Trichophyton indotineae – Facts and controversies – A review

BACKGROUND

Dermatologists across India have been challenged by the intriguing epidemic of recurrent, slow-responsive, or non-responsive dermatophytosis in India since 2014, which has been lasting for many years. Research went on all across India, where dermatologists and microbiologists across India worked hand in hand. Many underlying factors were attributed to this epidemic in different studies, which include the rampant use of topical steroids, non-compliance with treatment, and inadequate dosage of antifungals. There has been a shift in the predominant species causing dermatophytosis from Trichophyton rubrum to Trichophyton mentagrophytes in recent years.^[1] In addition to this, antifungal resistance was also reported.

Rudramurthy et al. for the first time reported terbinafine resistance in Trichophyton species in India due to a mutation in the squalene epoxidase gene.^[2] Singh et al. in 2018 reported terbinafine resistance in recalcitrant cases of dermatophytosis and mutation in the squalene epoxidase gene from Delhi.^[3] Till then, species identification of causative agents was done by morphological characteristics and microscopy. In 2018, Nenoff et al. did DNA sequencing of the fungal isolates in Germany from the samples collected from India and demonstrated the predominant species as T. mentagrophytes internal transcribed spacer region (ITS) type 8.^[4] In 2020, this new species of fungus, which was different from other species genotypically and clinically, and caused an epidemic in India, was named Trichophyton indotineae by Kano et al.^[5] The new species has largely displaced other previously prevalent dermatophytes on the Indian subcontinent.^[6] In this review, we discuss the evolution of T. indotineae, various taxonomical classifications, and factors that led to nomenclature, salient features of the species, and how to tackle the infection. Even though controversies are there regarding the nomenclature of this species as indotineae, in this article, we use the same term to avoid confusion.

INTRODUCTION

T. indotineae is a newly named dermatophyte species in 2020, which is the same as T. mentagrophytes ITS genotype 8 within the T. mentagrophytes/Trichophyton interdigitale species complex.^[6] Now, it is prevalent in different countries across continents due to travel and migration, including India, Canada, Australia, Iran, Germany, Japan, France, etc. It is the causative agent for the epidemic of dermatophytosis in India.^[6] Even though the controversial name points to its origin in India, the country where it originated is still not known.^[7] This anthropophilic fungus is genotypically and clinically different from other types.^[8] Point mutations in the squalene epoxidase (SQLE) gene confer terbinafine resistance in them, which adds to the burden of disease.^[8,9] The species is sensitive to itraconazole, but azole resistance is emerging.^[10] Hence, mycologists across the world consider this a threat and are closely monitoring the mutations and resistance patterns in them. Morphologically and microscopically, it is similar to the T. mentagrophytes/interdigital complex, and species identification is possible only through DNA sequencing of the ITS region and comparing with sequences in the gene bank.^[11] At present, most isolates have low minimal inhibitory concentration (MIC) to itraconazole (ITZ), and conventional ITZ (C-ITZ) may have to be given for a longer duration, as long as 12 weeks, for complete remission in some patients.^[12]

ORIGIN AND EPIDEMIOLOGY OF T. indotineae

T. mentagrophytes was a distinct species till 2007 and included many subtypes like zoophilic variants T. mentagrophytes variation (var.) granulosum, var. asteroides, var. erinacei, var. quinckeanum, and anthropophilic species like T. mentagrophytes var. interdigitale, var. goetzii (synonym T. krajdenii), and var. nodulare. In 2008, based on genetic markers, T. interdigitale species was separated from T. mentagrophytes, and T. interdigitale species included both zoophilic and anthropophilic species such as T. mentagrophytes sarkisovii, T. mentagrophytes var granulosum, T. mentagrophytes var interdigitale, T. mentagrophytes goetzii, and T. mentagrophytes var nodulare.^[13] In 2017, again, T. interdigitale species was reclassified as T. mentagrophytes zoophilic species and T. interdigitale anthropophilic species, and it was based on multiple genetic markers, ecological niche, and propagation form.^[14] In 2018, a genotypic classification based on DNA sequencing of the ITS region was put forth. As per this, single-nucleotide polymorphisms (SNPs) in the ITS region of ribosomal DNA (rDNA) were a decisive factor in classification at the species level. Among over 10 different ITS types of T. mentagrophytes, type 8 was a clinically significant clonal offshoot. It was clinically and genotypically different from others and showed terbinafine resistance.^[7]

Figure 1 depicts the taxonomical evolution of T. indotineae.

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Figure 1:

Temporal course and changes in taxonomy and nomenclature of T. mentagrophytes and T. interdigitale.

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The ongoing epidemic in India heralds new research in dermatophytosis all across the country, and these studies highlighted various factors for the occurrence of recurrent and chronic dermatophytosis. Rudramurthy et al.^[2] in their study for the first time identified terbinafine resistance in T. interdigitale species isolated from cases of recurrent dermatophytosis in India, which showed a missense mutation in the SQE gene, leading to a change of phenylalanine at the 397^th position to leucine (Phe397Leu). In 2018, Singh et al. in their study, found T. interdigitale in 94% of resistant cases in Delhi. Out of this, 32% were terbinafine resistant, and they harbored single-point mutations in Leu393Phe or Phe397Leu in the squalene epoxidase (SQLE) gene.^[3] This was a warning signal for Indian dermatologists and microbiologists to do more studies in this regard.

In a study on dermatophytosis by Nenoff et al. published in 2019, skin scraping samples were collected from 7 centers across India, and molecular work was done in Germany.^[4] Fungal culture and the polymerase chain reaction (PCR) – enzyme-linked immunosorbent assay, followed by DNA sequencing, were done and compared with the gene bank in an online database. Interestingly, all species belonged to T. mentagrophytes ITS type 8, zoophilic species.

Nenoff et al. proposed that a zoophilic species of fungi cannot cause an epidemic in India, as there was a paucity of contact with animals. Hence, the concept of anthropization was put forth, wherein a previously zoophilic or geophilic fungus becomes anthropophilic under favorable circumstances.^[15]

T. mentagrophytes isolated from cases of recalcitrant dermatophytosis were ITS type 8, and it differed from other genotypes in the same species by the presence of 2–3 SNPs. Furthermore, multigene polyphasic analysis revealed the presence of distinct sequences of the high mobility group (HMG) gene in them compared to other ITS subtypes of T. mentagrophytes.^[5,16] Whole genome sequencing identified this species as a distinct clonal offshoot of T. mentagrophytes.^[8]

This genotype was later isolated in Japan by Kano et al. in a Nepali patient and an Indian patient and was named T. indotineae in 2020, which is the same T. mentagrophytes ITS type 8.^[5] It was named T. indotineae as rDNA ITS region sequences of the isolates by Kano et al. were 100% identical to terbinafine (TRB) resistant strains of T. interdigitale, which were isolated in Delhi, India, in 2018 and harbored alterations in SQLE. Like Indian strains, the isolates exhibited high MICs (32 mg/L) for TRB and contained an amino acid substitution (Phe397Leu) in SQLE. To avoid confusion in the taxonomy, the highly TRB-resistant Indian strains were designated as a new species independent of T. interdigitale/T. mentagrophytes.^[5] This is the origin of T. indotineae.

Many disagree regarding this nomenclature of T. indotineae, which is country-specific, when the origin of the fungus is disputed, and the name is prejudicial. The WHO and the American College of Microbiology also exhort against such prejudicial naming.^[7] Retrospectively, this species was identified in the gene bank in Australia (2008), Oman (2010), Iran (2016), but as per taxonomical nomenclature then, they were called T. interdigitale.^[17] Hence, the country of origin of this fungus is also unknown. Probably globalization led to its spread across various countries, including India. The new species is now the main etiological agent and is considered to be responsible for recalcitrant dermatophytosis in India, Iran, Bangladesh,^[18] etc. There are reports of isolation of T. indotineae from different countries like the United Arab Emirates, Oman, Bahrain,^[19] and Iran.^[20] In Europe, many reports are coming up from Germany.^[21] Other countries from where T. indotineae has been described include France,^[22] Belgium,^[23] Switzerland, Greece, Denmark, China, Australia, Canada,^[24] and recently, from Vietnam.^[25]

The emergence of this new species could be attributed to the irrational use of topical steroids and steroid-containing fixed drug combinations, as well as the rampant use of antifungals and antibiotics, which might have led to changes in the skin microbiome and local immune system. Anthropization favored the easy transmission and rapid spread of infection, and resistance to terbinafine added to the malady.^[6]

MOLECULAR IDENTIFICATION OF T. indotineae

Cultural characteristics and microscopy of T. indotineae are similar to T. mentagrophytes/interdigitale.^[16] These species cannot be distinguished by phenotypic tests. The colony reverse of T. indotineae is most often pale-brown to yellow-orange in color, and they are less often positive in Tween-80 opacity, urea hydrolysis, and hair perforation tests than T. mentagrophytes and T. interdigitale.^[16] T. indotineae is identified by the unique ITS sequences. For identification, DNA extraction of the fungal isolate is done, followed by PCR amplification of ITS and DNA sequencing. Identification is made by comparison with either ISHAM ITS, National Center for Biotechnology Information (NCBI), or Japanese Gene Bank. As the ITS sequences of T. indotineae on the NCBI database still show 99% sequence similarity with T. mentagrophytes, T. interdigitale, for accurate identification ITS sequences of well-defined reference strains described by Tang et al.,^[16] importantly, primary T. indotineae strains (NUBS19006 and NUBS19007), should be included in the analysis. HMG sequences of T. indotineae differ from T. interdigitale/T. mentagrophytes at 4 and 1 nucleotide positions, respectively.^[16] Frequent updation of the database of Trichophyton species is required to avoid misidentification.^[26] Other techniques include quantitative PCR using other unique sequences of T. indotineae identified and validated through Basic Local Alignment Search Tool analysis. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry is another diagnostic method achieving an accuracy of up to 97.4% across various culture conditions.^[27]

CLINICAL CHARACTERISTICS

T. indotineae causes chronic and recurrent dermatophytosis and recalcitrant dermatophytosis. Extensive areas of involvement, lack of classical appearance, rapid spread to multiple family members, sexual transmission due to close contact, involvement of genitals, and severe pruritus were the main features of the disease during the epidemic [Figures 2 and 3].^[6] Most of these patients will be on topical steroids due to severe pruritus. Mostly, these cases do not respond to terbinafine, fluconazole, and griseofulvin, and a few cases respond to higher doses of terbinafine. It is easily transmissible from person to person and also through inanimate surfaces.^[6]

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Figure 2:

Tinea corporis caused by Trichophyton indotineae, hyperpigmented plaque with pustules and crusting due to topical steroid use.

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Figure 3:

Tinea genitalis.

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ANTIFUNGAL RESISTANCE IN T. indotineae

The rampant use of topical steroids, antifungals, and antibiotics led to the development of antifungal resistance in this species. Point mutations in the SQLE gene lead to amino acid substitutions at Leu393, Phe397, Phe415, and His440.^[27] Some isolates are inherently resistant to TRB, some are sensitive to TRB at higher doses, some double mutants are resistant to TRB and azoles, and griseofulvin resistance has also been reported. The most common point mutation is one leading to amino acid substitutions at the Phe397Leu region of the SQLE gene, followed by Leu393-Phe and Leu393-Ser.^[28] Those with mutations at Ala 448 Thr are sensitive to terbinafine at higher doses. Double mutations at Phe 397 Leu and Ala 448 Thr lead to reduced susceptibility to azole and terbinafine.^[29] Azole resistance in T. indotineae is due to overexpression of the TinCYP51B gene encoding sterol 14a-demethylases enzyme. Upregulation of ATP–binding cassette (ABC) transporters or mutation of the ERG 11/CYP51 gene also confers azole resistance.^[30]

CHALLENGES IN THE MANAGEMENT

From different studies, it is clear that T. indotineae may be resistant to the usual dose of terbinafine or sensitive to terbinafine at higher doses. TRB resistance rates ranging from 17% to 75% and varying levels of ITZ resistance up to 25% have been reported in T. indotineae strains from India.^[3,31] High in vitro MIC values of voriconazole and griseofulvin are also reported.^[32] Double mutants will be resistant to azole, terbinafine, and griseofulvin. The MIC is still lower for ITZ, and it is the drug of choice in these cases. ITZ 100 mg twice daily should be given for 4–6 weeks or longer.^[32] Higher doses of ITZ are not of added benefit due to congealing. This is because the excess pellets (in a 200 mg capsule) tend to congeal, and bioavailability will not be increased due to dynamic transit time in the stomach.^[33] Hepatic saturation of the drug occurs beyond a 200 mg dose, and thus, the levels do not rise consistently.^[33]

Oral drugs can be combined with topical antifungals like topical luliconazole , one with least MIC against T. indotineae. Ciclopirox olamine, sertaconazole, miconazole, and ketoconazole are also effective.^[34] However, it may be economically unfeasible due to extensive skin involvement, as often seen with T. indotineae. Terbinafine may be given at higher doses for a longer duration.^[35] A combination of terbinafine and ITZ does not add any benefits but may increase side effects.^[36] ITZ remains the most effective antifungal for dermatophyte infections, with rising resistance to TRB. However, wide fluctuations in serum concentrations of ITZ are common due to erratic absorption patterns.^[37] Super bioavailable ITZ (SUBA ITZ) is a uniform non-pellet solid formulation with Ph-dependent hypromellose phthalate polymeric matrix. This feature helps in enhanced dissolution and absorption of the drug. SUBA ITZ gets absorbed in the intestine and food intake does not alter absorption. Proton pump inhibitors increase its plasma levels by 22%. There is less intersubject variability, and bioavailability is also higher.^[37] But presently, we are lacking good quality evidence at present for the superiority or non-inferiority of SUBA-ITZ over C-ITZ.^[38,39] In India, still patients with dermatophytosis respond to C-ITZ. However, recurrence is a major therapeutic challenge for most dermatologists in India^[38] [Table 1].^{[20,21,25,40-49]} Newer triazoles, voriconazole, and posaconazole have proven efficacy in ITZ-resistant cases, voriconazole at a dose of 200 mg/day for 4–8 weeks and posaconazole at a dose of 300 mg/day for 4 weeks. Achieving a sustainable cure in T. indotineae infection is challenging and may require maintenance therapy and relapses are also seen, which may be attributed to re-infection caused by fomites or contact with infected family members. Hence, environmental disinfection has become important and regardless of whether the strain has acquired terbinafine resistance, soaking infected clothing in quaternary ammonium detergent for 24 h could effectively eliminate T. rubrum, T. mentagrophytes complex, and T. indotineae.^[50]

CONCLUSION

T. indotineae is a new species of dermatophytes with unique genotypic, clinical, and resistance patterns, which is the same T. mentagrophytes ITS genotype 8. Its naming imparts stigma, which is against existing conventions. Anthropization of the fungus due to various factors led to ease of spreadability, which attained an epidemic form in India. It is inherently resistant to terbinafine due to different amino acid substitutions in the SQLE gene, but also resistant to commonly used antifungals such as griseofulvin and fluconazole. ITZ still remains to be the effective drug against this species, along with other topical antifungals. Species identification is difficult, as morphologically it is similar to T. mentagrophytes and T. interdigitale, and DNA sequencing is required for identification. The emergence of this new species with favorable features for transmission and resistance should be a warning sign to physicians worldwide that it is high time to revamp our policies and practices, and an effective antifungal stewardship program should be initiated.