drugs have given way to newer derivatives and their discussion has been greatly abbreviated in this edition of the book. Older editions may be consulted for more detailed and historical information.

Chloramphenicol

Chemical Features

Chloramphenicol chemically is D-(-)-threo-1-p-nitrol-phenyl-2-dichloroacetamido 1,3-propanediol (Figure 36.1), has a pK_a of 5.5, and was first isolated from the soil organism Streptomyces venezuelae in 1947. The chloramphenicol used today is manufactured synthetically. Chloramphenicol is slightly soluble in water and freely soluble in propylene glycol and organic solvents. Chloramphenicol is a broad-spectrum antibiotic, inhibiting gram-positive and gram-negative organisms, aerobic and anaerobic bacteria, and many intracellular organisms. Chloramphenicol has three functional groups that largely determine its biological activity: the p-nitrophenol group, the dichloroacetyl group, and the alcoholic group at the third carbon of the propanediol chain (Yunis, 1988). Replacement of the p-NO₂ group by a methylsulfonyl (HC₃-SO₂) moiety produces thiamphenicol and a substantial change in biological activity, while modification of the propanediol group by the addition of a fluorine atom produces florfenicol. Both of these are discussed in more detail in Sections Thiamphenicol and Florfenicol. Loss of the dichloroacetyl group altogether results in loss of biological activity (Yunis, 1988; Hird and Knifton, 1986).

Image described by caption and surrounding text — **Figure 36.1** The chemical structure of chloramphenicol and modifications to form florfenicol.

After the discovery of chloramphenicol in 1947 it was in popular use decades ago, but has been gradually replaced by safer alternatives. The small animal formulation is approved by the FDA (Chloromycetin) but is not actively marketed. The use of chloramphenicol diminished in the 1970s and 80s because other active and safer drugs became available. Today, chloramphenicol has experienced a bit of a resurgence in companion animal medicine. Multidrug resistant bacteria, particularly methicillin-resistant Staphylococcus spp., are usually susceptible to chloramphenicol and this is one of the most common drugs selected for use by small animal veterinarians (Papich, 2012; Bryan et al., 2012; Frank and Loeffler, 2012). Antibiotic-resistant Enterococcus spp. are also often susceptible. Chloramphenicol has the disadvantage of a narrow margin of safety in dogs and cats, and necessity of frequent administration in dogs to maintain adequate concentrations (three or four times daily oral administration). These disadvantages still exist, but the activity of chloramphenicol against bacteria (e.g., staphylococci) that are resistant to other oral drugs has created increased use of chloramphenicol in recent years.

Drug Formulations

Many formulations have been removed from the commercial market because chloramphenicol no longer is in wide use for humans. Chloramphenicol has FDA approval for use in dogs, and is available in 100, 250, and 500 mg tablets (Chloromycetin). The oral suspension of chloramphenicol palmitate is rarely available. Chloramphenicol is not soluble and injectable formulations include esters such as succinate and palmitate, glycinate, or undecylenate. There also has been a propylene glycol solution. None of the injectable formulations are used today. Although chloramphenicol is poorly soluble (<5 mg/ml), the poor solubility does not interfere with oral absorption. Chloramphenicol is absorbed orally with or without food (except some formulations in cats). Tablets and capsules have similar oral absorption in dogs. Topical formulations of chloramphenicol have been used for otic and ophthalmic use, but the otic formulations have been replaced by newer forms containing florfenicol.

Mechanism of Action

Chloramphenicol inhibits protein synthesis. Its biological activity is due to interference with peptidyltransferase activity at the 50S ribosomal subunit, which is near the site of action of macrolide antibiotics and for which there can be competition (Yunis, 1988). Because of the interaction with peptidyltransferase, binding with the amino acid substrate cannot occur, and peptide bond formation is inhibited. Chloramphenicol affects mammalian protein synthesis to some degree, especially mitochondrial protein synthesis. Mammalian mitochondrial ribosomes have a strong resemblance to bacterial ribosomes (both are 70S), with the mitochondria of the bone marrow especially susceptible. Prolonged administration to animals has been associated with a dose-related bone marrow suppression, especially in cats (Watson, 1980).

The action of chloramphenicol (and florfenicol) is regarded as bacteriostatic, rather than bactericidal (Maaland et al., 2015). There are isolated examples in which bactericidal effects have been observed, but chloramphenicol and similar drugs in this class usually behave as bacteriostatic agents and the drug concentration in animals should be maintained above the MIC for as long as possible during the dose interval.

Spectrum of Activity

Chloramphenicol has a broad spectrum of activity. It is active against Staphylococcus pseudintermedius, S. aureus, streptococci, and some gram-negative bacteria, such as Pasteurella multocida, Mannheimia haemolyticia, and Histophilus somni. Escherichia coli, Proteus spp., and Salmonella spp. may be susceptible, but resistance can occur with many gram-negative bacteria, especially the Enterobacteraceae. One reason for the increased use of chloramphenicol, especially in dogs, is that it has retained activity against Staphylococcus pseudintermedius, including methicillin-resistant strains (Perreten et al., 2010). However, resistance by staphylococci may occur from repeated administration. Anaerobic bacteria, Mycoplasma spp., and many Rickettsiae also are susceptible. The Clinical Laboratory Standards Institute (CLSI, 2015) approved break point for susceptibility is ≤4 µg/ml for streptococci and ≤8 µg/ml for other organisms (Watts et al., 1999).

Bacterial Resistance

Four mechanisms of resistance to chloramphenicol have been described (Yunis, 1988; Schwarz et al., 2004). The most important is plasmid mediated due to the presence of the chloramphenicol acetyltransferase enzyme, which catalyzes a reaction that causes enzymatic inactivation by acetylation of the drug. This can occur through different types of chloramphenicol acetyltransferases (Schwarz et al., 2004). The acetyltransferases that cause resistance to chloramphenicol are less likely to affect florfenicol, making florfenicol more active against some pathogens (discussed in Section Florfenicol). Other mechanisms of resistance include efflux systems, inactivation by phosphotransferases, decreased bacterial cell wall permeability, altered binding capabilities at the 50S ribosomal subunit, and inactivation by nitroreductases.

Pharmacokinetics

Absorption and Distribution

The pharmacokinetic parameters of chloramphenicol have been studied in several animal species and are summarized in Tables 36.1 and 36.2. Chloramphenicol in animals is well absorbed via both oral and parenteral routes, with a few notable species exceptions. Plasma half-lives vary, ranging from 0.9 hours in ponies to 5.1 hours in the cat (Davis et al., 1972). Fasted cats showed differences in absorption between the chloramphenicol tablets and the chloramphenicol palmitate suspension (Watson 1992). The liquid formulation showed a lower systemic drug availability, indicating that hydrolysis of the palmitate form is necessary and that there is a higher risk of drug failure when the palmitate suspension is used to treat sick cats that are also not eating. In ruminants, microflora present in the ruminant forestomach tend to metabolize chloramphenicol faster than it can be absorbed, making chloramphenicol administered orally of little use in ruminant animals. This point is rather moot since administration of chloramphenicol to food animals in the United States is currently illegal (discussed in more detail in Chapter 55). In most animals, 30–46% of chloramphenicol is bound to plasma proteins, leaving much of the drug in the free and active form. Chloramphenicol is widely distributed to many tissues of the body due to its nonionized state and high lipophilicity, enabling it to cross lipid bilayers quite easily. The volume of distribution (Vd) is typically greater than 1.0 l/kg and has been measured at 1–2.5 l/kg (Tables 36.1 and 36.2). Chloramphenicol reaches sufficient concentrations in most tissues of the body, including the eye, central nervous system (CNS), heart, lung, prostate, saliva, liver, and spleen, among others (Ambrose, 1984; Hird and Knifton, 1986). Chloramphenicol concentrations in cerebrospinal fluid (CSF) are approximately 50% of corresponding plasma concentrations. Chloramphenicol can also cross the placental barrier in pregnant animals and can diffuse into the milk of nursing animals.

Table 36.1 Chloramphenicol pharmacokinetics in dogs (compilation of nine studies)

		Data set
		1	2	3	4	5	6	7	8	9	Mean	SD
Breed		Mixed	Beagle	Ghound	large dog	small dog	Ghound	Ghound	Ghound	Ghound
n		4	6	6	6	6	4	5	4	6
Formulation (50 mg/kg)		capsule	capsule	capsule	capsule	capsule	capsule	capsule	tablet	capsule
Elimination rate	1/hour	0.42	0.52	0.40	0.18	0.54	0.14	0.23	0.22	0.40	0.34	0.15
Half-life	hour	1.64	1.35	1.75	3.85	1.29	4.82	2.99	3.19	1.75	2.51	1.25
T_max	hour	4.00	2.00	3.00	3.00	1.50	2.00	3.00	1.50	3.00	2.56	0.85
C_max	μg/ml	16.70	19.65	18.60	27.50	20.00	16.50	18.50	23.80	18.60	19.98	3.54
AUC	h^*μg/ml	97.79	69.85	109.91	191.15	82.44	89.52	114.95	110.62	109.91	108.46	34.51
V/F	ml/kg	1212.94	1389.17	1149.22	1454.03	1124.29	3884.62	1874.17	2081.52	1149.22	1702.13	886.07
Cl/F	ml/h/kg	511.33	715.87	454.90	261.57	606.47	558.53	434.95	451.99	454.90	494.50	126.82
MRT	h	4.63	3.00	4.28	5.94	3.42	5.73	5.75	5.43	4.28	4.72	1.07

Ghound, Greyhound; T_max, time to peak concentration; C_max, peak concentration; AUC, area under the curve; V/F, volume of distribution per fraction absorbed orally; CL/F, clearance per fraction absorbed; MRT, mean residence time, SD, standard deviation.

Data set sources: (1) Eads, 1952; (2) Mercer, 1971; (3, 4, 5) Watson, 1974; (6) Watson, 1972a; (7) Watson, 1972b; (8) Watson and McDonald, 1976; (9) Watson, 1973.

Table 36.2 Selected serum pharmacokinetic parameters of chloramphenicol in animals

					Volume of
	Dose			Half-life	distribution
Species	(mg/kg)	Route	Formulation	(hr)	(l/kg)	Comments	Reference
Cats	22	IV	Base	5.1	2.36	Dissolved in 50% aqueous solution of N,N,di-methylacetamide	Davis et al., 1972
Sheep	30	IV	Base	1.702	0.691		Dagorn et al., 1990
	30	SC	Base	17.93	NA		Dagorn et al., 1990
	30	IM	Base	2.71	NA		Dagorn et al., 1990
Adult swine	22	IV	Base	1.3	1.05	Dissolved in 50% aqueous solution of N,N,di-methylacetamide	Davis et al., 1972
Piglets	25	IV	Base	12.7	0.9411	Normal piglets	Martin and Wiese, 1988
	25	IV	Base	17.2	0.9549	Colostrum-deprived piglets	Martin and Wiese, 1988
Goats	25	IV	Succinate	1.22	1.683	Nonfebrile animals	Kume and Garg, 1986
	25	IV	Succinate	1.29	1.962	Febrile animals	Kume and Garg, 1986
	25	IM	Succinate	1.46	3.019	Nonfebrile animals	Kume and Garg, 1986
	25	IM	Succinate	1.45	2.769	Febrile animals	Kume and Garg, 1986
	22	IV	Base	2.0	1.33	Dissolved in 50% aqueous solution of N,N,di-methylacetamide	Davis et al., 1972
	10	IV	Succinate	1.47	0.312	Normal animals	Abdullah and Baggot, 1986
Goats	10	IV	Succinate	3.97	0.287	Starved animals	Abdullah and Baggot, 1986
	22	IV	Base	2.0	1.33	Dissolved in 50% aqueous solution of N,N,di-methylacetamide	Davis et al., 1972
	10	IV	Succinate	1.47	0.312	Normal animals	Abdullah and Baggot, 1986
	10	IV	Succinate	3.97	0.287	Starved animals	Abdullah and Baggot, 1986
Cattle	40	IV	Base	2.81	0.351		Sanders et al., 1988
	90	IM	Base	1.345	NA	2 doses 48 hours apart	Sanders et al., 1988
	90	SC	Base	1.153	NA	2 doses 48 hours apart	Sanders et al., 1988
Calves	30	IV	Base	3.98	1.208	Age not reported; average weight 73 kg	Guillot and Sanders, 1991
1-day-old calves	25	IV	Base in PG vehicle	7.56	1.031		Burrows et al., 1983
7-day-old calves	25	IV	Base in PG vehicle	5.96	0.808		Burrows et al., 1983
14-day-old calves	25	IV	Base in PG vehicle	4.0	0.903		Burrows et al., 1983
28-day-old calves	25	IV	Base in PG vehicle	3.69	0.69		Burrows et al., 1983
9-month-old calves	25	IV	Base in PG vehicle	2.47	1.38		Burrows et al., 1983
Horses	22	IV	Base in PG vehicle	0.51–0.78	0.86–1.26		Varma et al., 1987
Ponies	22	IV	Base	0.9	1.02	Dissolved in 50% aqueous solution of N,N,di-methylacetamide	Davis et al., 1972
Foals
1 day old	25	IV	Succinate	5.29	1.1		Adamson et al., 1991
3 days old	25	IV	Succinate	1.35	0.759		Adamson et al., 1991
7 days old	25	IV	Succinate	0.61	0.491		Adamson et al., 1991
14 days old	25	IV	Succinate	0.51	0.426		Adamson et al., 1991
42 days old	25	IV	Succinate	0.34	0.362		Adamson et al., 1991
1–9 days old	50	IV	Succinate	0.95	1.6	After oral suspension administered oral, availability was 83% and half-life of 2.54 hours	Brumbaugh et al., 1983
Rabbits	100	IV	Succinate	1.1575	NA		Mayers et al., 1991
Chickens	20	IV	Succinate	8.32	0.24	Normal animals	Atef et al., 1991a
	20	IV	Succinate	26.21	0.3	E. coli-infected animals	Atef et al., 1991a
	20	IM	Succinate	7.84	0.44		Atef et al., 1991a
	20	PO	Succinate	8.26	0.41		Atef et al., 1991a

NA, data not available; PG, propylene glycol.

Metabolism and Excretion

Chloramphenicol is metabolized by the liver after absorption into the systemic circulation. One of the largest drawbacks to chloramphenicol administration is the rapid metabolic clearance, producing short half-lives in many species and necessity for frequent administration. As shown in Table 36.1 for dogs, the short half-life translates to a need to be administered three times daily. In horses, because of rapid elimination rates, tissue fluid concentrations persisted for only 3 hours after IV administration of chloramphenicol sodium succinate (Brown et al., 1984). Phase II glucuronidation is the principal pathway for the hepatic biotransformation of chloramphenicol, with the principal metabolite being chloramphenicol glucuronide. A few hydrolysis products have also been identified. Cats excrete chloramphenicol more slowly than other animals, perhaps owing to the cat’s deficiency in some glucuronidase enzymes. One report notes that 25% of the total dose of chloramphenicol is excreted in the urine in the active form in cats compared to 6% in dogs (Hird and Knifton, 1986). Most of the absorbed chloramphenicol (approximately 80%) is excreted into the urine as inactive metabolites via tubular secretion.

The effect of age on clearance of chloramphenicol is inconsistent. Calves showed differences compared to older cattle, but this is probably irrelevant because it should not be used in food animals (Burrows et al., 1984). Brumbaugh et al. (1983) found that in neonatal horses, elimination and Vd did not differ from adults. Bioavailability in foals was 83%, with an oral half-life of 2.54 hours.

Adverse Effects and Precautions

Bone marrow suppression has been the most important adverse effect associated with chloramphenicol administration to people. Bone marrow injury from chloramphenicol takes two forms (Yunis, 1988). The first type is the most common and involves a dose-related suppression of the bone marrow precursor erythroid series. This toxicosis is reversible. The evidence suggests that this bone marrow suppression is the result of suppression of mitochondrial protein synthesis in bone marrow cells. In bone marrow cells there is vacuolation of the myeloid and erythroid series precursor cells, and inhibition of erythroid and granulocytic colony forming units (IARC, 1976, 1990). In both the dog and the cat, dose-related bone marrow suppression is possible. However, signs of toxicity reverse when chloramphenicol therapy is discontinued.

The second type of bone marrow toxicity, aplastic anemia, has been described in people but not in animals. In people, it is rare and independent of dose and treatment duration. This toxicity results in bone marrow aplasia, chiefly characterized by a profound and persistent pancytopenia. Aplastic anemia occurs in approximately 1 : 10,000 to 1 : 45,000 humans who receive chloramphenicol. It appears that the para-nitro group of the chloramphenicol molecule is responsible for this more serious form of bone marrow toxicity (Figure 36.1). The para-nitro group undergoes nitroreduction, leading to the production of nitrosochloramphenicol and other toxic intermediates, which trigger the stem cell damage in humans (IARC, 1976, 1990; Yunis, 1988). Modification of the molecule to eliminate the para-nitro group to produce either thiamfenicol or florfenicol reduces the risk of chloramphenicol-associated aplastic anemia (Figure 36.1).

Chloramphenicol-induced aplastic anemia in humans is important from a food-animal residue standpoint. If chloramphenicol is used to treat infections in food animals, it is possible that low concentrations of chloramphenicol in milk, meat, and other edible tissues from the animals will be consumed by people and cause aplastic anemia in susceptible individuals. Chloramphenicol residues have been known to persist for prolonged periods in food animals (Korsrud et al., 1987). Even though the amount consumed may be small, reaction that may occur in people are not dependent on dose. Thus, there is a public health risk for individuals consuming these products. For this and other reasons, the use of chloramphenicol in food-producing animals has been banned in the United States. The hazards of using chloramphenicol in food animals have also been reviewed by others (Settepani, 1984; Lacey, 1984).

Other adverse effects caused by chloramphenicol in animals have been observed since the drug is used more in recent years to treat drug-resistant bacteria. Young animals and cats are sensitive to intoxication due to impaired glucuronidation pathways. Cats given 60 and 120 mg/kg/day PO every 8 hours for 21 and 14 days (respectively) showed clinical signs of depression, dehydration, reduced fluid intake, weight loss, emesis, and diarrhea. Bone marrow hypoplasia was also documented in addition to pancytopenia (Watson, 1980). Other investigators (Penny et al., 1967, 1970) administered to cats 50 mg/kg/day IM, with the cats showing marked depression and inappetence by day 7 of administration, severe bone marrow changes by day 14, and becoming extremely ill by day 21.

Gastrointestinal (GI) disturbances are among the most common in dogs (Short et al., 2014; Bryan et al., 2012). Dogs may exhibit events such as vomiting, diarrhea, anorexia, drooling, gagging or any combination of these clinical signs. They tend to resolve when the medication is discontinued. These effects may be related to GI injury as oral administration resulted in intestinal mucosal damage and diarrhea in calves and reduced glucose absorption (Rollin et al., 1986). Another issue that has emerged now that chloramphenicol is used more often is peripheral neuropathy. In one report, this adverse effect was almost as common as the gastrointestinal problems (Short et al., 2014). Signs that may be observed are ataxia, rear limb weakness, trouble standing, and/or jumping, or trembling on the back legs. This is believed to be caused by peripheral neuropathy, but the cellular mechanism is unknown. One microscopic study in three dogs (Kuroda et al., 1974) identified degenerative changes in peripheral nerves. Larger-breed dogs may be at higher risk for the neuropathy based on anecdotal accounts. Most dogs recover when the medication is discontinued.

Drug Interactions

Chloramphenicol is an inhibitor of the cytochrome P450 (CYP) drug-metabolizing enzymes. Enzyme specificity has not been fully characterized, but there is evidence that one of the enzymes inhibited is canine CYP2B11 (Martinez et al., 2013). Among the drugs substrates that may be affected by inhibition from chloramphenicol are anticonvulsants (e.g., phenobarbital), propofol, benzodiazepines, and other anesthetics. For example, chloramphenicol significantly affected metabolism of methadone in dogs (KuKanich and KuKanich, 2015).

Clinical Use

The FDA-approved dose for dogs is 55.5 mg/kg oral, every 6 hours. This dose is likely to increase the risk of adverse effects and the most common clinical dose, based on pharmacokinetic studies and evidence of efficacy is 50 mg/kg every 8 hours oral. Chloramphenicol has been used for treatment of a wide range of susceptible microbial infections, including those caused by salmonellae, intracellular and extracellular bacteria, rickettsiae, and mycoplasmata; infections of the eyes and CNS; and infections due to anaerobic organisms (IARC, 1976, 1990). One of the reasons for its popularity has been the high lipophilicity. Chloramphenicol readily penetrates cells, making it active against intracellular bacteria. It can penetrate tissues that otherwise are difficult to treat, such as the CNS, which is further discussed below. Chloramphenicol was shown in one study to be equally effective for treatment of Rocky Mountain spotted fever in dogs as enrofloxacin and tetracyclines (Breitschwerdt et al., 1990). Chloramphenicol has been used to treat infections caused by Staphylococcus spp., streptococci, Brucella spp., Pasteurella spp., E. coli, Proteus spp., Salmonella spp., Bacillus anthracis, Arcanobacterium pyogenes, Erysipelothrix rhusiopathiae, and Klebsiella pneumoniae. It is consistently active against anaerobic bacteria.

Chloramphenicol has been suggested for treatment of infections of the CNS (encephalitis, meningitis) because it is able to cross the inflamed or uninflamed blood–brain barrier and attain therapeutic concentrations in the CSF and the brain. Despite the rationale for this use, some experts have suggested that since chloramphenicol is merely bacteriostatic against gram-negative pathogens, and there is a lack of phagocytes or immunoglobulins in CSF, chloramphenicol is not well suited to treat serious infections of the CNS (Rahal and Simberkoff, 1979).

Chloramphenicol attains high concentrations in the eye when given systemically or after topical application on the cornea and is useful in treating susceptible bacterial conjunctivitis, panophthalmitis, endophthalmitis, and bacterial diseases of the cornea (Conner and Gupta, 1973). Topical formulations are not as readily available owing to the risk of aplastic anemia (discussed in Section Adverse Effects and Precautions), which can be caused by topical exposure.

Chloramphenicol has been used to treat bacterial infections of the respiratory tract because it may have better penetration across the blood–bronchus barrier into respiratory secretions and respiratory lining fluid than more polar or less lipophilic antibiotics. Respiratory infections in horses, dogs, cats, and exotic animals are among the uses of oral chloramphenicol.

Chloramphenicol is among the few drugs that can be administered orally to horses with safety. It achieves moderate systemic absorption of 21–40% (Gronwall et al., 1986) and has no serious adverse effects on the equine digestive system. For treatment in horses, tablets or capsules are mixed with vehicles such as molasses or corn syrup to facilitate oral administration. Chloramphenicol has been administered to horses for respiratory infections, pleuritis, CNS infections, and joint infections. Because there are other active drugs available, it is most often considered as an option when bacteria are resistant to other antibiotics. The recommended doses are based on specific pharmacokinetic studies in adults and foals (Gronwall et al., 1986; Brumbaugh et al., 1983).

Chloramphenicol has been administered to exotic animals, especially reptiles and amphibians, to treat a variety of infections (Clark et al., 1985); although florfenicol (see Section Florfenicol) has taken over some of these uses. Chloramphenicol administration in 15 species of birds was examined, and the investigators concluded that after IM injections of 50 mg/kg, chloramphenicol would produce adequate concentrations to treat susceptible bacteria for 8–12 hours, except in pigeons, macaws, and conures because effective concentrations could not be achieved in these birds (Clark et al., 1982). However, oral absorption was poor, and this route of administration was discouraged for all birds.

Chloramphenicol Derivatives

The ban on the use chloramphenicol in food-producing animals in the mid-1980s left a gap in the veterinarian’s armamentarium of effective antimicrobial drugs. Because the idiosyncratic aplastic anemia is associated with the presence of the para-nitro group on the chloramphenicol molecule, attempts were made to modify the chloramphenicol structure to simultaneously retain chloramphenicol’s broad spectrum of antimicrobial activity and eliminate the induction of aplastic anemia in people. Compounds synthesized in attempts to accomplish this goal are thiamphenicol and florfenicol. Thiamfenicol is not approved in the United States and will be discussed here only briefly. However, florfenicol has been approved for use in pigs, cattle, and fish (in some countries) and has been effective for treatment of various infections, especially bovine respiratory disease in cattle and swine respiratory disease in pigs intended for human consumption.

Thiamphenicol

Thiamphenicol is a semisynthetic structural analog of chloramphenicol. It is not available in North America; therefore, all of the experience has been learned from research studies or use in other countries. The major structural difference between chloramphenicol and thiamphenicol is that the para-nitrophenol group has been replaced by the methyl sulfonyl moiety (Figure 36.1). The mechanism of action and spectrum are similar to that of chloramphenicol. However, its structural differences result in different pharmacokinetic properties and decreased potency. Thiamphenicol is more water soluble and less lipid soluble and therefore diffuses more slowly through lipid membranes. It is not metabolized to a significant extent in the liver (Ferrari et al., 1974) and most of the dose is excreted in the urine as the unchanged active compound (Yunis, 1988; Lavy et al., 1991a; Gamez et al., 1992). Resistance to thiamphenicol is also similar to that of chloramphenicol, with bacterial acetylation of the thiamphenicol molecule, but at a rate approximately 50% less than that of chloramphenicol.

Few pharmacokinetic studies have been performed on food-producing animals, but thiamphenicol pharmacokinetics has been studied in veal calves (Gamez et al., 1992) and lactating goats (Lavy et al., 1991a). Both studies found thiamphenicol to have a large volume of distribution and rapidly eliminated in the urine. In dogs, thiamfenicol had a half-life of 1.7 hours and a volume of distribution of 0.66 l/kg (Castells et al., 1998). In dogs the injection of thiamfenicol was well absorbed, with availability of 97%, but the terminal half-life was longer (5.6 hours), suggesting slow release from the injection site.

Thiamphenicol is considered to be less toxic than chloramphenicol, yet a reversible bone marrow suppression has been reported in humans. However, millions of people have been treated with thiamphenicol in countries in which it is approved, with no reports linking its use to aplastic anemia (Adams et al., 1987). In a thiamphenicol toxicity study in rabbits (Kaltwasser et al., 1974), no changes attributed to thiamphenicol in erythrocyte, reticulocyte, or plasma iron parameters were noted after long-term treatments of up to 90 mg/kg/day.

Florfenicol

Florfenicol is structurally related to thiamphenicol; however, florfenicol contains fluorine at the 3′ carbon position (Figure 36.1). The fluorine molecule substitution at this position also reduces the number of sites available for bacterial acetylation reactions to occur, possibly making the antibiotic more resistant to bacterial inactivation. Florfenicol is as potent, or more potent, than either chloramphenicol or thiamphenicol against many organisms in vitro. The study by Maaland et al. (2015) using Staphylococcus pseudintermedius and E. coli isolates, showed that there were fewer nonwild-type isolates for florfenicol than chloramphenicol. There were no nonwild-type isolates of Staph. pseudintermedius for florfenicol. These results agree with previous studies that show that resistance mechanisms may be less likely for florfenicol compared to chloramphenicol (Schwarz et al., 2004).

The list of susceptible bacteria for florfenicol is the same as listed previously for chloramphenicol. However, as mentioned earlier, some bacteria resistant to chloramphenicol because of inactivation by acetylation may be susceptible to florfenicol. The CLSI (CLSI, 2015) quality control ranges of MIC for florfenicol are 2–8 µg/ml (Marshall et al., 1996). Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni are several-fold more susceptible in vitro than bacteria of the Enterobacteriaceae, with MIC₉₀ for Pasteurella and Histophilus somni in the range of 0.5–1.0 µg/ml. The CLSI breakpoints are ≤2 µg/ml (susceptible) 4 µg/ml (intermediate), and ≥8 µg/ml (resistant) for isolates of bovine and swine respiratory disease (CLSI, 2015). By comparison, the susceptible breakpoint for chloramphenicol is ≤8 µg/ml for organisms other than streptococci, and ≤4 µg/ml for Streptococcus spp. Florfenicol breakpoints for other animals and other bacteria have not been determined.

The advantage of florfenicol for administration to food animals is that it lacks the para-nitro group (Figure 36.1) that could contribute to the induction of aplastic anemia associated with chloramphenicol use in humans. Therefore, if residues were to occur in animals treated with florfenicol, no dangerous public health risk would ensue. However, it is possible that florfenicol can still produce a dose-related form of reversible bone marrow suppression with prolonged use or high doses. Such reactions have not been reported from routine use of florfenicol in animals, probably because it is rarely used for a long time. However, in one clinical account in a zoo animal, high doses induced bone marrow suppression (Tuttle et al., 2006).

Pharmacokinetics

The pharmacokinetics of florfenicol are summarized in Table 36.3 .

Table 36.3 Selected pharmacokinetic parameters of florfenicol in animals

	Dose	Half-life	Absorption (%)	Volume of	C_max
Species	mg/kg	(h)	(%)	distribution (l/kg)	(µg/ml)	Reference
Cats	22 (all routes)	4 (IV)	–	0.61	57 (IV)	Papich, 1999
		7.8 (oral)	>100 (oral)	–	28 (oral)
		5.6 (IM)	>100 (IM)	–	20 (IM)
Dogs	20 mg (all routes)	2 (IV)	28 (SC)	1.2	44 (IV)
		18 (SC)	16 (IM)	–	0.93 (SC)
		9 (IM)	–	–	1.64 (IM)
		3 (oral)	>100 (oral)	–	17 (oral)
	20 IV	1.11	–	1.45	–	Park et al., 2008
	20 oral	1.24	95.43	–	6.18	Park et al., 2008
Sea turtles	20 IM, IV	2–7.8 h (IM)	67 (IM)	10–60	0.5–0.8 (IM)	Stamper et al., 2003
Sharks	40 IM	269	ND	2.9	10.5	Zimmerman et al., 2006
Fish (red pacu)	10 IM	4.25	ND	5.69	1.09	Lewbart et al., 2005
Horses	22 IV	1.83	81 (IM)	0.72	4 (IM)	McKellar and Varma, 1996
	22 oral	ND	83 (oral)	ND	13 (oral)
Cattle	50 IV	3.2	ND	0.67	157.7	Bretzlaff et al., 1987
Feeder calves	20 IV	2.65	ND	0.88	73	Lobell et al., 1994
	20 IM	18.3	78.5	ND	3.07	Lobell et al., 1994
Veal calves	22 oral	ND	88	ND	11.3	Varma et al., 1986
	22 IV	2.87	ND	0.78	66	Varma et al., 1986
	11 IV	3.71	ND	0.91	26.35	Adams et al., 1987
	11 oral	3.7	89	ND	5.7	Adams et al., 1987
Angus calves	40 SC	27.5	ND	ND	6.04	Sidhu et al., 2014
Dairy calves	40 SC	28.44	ND	ND	3.42	Foster et al., 2016
Lactating cows	20 IV	2.9	–	0.35	12.4	Soback et al., 1995
	20 IM	12	38	ND	3.6
Alpaca	20 IM	17.59	ND	11.07 (Vd/F)	4.31	Holmes et al., 2012
	40 SC	99.67	ND	55.74 (Vd/F)	1.95
Llama	20 IV	2.2	63	0.96	–	Pentecost et al., 2013
	20 IM	11.6	–	–	3.2
Camels	20 IV	1.49	69.2	0.89	–	Ali et al., 2003
	20 IM	2.52	–	–	0.84
Sheep	20 IV	1.31	65.8	0.69	–
	20 IM	2.28	–	–	1.04
Goats	20 IV	1.19	60.9	0.57
	20 IM	2.12	–	–	1.21

Route of administration used is listed with dose. C_max is the maximal concentration after administration. ND, not determined. Empty cells indicate that the parameter is not relevant for the route administered.

Absorption:

Studies in calves and other species listed in Table 36.3 show that absorption from all routes after either IM or SC injection is generally high. Oral absorption in horses and dogs was also high. After IM or SC injection, the absorption is slow and often prolonged in animals because of the vehicle in the solution. Therefore, the IM or SC injection produces a flip-flop effect, in which the terminal half-life is determined by the slow absorption. This can be seen in Table 36.3 in which the IM and SC half-life is generally much longer than the IV half-life. This effect prolongs the duration of effective concentrations.

Distribution:

Like chloramphenicol, florfenicol has a wide distribution in most tissues of the body (Adams et al., 1987), with a volume of distribution approaching 1 l/kg (0.7–0.9 l/kg in most cattle studies) (Table 36.3 ). Protein binding is low in cattle with values of 13–19% reported (Bretzlaff et al., 1987; Lobell et al., 1994), but in other studies it was 5% at high concentrations and negligible at low concentrations in cattle plasma (Foster et al., 2016). Protein binding has not been reported for other animals. High concentrations are detected in the kidney, urine, bile, and small intestine, but less penetration in the CSF, brain, and aqueous humor of the eye than that attained with chloramphenicol. Concentrations in brain and CSF are one-quarter and one half the corresponding concentrations in plasma, respectively. Although in one study the distribution into CSF was only 46% relative to plasma, these levels were high enough to produce concentrations in CSF of cattle to inhibit Histophilus somni for over 20 hours (De Craene et al., 1997). Florfenicol reached high concentrations in the synovial fluid of cattle following regional limb perfusion (Gilliam et al., 2008). Florfenicol also penetrated well into the milk of lactating goats after IV and IM administration; therefore, it could be used to treat microbial infections in the udder of lactating animals (Lavy et al., 1991b) if appropriate milk withdrawal times are available. The penetration into interstitial fluid was almost 98%, a reflection of the low protein binding. In the same study, the penetration into the pulmonary epithelial lining fluid of calves was over 200% and produced high concentrations for treating respiratory infections (Foster et al., 2016).

Metabolism and Elimination:

The elimination half-life in various species and for different routes is shown in Table 36.3. Most of the dose administered to cattle is excreted as the parent drug (64%) in the urine, with the remaining excreted as urinary metabolites. Florfenicol amine is the metabolite that persists longest in tissues of cattle and is used as the marker residue for withdrawal determination.

Pharmacokinetic studies in other species:

As seen in Table 36.3, there have been pharmacokinetic studies in small animals and some exotic and zoo animals. As in cattle, it has rapid clearance when injected IV, but more prolonged terminal half-life if administered by other routes.

Pharmacokinetic–pharmacodynamic properties:

The Pharmacokinetic–pharmacodynamic (PK-PD) properties for florfenicol may be dependent on the organism studied. There is evidence for a bactericidal effect against some bacteria but not others (Maaland et al., 2015; Sidhu et al., 2014). Florfenicol may be bactericidal against isolates of Staph. pseudintermedius but not E. coli (Maaland et al., 2015). Against bovine isolates of Mannheimia haemolytica and Pasteurella multocida, florfenicol appears to have bactericidal activity. It is not established if the parameter for predicting efficacy is time above MIC (T>MIC) or area-under-the-curve (AUC) / MIC. It is likely that as for most other protein synthesis inhibiting agents with little or no postantibiotic effect, the AUC/MIC would be the best parameter to predict clinical efficacy. In a study in calves (Sidhu et al., 2014) a AUC/MIC ratio of approximately 18–27 was identified from modeling experiments.

Clinical Use

Florfenicol is available in three injectable solutions: 300 mg/ml solution for injection (Nuflor, or Nuflor gold), and a solution combined with flunixin meglumine (Resflor Gold, 300 mg/ml florfenicol plus 16.5 mg/ml flunixin). There is also a solution to be added to drinking water for swine (23 mg/ml, to be added as 400 mg per gallon) and a Type A medicated feed. For fish there is a 500 gram per kilogram premix for fish (Aqua-Flor).

Cattle and pigs:

Several studies in cattle have been conducted to support the use of florfenicol for treating bovine respiratory disease caused by Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni. Florfenicol penetrates well into the epithelial lining fluid of the airways of cattle (Foster et al., 2016), and has been effective for treating undifferentiated bovine respiratory disease (Hoar et al., 1998; Jim et al., 1999). There are two doses approved for cattle: 20 mg/kg SC or IM, given every 48 hours and injected in the neck, or a single dose for cattle at 40 mg/kg SC in the neck. Florfenicol is also approved for treatment of bovine interdigital phlegmon (foot rot, acute interdigital necrobacillosis, infectious pododermatitis) associated with Fusobacterium necrophorum and Bacteroides melaninogenicus.

Florfenicol has been effective in calves for treating experimentally induced infections and naturally occurring infectious bovine keratoconjunctivitis (Dueger et al., 1999; Angelos et al., 2000). In the naturally occurring case, florfenicol was administered one dose SC at 40 mg/kg or IM two doses 48 hours apart at 20 mg/kg. Concentrations persist in CSF for a long enough period after administration of 20 mg/kg in cattle that concentrations are above MIC of susceptible pathogens for at least 20 hours.

The swine dose is 15 mg/kg IM in the neck, every 48 hours. For pigs, florfenicol can also be added to the feed (182 g per ton of feed), or drinking water (400 mg per gallon) for 5 days for the control of swine respiratory disease associated with Actinobacillus pleuropneumoniae, Pasteurella multocida, Streptococcus suis, and Bordetella bronchiseptica.

Small animals:

Although some pharmacokinetic studies have been conducted in small animals and exotic animals, there are no reports of efficacy. Pharmacokinetic studies in reptiles and dogs suggest that frequent dosing with high doses would be necessary to maintain plasma concentrations above the MIC for susceptible organisms throughout the dosing interval. By contrast, florfenicol solution in cats was absorbed well from both routes, with peak concentrations of 20 µg/ml and 27 µg/ml after IM and oral dose, respectively. Absorption was high from both routes (greater than 100% from IM and oral). The half-life was 5.6 hours and 7.8 hours for IM and oral dose, respectively. In cats, florfenicol produced inhibitory concentrations for 12 hours. These studies indicate that a dose of 22 mg/kg administered every 12 hours orally or parenterally would be adequate to produce sustained plasma concentrations for treatment of susceptible bacteria. Safety of these doses for small animals have not been established.

Topical forms:

Two topical formulations were approved by the FDA in 2014 and 2015. The product Osurnia^® contains 10 mg florfenicol, 10 mg terbinafine, and 1 mg betamethasone acetate per ml in a gel for topical administration. The product Claro™ contains florfenicol 15 mg/ml, terbinafine 13.3 mg/ml, and mometasone 2 mg/ml. The indication for each product is for the treatment of otitis externa in dogs associated with susceptible strains of bacteria (Staphylococcus pseudintermedius) and yeast (Malassezia pachydermatis).

Use in fish:

Florfenicol has been administered orally for treatment of infections in captive fish and is approved in some countries for this use (Aqua-Flor^®). Florfenicol is effective for treating bacterial infections in fish, such as trout and salmon (Fukui et al., 1987). Florfenicol premix is approved in some countries for treatment of furunculosis in salmon caused by Aeromonas salmonicida. Florfenicol has been administered orally for treatment of furunculosis caused by susceptible strains of Aeromonas salmonicida in captive fish and is approved in other countries. The premix (Aqua-Flor^®) is approved for use in catfish and salmonids at a dose of 10 mg/kg for 10 days to treat susceptible fish pathogens.

The typical dose for fish is 10 mg/kg. At this dose, the half-life is 12–16 hours in most fish, with a peak (C_max) concentration of approximately 3–10 µg/ml (Pinault et al., 1997; Martinsen et al., 1993; Horsberg et al., 1994). Red pacu had a shorter half-life and lower C_max after 10 mg/kg IM (Lewbart et al., 2005). In sharks at a dose of 40 mg/kg IM, florfenicol produced effective levels for 120 hours (Zimmerman et al., 2006). Admnistration to sharks every 3–5 days will produce concentrations in a therapeutic range.

Other species:

Pharmacokinetic studies in horses show that florfenicol has longer half-life than chloramphenicol, good distribution, and good absorption. However, in experimental horses there were consistent loose stools and elevated bilirubin (McKellar and Varma, 1996). Until additional studies are available to establish safe doses, florfenicol cannot be recommended for horses. Adverse effects were observed in alpaca at repeated doses of 40 mg/kg SC, but not in llamas administered a single dose of 20 mg/kg (Holmes et al., 2012; Pentecost et al., 2013). The authors recommended a dose in llamas of 20 mg/kg once daily, IM. After a single injection of 20 mg/kg (IM and IV) there were no adverse effects identified clinically or in blood tests in camels, sheep, or goats (Ali et al., 2003). However, the low concentrations achieved and the short half-life in the study by Ali et al., raises questions about whether or not it would be clinically effective at this dose.

In snakes (boas), the half-life was 28 hours from IM injection. It was estimated that 50 mg/kg once daily for boas is the best dose to produce therapeutic plasma concentrations, even though efficacy studies are not available. In sea turtles the clearance was rapid (60–100 ml/kg/h) and the half-life was short (Stamper et al., 2003). It was concluded that florfenicol was not a practical drug for treatment of infections in sea turtles.

Adverse Effects

Effect of florfenicol on bovine pregnancy, reproduction, and lactation have not been determined. Mild diarrhea and elevated bilirubin have been reported from administration to horses (McKellar and Varma, 1996). Reversible, dose-related bone marrow suppression is possible but not reported, except for a reaction reported in a zoo animal that was mentioned previously (Tuttle et al., 2006). In cattle, diarrhea and decreased feed consumption have been observed, which are transient. A local tissue reaction from IM or SC injection is possible. When toxic overdoses were administered to calves (200 mg/kg) there was marked anorexia, decrease in body weight, ketosis, and elevated liver enzymes. In dogs administered high doses for prolonged periods there was CNS vacuolation, hematopoietic toxicity, and renal tubule dilation. Adverse effects were detected in alpacas after a dose of 40 mg/kg SC that may be related to the prolonged concentrations at this dose (Table 36.3 ) (Holmes et al., 2012). These effects included significant hematological abnormalities and protein decrease. Caution should be used if administering to these animals for repeated doses.

Regulatory Information

The tolerance for florfenicol is 3.7 ppm for florfenicol amine (the marker residue) in liver and 0.3 ppm in muscle. Withdrawal time for use in salmon is 12 days. Withdrawal time for oral administration to pigs in feed is 13 days and for administration in water 16 days. After injection to cattle, the withdrawal time for slaughter is 28 days if injected at a dose of 20 mg/kg IM (36 days in Canada). If injected at a dose of 40 mg/kg SC, the withdrawal time for slaughter is 38 days. A formulation with different excipients (Nuflor Gold) has a withdrawal time of 44 days in cattle when injected at 40 mg/kg SC, once. More than 10 ml should not be injected at each site to avoid tissue reactions and injections should be administered in the neck (both SC and IM). Do not administer to dairy cows older than 20 months, to calves under 1 month of age, or to calves on an all-milk diet.

Macrolide Antibiotics

Source and Chemistry

The macrolide antibiotics are a group of structurally similar compounds, most of which are derived from various species of Streptomyces soil-borne bacteria. Chemically, all the drugs in this group are classified as macrocyclic lactones, with members containing 12–20 atoms of carbon in the lactone ring structure (Table 36.4 ). Attached to this lactone ring are various combinations of deoxy sugars held to the lactone ring by glycosidic linkages. Since erythromycin’s discovery in the early 1950s from the soil organism Streptomyces erythreus, numerous other macrolides have been isolated or synthesized from the parent molecule erythromycin, including tylosin, roxithromycin, erythromycylamine, tilmicosin, dirithromycin, azithromycin, tulathromycin, clarithromycin, spiramycin, and flurithromycin (Kirst and Sides, 1989). The most common agents used clinically in veterinary medicine are listed in Table 36.4 .

Table 36.4 Macrolides used in animals

Drug	Structure	Brand Name
Erythromycin	14 member ring	Gallimycin (and generic)
Tilmicosin	16 member ring	Micotil, Pulmotil
Azithromycin	15 member ring	Zithromax (human drug)
Gamithromycin	15 member ring	Zactran
Tylosin	16 member ring	Tylan
Tildipirosin	16 member ring	Zuprevo
Tulathromycin	15 member ring	Draxxin

Erythromycin and tylosin (Figure 36.2) are traditional macrolides. The newer drugs (Table 36.4 and Figure 36.3) were either developed for use in people (azithromycin) or specifically for use in cattle and/or pigs. These newer drugs differ from erythromycin in that they have a prolonged action and can be administered intermittently, or for just a single injection. Other macrolides such as oleandomycin and carbomycin have been used as feed additives for growth promotion in food animals and will not be discussed in detail here.

Macrolides are composed of a macrolactone ring of 12, 14, 15, or 16 carbon atoms, substituted with sugar moieties. Erythromycin, the prototype of this class, consists of a 14-atom polyhydroxylactone erythronolide ring and the two sugars clandinose and desosamine. Similarly, tylosin is composed of a 16-atom lactone ring (a tylonolide) to which three sugars – mycinose, mycaminose, and mycarose – are attached (Wilson, 1984; Kirst et al., 1982). Azithromycin is the first drug in the group of azalides, which are semisynthetic derivatives of erythromycin (Lode et al., 1996). Azithromycin has a 15-member ring structure. Tulathromycin, resembles azithromycin (Figure 36.3) and also has a 15-member ring structure. The structure of the newer agents includes basic nitrogen groups. All macrolides are weak bases, with pK_a ranges from 6 to 9. They can have either two (di-basic) or three (tri-basic) nitrogen groups. For example, tulathromycin has three (tribasic) and has been referred to as a “trimilide”. Tildipirosin also has three basic groups. The basic nitrogen groups on these newer agents (Figure 36.3) produces a positive charge in an acidic environment below their pKa. The positive charge increases the affinity for intracellular sites caused by ion trapping. It is this property that gives these agents such large intracellular distribution and a high volume of distribution.

Effect on Antibacterial Activity:

The basic nature of the macrolides also influences the antibacterial activity. The in vitro antibacterial activity of macrolides varies according to the pH of the culture medium and the pH at the site of infection. Subsequently, antibacterial activity decreases in acid pH and increases in alkaline conditions. A change in pH of only 0.2 units has been known to change the MIC by a full Log-2 dilution step. These changes become important when CO₂ is used during culture incubation because it lowers the pH of the medium.

Mechanism of Action

The antibacterial action of macrolides is produced by an inhibition of protein synthesis by binding to the 50S ribosomal subunit at the 23S rRNA site of prokaryote organisms. The binding site on the ribosome is near, but not identical to, that of chloramphenicol, and antagonism of effect is possible if macrolides are administered with chloramphenicol. By binding to the 50S ribosomal site, macrolides cause dissociation of peptidyl-tRNA from the ribosomes during the elongation phase, which disrupts addition of new peptide bonds and thus prevents synthesis of new proteins. Although macrolides can bind to mitochondrial ribosomes, they are unable to cross the mitochondrial membrane (in contrast to chloramphenicol) and do not produce bone marrow suppression in mammals. Macrolides do not bind to mammalian ribosomes, making them a relatively safe group of drugs for veterinary use.

Although most authors have listed macrolides as bacteriostatic at therapeutic concentrations (Wilson, 1984), this effect may be both bacterial species, concentration, and drug dependent. For example, the agents developed for pigs and cattle (Table 36.4 ) can have bactericidal activity against bovine and swine respiratory pathogens, including Mannheimia haemolytica, Pasteurella multocida, Histophilus somni, and Actinobacillus pleuropneumoniae. As discussed above, the antimicrobial action of macrolides is enhanced by a high pH (Sabath et al., 1968), with the optimum antibacterial effect at a pH of 8. Therefore, in an acidic environment, such as in an abscess, necrotic tissue, or urine, the antibacterial activity is suppressed.

Resistance Mechanisms

Resistance to macrolides is usually plasmid mediated, but modification of ribosomes may occur through chromosomal mutation. Resistance can occur from: (i) decreased entry into bacteria (most common with the gram-negative organisms), and also mediated by mef-efflux genes, (ii) synthesis of bacterial enzymes that hydrolyze the drug, and (iii) modification of target (the ribosome in this instance) by RNA methylation or RNA sequence changes through mutation. The ribosomal attenuation (most common mechanism) involves methylation of the 50S drug receptor site. This resistance, coded by erm genes (e.g., ermA, ermB, ermC) may also lead to cross-resistance with other antibiotics that preferentially bind to these sites, such as other macrolides and lincosamides (Wilson, 1984). Resistance to erythromycin in animals in several microorganisms has been discussed in more detail elsewhere (Maguire et al., 1989; Dutta and Devriese, 1981, 1982a, 1982b; Leclercq and Courvalin, 1991; Devriese and Dutta, 1984). In small animals with staphylococcal infections, resistance was more likely if antibiotics had previously been prescribed, especially in cases of recurrent pyoderma (Lloyd et al., 1996; Medleau et al., 1986; Noble and Kent, 1992). Seven to 22% of small animal isolates of Staphylococcus spp. can exhibit resistance (depending on region and use), and in some countries this has remained relatively stable at around 22–24%.

Spectrum of Activity

Erythromycin is mainly effective against gram-positive organisms such as streptococci, staphylococci, including staphylococci that may be resistant to β-lactams because of β-lactamase synthesis or modification of the penicillin-binding protein target. Other organisms that show in vitro susceptibility to macrolides include Mycoplasma, Arcanobacterium, Erysipelothrix, Bordetella, and Bartonella. Although the spectrum favors the gram-positive group, a few gram-negative bacteria are susceptible, especially Pasteurella spp. Activity against anaerobic bacteria is only moderate. Most other gram-negative bacteria, such as those of the Enterobacteriaceae or Pseudomonas spp., are resistant. Azithromycin is an exception among the macrolides and can exhibit more activity against gram-negative bacteria. In addition to better activity against Enterobacteriaceae, it also has activity against other enteric pathogens, such as Campylobacter spp. (Gordillo et al., 1993).

The activity of newer derivatives (Table 36.4 and Figure 36.3) is similar to that of erythromycin, but these agents have better activity against respiratory pathogens, including Pasteurella, Mannheimia haemolytica, and Histophilus somni, which corresponds to their use for treating respiratory tract infections in pigs and cattle. These macrolides also have activity against Mycoplasma spp. The activity of macrolides against Rhodococus equi is important for treating lung infections caused by this organism in horses, particularly foals (Jacks et al., 2003). The MIC values for 32 antimicrobials against Rhodococcus equi were compared by Riesenberg et al. (2014). In decreasing order of activity, the MIC₉₀ values for clarithromycin, erythromycin, azithromycin, tilmicosin, tylosin, tulathromycin were 0.06, 0.5, 1, 32, 32, and 64 µg/ml, respectively. In a separate study, gamithromycin had a MIC₉₀ of 1 µg/ml (Berghaus et al., 2012). Therefore, all macrolides are not alike with respect to their activity against this equine pathogen.

The CLSI (CLSI, 2015) standards and interpretive categories are shown in Table 36.5 for susceptibility testing. As shown in this table, drugs in this class vary in their potency and activity against various pathogens. Because of their targeted use, most of this data were generated for respiratory pathogens (Watts, 1999).

Table 36.5 Interpretive criteria for macrolides and lincosamides used in animals. Source: Data from CLSI, 2015.

		MIC Interpretive Category (µg/ml)
Drug	Species	S	I	R	Comments / Pathogens
Erythromycin	Humans	≤ 0.5	1-4	≥ 8	Human Staphylococcus. No criteria available for animals.
	Humans	≤ 0.25	0.5	≥ 1	Human Streptococcus. No criteria available for animals.
Azithromycin	Humans	≤ 0.5	1	≥ 2	Human only. No criteria available for animals.
Tilmicosin	Bovine	≤ 8	16	≥ 32	Bovine respiratory pathogens.
	Swine	≤ 16	–	≥ 32	Swine respiratory pathogens
Tulathromycin	Bovine	≤ 16	32	≥ 64	Bovine respiratory pathogens (Mannheimia, Pasteurella, Histophilus)
	Swine	≤ 16	32	≥ 64	Pasteurella multocida, Bordetella bronchiseptica
		≤ 64	–	–	Actinobacillus pleuropneumoniae
Tildipirosin	Bovine	≤ 8	16	≥ 32	Bovine respiratory pathogens (Histophilus, Pasteurella)
		≤ 4	8	≥ 16	Bovine respiratory pathogens (Mannheimia)
	Swine	≤ 8	–	–	Bordetella bronchiseptica
		≤ 4	–	–	Pasteurella multocida
		≤ 16	–	–	Actinobacillus pleuropneumoniae
Gamithromycin	Bovine	≤ 4	8	≥ 16	Bovine respiratory pathogens. (Mannheimia, Pasteurella, Histophilus)
Clindamycin	Canine	≤ 0.5	1-2	≥ 4	Staphylococcus spp. Streptococcus spp.

S, susceptible; R, resistant; I, intermediate.

Pharmacokinetic–Pharmacodynamic Properties

The PK/PD properties of macrolides have been more difficult to define compared to other antimicrobials. Plasma concentrations, especially for the newer, long-acting agents (Table 36.6 ) are often below the MIC of pathogens for most, or all of the dose interval. Therefore, parameters such as peak above MIC (C_max/MC) or time above MIC (T>MIC) cannot be used to predict efficacy. Efficacy is probably best attributed to the concentrations at the site of infection – the pulmonary epithelial lining fluid (PELF). Although concentrations in the PELF have been reported from many studies in research animals (Giguère and Tessman, 2011; Villarino et al., 2013), this fluid is difficult to sample routinely in clinical cases. Therefore, the plasma drug concentration has been examined as a surrogate marker for efficacy from administration of macrolides and their derivatives. The parameter that is best suited to predict efficacy is the AUC of the plasma drug concentration to MIC (AUC/MIC) ratio (Drusano, 2005; Toutain et al., 2017). It has been suggested that the high concentrations in inflammatory cells deliver high concentrations to infected site and this effect is responsible for the efficacy in infected tissues. However, as summarized in the review by Villarino et al. (2013), citing studies by their laboratory and others, the concentrations contained in these cells are not likely high enough to contribute significantly to the PK/PD properties of the macrolides. This view was supported by the analysis by Toutain and associates (Toutain et al., 2017).

Table 36.6 Pharmacokinetics of macrolide derivatives, including azalides, in animals

				Volume of	Peak
		Dose	Half-life	distribution	Concentration
Drug	Species	(mg/kg)	(hour)^a	(Vd) (l/kg)^b	(C_max) (µg/ml)	Reference
Azithromycin	Foals	10 IV	20.3	18.6	–	Jacks et al., 2003
	Foals	10 oral	44 (MRT)	–	0.57
	Foals	5 IV	16	12.4	–	Davis et al., 2002
	Foals	10 oral	18.32	–	0.72
	Dogs	20 oral	35	–	4.2	Shepard and Falkner, 1990
	Dogs	20 IV	29	12	–
	Cats	5 IV	35	23	–	Hunter et al., 1995
	Cats	10 oral	30 (MRT)	–	0.97
Clarithromycin	Foals	10 oral	4.81	–	0.92	Jacks et al., 2002
	Foals	7.5 IV	5.4	10.4	–	Womble et al., 2006
	Foals	7.5 oral	7.1 (MRT)	–	0.52
	Foals^c	7.5 oral	6.11	–	0.614	Peters et al., 2011
	Foals^c	7.5 oral	7.17	–	0.61	Peters et al., 2012
	Foals^c	7.5 oral	5.62	–	0.27	Berlin et al., 2016
	Foals^c	7.5 IV	5.91	–	1.71	Berlin et al., 2016
	Dogs	10 IV	3.9	1.4	–	Vilmànyi et al., 1996
	Dogs	10 (oral tablet)	4.6-5.9	–	3.3-3.5
Gamithromycin	Calves	6 SC	62	–	0.43	Giguére et al., 2011
	Calves	6 SC	50.8	24.9	0.75	Huang et al., 2010
	Feeder cattle	6 SC	52.8 (MRT)	97.4 (V/F)	0.13	DeDonder et al., 2016
	Foals	6 IM	39.1	–	0.33	Berghaus et al., 2012
	Sheep	6 SC	34.5	35.5 (V/F)	0.573	Kellermann et al., 2014
Tildipirosin	Calves	4 SC	210	–	0.71	Menge et al., 2012
	Calves	6 IV	204	49.4	0.64
	Pigs	4 IM	106	–	0.895	Rose et al., 2013
Tulathromycin	Calves	2.5 SC	81.24	–	1.82	Foster et al., 2016
	Calves	2.5	79.5	11	0.39	Villarino et al., 2013^d
	Pigs	2.5	73.95	28.9	0.75
	Horses	2.5	122	–	0.57
	Goats	2.5	76.7	29.3	0.94
	Goats	2.5 SC	45.7	7.0 (V/F)	1.0	Romanet et al., 2012
	Sheep	2.5 SC	118.4	–	3.6	Washburn et al., 2014
Tilmicosin	Calves	20 SC	33.34	5.5 (V/F)	3.48	Foster et al., 2016
	Dairy cow	10 IV bolus	0.76	2.14	–	Ziv et al., 1995
	Dairy cow	10 SC	4.18	–	0.13
	Beef cattle	10 IV	28	28.2	1.56	Lombardi et al., 2011
	Beef cattle, light	10 SC	31.15	–	0.71
	Beef cattle, light	20 SC	31.13	–	1.06
	Beef cattle, heavy	10 SC	30.83	–	0.55
	Beef cattle, heavy	20 SC	30.98	–	1.07
	Pigs	20 oral	25.3	–	1.19	Shen et al., 2005
	Pigs	40 oral	20.7	–	2.03

^aFor some studies, half-life was not reported and mean residence time (MRT) is listed in the table.

^bFor some studies, the volume of distribution was from a nonintravenous route and is listed as Vd/F.

^cData listed for clarithromycin by Peters et al., (2011, 2012), and Berlin et al., (2016) is without coadministration of rifampin. Administration of rifampin with clarithromycin lowers the concentration by 70% to over 90%. These values are provided in detail in those papers.

^dData referenced for Villarino et al. (2013) are an average of multiple studies reported in their paper.

The magnitude of the AUC/MIC target has emerged from laboratory animal studies and analysis of clinical results. The azithromycin free serum AUC/ MIC for a 24-hour interval (AUC₂₄) ratio of >25 has been suggested from a mouse thigh infection model reported by Craig et al. (2002). However, this ratio is likely lower in nonneutropenic animals and Rodvold et al. (2003) suggested plasma AUC₂₄/ MIC ratios of at least 10 in nonneutropenic hosts with pneumonia, and higher AUC₂₄/ MIC ratios of 25–30 for worst case scenarios with experimental neutropenia. The study by Sevillano et al. (2006) showed that a serum azithromycin AUC₂₄/MIC ratio of approximately 27 was adequate for sustained bactericidal activity against susceptible strains. The analysis by Toutain and colleagues cited earlier supported a AUC₂₄/MIC value of approximately 24 for tulathromycin treatment of pneumonia in calves (Toutain et al., 2017).

Because most of the newer macrolides have long half-lives and produce concentrations for much longer than 24 hours, some investigators have considered the AUC values for the duration of treatment, rather than limited to 24 hours. In the study by Muto et al. (2011), using this approach, the AUC/MIC ratio >5 for azithromycin was associated with successful clinical outcome. If one examines the pharmacokinetics from studies of the long-acting macrolides listed in Table 36.6, AUC/MIC ratios of 5–10 for gamithromycin, azithromycin, and tildipirosin have been associated with clinical success. Tildipirosin, which has a longer half-life, produces a ratio of approximately 24. The study by DeDonder et al. (2016) showed that for gamithromycin administration to feeder cattle with bovine respiratory disease associated with Mannheimia haemolytica or Pasteurella multocida, the AUC infinity/MIC ratio associated with clinical success in these cases was 3.49 (Mannheimia haemolytica) and 3.21 (Pasteurella multocida).

Immunomodulatory Effects

Virulence properties of some bacteria may be inhibited by macrolides at concentrations that are less than the MIC required for inhibition or killing. This property, along with the effects on immunomodulation described in more detail below, may explain many of the benefits of macrolides for treating pneumonia (Kovaleva et al., 2012).

The macrolides, particularly the ones that concentrate in immune cells (Figure 36.3) have multiple immunomodulatory effects that likely contribute to the therapeutic response in respiratory infections, and perhaps other diseases. Beneficial effects may be produced by enhanced degranulation and apoptosis of neutrophils and inhibition of inflammatory cytokine production. Enhanced macrophage functions may also may help clear infections faster. These properties have been studied for azithromycin (Parnham et al., 2014) and for the veterinary drugs tilmicosin and tulathromycin (Chin et al., 2000; Duquette et al., 2015). As these reviews and studies point out, there is likely an immunomodulatory effect of these agents that contributes to the therapeutic benefits that is independent of the direct effect on bacteria. These drugs have been known to produce therapeutic benefits in patients even when the bacteria have MIC values in the range that is considered resistant, and above achievable concentrations in plasma or the epithelial lining fluid of the respiratory tract. The authors of these studies are careful to point out that the effect of macrolides is best termed immunomodulatory rather than immunosuppressive, which implies that it may modify or regulate functions of the immune system without impairing normal responses to combat bacterial infection (Kanoh and Rubin, 2010). According to Kanoh and Rubin (2010) the 14- and 15- membered macrolides exert these immunomodulatory effects, but not 16-membered macrolides. However, tilmicosin, a 16-membered ring macrolide (Figure 36.3, Table 36.4 ) also exhibits some of these properties (Chin et al., 2000).

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Tags: Veterinary Pharmacology and Therapeutics

Feb 8, 2018 | Posted by admin in PHARMACOLOGY, TOXICOLOGY & THERAPEUTICS | Comments Off

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