Metronidazole

Metronidazole was first synthesized in the 1950s when the pharmaceutical company Rhône-Poulenc was searching for an effective anti­trichomonal drug for the treatment of vaginal trichomoniasis.¹ Initially, a crude extract of a Streptomyces bacterium was found to kill Trichomonas vaginalis and the active component was determined to be azomycin, a previously characterized nitroimidazole antibiotic.² Metronidazole was a synthetic analogue of azomycin that was both more active against T. vaginalis and less toxic.^1,3 It was initially called 8823 R.P.¹ This discovery soon led to its successful use in human clinical studies of trichomoniasis.^4,5 The presumed antibacterial activity of metronidazole was discovered incidentally by Shinn in 1962, who reported improvement in ulcerative gingivitis in a patient treated for a concomitant T. vaginalis infection, a result he confirmed in a series of patients with ulcerative stomatitis.⁶

Mechanism of Action

Metronidazole and other nitroimidazoles (e.g., nimorazole, ornidazole, ronidazole, secnidazole, tinidazole) are inert, and their spectrum of antimicrobial activity is determined by the capacity of susceptible organisms to activate the drugs once they enter the cell via passive diffusion. The structures of metronidazole and related compounds are illustrated in Figure 28-1. Their bactericidal and parasiticidal activities are rapid and proportional to the concentration of the activated drugs within the target cell.⁷ The members of this class of antibiotics are therefore best considered as prodrugs, which are activated through a reduction step to form highly reactive products that interact with intracellular targets (see later).⁸ Metronidazole's mechanism of action describes that of the other nitroimidazoles.

The nitroimidazoles share a heterocyclic structure consisting of an imidazole-based nucleus with a nitro group, NO₂, in position 5. There are four major steps involved in the mechanism of action of metronidazole that result in the intracellular formation of critical redox intermediate metabolites.⁹ In the first two steps, the drug enters cells by passive diffusion and an electron is transferred to the nitro group of metronidazole, resulting in the production of a short-lived nitroso free radical, which is cytotoxic and can interact with cellular DNA.¹⁰ This process of activating the prodrug creates a concentration gradient that augments the increased uptake of the drug by the organism, further increasing its antimicrobial effect. The third step in metronidazole's action relates to the cytotoxic effect of the reduced product because the activated metronidazole compound can inhibit DNA synthesis and induce DNA damage via oxidation, resulting in single-strand and double-strand breaks.¹⁰ Thus, metronidazole induces DNA degradation and cell death.¹⁰ Finally, there is the release of inactive end products of the drug.¹¹

The microbial selectivity of metronidazole reflects the inability of aerobic bacteria to activate the prodrug because they lack the necessary electron transport proteins with sufficient negative redox potential.¹⁰ However, in susceptible anaerobic bacteria, the redox potential of components of the electron transport chain is sufficiently negative to reduce the nitro group of metronidazole. The drug is activated in anaerobic bacteria when it receives an electron from ferredoxin or flavodoxin, which are themselves reduced by iron-sulfur proteins called pyruvate:ferredoxin oxidoreductases (PFORs).¹² The exact electron donors involved in nitroimidazole reduction vary, depending on the organism.¹⁰ In the microaerophile Helicobacter pylori, for example, a separate mechanism appears to be involved in metronidazole susceptibility, involving a 2-electron transfer step mediated by an oxygen-insensitive nitroreductase (RdxA).¹⁰ Several microaerophilic protists (Giardia lamblia, Entamoeba histolytica, and T. vaginalis) have bacteria-like enzymes (nitroreductases) capable of activating metronidazole.¹³

Spectrum of Activity

Metronidazole and related nitroimidazoles are active against a variety of anaerobic bacteria, as well as microaerophilic bacteria and protozoa. Resistance, as detailed later, has been increasingly detected in certain organisms, though this may not be identified easily because sensitivity testing for anaerobes is not performed routinely. However, the emergence of resistance suggests that ongoing surveillance is important.¹⁰

Many gram-negative anaerobes are susceptible to metronidazole.^14,15 As a rule, members of the genera Bacteroides and Parabacteroides are susceptible to metronidazole, with resistance generally detected in less than 5% of isolates.^16-19 Higher rates of resistance have been reported among Bacteroides isolates in South Africa.²⁰ Desulfovibrio species are also highly susceptible,²¹ and clinically relevant members of the Fusobacterium, Porphyromonas, Prevotella, and Bilophila genera are usually sensitive.^14,22 However, nonsusceptible Prevotella strains have been described.¹⁴ Recently, metronidazole resistance has been reported in oral isolates of Porphyromonas gingivalis.²³ Reduced susceptibility has also been noted in isolates of Sutterella¹⁴ and in the gram-negative coccus Veillonella.^15,24 The bacterial vaginitis-associated bacteria of the genus Mobiluncus are usually not susceptible to metronidazole.^25,26

Facultative anaerobes have variable susceptibility to metronidazole and are generally not empirically treated with this agent. Aggregatibacter actinomycetemcomitans is occasionally susceptible, but resistance is sufficiently common that empiric metronidazole should not be used for these infections.^23,27 Another oral, facultative anaerobe, Eikenella corrodens, is generally resistant to the nitroimidazoles.²⁸ The CO₂-requiring members of the genus Capnocytophaga are generally resistant to metronidazole.²⁹ Although Campylobacter jejuni and Campylobacter coli isolates may be susceptible in vitro to metronidazole, the drug is not recommended for therapy against these pathogens.³⁰ Gardnerella vaginalis, which is associated with bacterial vaginosis, is variable in its sensitivity, with nearly 30% of isolates in one series demonstrating resistance, suggesting that resistance should be considered in cases of treatment failure.³¹ A more recent survey from India documented nearly 70% of G. vaginalis strains were metronidazole resistant, but the cases included metronidazole-exposed women exhibiting recurrent infection.³² The clinical relevance of in vitro resistance of Gardnerella to metronidazole is difficult to interpret because a hydroxy metabolite of metronidazole is actually more active against G. vaginalis than the parent compound.^33,34

H. pylori is a facultative anaerobe that was initially sensitive to metronidazole but has increasingly developed clinically important resistance. A recent survey of 10,670 clinical isolates obtained from H. pylori–treated and H. pylori–untreated individuals revealed metronidazole resistance in 17.4% of isolates from previously untreated children and 26.1% of strains from previously untreated adults. Resistance rates in strains obtained from antibiotic-exposed children and adults were higher, at 37.7% and 49.0%, respectively.³⁵ Treatment failure for H. pylori was a significant risk factor for harboring a metronidazole-resistant isolate.³⁵

Among the gram-positive anaerobes, the clostridia remain quite susceptible to metronidazole.³⁶ However, reduced susceptibility to metronidazole has been noted among a limited number of Clostridium difficile isolates.^37-39 Notably, the Etest overestimates metronidazole susceptibility in C. difficile isolates, compared with the more involved agar-incorporation-based methods for determining minimal inhibitory concentrations (MICs).^37,39 Whether the clinical response to metronidazole is affected by reduced in vitro susceptibility remains to be determined. However, this deserves closer attention in light of the highly variable concentrations of metronidazole in the stools of treated patients^40,41 and increasing reports of treatment failure and recurrence of C. difficile infection (CDI) in metronidazole-treated patients.⁴²

An important hole in the anaerobic spectrum of metronidazole is found in its lack of activity against a number of non–spore-forming, gram-positive anaerobic bacteria that possess intrinsic resistance to the drug. These include isolates of Actinomyces, Bifidobacterium, Lactobacillus, and Propionibacterium.^10,36,43 Propionibacterium acnes is highly resistant.^15,36,44 Metronidazole should not be routinely used to treat infections with these organisms unless susceptibility is confirmed. In contrast, the genus Eubacterium is generally sensitive to metronidazole in vitro.³⁶

The nitroimidazoles possess good activity against several protozoa. Apart from T. vaginalis, metronidazole has activity against Giardia (syn. G. duodenalis, G. lamblia, G. intestinalis) and Entamoeba histolytica. Resistance is uncommon in Giardia, and clinical efficacy is generally greater than 90%, but in vitro testing has revealed reduced susceptibility to metronidazole in clinical isolates, causing concern.^45,46 Nitroimidazoles exhibit in vitro activity against Dientamoeba fragilis, which is a trichomonad known to cause gastroenteritis.⁴⁷

Apart from its antimicrobial actions, metronidazole exhibits immunosuppressive and anti-inflammatory actions⁴⁸ and has been used effectively in the treatment of rosacea,⁴⁹ although the extent to which this relates to metronidazole's antibacterial properties is unclear.

Effects on the Human Microbiome

Metronidazole, like many antimicrobials, affects the human microbiome, although because this has mostly been examined in subjects exposed to metronidazole in combination with other antimicrobials, it is difficult to fully understand the impact of metronidazole itself.^10,50 Early culture-based studies of the impact of metronidazole monotherapy on bacterial communities in the human gastrointestinal tract described little impact of the drug on microbial populations.^41,51 However, some investigators reported a suppression of anaerobes and a relative increase in the abundance of certain aerobic bacteria (Escherichia coli and fecal streptococci).⁵² The reasons for the relatively low impact on normal gut microbes are not well understood but may relate to the pharmacology of metronidazole, which achieves low concentrations in the feces of healthy adults.^40,53 At present there are few studies of metronidazole's impact on the gastrointestinal microbiome that have used culture-independent techniques such as DNA pyrosequencing, apart from studies involving antibiotic combinations.⁵⁰ This is an area in need of new research.

Nucleic acid sequencing methods have been applied to understand the impact of metronidazole on the microbiome of the female reproductive tract, particularly in the context of bacterial vaginosis. Topical metronidazole has recently been evaluated for its impact on the vaginal microbiome in women with bacterial vaginosis.^54-56 Five to seven days of metronidazole consistently reduced the diversity of bacterial communities in these studies of bacterial vaginosis, compared with no treatment, and for many women it restored a more normal, Lactobacillus-dominated mucosal microbiome.^54-56

Pharmacology

Metronidazole is commercially available in a variety of formulations: oral capsules and tablets (immediate and extended release); intravenous solution; topical gels, creams, and lotions; and vaginal gels.^7,57 Although oral metronidazole suspension is not commercially available, it is commonly compounded in pharmacies by crushing immediate release tablets and mixing with a 1 : 1 ratio of an aqueous suspending solution and buffered oral syrup.⁵⁸ The dose and duration of metronidazole is dependent on the specific product and indication (Table 28-1). An IV loading dose of 15 mg/kg, followed by 7.5 mg/kg every 6 to 8 hours, is recommended in the package insert, with a maximum daily dose limit of 4 g.⁷ A fixed dose of 500 mg IV every 8 hours maintains concentrations above typical MICs for Bacteroides species and is effective for treatment of intra-abdominal infections.^59-61 An infusion time of 1 hour is traditionally recommended, but 20- to 30-minute infusions have been used.⁶² Given the long half-life and concentration-dependent activity, high-dose metronidazole, administered as 1 to 1.5 g every 24 hours, may be a safe and effective alternative to 500 mg every 6 to 8 hours.^63,64 The typical duration of oral or intravenous metronidazole courses ranges from 1 to 10 days depending on the indication and patient condition. Longer durations may be prescribed, but caution should be exercised with durations greater than 1 month due to increased risk of peripheral neuropathy and central nervous system adverse effects.^65-67

Oral metronidazole is rapidly and almost completely absorbed, with bioavailability approaching 100%.^7,57 When rectally administered, metronidazole is also well absorbed with reported bioavailability of 59% to 94%; topical and vaginal metronidazole achieves detectable systemic concentrations with bioavailability ranging from 2% to 25%.^7,57,68 Administration of oral metronidazole with food is encouraged to minimize gastrointestinal adverse effects and does not affect bioavailability but may delay the time to peak serum concentrations. Peak serum concentrations range from 12 to 40 µg/mL and occur 1 to 2 hours after oral administration and approximately 3 hours after rectal administration.^7,57

Metronidazole is a lipophilic molecule with low protein binding and moderate to large volume of distribution, allowing extensive distribution into various tissues (Table 28-2).^7,57 Penetration into inflamed cerebrospinal fluid, epithelial lining fluid, saliva, and bile is excellent and concentrations are similar to serum.^7,57,69 Patients with noninflamed meninges still achieve therapeutic concentrations at approximately 43% of serum.⁷⁰ Additionally, penetration into abscesses, appendix tissue, peritoneal fluid, and pancreatic tissue is very good, ranging from 2.3 to 7.2 µg/mL.^7,57,71 However, patients with obstructive cholecystitis have negligible amounts of drug detected in the bile.^7,57 Metronidazole crosses the placental barrier and penetrates into breast milk and may be teratogenic during the first trimester (see “Precautions”).⁷² Stool concentrations during C. difficile colitis are highest at the beginning of infection and taper as inflammation subsides and stool is formed, but concentrations generally remain well above reported MICs.⁴⁰ This effect of higher stool concentrations when diarrhea is present is also noted during flares of Crohn's disease.⁴¹

Metronidazole undergoes oxidation as the primary step in eliminating the drug from the body, and 6% to 18% of active unchanged drug is found in the urine.^7,57,73 Oxidation, glucuronidation, and metabolism by cytochrome P-450 system yield five major metabolites, including 1-2 hydroxyethyl-2-hydroxy-methyl-5-nitroimedazole (hydroxy metabolite), which maintains antimicrobial activity; 2-methyl-5-nitroimidazole-1-acetic acid (acetic acid metabolite) is another major metabolite but has no antimicrobial activity.^7,57,73 All metabolites are extensively excreted in the feces or urine.⁷ The half-life of metronidazole is approximately 8 hours in healthy patients and 18 to 20 hours with end-stage hepatic failure.⁷⁴ Patients with moderate-to-severe hepatic diseases should receive a 50% dose reduction.⁷ Additionally, patients with end-stage renal disease (creatinine clearance <10 mL/min) will have slightly longer half-life of metronidazole and significantly impaired clearance and accumulation of the hydroxy and acetic metabolites.^7,75-81 Metronidazole and metabolites are removed by conventional, continuous, and peritoneal hemodialysis. During a 4-hour conventional hemodialysis session, 45% of drug is removed, and patients should receive a supplemental dose post dialysis. Patients receiving continuous renal replacement therapy also experience significant removal of metronidazole and its metabolites and do not require dose adjustment.⁷⁴ Peritoneal dialysis removes approximately 10% of drug.⁸² Patients on peritoneal dialysis and those with creatinine clearance less than 10 mL/min not receiving conventional or continuous hemodialysis will have accumulation of active metabolites.^{74-76,78,79,81} Renal dose adjustment is currently not recommended because the ramifications of metabolite accumulation are not well understood, but caution should be used in long-term therapy.⁵⁷ Finally, preterm infants 32 weeks' gestational age or younger will have impaired clearance and may require dose adjustment depending on chronologic age.^83,84

Metronidazole is associated with carcinogenic activity in rats and mice. It should be avoided during the first trimester of pregnancy and used during the second and third trimester only if clearly necessary.^7,57 There are case reports of fetal malformation occurring with metronidazole exposure during pregnancy, but three large studies failed to demonstrate a link of fetal malformations compared with the general population.^72,85,86 Metronidazole is also excreted in breast milk⁸⁷ and has been shown to achieve infant plasma concentrations approximately one fifth of those observed in the mother's plasma.⁸⁸ It has been recommended to withhold nursing during metronidazole therapy for 12 to 24 hours following oral single-dose regimens.⁸⁸

Whenever possible, metronidazole should be avoided during lactation because of its effects on the developing microbiota. Infants may have increased risk of diarrhea, but there is a lack of evidence linking breast milk with carcinogenic effectors or developmental disorders.⁸⁸

Mechanisms of Resistance

Resistance to metronidazole (and other nitroimidazoles) in strict anaerobes remains unusual, and not all mechanisms that reduce susceptibility to the nitroimidazoles have been characterized.¹⁰ On the basis of critical steps in its mechanism of action, several models of drug resistance have been proposed, including reduced antibiotic uptake, active drug efflux, reduced drug activation (e.g., by decreased expression of activating nitroreductase enzymes), drug inactivation (e.g., nim-encoded nitroimidazole reductase), and altered DNA repair.¹⁰

Metronidazole resistance (MIC ≥32 µg/mL) among Bacteroides strains is uncommon, generally occurring in less than 5% of isolates.^15,16 Although many mechanisms of resistance have been induced in vitro,⁹⁰ the best-characterized mechanism in clinical isolates is encoded by the nim (5-nitroimidazole reductase) genes (nimA–F).¹⁹ The nim genes induce the reduction of the nitrate residue of metronidazole (and related compounds) into an inert amino derivate without any toxicity for the bacterial chromosome.¹⁶ These genes may occur in all Bacteroides species and are either located on plasmids or on the chromosome.¹⁹ In a recent survey of 640 Bacteroides isolates obtained from across Europe, 21 strains (3.3%) had MIC values greater than or equal to 4 µg/mL. Notably, of only three isolates (two Bacteroides fragilis strains and one Bacteroides thetaiotaomicron isolate) harboring nim genes, one was resistant to metronidazole.¹⁹ The two metronidazole-susceptible B. fragilis isolates had chromosomal nimA and nimC genes, while a nimE gene was identified on a plasmid in the resistant B. thetaiotaomicron strain.¹⁹ It was speculated that a small number of nim-negative, but metronidazole-resistant, Bacteroides strains identified in this study might have used diverse resistance strategies, including reduced uptake, nitroreductase and/or PFOR activities, increased lactate dehydrogenase activity, or mutations that alter the carbohydrate utilization affecting the redox state.¹⁹ Recent studies suggest that alterations in the recA gene encoding the RecA protein involved in DNA damage repair might be a resistance strategy in Bacteroides.⁹¹

The high prevalence of metronidazole resistance among H. pylori strains appears to be due primarily to decreased activation of the drug, which would also be expected to affect its accumulation within the bacterium. Mutations in the oxygen-insensitive reduced nicotinamide adenine dinucleotide phosphate (NADPH) nitroreductase RdxA or the NADPH-flavin-oxidoreductase FrxA confer resistance to metronidazole in H. pylori.⁹² H. pylori might employ other strategies to protect itself against nitroimidazole-induced damage, but its relevance in vivo remains speculative.^93,94

Resistance to metronidazole has emerged in protozoal pathogens. Trichomonal resistance was noted as early as 1962 in clinical isolates,⁹⁵ but prevalence has generally remained below 10%.⁹⁶ A recent survey from six U.S. cities tested 538 T. vaginalis isolates for nitroimidazole resistance (aerobic minimum lethal concentration [MLC] > 50 µg/mL) and found that 23 (4.3%) exhibited low-level in vitro metronidazole resistance (MLC 50 to 100 µg/mL).⁹⁶ However, there were no isolates identified with moderate- to high-level nitroimidazole resistance.⁹⁶ An unsettling report from New Guinea revealed 17.4% of T. vaginalis strains (4 out of 23 isolates) exhibited metronidazole resistance under aerobic conditions (MIC > 200 µM), but the number of isolates tested was relatively small and the organisms were sensitive under anaerobic conditions.⁹⁷

The mechanisms of resistance in T. vaginalis appear to differ between laboratory-generated nitroimidazole resistance and those found in clinical isolates.⁹⁸ Laboratory-induced resistance manifests in anaerobic conditions and results from a loss of the aforementioned drug-activating pathways that reduce the inert prodrug metronidazole to active metabolites.⁹⁸ Clinical resistance, on the other hand, typically occurs under aerobic conditions due to actions of oxygen itself.⁹⁸ For example, a study of resistance in clinical strains of T. vaginalis found that flavin reductase activity was downregulated, or even absent, in metronidazole-resistant strains.⁹⁸ It was postulated that because flavin reductase can reduce oxygen to hydrogen peroxide, its downregulation might impair oxygen scavenging.⁹⁸ This would result in resistance because oxygen interferes with the activation of nitroimidazoles by either inhibiting drug-activating pathways or by re-oxidizing a critical, toxic, nitroradical anion intermediate, also resulting in reduced metronidazole uptake.⁹⁸

As noted, in Giardia, clinical resistance occurs in approximately 20% of cases.⁴⁶ Microbiologic resistance to metronidazole is complex but appears to be due to a lack of activation of the prodrug to the active nitroso free radical.⁹⁹ Resistance to metronidazole has traditionally been explained by a loss of ferredoxin and PFOR activities.¹⁰⁰ However, recent studies suggest that some metronidazole-resistant G. lamblia strains have normal activity levels of these redox proteins and metronidazole can be activated by a flavin adenine dinucleotide (FAD)-dependent G. lamblia thioredoxin reductase.⁹⁹ Although not entirely clear, drug resistance in those isolates appeared to be related to lower availability of reduced FAD.⁹⁹ Resistance to metronidazole in amoebae has been associated with an increase in iron-containing superoxide dismutase, without a significant decrease of the PFOR activity.¹⁰⁰

Drug Interactions and Interference with Laboratory Tests

The major interactions between metronidazole and other pharmaceuticals or food are listed in Table 28-3. An important and serious interaction exists between warfarin and metronidazole, as metronidazole increases the blood levels and hypothrombotic effects of warfarin through inhibition of enzymes responsible for oxygenation of S-warfarin.¹²³ Preemptive dose reduction of warfarin and close monitoring of prothrombin activity have been recommended if the two drugs require concomitant administration.¹²⁴ An uncommonly reported interaction suggests that metronidazole reduces the clearance of the alkylating chemotherapy agent busulfan, increasing the levels of the latter drug.¹²⁵ Metronidazole therapy should be avoided with concomitant busulfan whenever possible.

Several case reports have proposed that metronidazole use can increase the systemic concentration of concomitant CYP3A substrates, including amiodarone, carbamazepine, quinidine, tacrolimus, and cyclosporine.¹²⁶ However, in vitro studies have not consistently supported CYP3A inhibition as a mechanism of metronidazole drug interaction.¹²⁶ Empiric dose adjustments are not required for potential CYP3A interactions, but increased monitoring and patient education are prudent.

Although it is not commonly thought of as a drug that poses a risk for inducing QT interval prolongation or arrhythmias, recent reports have linked metronidazole to long QT and torsades de pointes.^127,128 In most cases, this appeared to be due to metronidazole impairing the cytochrome P450 metabolism of other agents that were responsible for lengthening the QT interval.¹²⁷ Anecdotal reports suggest that metronidazole itself can prolong the QT interval, but this is likely rare.¹²⁸

Metronidazole is largely believed to cause a disulfiram-like effect on ethanol metabolism, leading to symptoms such as severe nausea and vomiting.¹²⁹ Adverse effects similar to disulfiram-like reactions have been reported with topical metronidazole administration, including vaginal, so ethanol should be avoided while on therapy.¹²⁹ However, there is controversy about the actual risk for and nature of ethanol-metronidazole interactions.^129,130 Although disulfiram inhibits hepatic aldehyde dehydrogenase, resulting in the accumulation of blood acetaldehyde concentrations following ethanol consumption, metronidazole has not been demonstrated to share this ability.¹²⁹ Thus, although metronidazole is associated with disulfiram-like symptoms when administered with ethanol, the mechanism is poorly defined and the incidence is not well understood.¹²⁹

Other Nitroimidazole Antimicrobials

Tinidazole, secnidazole, and ornidazole are other members of the 5-nitroimidazole class. Trindiazole, which has been widely prescribed in Europe and developing countries, was approved for used in the United States in 2004. All agents in the class exhibit similar mechanism of action, spectrum activity, toxicity, and adverse effects.¹³¹ However, the distinguishing feature among agents is the half-life and need for less frequent dosing compared with metronidazole.¹³¹ The half-life for tinidazole, secnidazole, and ornidazole is 10 to 15 hours, 17 to 28.8 hours, and 11 to 14 hours, respectively, which allows for once-daily dose (see Table 28-1).¹³¹ These agents offer a potential advantage over metronidazole, as a single-dose option for the treatment of intestinal amebiasis, giardiasis, and bacterial vaginosis. However, metronidazole should be considered the drug of choice for life-threatening anaerobic infections because there are limited data evaluating the efficacy and safety of other nitromidazole agents.^132,133