Maria Sofia Núncio

Genotypic evaluation of rickettsial isolates recovered from various species of ticks in Portugal

Twelve rickettsial isolates, from Rhipicephalus sanguineus, R. turanicus, Dermacentor marginatus and Hyalomma marginatus, were subjected to genotypic analysis. Amplification of specific DNA sequences, restriction endonuclease digestion of amplified DNA products, and gel electrophoresis were used to identify specific DNA fragment-banding patterns. Five patterns were resolved. Four were homologous with those of previously described rickettsial genotypes, R. conorii, R. slovaca, R. rhipicephali and R. massiliae. The fifth pattern differed by only a single altered restriction endonuclease cleavage site. For the first time in Portugal a widely distributed spectrum of spotted fever group rickettsia was found among potential vector species stressing the need to determine their potential for human and domestic animals infection.

Francisella-Like Endosymbiont in Dermacentor reticulatus Collected in Portugal

In Portugal, recent studies have confirmed the presence of Francisella tularensis in Dermacentor reticulatus. Bacterial endosymbionts with significant homology to F. tularensis have been described in several species of ticks. In this work we identified Francisella-like endosymbionts in D. reticulatus ticks (39%), confirming the presence of these bacteria in Portugal. This finding should be considered in future studies using molecular approaches to detect Francisella prevalence in ticks and environmental samples.

Rickettsia lusitaniae sp. nov. isolated from the soft tick Ornithodoros erraticus (Acarina: Argasidae)

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.

Palma Virus, a New Bunyaviridae Isolated from Ticks in Portugal

An agent pathogenic for laboratory albino Swiss mice was isolated from a pool of Haemaphysalis punctata ticks collected from cattle on a farm located in Alcácer do Sal county, southern Portugal. The isolated virus was shown to be distinct from but serologically related to virus members of the Bhanja antigenic group. This new virus in the family Bunyaviridae was named Palma for the farm where ticks have been collected for several studies.

Molecular Characterization of a New Isolate of Borrelia lusitaniae Derived from Apodemus sylvaticus in Portugal

A total of 196 small mammals were collected in Portugal and tested for Borrelia burgdorferi sensu lato. Tissue samples were taken from each animal and cultured in Barbour-Stoenner-Kelly (BSK)-II medium. The single strain of spirochete isolated was confirmed as Borrelia lusitaniae by genetic analyses. This is the first report of B. lusitaniae isolated from Apodemus sylvaticus.

Portuguese Hosts for Ornithodoros erraticus Ticks

The hematophagous soft tick Ornithodoros erraticus feeds nocturnally on multiple warm-blooded vertebrate hosts. This tick is often found living buried in the soil of traditional pigpens. O. erraticus is an important infectious disease vector both for humans and animals. In the Iberian Peninsula, this tick serves as the vector of human tick-borne relapsing fever caused by the spirochete Borrelia hispanica. The natural ecosystems maintaining this spirochete are not well understood, with details of competent vertebrate reservoirs and tick-host interactions poorly understood. Investigation of arthropod blood meal composition provides evidence linking the vector to specific hosts, providing insights into possible disease reservoirs. Ticks collected from two pigpens located in southern Portugal were subjected to blood meal analysis. PCR amplification of vertebrate cytochrome b was used to disclose the original host from which 349 ticks had derived their previous blood meal. Host origins for blood meal analysis from 79 of 349 ticks revealed that 46.8% had previously fed from pigs, 35.4% human, 13.9% bovine, 5.1% sheep, 1.3% rodent, and 1.3% from birds. Three samples revealed mixed blood meals, namely, human-pig (1.3%), sheep-pig (1.3%), and bovine-pig (1.3%). The major role of pigs as hosts is consistent with fieldwork observations and underlines the importance of pigs for maintaining O. erraticus tick populations. Humans serve as accidental hosts, frequently confirmed by reports from both producers and veterinarians. Other livestock species and wildlife prevalent in the region appear only to have a minor role in maintaining this tick. The results demonstrate the importance of blood meal analysis to determine tick hosts providing a tool for investigation of sylvatic cycle for Borrelia hispanica.

Implementation of bioterrorism agents diagnosis by electron microscopy in Portuguese reference laboratory

between the duration of intubation and biofilm stage. Of the 17 positive cases with biofilm formation, colonization of the inner ETT surface occurred in 14 (70%) cases and 17 isolates were recovered. Five isolates were Gram positive, whereas the majority of isolates were Gram-negative bacilli. Seventeen patients developed pneumonia. All patients who developed biofilm also developed VAP. The occurrence of multidrug resistance among detected microorganisms was high. We concluded that the density of ETT biofilm increased with increased duration of intubation. We also concluded that ETT colonization with biofilm-producing organisms increased the risk of developing VAP with highly resistant microbes.