Photodynamic therapy as a new approach to Trichomonas vaginalis inactivation

https://doi.org/10.1016/j.pdpdt.2018.02.006Get rights and content

Highlights

Abstract

The emergence of nitroimidazole resistant isolates has been an aggravating factor in the treatment of trichomoniasis, the most common non-viral sexually transmitted disease in the world. This highlights the importance of new technologies that are safe, effective, and have minor side effects or resistance. Hence, we evaluated the effectiveness of photodynamic therapy on the inactivation of Trichomonas vaginalis in vitro. We used methylene blue as a photosensitizing substance, and a light-emitting diode (LED) for irradiation of metronidazole sensitive and resistant strains. Our results showed that only the presence of light did not interfere with parasite growth; however, methylene blue isolated or associated with light inhibited 31.78% ± 7.18 and 80.21% ± 7.11 of the sensitive strain, respectively, and 31.17% ± 4.23 and 91.13% ± 2.31 of the resistant strain, respectively. The high trichomonicidal activity of the photodynamic therapy, associated with low cost and ease of application, signalize its great therapeutic potential not only when conventional treatment fails, but also routinely in women with trichomoniasis.

Introduction

Trichomonas vaginalis is the protozoan responsible for trichomoniasis, the most common curable sexually transmitted infection in the world [1,2]. Based on estimates from the World Health Organization (WHO), the disease affects more than 276 million people per year, with almost 90% of these infections occurring among people living in resource-limited environments [3]. The main concern with T. vaginalis is its predisposing nature to other diseases or sequelae. In the past 20 years, many studies have shown that secondary infections to trichomoniasis increased both the risk of adverse conditions during pregnancy and the transmission of human immunodeficiency virus (HIV). In addition, T. vaginalis infection may be associated with a higher risk of prostate cancer [4]. Despite these facts, little attention is given to trichomoniasis and it is currently considered a neglected disease [5].

Sexually transmitted infections have been a significant burden on global public health, not only because they present high morbidity and mortality, but also because of the costs associated with the treatment and management of these infections [6]. Given the high prevalence of T. vaginalis and its potential to promote HIV transmission, a greater investment in infection diagnosis, treatment, and control is clearly needed [7].

Nitroimidazoles such as metronidazole (MTZ) have been the therapeutic choice for the treatment of trichomoniasis [8]. Despite this, patients treated with MTZ have presented several undesirable side effects that limit treatment success due to discontinuation of the same [9]. This fact can directly influences the emergence of resistant clinical isolates, representing an aggravating factor in disease treatment [10,11]. In this context, new therapeutic approaches, such as photodynamic therapy (PDT), may be a promising alternative to conventional treatments. PDT is a minimally invasive technique and can be applied repeatedly at the same site. Pain during PDT is the most commonly reported adverse effect and may prevent successful treatment delivery, although low-irradiance regimes and use of nerve blocks or cooling air can be used to reduce pain. Other acute and chronic adverse effects are reported but do not seem to be limiting factors in the use of topical PDT [12].

PDT uses irradiated photosensitizing substances at an appropriate wavelength in the presence of oxygen. Once exposed to light, the photosensitizers are activated to a short-lived excited state (singlet state), which is then converted to a long-lived triplet state [13]. The photosensitizer in the triplet state reacts with biomolecules by two mechanisms. The Type I reaction involves electron transfer directly from the photosensitizing substance, producing ions or removal of electrons to form free radicals. These radicals react rapidly with oxygen, resulting in the production of reactive oxygen species (superoxide, hydroxyl radicals, and hydrogen peroxide). In the type II mechanism, there is an energy transfer of the photosensitizing substance to the oxygen molecule, forming highly reactive singlet oxygen [14]. These two reactions consolidate the mechanisms of tissue and cellular injury, which depends both on oxygen tension and on photosensitizer concentration [15].

In the present study, we evaluated the efficacy of PDT using methylene blue (MB) as a photosensitizer and a light-emitting diode (LED) in the inactivation of MTZ susceptible and resistant strains of the T. vaginalis parasite in vitro.

Section snippets

Organism

This study used the JT strain, isolated in Brazil from a symptomatic 40-year-old woman, and CDC 085 strain (ATCC 50143), considered a standard strain resistant to MTZ isolated in the United States (USA) from a patient, whose condition is refractory to treatment, were used [8,16]. Both were maintained under axenic conditions in YI-S medium [17] at 37 °C.

Photosensitizer

The photosensitizer used in this study was methylene blue (Sigma, St. Louis, MO, USA), solubilized in sterile Milli-Q water and used at 0.25%.

Inactivation of T. vaginalis trophozoites after treatment with MB and PDT in vitro

First, we evaluated the effects of MB and LED light separately on the sensitive and resistant trophozoites, compared to the control without intervention. No significant difference was observed between the LC and CTR groups, showing that the presence of light did not influence the growth of the trophozoites of either strain (Fig. 1). However, the MB maximal inhibitory effect on trophozoites after 24 h of incubation reached 32.29% ± 4.54 for the JT strain and 31.17% ± 4.23 for the CDC 085 strain (

Discussion

According to WHO, trichomoniasis accounts for almost 50% of all curable sexually transmitted diseases worldwide [18], and is more prevalent than important sexually transmitted infections such as syphilis, gonorrhea, and chlamydia [19]. Many isolates with clinical resistance to standard treatment regimens and with evidence of resistance to MTZ in aerobic resistance tests, respond only to higher doses of this drug (1 g three times per day orally along with 500 mg topical use). Nevertheless, at

Declaration of interest

Authors declare that there is no conflict of interest. Authors are responsible for the content and writing of the manuscript.

Funding

This work was supported by Fundação de Amparo à Pesquisa do Estado de Minas Gerais – FAPEMIG (grant PPM00140-14), Pró-Reitoria de Pesquisa da UFMG and Coordenação de Aperfeiçoamento Pessoal de Nível Superior − CAPES.

Acknowledgment

We thank Dr. Fernando Costa e Silva Filho for providing the JT strain of T. vaginalis.

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