Dihydropteroate synthase gene mutations in Pneumocystis and sulfa resistance

Emerging Infectious Diseases, Oct, 2004 by Laurence Huang, Kristina Crothers, Chiara Atzori, Thomas Benfield, Robert Miller, Meja Rabodonirina, Jannik Helweg-Larsen

Pneumocystis pneumonia (PCP) remains a major cause of illness and death in HIV-infected persons. Sulfa drugs, trimethoprim-sulfamethoxazole (TMP-SMX), and dapsone are mainstays of PCP treatment and prophylaxis. While prophylaxis has reduced the incidence of PCP, its use has raised concerns about development of resistant organisms. The inability to culture human Pneumocystis, Pneumocystis jirovecii, in a standardized culture system prevents routine susceptibility testing and detection of drug resistance. In other microorganisms, sulfa drug resistance has resulted from specific point mutations in the dihydropteroate synthase (DHPS) gene. Similar mutations have been observed in P. jirovecii. Studies have consistently demonstrated a significant association between the use of sulfa drugs for PCP prophylaxis and DHPS gene mutations. Whether these mutations confer resistance to TMPSMX or dapsone plus trimethoprim for PCP treatment remains unclear. We review studies of DHPS mutations in P. jirovecii and summarize the evidence for resistance to sulfamethoxazole and dapsone.

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Although decreasing in incidence as a result of combination antiretroviral therapy and effective prophylaxis, Pneumocystis pneumonia (PCP), caused by Pneumocystis jirovecii (formerly P. carinii f. sp. hominis), remains the most common AIDS-defining opportunistic infection, as well as the most frequent serious opportunistic infection in HIV-infected persons, in the United States and Europe. Despite the fact that this infection can be prevented, certain patients continue to be at increased risk for PCR Specifically, PCP frequently signals H1V infection in patients not previously known to be HIV-infected (1). Patients who are not receiving regular medical care, as well as those who are not receiving or responding to antiretroviral therapy or prophylaxis, are also at increased risk for PCP (2). PCP may also develop in other immunosuppressed populations, such as cancer patients and transplant recipients. Furthermore, PCP remains a leading cause of death among critically ill patients, despite advances in treatment and management (3).

The first-line treatment and prophylaxis regimen for PCP is trimethoprim-sulfamethoxazole (TMP-SMX) (4). While prophylaxis has been shown to reduce the incidence of PCP, the widespread and long-term use of TMP-SMX in HIV patients has raised concerns regarding the development of resistant organisms. Even short-term exposure to TMP-SMX can be associated with the emergence of TMPSMX resistance, as has been demonstrated in patients with acute cystitis caused by Escherichia coli (5). Indeed, an increased number of sulfa-resistant bacteria have been isolated in HIV patients, which coincides with the rise in TMP-SMX prophylaxis for PCP (6,7). In one study, the prevalence of TMP-SMX-resistant Staphylococcus aureus and Enterobacteriaceae species isolated in all hospitalized patients increased significantly from <5.5% of isolates before 1986 to 20% in 1995, during which time TMPSMX prophylaxis was increasing in HIV-infected patients (6). In addition, the rise in resistant organisms was significantly more prominent in samples obtained from HIV-infected patients, in whom resistant isolates increased from 6.3% in 1988 to 53% in 1995. Another study found that significantly more TMP-SMX-resistant organisms were isolated from HIV-infected patients who had received TMP-SMX than from patients who had not received TMPSMX (7).

Given the emergence of resistance to TMP-SMX among many bacteria (8), concern has focused on the potential development of resistant Pneumocystis. Based on animal studies, nearly all of the anti-Pneumocystis activity of TMP-SMX is due to sulfamethoxazole (9). The development of sulfonamide resistance could result in the failure of sulfamethoxazole as well as dapsone, a sulfone antimicrobial agent also used in the treatment and prophylaxis of PCP. While separate lines of investigation also suggest that Pneumocystis may be developing resistance to atovaquone, a second-line PCP treatment and prophylaxis regimen (10), we concentrate our review on the evidence for the development of sulfonamide-resistant Pneumocystis.

Mechanisms of Sulfonamide Resistance

Sulfonamides act by interfering with folate synthesis. Since many microorganisms cannot transport folate into cells as mammalian cells can, most prokaryotes and lower eukaryotes must synthesize folates de novo (11). Sulfonamides inhibit one of the integral enzymes in folate synthesis, dihydropteroate synthase (DHPS), which catalyzes the condensation of para-aminobenzoic acid and pteridine to form dihydropteroic acid (Figure). Since mammalian cells lack DHPS, sulfonamides can selectively inhibit the growth of various microorganisms. Trimethoprim, part of the fixed combination TMP-SMX, inhibits another of the integral enzymes, the dihydrofolate reductase (DHFR).

Resistance to sulfonamides can emerge by means of a number of mechanisms (12). In most gram-negative enteric bacteria, sulfonamide resistance is largely plasmidborne and related to drug-resistant DHPS variants with substantial sequence divergence (12). Chromosomal mutations in the DHPS locus--such as point mutations, insertions of duplicate amino acids, or larger sequence alterations as a result of recombination--can also lead to resistance (8). In some organisms, several different mechanisms of resistance have been identified in different strains. For example, some strains of Neisseria meningitidis have acquired a DHPS gene with 10% sequence divergence, postulated by others to be due to recombination (12), whereas other Neisseria strains have acquired a chromosomal insertion, resulting in the addition of two amino acids to DHPS (13). In other organisms, such as E. coil and Plasmodiumfalciparum, nonsynonymous point mutations resulting in amino acid substitutions in DHPS can confer sulfa resistance (14,15). Furthermore, the accumulation of additional mutations over time can confer increasing levels of sulfa resistance, as has occurred in P. falciparum (16).