Browsing by Author "Milhano, N."
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- Detection of Borrelia lusitaniae, Rickettsia sp. IRS3, Rickettsia monacensis and Anaplasma phagocytophilum in Ixodes ricinus collected in Madeira Island, PortugalPublication . Lopes de Carvalho, Isabel; Milhano, N.; Santos, A.S.; Almeida, V.; Barros, S.C.; De Sousa, R.; Núncio, M.S.A total of 300 Ixodes ricinus ticks were tested by polymerase chain reaction (PCR) for the presence of Borrelia spp., Rickettsia spp., and Anaplasma phagocytophilum. Sequence analysis demonstrated 8 (2.7%) ticks infected with B. lusitaniae, 60 (20%) with Rickettsia spp., and 1 (0.3%) with A. phagocytophilum. Seven (2.3%) ticks were coinfected with B. lusitaniae and Rickettsia spp., 2 (0.6%) with R. monacensis, and 5 (1.7%) with Rickettsia sp. IRS3. The results of this study suggest simultaneous transmission of multiple tick-borne agents on Madeira Island, Portugal.
- Distribution, abundance and ecology of ticks in Portugal mainland: data from five years of a surveillance program REVIVEPublication . Santos-Silva, M.M.; Santos, A.; Lopes de Carvalho, I.; Sousa, R.; Luz, T.; Parreira, L.; Chainho, L.; Gomes, M.S.; Milhano, N.; Osório, H.; Alves, M.J.; Núncio, M.S.; REVIVE WorkgroupREVIVE (National Network for Vector Surveillance) aims to: i) Monitor the activity of hematophagous arthropods; ii) Characterize the species and its seasonal occurrence; iii) Identify important pathogens in Public Health, depending on the density of the vectors, the level of infection or the introduction of exotic species to alert for control measures.
- PCR screening of tick-borne agents in sensitive conservation areas, Southeast PortugalPublication . Santos-Silva, M.M.; Melo, P.; Santos, N.; Antunes, S.; Duarte, L.R.; Ferrolho, J.; Milhano, N.; Santos, P.T.; Domingos, A.; Santos, A.S.The Southeast region of Portugal, particularly the Guadiana valley, is currently the reintroduction territory of Lynx pardinus (Iberian lynx), one of the most endangered felids in the world that is only found in the Iberian Peninsula. Over the last century, populations have declined, placing L. pardinus at extremely high risk of extinction in the wild and relying on reintroduction projects. Among the aspects taken into account in the establishment of new populations is the sanitary status of the selected habitats, especially concerning infectious diseases, including tick-borne pathogens (TBPs). This study presents the results of TBPs survey on ticks collected at sensitive conservation areas of Southeast Portugal. From 2012 to 2014, 231 ticks obtained from vegetation, sympatric domestic and wild animals were submitted for analysis. The presence of Babesia spp., Cytauxzoon spp., Theileria spp., Hepatozoon spp., Anaplasma spp., Ehrlichia spp., Candidatus Neoehrlichia mikurensis, among other Anaplasmataceae, and Coxiella burnetii were investigated by PCR. Six tick species were recorded, Dermacentor marginatus (n = 13/5.6%), Hyalomma lusitanicum (n = 175/75.8%), Ixodes ricinus (n = 4/1.7%), Rhipicephalus bursa (n = 7/3.0%), R. pusillus (n = 21/9.1%) and R. sanguineus sensu lato (n = 11/4.8%). The molecular screening confirmed the presence of two tick-borne pathogens, C. burnetii (N = 34) and Anaplasma platys (N = 1), and one tick-endosymbiont, Candidatus Midichloria mitochondrii (N = 45). The results obtained provide new information on the circulation of ticks and TBPs with potential veterinary importance in Iberian lynx habitat.
- REVIVE, a surveillance program on vectors and vector-borne pathogens in Portugal - four year experience on ticksPublication . Santos, A.S.; Santos Silva, M.; Lopes de Carvalho, I.; Milhano, N.; Chaínho, L.; Luz, T.; Parreira, P.; Gomes, S.; De Sousa, R.; Núncio, M.S.; REVIVE WorkgrupREVIVE is a national wide surveillance program on vector and vector-borne agents implement and coordinate by the National Institute of Health (CEVDI/INSA) in collaboration with other institutions of the Health Ministry. The programme started in 2008 with the surveillance of mosquitoes and later in 2011 was extended to ticks. The main goals of this project are to collect and identify vectors, updating our knowledge in the distribution, hostassociations, seasonality and abundance of the Portuguese species. Additionally this project contributes for monitoring the introduction of exotic vector species. This work regards the 4-year REVIVE studies on ticks and Borrelia/Rickettsia surveillance, among other tick-borne agents, discussing the established circuits, obtained results and practical interventions. Over 29.000 ticks were collected on hosts or by flagging vegetation from 168 (60.4%) municipalities of mainland Portugal. Collection in humans reached the 583 specimens. In total, 13 autochthonous tick species were identified, including Dermacentor marginatus; D. reticulatus; Haemaphysalis punctata; Hyalomma lusitanicum; H. marginatum; Ixodes canisuga; I. hexagonus; I. ricinus; I. ventalloi; Rhipicephalus annulatus; R. bursa; R. pusillus; R. sanguineus. Of note is the identification of an exotic species, Amblyomma sp., attached to a Portuguese emigrant arriving from USA. The top three species collected during this surveillance program were R. sanguineus (69%), followed by R. pusillus (16.4%) and H. marginatum (9.7%). However regarding antropofilic behaviour, from the 11 species found in humans the most prevalent were I. ricinus (35%), followed by R. sanguineus (34%), and H. marginatum (14%). The abundance, distribution, host association and other relevant patterns are compared with previous existing records. Regarding the tick-borne agents, all ticks collected from humans and about 10% of the questing/host-attached ticks were tested for Borrelia and Rickettsia spp., among other agents. Ten bacteria were identified so far in single or multiple infection, including Borrelia afzelii, B. garinii, B. lusitaniae, Rickettsia aeschlimannii, R. conorii, R. helvetica, R. massiliae, R. monacensis, R. raoulti, and R. slovaca. The importance of including other tick-borne agents in routine screening is also discussed. The presented data reinforces the importance of the REVIVE. The program has contributed to call attention to tick-borne diseases not only among healthcare providers but also in the populations. The workflow established, has also enabled timely screeningof ticks removed from humans, animals or in a given environment, allowing the implementation of informed prevention/control strategies and directly contributing to improve Public Health in Portugal.
- Rickettsia lusitaniae sp. nov. isolated from the soft tick Ornithodoros erraticus (Acarina: Argasidae)Publication . Milhano, N.; Palma, M.; Marcili, A.; Núncio, M.S.; Lopes de Carvalho, I.; Sousa, R.In this study a novel Rickettsia from the spotted fever group, isolated from Ornithodoros erraticus soft ticks collected from pigpens in the south of Portugal, is described. After initial screening revealed Rickettsia-positive ticks, isolation attempts were then performed. Successful isolates were achieved by shell-vial technique using Vero E6 cells at 28°C. Molecular characterization of the isolate was performed based on analysis of five rickettsial genes gltA, ompA, ompB, sca1 and htr with their subsequent concatenation along with other rickettsial species resulting in a clustering of the new isolate with Rickettsia felis and Rickettsia hoogstraalii. The degree of nucleotide sequence similarity with other rickettsiae fulfills the criteria for classification of our isolate as a novel species. The name Rickettsia lusitaniae sp. nov. (=CEVDI PoTiRo) is proposed for this new species found in O. erraticus.
- The role of Rhipicephalus sanguineus sensu lato saliva in the dissemination of Rickettsia conorii in C3H/HeJ micePublication . Milhano, N.; Saito, T.B.; Bechelli, J.; Fang, R.; Vilhena, M.; De Sousa, Rita; Walker, D.H.Animal models have been developed for the study of rickettsial pathogenesis. However, to understand what occurs during the natural route of rickettsial transmission via the tick bite, the role of tick saliva should be considered in these models. To address this, we analysed the role of tick saliva in the transmission of Rickettsia conorii (Rickettsiales: Rickettsiaceae) in a murine host by intradermally (i.d.) inoculating two groups of susceptible C3H/HeJ mice with this Rickettsia, and infesting one group with nymphal Rhipicephalus sanguineus sensu lato (Ixodida: Ixodidae) ticks. Quantification of bacterial loads and mRNA levels of interleukin-1β (IL-1β), IL-10 and NF-κB was performed in C3H/HeJ lung samples by real-time quantitative polymerase chain reaction (PCR) and real-time reverse transcriptase PCR, respectively. Lung histology was examined to evaluate the pathological manifestations of infection. No statistically significant difference in bacterial load in the lungs of mice was observed between these two groups; however, a statistically significant difference was observed in levels of IL-1β and NF-κB, both of which were higher in the group inoculated with rickettsiae but not infected with ticks. Lung histology in both groups of animals revealed infiltration of inflammatory cells. Overall, this study showed that i.d. inoculation of R. conorii caused infection in the lungs of C3H/HeJ mice and tick saliva inhibited proinflammatory effects.