Infinity war : Trichomonas vaginalis and interactions with host immune response

Trichomonas vaginalis is the pathological agent of human trichomoniasis. The incidence is 156 million cases worldwide. Due to the increasing resistance of isolates to approved drugs and clinical complications that include increased risk in the acquisition and transmission of HIV, cervical and prostate cancer, and adverse out-comes during pregnancy, increasing our understanding of the patho-gen’s interaction with the host immune response is essential. Produc-tion of cytokines and cells of innate immunity: Neutrophils and mac-rophages are the main cells involved in the fight against the parasite, while IL-8, IL-6 and TNF-α are the most produced cytokines in re-sponse to this infection. Clinical complications: T. vaginalis increases the acquisition of HIV, stimulates the invasiveness and growth of prostate cells, and generates an inflammatory environment that may lead to preterm birth. Endosymbiosis: Mycoplasma hominis increased cytotoxicity, growth, and survival rate of the parasite. Purinergic sig-naling: NTPD-ases and ecto-5’-nucleotidase helps in parasite survival by modulating the nucleotides levels in the microenvironment. Anti-bodies: IgG was detected in serum samples of rodents infected with isolates from symptomatic patients as well as patients with symp-toms. However, antibody production does not protect against a rein-fection. Vaccine candidate targets: The transient receptor potential- like channel of T. vaginalis (TvTRPV), cysteine peptidase, and α-actinin are currently cited as candidate targets for vaccine develop-ment. In this context, the understanding of mechanisms involved in the host-T. vaginalis interaction that elicit the immune response may contribute to the development of new targets to combat trichomoni-asis

Hidrólise extracelular de nucleotídeos em Trichomonas vaginalis : caracterização bioquímica e localização enzimática

Trichomonas vaginalis é o protozoário flagelado causador da tricomonose, a doença sexualmente transmissível (DST) não virai mais comum no mundo. O T. vaginalis tem se destacado como um importante patógeno e está associado a graves complicações de saúde. Considerando o sério impacto da tricomonose na saúde pública, é importante estudar os aspectos bioquímicos do T. vaginalis que contribuem para infecção do hospedeiro e patogênese. Nucleotídeos extracelulares estão envolvidos em uma variedade de funções fisiológicas e patológicas, além de apresentarem propriedades citotóxicas. Os nucleotídeos extracelulares podem ser hidrolisados via ectonucleotidases, incluindo a família E-NTPDase (ectonucleosídeo trifosfato difosfoidrolase) e a ecto-5'-nucleotidase. Uma atividade de NTPDase1 foi previamente caracterizada em T. vaginalis. Nossos resultados mostraram a caracterização de uma ecto-5'-nucleotidase dependente de magnésio e cálcio e com atividade máxima em pH alcalino. Os valores de Km (Constante de Michaelis) situaram-se na faixa micromolar, com ampla especificidade a nucleosídeos monofosfatados. A atividade de ecto-5'-nucleotidase em T. vaginalis foi inibida pelo 5'-[α,ß-metileno]difosfato (AMPCP), inibidor específico da enzima em vertebrados. O metronidazol e o tinidazol apresentaram efeitos diversos na atividade da NTPDase1 e da ecto-5'nucleotidase de um isolado de T. vaginalis proveniente da American Type Culture Collection (ATCC) cultivado por longo período no laboratório e de um isolado clínico fresco. A hidrólise de ATP foi aumentada na presença de metronidazol no isolado ATCC. Por outro lado, ela foi inibida pelo tinidazol no isolado clínico fresco. O tratamento dos parasites na presença de metronidazol por 2h inibiu a hidrólise de ATP e ADP, enquanto o tratamento com tinidazol inibiu as atividades ATPásica e ADPásica somente no isolado clínico fresco. Os fármacos não produziram efeitos significativos na atividade da ecto-5'-nucleotidase de ambos os isolados de T. vaginalis testados. Os dados sugerem a função da NTPDase1 como moduladora da concentração de nucleotídeos extracelulares na presença de metronidazol e tinidazol, uma condição adversa para o parasito. Através de citoquímica enzimática, ambas as enzimas demonstraram ecto-localização, confirmando os mecanismos presentes no parasito capazes de realizar hidrólise extracelular de nucleotídeos. Diferentes concentrações de nucleotídeos da adenina e de adenosina não exerceram efeitos citolíticos nos trofozoítos. As atividades ATPásica e ADPásica apresentam-se mais elevadas em isolados clínicos frescos em comparação a isolados cultivados por longos períodos. A heterogeneidade foi demonstrada entre os diferentes isolados de T. vaginalis com relação à razão de hidrólise ATP:ADP. Alguns isolados apresentaram razão de hidrólise ATP:ADP igual a 1:0,8, característica de NTPDase1, enquanto outros apresentaram uma razão de hidrólise ATP:ADP de aproximadamente 2:1. Os isolados de T. vaginalis apresentaram diferenças dramáticas nos níveis de atividade da ecto-5'-nucleotidase e, surpreendentemente, alguns isolados apresentaram muito baixa ou nenhuma atividade. Diferentes condições de cultivo foram testadas e não apresentaram efeitos na elevação dos níveis da atividade enzimática. A ausência de atividade de hidrólise de AMP pelo T. vaginalis pode acarretar importantes conseqüências para ambos hospedeiro e parasito durante a infecção. O cultivo dos parasites em meio de cultura com e sem a adição de ferro não apresentou efeitos dramáticos na atividade da NTPDase dos diferentes isolados. Ao contrário, concentrações baixas e elevadas de ferro apresentaram decréscimo e aumento, respectivamente, na atividade da ecto-5'-nucleotidase daqueles isolados que apresentaram atividade sob condições normais de cultivo. Considerando os altos níveis de nucleotídeos púricos no sítio do T. vaginalis, a ausência de efeitos citolíticos nos parasites e a ecto-localização das enzimas envolvidas na hidrólise de nucleotídeos, sugere-se que a NTPDase1 e a ecto-5'-nucleotidase modulam as concentrações extracelulares de ATP, ADP e AMP. O produto final dessas reações, o nucleosídeo adenosina, é recaptado pelas vias de salvação.The flagellated protozoan, Trichomonas vaginalis, causes trichomonosis, the most prevalent non-viral sexually transmitted disease (STD) worldwide. lnfection with T. vaginalis has major health consequences for women. Taking in account the serious impact caused by trichomonosis on public health, it is important to study the biochemical aspects of T. vaginalis that contribute with host infection and pathogenesis. Extracellular nucleotides are involved in several physiologic and pathologic functions, including cytotoxic properties. Nucleotides are hydrolyzed by ectonucleotidases, including E-NTPDase (ectonucleoside triphosphate diphosphohydrolase) family and ecto-5'-nucleotidase. A NTPDase1 activity was previously characterized in T. vaginalis. Our results show the characterization of an ecto-5'nucleotidase activity, magnesium- and calcium-dependent and with maximum activity in alkaline pH. Km (Michaelis Constant) values are in the micromolar range, with broad monophosphate nucleosides specificity. The ecto-5'-nucleotidase activity from T. vaginalis was inhibited by 5'-[α,ß-methylene]diphosphate (AMPCP), the specific inhibitor of this enzyme in vertebrates. Metronidazole and tinidazole presented different effects on NTPDase1 and ecto-5'-nucleotidase activities from T. vaginalis long-term-grown isolate from American Type Culture Collection (ATCC) and from a fresh clinical isolate. ATP hydrolysis was activated in presence of metronidazole in the ATCC strain, whilst it was inhibited by tinidazole in a fresh clinical isolate. The treatment of cells in presence of metronidazole for two hours inhibited ATP and ADP hydrolysis, whilst treatment with tinidazole inhibited ATP and ADP hydrolysis only in the fresh clinical isolate. The drugs did not change the ecto-5'nucleotidase activity for both strains. Our results suggest that the modulation of extracellular A TP and ADP levels during treatment with these drugs could be a parasitic defense strategy as a survival mechanism in an adverse environment. Cytochemical localization showed that both NTPDase1 and ecto-5'-nucleotidase presented ecto-localization, reinforcing the mechanisms present in T. vaginalis capable to hydrolyze extracellular nucleotides. Different adenine nucleotides and adenosine concentrations did not produce any cytolytic effects on trophozoites. ATPase and ADPase activities were higher in fresh clinical isolates when compared to long-term-grown isolates. Heterogeneity on ATP:ADP hydrolysis ratio was shown among different T. vaginalis isolates. Some isolates had ATP:ADP hydrolysis ratio of 1:0.8, characteristic of NTPDase1, whilst other isolates presented ATP:ADP hydrolysis ratio of approximately 2:1. T. vaginalis isolates had dramatic differences in levels of ecto-5'nucleotidase activity, and surprisingly, some isolates had little or no enzymatic activity. Different growth conditions were without effect in elevating levels of ecto-5'-nucleotidase activity. This lack of ecto-5'-nucleotidase activity by T. vaginalis may have important consequences for both host and parasite during infection. Growth of trichomonads in ironreplete and iron-depleted medium had overall little dramatic effect in distinct pattems in NTPDase activity among fresh isolates. ln contrast, high- and low-iron trichomonads had increased and decreased ecto-5'-nucleotidase activity, respectively, among fresh isolates with activity detectable under normal medium conditions. Taking in account the high purine nucleotides levels present in vagina, the T. vaginalis site, the lack of cytolytic effects on parasites and the ecto-localization of enzymes involved on nucleotide hydrolysis, it is suggested that NTPDase1 and ecto-5'-nucleotidase modulate extracellular ATP, ADP and AMP concentrations. The final product, adenosine, is reuptake by salvage pathways

Trichomoniasis : are we giving the deserved attention to the most common non-viral sexually transmitted disease worldwide?

Etiology: Trichomonas vaginalis is the etiologic agent of trichomo-niasis, the most common non-viral sexually transmitted disease (STD) in the world. Transmission: Trichomoniasis is transmitted by sexual intercourse and transmission via fomites is rare. Epidemiology, incidence and prevalence: The WHO estimates an incidence of 276 million new cases each year and preva-lence of 187 million of infected individuals. However, the infection is not noti-fiable. Pathology/Symptomatology: The T. vaginalis infection results in a vari-ety of clinical manifestations - in most cases the patients are asymptomatic, but some may develop signs typically associated to the disease. Importantly, the main issue concerning trichomoniasis is its relationship with serious health consequences such as cancer, adverse pregnancy outcomes, infertility, and HIV acquisition. Molecular mechanisms of infection: To achieve success in parasitism trichomonads develop a complex process against the host cells that includes dependent- and independent-contact mechanisms. This multi-factorial pathogenesis includes molecules such as soluble factors, secreted proteinases, adhesins, lipophosphoglycan that culminate in cytoadherence and cytotoxicity against the host cells. Treatment and curability: The treat-ment with metronidazole or tinidazole is recommended; however, cure fail-ures remain problematic due to noncompliance, reinfection and/or lack of treatment of sexual partners, inaccurate diagnosis, or drug resistance. There-fore, new therapeutic alternatives are urgently needed. Protection: Strategies for protection including sexual behavior, condom usage, and therapy have not contributed to the decrease on disease prevalence, pointing to the need for innovative approaches. Vaccine development has been hampered by the lack of long-lasting humoral immunity associated to the absence of good animal models

COMPARISON OF PERMANENT STAINING METHODS FOR THE LABORATORY DIAGNOSIS OF TRICHOMONIASIS

Trichomonas vaginalis is the etiologic agent of trichomoniasis, the most common non-viral sexually transmitted disease (STD) in the world. The diagnosis is based on wet mount preparation and direct microscopy on fixed and stained clinical specimens. The aim of this study was to compare the performance of different fixing and staining techniques used in the detection of T. vaginalis in urine. The smears were fixed and submitted to different methods of permanent staining and then, the morphological aspects of the parasites were analyzed and compared. The Papanicolaou staining with ethanol as the fixative solution showed to be the best method of permanent staining. Our data suggest that staining techniques in association with wet mount examination of fresh specimens contribute to increase the sensitivity in the diagnosis of trichomoniasis

Parasitismo intestinal e fatores socioambientais de indígenas Mbyá-Guarani, Porto Alegre, Rio Grande do Sul, Brasil

Disturbing data reveal the prevalence of intestinal parasites and their relationship with socio-environmental factors among Mbyá-Guarani Indians. The prevalence was determined by spontaneous sedimentation in water, centrifugation-floatation, and Kato-Katz. A socioeconomic questionnaire was submitted to each family. The overall prevalence of intestinal parasites was 88.7%, and 45.5% were polyparasitized. There was 90.5% prevalence of enteric parasites in children (1-12- year-old), and 85% among 13-65-year-old individuals, indicating that both age groups are extensively parasitized. The parasite load was low to moderate for geohelminths and 75% of the families did not have latrine, thus the practice of defecation occurred outdoors. These findings suggest that the multiple intestinal parasitism in the Mbyá-Guarani community is high to the point of being the rule, and that it relates essentially to the traditional lifestyle and health habits. It is urgently necessary to implement the association of anti-parasitic treatment with sanitation improvement. This should be done simultaneously with health education activities for this population.Dados preocupantes demonstram a prevalência de parasitos intestinais e sua relação com fatores socioambientais entre indígenas Mbyá-Guarani. A prevalência foi determinada pelas técnicas de sedimentação espontânea em água, centrífugo-flutuação e Kato-Katz. Para cada família, um questionário socioeconômico foi aplicado. A prevalência geral de parasitos intestinais foi de 88,7% e 45,5% estavam poliparasitados. A prevalência em crianças (1-12 anos) foi 90,5% e em indivíduos entre 13-65 anos foi 85%, indicando que ambas as faixas etárias são amplamente parasitadas. A carga parasitária foi baixa a moderada para geohelmintos e 75% das famílias não possuem banheiro, sendo a prática da defecação feita ao ar livre. Estes achados sugerem que o poliparasitismo intestinal na comunidade Mbyá-Guarani é altíssimo, a ponto de ser a regra, e que se refere essencialmente ao estilo de vida tradicional e hábitos de saúde. Há a necessidade de aplicar urgentemente a associação de tratamento antiparasitário às melhorias sanitárias. Isto deve ser feito simultaneamente com atividades de educação em saúde para esta população

Specific pathogenic mechanisms of mucosal protozoans : Entamoeba histolytica, Giardia lamblia and Trichomonas vaginalis

Entamoeba histolytica e Giardia lamblia são protozoários que podem parasitar a mucosa intestinal, causando principalmente diarreia. Trichomonas vaginalis coloniza a mucosa vaginal causando tricomonose, a doença sexualmente transmissível não viral mais comum no mundo. Embora coletivamente estes parasitos infectem mais de um bilhão de pessoas a cada ano, seus mecanismos de patogenicidade ainda não estão totalmente esclarecidos. Assim, esta revisão reúne os principais mecanismos envolvidos na patogenicidade destes protozoários, bem como os fatores do microambiente que podem interferir no sucesso da colonização. A patogênese da E. histolytica envolve adesão, lise, fagocitose de células epiteliais e bactérias, invasão tecidual por ação de enzimas e evasão da resposta imune do hospedeiro. A lectina Gal/GalNAc, os amebaporos e as cisteína proteases são as principais moléculas envolvidas nesses processos. O estabelecimento da giardiose depende de diversos mecanismos patogênicos e de virulência desenvolvidos pela G. lamblia, tais como as moléculas envolvidas na adesão, encistamento e variação antigênica. Para o sucesso da colonização da mucosa vaginal, o T. vaginalis expressa moléculas como as adesinas de superfície, lipofosfoglicanos e galectina, envolvidas na adesão às células epiteliais vaginais e alteração da expressão gênica, tanto do parasito como do hospedeiro.Entamoeba histolytica and Giardia lamblia are protozoans that may parasitize the intestinal mucosa, mainly causing diarrhea. Trichomonas vaginalis colonizes the vaginal mucosa causing trichomonosis, the most common non-viral sexually transmitted disease in the world. Although collectively these parasites infect over a billion people each year, their pathogenic mechanisms have not been completely understood so far. Hence, this review of the literature demonstrates the main mechanisms involved in the pathogenicity of these protozoans, as well as the microenvironmental factors that can interfere with successful colonization. The pathogenesis of E. histolytica involves adhesion, lysis, phagocytosis of epithelial cells and bacteria, tissue invasion by enzymatic action, and evasion of host immune response. Lectin Gal/GalNac, amoebapores, and cysteine proteases are the main molecules involved in these processes. The establishment of giardiosis depends on several pathogenic mechanisms and virulence developed by G. lamblia, such as molecules involved in adhesion, encystation and antigenic variation. For successful colonization of vaginal mucosa, T. vaginalis express molecules like adhesins on the surface and galectin and lipophosphoglycan, involved in the adherence to vaginal epithelial cells and altered gene expression of both the parasite and the host

Resistência bacteriana no meio ambiente e implicações na clínica hospitalar

O aumento da incidência de infecções microbianas resistentes a antibióticos adquiridas tanto na comunidade quanto nos hospitais tem chamado a atenção da comunidade de saúde. Neste contexto, muitos estudos têm demonstrado que o próprio meio ambiente funciona como um grande reservatório de genes de resistência a antimicrobianos. A resistência a antibióticos tem sido observada em vários ambientes aquáticos incluindo rios e áreas costeiras, esgoto doméstico, esgoto hospitalar, sedimentos, águas superfíciais, lagos, oceanos e água potável, bem como em solos. Estudos mostram que mais de 90% dos isolados bacterianos originados da água do mar são resistentes a pelo menos um antibiótico e 20% são resistentes a pelo menos cinco. Com base nos resultados levantados na literatura envolvendo amostras de solo e água, torna-se evidente a relevância da detecção de genes de resistência a partir de bactérias presentes na natureza, pois podem refl etir parte da origem dos mecanismos de resistência observados no ambiente hospitalar.The increasing bacterial infections caused by antibiotic resistant microorganisms acquired either in the community or at the hospitals has concerned the health staff. In this context, many studies have demonstrated that the environment acts like a reservoir of antibiotic resistant genes. The antimicrobial resistance has been found in several environments, such as river, domestic waste and hospital waste, sediments, lakes, ocean, potable water, and soils. Studies indicate that more than 90% of bacterial isolates from ocean are resistant to at least one antibiotic and 20% are resistant to at least fi ve. In summary, the results present in the literature with soil and water, makes evident the relevance of resistance genes detection in bacteria found in the nature, because it might refl ect part of the origin of the resistance mechanisms found at the hospital

Resistência bacteriana no meio ambiente e implicações na clínica hospitalar

O aumento da incidência de infecções microbianas resistentes a antibióticos adquiridas tanto na comunidade quanto nos hospitais tem chamado a atenção da comunidade de saúde. Neste contexto, muitos estudos têm demonstrado que o próprio meio ambiente funciona como um grande reservatório de genes de resistência a antimicrobianos. A resistência a antibióticos tem sido observada em vários ambientes aquáticos incluindo rios e áreas costeiras, esgoto doméstico, esgoto hospitalar, sedimentos, águas superfíciais, lagos, oceanos e água potável, bem como em solos. Estudos mostram que mais de 90% dos isolados bacterianos originados da água do mar são resistentes a pelo menos um antibiótico e 20% são resistentes a pelo menos cinco. Com base nos resultados levantados na literatura envolvendo amostras de solo e água, torna-se evidente a relevância da detecção de genes de resistência a partir de bactérias presentes na natureza, pois podem refl etir parte da origem dos mecanismos de resistência observados no ambiente hospitalar.The increasing bacterial infections caused by antibiotic resistant microorganisms acquired either in the community or at the hospitals has concerned the health staff. In this context, many studies have demonstrated that the environment acts like a reservoir of antibiotic resistant genes. The antimicrobial resistance has been found in several environments, such as river, domestic waste and hospital waste, sediments, lakes, ocean, potable water, and soils. Studies indicate that more than 90% of bacterial isolates from ocean are resistant to at least one antibiotic and 20% are resistant to at least fi ve. In summary, the results present in the literature with soil and water, makes evident the relevance of resistance genes detection in bacteria found in the nature, because it might refl ect part of the origin of the resistance mechanisms found at the hospital

Novel treatment approaches to combat trichomoniasis, a neglected and sexually transmitted infection caused by trichomonas vaginalis : translational perspectives

The multistep translational science behind new drugs comprehends the entire process through laboratory, clinical, and community observations turned into health interventions. The development of new drug options from discovering targets and leading compounds in basic research for implementing therapeutic guidelines contributes to the emergence of health policies essential for infection control. This review updates the translational research in the scenario of the most common non-viral sexually transmitted infection (STI), trichomoniasis. Paradoxically to its high occurrence, it is considered neglected since notification is not mandatory. It turns into a stable disease with health complications, and receives little emphasis from public health programs to control STI. Although related to curable STIs, the current drugs, metronidazole and tinidazole, present therapeutic failures. The need for new options to treat trichomoniasis is established by basic research studies and patents revealing novel synthetic compounds and natural products presenting anti-Trichomonas vaginalis activities, mainly based on in vitro findings. Clinical trials are still focused on new routes of administration for conventional drugs. In addition, nanotechnology approaches are in their infancy, shedding light on potential possibilities for creating more effective, targeted, and safe delivery systems. Overall, the novel proposed approaches need, in addition to pharmaceutical development and efficacy assessments, to ensure that the quality requirements for their use as medicines are met. It is essential to overcome these issues to cross the “Death Valley” of drug discovery and to advance in the translational science criteria in the trichomoniasis drug development field

Peptidases Are Potential Targets of Copper(II)-1,10-Phenanthroline-5,6-dione Complex, a Promising and Potent New Drug against Trichomonas Vaginalis

Trichomonas vaginalis is responsible for 156 million new cases per year worldwide. When present asymptomatically, the parasite can lead to serious complications, such as development of cervical and prostate cancer. As infection increases the acquisition and transmission of HIV, the control of trichomoniasis represents an important niche for the discovery and development of new antiparasitic molecules. This urogenital parasite synthesizes several molecules that allow the establishment and pathogenesis of infection. Among them, peptidases occupy key roles as virulence factors, and the inhibition of these enzymes has become an important mechanism for modulating pathogenesis. Based on these premises, our group recently reported the potent anti-T. vaginalis action of the metal-based complex [Cu(phendione)3](ClO4)2.4H2O (Cu-phendione). In the present study, we evaluated the influence of Cu-phendione on the modulation of proteolytic activities produced by T. vaginalis by biochemical and molecular approaches. Cu-phendione showed strong inhibitory potential against T. vaginalis peptidases, especially cysteine- and metallo-type peptidases. The latter revealed a more prominent effect at both the post-transcriptional and post-translational levels. Molecular Docking analysis confirmed the interaction of Cu-phendione, with high binding energy (9.7 and 10.7 kcalmol1, respectively) at the active site of both TvMP50 and TvGP63 metallopeptidases. In addition, Cu-phendione significantly reduced trophozoite-mediated cytolysis in human vaginal (HMVII) and monkey kidney (VERO) epithelial cell lineages. These results highlight the antiparasitic potential of Cu-phendione by interaction with important T. vaginalis virulence factors