{"id":487,"date":"2013-11-11T19:01:59","date_gmt":"2013-11-12T00:01:59","guid":{"rendered":"http:\/\/www.axopub.com\/wp01\/?p=487"},"modified":"2013-11-11T19:01:59","modified_gmt":"2013-11-12T00:01:59","slug":"antifungal-resistance-is-not-always-great-for-the-fungus","status":"publish","type":"post","link":"https:\/\/www.axopub.com\/wp01\/2013\/11\/11\/antifungal-resistance-is-not-always-great-for-the-fungus\/","title":{"rendered":"Antifungal resistance is not always great for the fungus"},"content":{"rendered":"

The paper<\/em><\/strong>: Vincent BM, et al. (2013) Fitness trade-offs restrict the evolution of resistance to amphotericin B. PLoS Biol<\/em> 11(10): e1001692. doi:10.1371\/journal.pbio.1001692<\/p>\n

Subject areas<\/strong><\/em>: microbiology, evolution<\/p>\n

Vocabulary<\/strong><\/em>:<\/p>\n

thrush (oral thrush) – a condition in which the fungus Candida albicans infects the mouth and throat. \u00a0It presents as velvety white lesions or sores on the tongue and inner cheeks, but can spread down the throat and to the tonsils. \u00a0It can be painful and the lesions may bleed easily. \u00a0Candida is often present in the mouth without pathogenicity, as its presence is bounded by other microorganisms in the mouth and the immune system. \u00a0However, if that balance is upset due to illness or medication (e.g. antibiotics may kill the bacteria in your mouth that are preventing the Candida from proliferating), Candida can become pathogenic.<\/p>\n

—–
\nThis article is a summary of a recent primary research paper intended for high school teachers to add to their general knowledge of current biology, or to supplement their lessons by showing students the kinds of projects that current biological research addresses.
\n—–<\/p>\n

Here we have a paper that is interesting because the conclusion makes sense and seems intuitive, but at the same time, it probably does not occur to many lay people. The general idea is simple: any new trait that comes up from a mutated gene can simultaneously have positive effects on some parts of the organism’s life, while having negative effects on some other part or parts. What seem to be useful mutations may not take hold in a population if the “side effects” are too costly, especially with respect to reproductive fitness.<\/p>\n

Vincent et al noticed that populations of Candida albicans<\/em>, the most common fungal infection of people, very quickly develops effective resistance to two of the three commonly used classes of antifungal drugs, but not to amphotericin B. They investigated the reasons that this might be the case.<\/p>\n

What they knew<\/strong><\/em><\/p>\n

Candida albicans<\/em> is the most common fungal pathogen infecting human beings. It is also a common commensal organism living on humans without causing harm – pathogenicity usually occurs due to some change in the host, such as weakening of the immune system by another illness. Although some kinds of Candida<\/em> infections can be just minor irritations, it can also become a very serious, life-threatening disease. There are three major classes of drugs used to treat Candida<\/em>: triazoles, ehinocandins, and polyenes.<\/p>\n

Amphotericin B (AmB) is a member of the last group, the polyenes, and is the third choice drug primarily because it can cause cumulative kidney damage. However, AmB does have an important advantage in clinical use, because for some reason despite having been in use for half a century, there are still very few cases of AmB-resistant C. albicans<\/em>. By way of comparison, resistance to triazoles is frequent, and resistance to the newer echinocandin drugs is increasing steadily.<\/p>\n

Amphotericin B works by binding to ergosterol, which is a component of the Candida<\/em> cell membrane. When it does so, it causes a pore to be opened up, allowing ions to leak out of cell, quickly killing it. The mechanism of resistance to AmB is thought to be in mutations to the enzymes that make ergosterol, switching production to different sterols that are not recognized by AmB.<\/p>\n

What they did<\/em><\/strong><\/p>\n

The first order of business was to examine the genome sequence of AmB-resistant Candida<\/em> strains and compare them to AmB-sensitive strains. figure below shows a comparison of sequences from chromosome 2 of a wild-type strain and an AmB-resistant strain isolated from a clinical setting. The colored areas show differences in the sequences, many of which are in the Erg2 gene.<\/p>\n

\"ampho1\"<\/a><\/p>\n

Examining other non-essential genes in the ergosterol biosynthesis pathway, the graph below shows the sensitivity of different strains to increasing concentrations of AmB. The least sensitive\/most resistant strain is the erg3\/erg11 double-knockout, although the individual erg2 knockout and erg6 knockout are roughly as resistant as the clinically isolated AmB-resistant strain. Also shown for comparison at the bottom are wild-type and AmB-resistant strains of a different Candida<\/em> species, C. tropicalis<\/em>, which was also sequenced for comparison. The C. tropicalis<\/em> AmB-resistant strain had a normal Erg2 sequence, but its Erg3 gene is different.<\/p>\n

\"ampho2\"<\/a><\/p>\n

Previous work had shown the involvement for Hsp90 in drug resistance to triazoles and echinocandins. Hsp90 is so named as a member of a family of proteins called “heat shock proteins” because they were first discovered in prokaryotes as proteins that were made to help the organism survive environmental stresses such as acute temperature change. Hsp90 works as a molecular chaperone: often stress conditions cause important proteins to denature, or change their shape, which can inactivate them. Hsp90 binds to proteins like a reinforcement, helping to hold its active, native shape, or to help unfolded proteins fold in the correct orientation.<\/p>\n

Previous studies have shown that partially knocking down Hsp90 function can reverse drug-resistance to triazoles and echinocandins. The researchers asked if Hsp90 is similarly important for amphotericin B resistance. The figure below shows the effect of geldanamycin, an Hsp90 inhibitor, on either fluconazole resistance or amphotericin B resistance in C. albicans<\/em>.<\/p>\n

\"ampho3\"<\/a><\/p>\n

The top panels show growth on normal medium. The middle panels show growth on medium containing the triazole drug, fluconazole. The bottom panels are the same Candida<\/em> cultures, but grown on media containing AmB. The photos on the right side all also contain the Hsp90 inhibitor geldanamycin. Part of this figure is exactly as expected. In the middle rows of the two middle panels, we see that partially inhibiting Hsp90 destroys fluconazole resistance. Also, in the top two rows of the top panels, Hsp90 inhibition does not affect growth of either wild-type or fluconazole-resistant strains in the absence of any antifungal drug. In the bottom row of the bottom panel, they discovered what they expected: Hsp90 inhibition destroys AmB resistance also.<\/p>\n

However, there was also an unexpected finding. Looking at the bottom row of the top panels, what that tells us is that even under non-selective conditions, the AmB-resistant strains are very sensitive to inhibition of Hsp90. Using a structurally different Hsp90 inhibitor, radicicol, the results are the same as with geldanamycin for either the clinically isolated AmB-resistant Candida<\/em>, or the erg2, er6, or erg3\/11 knockouts.<\/p>\n

\"ampho4\"<\/a><\/p>\n

This suggests that for some reason, the stress response is actively working to maintain AmB-resistant cells keeping them alive. Knocking out the Hsp90 gets kills one of the mechanisms keeping the AmB-resistant cells alive, whereas in AmB-sensitive strains, knocking out Hsp90 just knocks out a safety net that is not yet engaged. To test this hypothesis, Vincent and colleagues tested different C. albicans<\/em> strains under a wide variety of environmental stressors. The figure below shows the effects of temperature, oxidizers, nutrient (iron) starvation, and the a peptide that signals the presence of neutrophils (immune system cells that would normally attack Candida<\/em>). There was no difference in the response to the neutrophil peptide, but in all of the other cases, the AmB-resistant cells were significantly more susceptible to these environmental stresses than AmB-sensitive\/wild-type C. albicans<\/em>.<\/p>\n

\"ampho5\"<\/a><\/p>\n

AmB-resistance also seems to negatively affect the ability of C. albicans<\/em> to invade a host. Candida<\/em> normally switches to a filamentous form (figure below) that penetrates more effectively into host tissue than its usual spherical morphology. Normally, exposing the Candida<\/em> to 10% fetal bovine serum at 37 \u00baC initiates the filamentation process. This was delayed in all of the ERG knockout mutants and the clinically isolated AmB-resistant strains, and even when it started in a few strains (erg3 and erg2 knockouts), the filaments were not properly formed.<\/p>\n

\"ampho6\"<\/a><\/p>\n

Since filamentation is directly correlated with virulence in many instances, AmB-resistant mutants were tested for their ability to infect mice after being injected at high dose into the bloodstream. The erg3 knockouts were able to infect at least as effectively as wild-type, but the other erg-knockouts and clinical AmB-resistant strains of C. albicans<\/em> were all significantly less effective at colonizing the kidneys of infected mice.<\/p>\n

What they showed<\/strong><\/em><\/p>\n

Taken together, the data here show that there are several significantly deleterious consequences to amphotericin-B resistance in Candida albicans<\/em>. Insofar as AmB resistance is established by altering ergosterol biosynthesis, mutations in several different enzymes of this pathway all lead to similar deficiencies, at least partly mediated by the stress protein, Hsp90. Since this mechanism is not a characteristic of resistance to the other two classes of antifungals, it effectively explains why AmB resistance has not developed as quickly as resistance to triazoles or echinocandins.<\/p>\n

 <\/p>\n","protected":false},"excerpt":{"rendered":"

The paper: Vincent BM, et al. (2013) Fitness trade-offs restrict the evolution of resistance to amphotericin B. PLoS Biol 11(10): e1001692. doi:10.1371\/journal.pbio.1001692 Subject areas: microbiology, evolution Vocabulary: thrush (oral thrush) – a condition in which the fungus Candida albicans infects the mouth and throat. \u00a0It presents as velvety white lesions […]<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"jetpack_post_was_ever_published":false,"_jetpack_newsletter_access":"","_jetpack_dont_email_post_to_subs":false,"_jetpack_newsletter_tier_id":0,"_jetpack_memberships_contains_paywalled_content":false,"_jetpack_memberships_contains_paid_content":false,"footnotes":"","jetpack_publicize_message":"","jetpack_publicize_feature_enabled":true,"jetpack_social_post_already_shared":true,"jetpack_social_options":{"image_generator_settings":{"template":"highway","enabled":false},"version":2}},"categories":[7],"tags":[],"class_list":["post-487","post","type-post","status-publish","format-standard","hentry","category-bionews"],"jetpack_publicize_connections":[],"jetpack_featured_media_url":"","jetpack_sharing_enabled":true,"jetpack_shortlink":"https:\/\/wp.me\/p22HFc-7R","_links":{"self":[{"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/posts\/487","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/comments?post=487"}],"version-history":[{"count":2,"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/posts\/487\/revisions"}],"predecessor-version":[{"id":495,"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/posts\/487\/revisions\/495"}],"wp:attachment":[{"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/media?parent=487"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/categories?post=487"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.axopub.com\/wp01\/wp-json\/wp\/v2\/tags?post=487"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}