Browsing by Author "Campino, Lenea"
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- 1,2,4-Trioxolane and 1,2,4,5-Tetraoxane endoperoxides against old-world Leishmania parasites: in vitro activity and mode of actionPublication . Mendes, Andreia; Armada, Ana; Cabral, Lília; Amado, Patrícia; Campino, Lenea; Cristiano, Maria de Lurdes; Cortes, SofiaLeishmaniasis remains one of the ten Neglected Tropical Diseases with significant morbidity and mortality in humans. Current treatment of visceral leishmaniasis is difficult due to a lack of effective, non-toxic, and non-extensive medications. This study aimed to evaluate the selectivity of 12 synthetic endoperoxides (1,2,4-trioxolanes; 1,2,4,5-tetraoxanes) and uncover their biochemical effects on Leishmania parasites responsible for visceral leishmaniasis. The compounds were screened for in vitro activity against L. infantum and L. donovani and for cytotoxicity in two monocytic cell lines (J774A.1 and THP-1) using the methyl thiazol tetrazolium assay. Reactive oxygen species formation, apoptosis, and mitochondrial impairment were measured by flow cytometry. The compounds exhibited fair to moderate anti-proliferative activity against promastigotes of the 2 Leishmania species, with IC50 values ranging from 13.0 ± 1.7 µM to 793.0 ± 37.2 µM. Tetraoxanes LC132 and LC138 demonstrated good leishmanicidal activity on L. infantum amastigotes (IC50 13.2 ± 5.2 and 23.9 ± 2.7 µM) with low cytotoxicity in mammalian cells (SIs 22.1 and 118.6), indicating selectivity towards the parasite. Furthermore, LC138 was able to induce late apoptosis and dose-dependent oxidative stress without affecting mithocondria. Compounds LC132 and LC138 can be further explored as potential antileishmanial chemotypes.
- Antileishmanial activity of meroditerpenoids from the macroalgae Cystoseira baccataPublication . Sousa, Carolina Bruno de; Gangadhar, Katkam N.; Morais, Thiago R.; Conserva, Geanne A. A.; Vizetto-Duarte, C; Pereira, H.; Laurenti, Marcia D.; Campino, Lenea; Levy, Debora; Uemi, Miriam; Barreira, Luísa; Custódio, L.; Passero, Luiz Felipe D.; Lago, Joao Henrique G.; Varela, JoãoThe development of novel drugs for the treatment of leishmaniases continues to be crucial to overcome the severe impacts of these diseases on human and animal health. Several bioactivities have been described in extracts from macroalgae belonging to the Cystoseira genus. However, none of the studies has reported the chemical compounds responsible for the antileishmanial activity observed upon incubation of the parasite with the aforementioned extracts. Thus, this work aimed to isolate and characterize the molecules present in a hexane extract of Cystoseira baccata that was found to be bioactive against Leishmania infantum in a previous screening effort. A bioactivity-guided fractionation of the C. baccata extract was carried out and the inhibitory potential of the isolated compounds was evaluated via the MIT assay against promastigotes and murine macrophages as well as direct counting against intracellular amastigotes. Moreover, the promastigote ultrastructure, DNA fragmentation and changes in the mitochondrial potential were assessed to unravel their mechanism of action. In this process, two antileishmanial meroditerpenoids, (3R)- and (3S)-tetraprenyltoluquinol (1a/1b) and (3R)- and (3S)-tetraprenyltoluquinone (2a/2b), were isolated. Compounds 1 and 2 inhibited the growth of the L. infantum promastigotes (IC50 = 44.9 +/- 4.3 and 94.4 +/- 10.1 mu M, respectively), inducing cytoplasmic vacuolization and the presence of coiled multilamellar structures in mitochondria as well as an intense disruption of the mitochondrial membrane potential. Compound 1 decreased the intracellular infection index (IC50 = 25.0 +/- 4.1 mu M), while compound 2 eliminated 50% of the intracellular amastigotes at a concentration > 88.0 mu M. This work identified compound 2 as a novel metabolite and compound 1 as a biochemical isolated from Cystoseira algae displaying antileishmanial activity. Compound 1 can thus be an interesting scaffold for the development of novel chemotherapeutic molecules for canine and human visceral leishmaniases studies. This work reinforces the evidence of the marine environment as source of novel molecules. (C) 2017 Elsevier Inc. All rights reserved.
- Bacterial and protozoal agents of feline vector-borne diseases in domestic and stray cats from southern PortugalPublication . Maia, Carla; Ramos, Claudia; Coimbra, Monica; Bastos, Filipa; Martins, Angela; Pinto, Pedro; Nunes, Monica; Vieira, Maria Luisa; Cardoso, Luis; Campino, LeneaBackground: Feline vector-borne diseases (FVBD) have emerged in recent years, showing a wider geographic distribution and increased global prevalence. In addition to their veterinary importance, domestic cats play a central role in the transmission cycles of some FVBD agents by acting as reservoirs and sentinels, a circumstance that requires a One Health approach. The aim of the present work was to molecularly detect feline vector-borne bacteria and protozoa with veterinary and zoonotic importance, and to assess associated risk factors in cats from southern Portugal. Methods: Six hundred and forty-nine cats (320 domestic and 329 stray), from veterinary medical centres and animal shelters in southern Portugal, were studied. Anaplasma spp./Ehrlichia spp., Babesia spp., Bartonella spp., Borrelia burgdorferi sensu lato, Hepatozoon spp. and Leishmania spp. infections were evaluated by polymerase chain reaction (PCR) in blood samples. Results: One hundred and ninety-four (29.9%) cats were PCR-positive to at least one of the tested genera or complex of FVBD agents. Sixty-four (9.9%) cats were positive to Leishmania spp., 56 (8.6%) to Hepatozoon spp., 43 (6.6%) to Babesia spp., 35 (5.4%) to Anaplasma spp./Ehrlichia spp., 19 (2.9%) to Bartonella spp. and 14 (2.2%) to B. burgdorferi s.l. Thirty-three (5.1%) cats were positive to two (n = 29) or three (n = 4) genera/complex. Babesia vogeli, Bartonella clarridgeiae, Bartonella henselae, Ehrlichia canis, Hepatozoon felis and Leishmania infantum were identified by DNA sequencing. Conclusions: The occurrence of FVBD agents in southern Portugal, some of them with zoonotic character, emphasizes the need to alert the veterinary community, owners and public health authorities for the risk of infection. Control measures should be implemented to prevent the infection of cats, other vertebrate hosts and people.
- Cystoseira algae (Fucaceae): update on their chemical entities and biological activitiesPublication . Bruno De Sousa, Carolina; Gangadhar N. Katkam, Dr.; Macridachis, Jorge; Pavao, Madalena; Morais, Thiago R.; Campino, Lenea; J. C. or Varela J. or Varela J.C.S., Varela; Lago, Joao Henrique G.Cystoseira (Sargassaceae) is a genus of marine brown algae composed of about 40 species, which is distributed along the Eastern Atlantic and Mediterranean coasts. The biological potential of the Cystoseira genus has been investigated and several activities have been reported. Chemically, this genus contains a wide variety of secondary metabolites, such as terpenoids, steroids, phlorotannins and phenolic compounds. Additionally, other chemical components as, for instance, carbohydrates, triacylglycerolsgatty acids, pigments as well as vitamins have been identified in the studied species. Some of the isolated compounds were associated with the reported pharmacological properties, as for example antioxidant, anti-inflammatory, cytotoxicity, anticancer, cholinesterase inhibition, anti-diabetic, activities but also antibacterial, antifungal and anti-parasitic activities. In this review, we provide a comprehensive overview of the compounds isolated and identified after 1995 from the different species of Cystoseira, compiling more than 200 compounds isolated, together with their therapeutic potential. (C) 2017 Elsevier Ltd. All rights reserved.
- Emergence of Thelazia callipaeda Infection in dogs and cats from east-central PortugalPublication . Maia, C.; Catarino, A. L.; Almeida, B.; Ramos, C.; Campino, Lenea; Cardoso, L.The eyeworm Thelazia callipaeda (Spirurida, Thelaziidae) infects domestic animals, wildlife and human beings, and is considered an emerging pathogen in Europe. This study aimed at investigating the prevalence and risk factors of T.callipaeda infection in dogs and cats from east-central Portugal, a region where the parasite was previously detected in two red foxes (Vulpes vulpes). Thelazia callipaeda was found in 22 (3.8%) of 586 dogs and in four (23.5%) of 17 cats. A total of 178 adult worms (71.9% of females and 28.1% of males) were collected from the conjunctiva of the infected dogs. The number of worms collected per dog ranged from 1 to 35 (average +/- standard deviation: 8.08 +/- 9.49), with four dogs (18.2%) harbouring only a single parasite. Worms were gathered from dogs throughout all months of the year. A total of 17 adult worms (64.7% of females and 35.3% of males) were obtained from cats. The number of worms per cat ranged from 1 to 14 (4.3 +/- 6.5), with three cats (75.0%) having a single parasite. Eyeworm infection was statistically more prevalent in pastoral and farm dogs, in those dogs with contact with other animals and in dogs with ocular manifestations. T. callipaeda is endemic in the east-central part of Portugal, reportedly infecting domestic (dogs and cats) and wild carnivores (red foxes) and evidencing a southerly dissemination. Future investigations should be focused on determining the local distribution and density of the insect vector (Phortica variegata) in this geographical area. This emergent zoonosis should be included by veterinarians, physicians and ophthalmologists in the differential diagnosis of ocular manifestations in their patients, particularly in areas where T. callipaeda is endemic.
- Improved phylogeny of brown algae Cystoseira (Fucales) from the Atlantic-Mediterranean region based on mitochondrial sequencesPublication . Bruno De Sousa, Carolina; Cox, Cymon; Brito, Luís; Pavão, Maria Madalena; Pereira, Hugo; Ferreira, Ana; Ginja, Catarina; Campino, Lenea; Bermejo, Ricardo; Parente, Manuela; J. C. or Varela J. or Varela J.C.S., VarelaCystoseira is a common brown algal genus widely distributed throughout the Atlantic and Mediterranean regions whose taxonomical assignment of specimens is often hampered by intra- and interspecific morphological variability. In this study, three mitochondrial regions, namely cytochrome oxidase subunit 1 (COI), 23S rDNA (23S), and 23S-tRNAVal intergenic spacer (mt-spacer) were used to analyse the phylogenetic relationships of 22 Cystoseira taxa (n = 93 samples). A total of 135 sequences (48 from COI, 43 from 23S and 44 from mt-spacer) were newly generated and analysed together with Cystoseira sequences (9 COI, 31 23S and 35 mt-spacer) from other authors. Phylogenetic analysis of these three markers identified 3 well-resolved clades and also corroborated the polyphyletic nature of the genus. The resolution of Cystoseira taxa within the three clades improves significantly when the inclusion of specimens of related genera was minimized. COI and mt-spacer markers resolved the phylogeny of some of the Cystoseira taxa, such as the C. baccata, C. foeniculacea and C. usneoides. Furthermore, trends between phylogeny, embryonic development and available chemotaxonomic classifications were identified, showing that phylogenetic, chemical and morphological data should be taken into account to study the evolutionary relationships among the algae currently classified as Cystoseira. The resolution of Cystoseira macroalgae into three well supported clades achieved here is relevant for a more accurate isolation and identification of natural compounds and the implementation of conservation measures for target species.
- In vitro susceptibility of Leishmania infantum to Artemisinin derivatives and selected trioxolanesPublication . Cortes, Sofia; Albuquerque, Andreia; Cabral, Lília; Lopes, Liliana; Campino, Lenea; Cristiano, Maria L. S.Leishmaniasis is among the world's most neglected diseases. Currently available drugs for treatment present drawbacks, urging the need for more effective, safer, and cheaper drugs. A small library of artemisinin-derived trioxanes and synthetic trioxolanes was tested against promastigote and intramacrophage amastigote forms of Leishmania infantum. The trioxolanes LC50 and LC95 presented the best activity and safety profiles, showing potential for further studies in the context of leishmanial therapy. Our results indicate that the compounds tested exhibit peroxide-dependent activity.
- Leishmania infection and host-blood feeding preferences of phlebotomine sandflies and canine leishmaniasis in an endemic European area, the Algarve Region in PortugalPublication . Maia, Carla; Dionisio, Lidia; Afonso, Maria Odete; Neto, Luis; Cristovao, Jose Manuel; Campino, LeneaThe Algarve Region (AR) in southern Portugal, which is an international tourist destination, has been considered an endemic region of zoonotic leishmaniasis caused by Leishmania infantum since the 1980s. In the present study, phlebotomine and canine surveys were conducted to identify sandfly blood meal sources and to update the occurrence of Leishmania infection in vectors and dogs. Four sandfly species were captured: Phlebotomus perniciosus, Phlebotomus ariasi, Phlebotomus sergenti and Sergentomyia minuta. In one P. perniciosus female, L. infantum DNA was detected. Blood meal tests showed that this species had no host preferences and was an opportunistic feeder. An overall canine leishmaniasis (CanL) seroprevalence of 16.06% was found; the seroprevalence was 3.88% in dogs housed in kennels and 40.63% in dogs that attended veterinary clinics. The simultaneous occurrence of dogs and P. perniciosus infected with L. infantum in the AR indicates that the region continues to be an endemic area for CanL. Our results reinforce the need for the systematic spatial distribution of phlebotomine populations and their Leishmania infection rates and the need to simultaneously perform pathogen monitoring in both invertebrate and vertebrate hosts to investigate the transmission, distribution and spreading of Leishmania infection.
- Molecular detection of Leishmania DNA and identification of blood meals in wild caught phlebotomine sand flies (Diptera: Psychodidae) from southern PortugalPublication . Maia, Carla; Parreira, Ricardo; Cristovao, Jose Manuel; Freitas, Ferdinando Bernardino; Afonso, Maria Odete; Campino, LeneaBackground: Zoonotic visceral leishmaniasis caused by Leishmania infantum which is transmitted by phlebotomine sand flies (Diptera, Psychodidae) is endemic in the Mediterranean basin. The main objectives of this study were to (i) detect Leishmania DNA and (ii) identify blood meal sources in wild caught female sand flies in the zoonotic leishmaniasis region of Algarve, Portugal/Southwestern Europe. Methods: Phlebotomine sand flies were collected using CDC miniature light traps and sticky papers. Sand flies were identified morphologically and tested for Leishmania sp. by PCR using ITS-1 as the target sequence. The source of blood meal of the engorged females was determined using the cyt-b sequence. Results: Out of the 4,971 (2,584 males and 2,387 females) collected sand flies, Leishmania DNA was detected by PCR in three females (0.13%), specifically in two specimens identified on the basis of morphological features as Sergentomyia minuta and one as Phlebotomus perniciosus. Haematic preferences, as defined by the analysis of cyt-b DNA amplified from the blood-meals detected in the engorged female specimens, showed that P. perniciosus fed on a wide range of domestic animals while human and lizard DNA was detected in engorged S. minuta. Conclusions: The anthropophilic behavior of S. minuta together with the detection of Leishmania DNA highlights the need to determine the role played by this species in the transmission of Leishmania parasites to humans. In addition, on-going surveillance on Leishmania vectors is crucial as the increased migration and travelling flow elevate the risk of introduction and spread of infections by Leishmania species which are non-endemic.
- Molecular detection of Leishmania infantum, filariae and Wolbachia spp. in dogs from southern PortugalPublication . Maia, Carla; Altet, Laura; Serrano, Lorena; Cristovao, Jose Manuel; Dolores Tabar, Maria; Francino, Olga; Cardoso, Luis; Campino, Lenea; Roura, XavierBackground: Leishmaniosis caused by the protozoan Leishmania infantum and dirofilariosis caused by the nematodes Dirofilaria immitis or Dirofilaria repens are vector-borne zoonoses widely present in the Mediterranean basin. In addition, some studies reported that the endosymbiont Wolbachia spp. play a role in the biology and pathogenesis of filarial parasites. The aim of this work was to evaluate the frequency of mono-and co-infections by L. infantum, filariae and Wolbachia spp. and their association with clinical signs in dogs from the south of Portugal. Leishmanial, filarial and Wolbachia spp. DNA were evaluated by specific real-time polymerase chain reaction (qPCR) assays in blood samples from 230 dogs.Findings: One hundred and thirty-nine (60.4 %) dogs were qPCR-positive for L. infantum and 26 (11.3 %) for filariae (24 for D. immitis only, one D. immitis and for Acanthocheilonema dracunculoides and another one for Acanthocheilonema reconditum only). Wolbachia spp. DNA was amplified from 16 (64.0 %) out of the 25 D. immitis-positive dogs. Nineteen (8.3 %) dogs were co-infected with L. infantum and D. immitis, including the one (0.4 %) A. drancunculoides-positive animal. In dogs without clinical signs consistent with leishmaniosis and/or dirofilariosis, L. infantum prevalence was 69 %, whereas in those dogs with at least one clinical manifestation compatible with any of the two parasitoses prevalence was 42.7 %. Leishmania prevalence was significantly higher in apparently healthy mongrels (77.2 %) and pets (76.9 %) than in defined-breed dogs (including crosses; 58.8 %) and in dogs with an aptitude other than pet (i.e. farm, guard, hunting, shepherd or stray), respectively, whereas in those dogs with at least one clinical sign, the detection of L. infantum DNA was higher in males (53.3 %) and in those dogs not receiving insect repellents (52.8 %).Conclusions: The molecular detection of canine vector-borne disease (CVBD) agents, some of which are zoonotic, reinforces the need to implement efficient prophylactic measures, such as insect repellents and macrocyclic lactones (including compliance to administration), in the geographical areas where these agents are distributed, with the view to prevent infection and disease among mammalian hosts including humans.