Effects of immunosuppressive drugs on COVID-19 severity in patients with autoimmune hepatitis.

BACKGROUND: We investigated associations between baseline use of immunosuppressive drugs and severity of Coronavirus Disease 2019 (COVID-19) in autoimmune hepatitis (AIH). PATIENTS AND METHODS: Data of AIH patients with laboratory confirmed COVID-19 were retrospectively collected from 15 countries. The outcomes of AIH patients who were on immunosuppression at the time of COVID-19 were compared to patients who were not on AIH medication. The clinical courses of COVID-19 were classified as (i)-no hospitalization, (ii)-hospitalization without oxygen supplementation, (iii)-hospitalization with oxygen supplementation by nasal cannula or mask, (iv)-intensive care unit (ICU) admission with non-invasive mechanical ventilation, (v)-ICU admission with invasive mechanical ventilation or (vi)-death and analysed using ordinal logistic regression. RESULTS: We included 254 AIH patients (79.5%, female) with a median age of 50 (range, 17-85) years. At the onset of COVID-19, 234 patients (92.1%) were on treatment with glucocorticoids (n = 156), thiopurines (n = 151), mycophenolate mofetil (n = 22) or tacrolimus (n = 16), alone or in combinations. Overall, 94 (37%) patients were hospitalized and 18 (7.1%) patients died. Use of systemic glucocorticoids (adjusted odds ratio [aOR] 4.73, 95% CI 1.12-25.89) and thiopurines (aOR 4.78, 95% CI 1.33-23.50) for AIH was associated with worse COVID-19 severity, after adjusting for age-sex, comorbidities and presence of cirrhosis. Baseline treatment with mycophenolate mofetil (aOR 3.56, 95% CI 0.76-20.56) and tacrolimus (aOR 4.09, 95% CI 0.69-27.00) were also associated with more severe COVID-19 courses in a smaller subset of treated patients. CONCLUSION: Baseline treatment with systemic glucocorticoids or thiopurines prior to the onset of COVID-19 was significantly associated with COVID-19 severity in patients with AIH.

Great tit response to decreasing industrial heavy metal emissions.

This study evaluated the effects of environmental pollution on decreasing great tit (Parus major) breeding parameters in an industrial area in the west coast of Portugal. Several great tit breeding parameters were monitored in the industrial area, as well as in a rural area not affected by industrial emissions. Low levels of air pollution in both industrial and rural areas were confirmed by assessing trace element concentrations in pine needles. Concentrations of Cd and Hg in pine needles from the industrial area (Cd = 0.05 ppm; Hg = 0.005 ppm) were significantly lower than those found in needles collected from the reference area (Cd = 0.07 ppm; Hg = 0.007 ppm). Additionally, the breeding success of great tits increased in the industrial area in comparison to the reference area (0.93 ± 0.08 and 0.62 ± 0.22, respectively). The changes on great tit breeding parameters were probably related with changes in the breeding density of other competitive species, together with a decreasing frass-fall biomass. Further long-term ecological studies in industrial areas are necessary to understand the changing breeding performance and strategies used by great tits in response to pollution shifts in the environment.

Assessing heavy metal pollution using Great Tits (Parus major): feathers and excrements from nestlings and adults.

Passerine species have been increasingly used as bioindicators of metal bioaccumulation especially by taking benefit of non-invasive procedures, such as collecting feathers and excrements. In 2009, metal (As, Cd, Cu, Hg, Ni, Pb, Se and Zn) concentrations were determined in feathers and excrements of nestling and adult female great tits (Parus major) in industrial (a paper mill) and rural sites in maritime pine forests on the west coast of Portugal. The aim of this study was to compare the levels of metals between the areas but also between sampling methods (feather vs. excrement) and age classes (nestling vs. adult). Although excrements and feathers of nestling great tits showed different concentrations, similar patterns of accumulation were detected in both study areas. There was a significantly higher concentration of mercury in the industrial area and significantly higher concentrations of arsenic in the rural area in both sample types. Metal levels in adult females had quite different results when compared to nestlings, and only nickel presented significantly higher levels near the paper mill. Since metal levels showed a consistent pattern in feathers and excrements of nestling great tits, we conclude that both represent good and non-invasive methods for the evaluation of these elements in polluted areas.

Trace elements in faeces of great tit nestlings in relation to breeding performance in coastal areas in central Portugal.

This long-term study (2003-2010) compared the breeding parameters of great tits living in a paper-and-pulp-industry area to those of great tits living in a rural area on the west coast of Portugal. We also measured the abundance of caterpillar biomass, an important food source and determinant of breeding success for tits. In 2009, we further analysed trace metal [arsenic (As), calcium (Ca), cadmium, copper, mercury (Hg), nickel, lead, selenium, and zinc] as well as Ca concentrations in excrement of 15-day-old great tit nestlings. Generally, for most trace metals, fecal concentrations were similar at both sites. Nonetheless, greater Hg levels and lower As levels were detected in the industrial area. Great tits laid more eggs and produced more fledglings in the industrial area than in the rural area. Caterpillar biomass was also greater in the industrial area, which likely explains the better breeding success. Our results suggest that there are no direct effects of emissions on the studied species.

Prevalence of entanglements of seabirds in marine debris in the central Portuguese coast.

Plastic pollution and the subsequent entanglement of marine animals is a global and increasing problem. In this study we present an analysis of the seabirds recorded as entangled by a rehabilitation centre and an associated marine animal stranding network, along the central coast of Portugal, between 2008 and 2018. Results show a high annual rate of entangled seabirds (average 6.9%) compared to other studies and fisheries related materials are a relevant cause of seabird entanglement (82%) compared to other debris. When comparing age classes, juveniles were more vulnerable to entanglement than other age classes in the species studied. Regarding the rehabilitation of entangled seabirds, the release rate was higher in non-fishing material entanglement cases. In conclusion, this study highlights the impact of fisheries related material on marine fauna and the need for reinforcement of the existing legislation for protecting seabirds and the implementation of mitigation measures associated with fishing activities.

Helminth species richness in wild wood mice, Apodemus sylvaticus, is enhanced by the presence of the intestinal nematode Heligmosomoides polygyrus.

We analysed 3 independently collected datasets of fully censused helminth burdens in wood mice, Apodemus sylvaticus, testing the a priori hypothesis of Behnke et al. (2005) that the presence of the intestinal nematode Heligmosomoides polygyrus predisposes wood mice to carrying other species of helminths. In Portugal, mice carrying H. polygyrus showed a higher prevalence of other helminths but the magnitude of the effect was seasonal. In Egham, mice with H. polygyrus showed a higher prevalence of other helminth species, not confounded by other factors. In Malham Tarn, mice carrying H. polygyrus were more likely to be infected with other species, but only among older mice. Allowing for other factors, heavy residual H. polygyrus infections carried more species of other helminths in both the Portugal and Egham data; species richness in Malham was too low to conduct a similar analysis, but as H. polygyrus worm burdens increased, so the prevalence of other helminths also increased. Our results support those of Behnke et al. (2005), providing firm evidence that at the level of species richness a highly predictable element of co-infections in wood mice has now been defined: infection with H. polygyrus has detectable consequences for the susceptibility of wood mice to other intestinal helminth species.

Trace element concentrations in Proteocephalus macrocephalus (Cestoda) and Anguillicola crassus (Nematoda) in comparison to their fish host, Anguilla anguilla in Ria de Aveiro, Portugal.

The use of some fish parasites as bioindicators of heavy metal pollution has been demonstrated as particularly adequate due to their capacity of bioconcentration. This study evaluated the effect of Proteocephalus macrocephalus on the accumulation of trace elements in the edible fish, Anguilla anguilla, in a contaminated area in Portugal (Ria de Aveiro). Also, the model P. macrocephalus/A. anguilla was assessed as a bioindicator system in the presence of the highly prevalent nematode Anguillicola crassus. Samples (kidney, liver, muscle, A. crassus and P. macrocephalus) of 20 eels harbouring A. crassus and another 20 harbouring both A. crassus and P. macrocephalus were selected for element analysis by ICP-MS. The highest concentrations of Cr, Ni and Zn were detected in P. macrocephalus. However, there was a higher liver and muscle Cr concentration in eels not infected by P. macrocephalus. Also, the nematode A. crassus presented higher Cr concentrations in those eels harbouring P. macrocephalus. Results suggest that P. macrocephalus individuals accumulate Cr and Ni while levels of Cr in eel livers and Ni levels in eel kidney are reduced. The system P. macrocephalus/A. anguilla yielded bioaccumulation factors for Cr, Ni, Pb and Zn, whereas bioaccumulation of Cu, Cr and Pb in A. crassus varied according to eel co-infection with P. macrocephalus, thus emphasising the possible role of cestode infection in metal metabolization/storage processes in host tissues. Results suggest that heavy metal pollution in Ria de Aveiro has been decreasing although it is still higher than in other contaminated areas in Europe. Nevertheless, eel consumption in Ria de Aveiro represents no risk for humans although they may represent a real contamination risk for wildlife. The system P. macrocephalus/A. anguilla is proposed as another promising bioindicator system to evaluate environmental Cr, Ni, Pb and Zn exposure in estuarine areas where both species co-occur.

Toxic element concentrations in the Razorbill Alca torda (Charadriiformes, Alcidae) in Portugal.

The present study provides the first data on inorganic element levels (As, Cd, Co, Cr, Cu, Hg, Mn, Pb, Se, and Zn) in juvenile, immature, and adult razorbills (Alca torda) collected along the central coast of Portugal. Element concentrations were assessed by ICP-MS in kidney, liver, muscle, and feathers of 28 razorbills, including 4 juveniles, 17 subadults, and 7 adults. The effect of age and tissue on element accumulation was also assessed. The detected levels in razorbills may indicate a possible contamination risk by Hg and Cr. With respect to bird tissues, higher accumulation of Se and Cd was detected in kidney, Zn and Pb in feathers, and As and Mn in liver. Age was found to affect the accumulation of Cd, Cr, Cu, Hg, and Mn, juveniles presenting higher levels of Cu and Mn than older individuals. In razorbill kidney, Zn-Hg constituted the most significant relationship among metal concentrations. Liver presented the highest number of significant relationships (mostly involving Zn and Co). With regard to feathers, the most significant relationships involved Se, Zn, Cr and Cu concentrations. Positive linear relationships were detected among kidney, liver, and muscle, with emphasis on relationships involving Se and Hg, which may be indicative of similar accumulation/regulation mechanisms in those organs. Element concentrations are discussed in view of possible detoxification mechanisms in seabirds.

Herpesvirus infection in marine mammals: A retrospective molecular survey of stranded cetaceans in the Portuguese coastline.

Herpesvirus (HVs) infection has already been reported in cetaceans, but available information on its epidemiology is scarce. In this study we surveyed a total of 179 cetaceans belonging to 6 different species. Samples were obtained from cetaceans stranded along the Portuguese coastline, belonging to populations that roam the north-east region of the Atlantic Ocean. Detection of HVs was performed by conventional nested PCR. Amplicons were sequenced by Sanger's method and sequences used to construct phylogenetic trees by Maximum Likelihood method. Our results show that prevalence of positive samples, among fresh carcasses, reached 14.3% (10/70) and both alpha and gammaherpesvirus were detected. Histopathology showed that herpesvirus infection varied from absence of signs compatible with disease, localized genital lesions and systemic disease. Phylogenetic analysis revealed three clusters within the alphaherpesvirus family; within the gammaherpesvirus no subdivision was detected. All clusters included animals from different species and geographic origins. In seven of the positive HVs samples, co-infections with other agents such as morbillivirus and toxoplasma gondii were detected. The viral nucleotide sequences were not assigned to a specific animal species, nor presented a given geographic distribution, which may imply a wider distribution of herpesvirus in these animal populations. Our results are also the first report of herpesvirus infection in common dolphins (Delphinus delphis), with both alpha and gammaherpesvirus detected.

Toxic element concentration in the Atlantic gannet Morus bassanus (Pelecaniformes, Sulidae) in Portugal.

The present study provides the first data on inorganic element levels in juvenile, subadult, and adult Atlantic gannets (Morus bassanus). Physiological and potentially toxic elements (As, Cd, Co, Cr, Cu, Hg, Mn, Pb, Se, and Zn) were assessed by ICP-MS in kidney, liver, muscle, and feathers of 31 gannets, including 18 juveniles, 7 subadults, and 6 adults. The effect of age and tissue on element accumulation was also assessed. Mercury was roughly above the minimum level for adverse effects in birds. A higher accumulation of Se and Cd was detected in kidney, Pb in feathers, and Mn in liver. Age was found to affect the accumulation of Cd, Co, Hg, Mn, Se, and Zn. Adults presented significantly lower levels of Mn, Se, and Zn than subadults. Linear positive relationships within tissues were detected involving Se-Cd and Se-Hg. Also, positive linear relationships were detected among kidney, liver, and muscle, with emphasis on relationships involving Cd, Hg, Se, and Zn, which may be indicative of analogous regulation mechanisms in those organs. Atlantic gannets occurring in the study area leave their reproduction sites as juveniles. During their development process, several molting cycles occur and thus the possible contamination risk by Hg should reflect levels in the development areas rather than contamination levels in reproduction areas. The present study provides basic information on multielement accumulation in Morus bassanus, which may help us to understand the behavior and toxicity of various elements in marine birds.

Cranial helminths of Mustela vison Schreber, 1777 in Spain.

A survey was carried out to investigate the presence of cranial helminths in 337 American minks (Mustela vison) from Spain. This information was obtained partly in order to evaluate potential conservation problems and sanitary risks to the congeneric European mink (Mustela lutreola), one of the most endangered carnivores in the world. Skulls and rectal faeces of each specimen were simultaneously analysed. Troglotrema acutum and Skrjabingylus nasicola were found in 5.6% of the M. vison analysed. No cranial lesions were seen in any of the examined skulls. The finding of both helminths in Spanish free-living M. vison specimens enlarges their natural definitive host spectrum in Western Europe. One relatively important focus of T. acutum in M. vison was detected (30.4%) in the Spanish Alava province while S. nasicola was found to be very infrequent. The suitability of both analytical methods was assessed in order to know to what degree coprological analysis reflects the real prevalence of cranial helminths in this host. It is possible to conclude that coprological analysis can be used instead of necropsies to analyse the possible incidence of pathogenic cranial helminths in mustelids. This aspect is very important and useful when trying to analyse the helminthological status of endangered species such as the native mink (M. lutreola) particularly in areas where both congeneric species are present and strict competition occurs.

Persistent organic pollutants and inorganic elements in the Balearic shearwater Puffinus mauretanicus wintering off Portugal.

This study presents the first data on trace element and organic pollutant concentrations in the Critically Endangered Balearic shearwater Puffinus mauretanicus collected in 2010 and 2011 in Portugal. Trace element levels were below the threshold levels for adverse effects on birds, despite the Hg concentrations in feathers (4.35µg·g-1ww). No significant differences were detected between individuals from 2010 and 2011 except for Se concentrations in liver, feathers and muscle (higher in 2010) and Ag in liver and muscle (higher in 2011). No significant differences were detected in total concentrations of organochlorine compounds in Balearic shearwaters between years, although PCB congeners -101 and -180 presented higher concentrations in individuals from 2010. The PCB congeners -138, -153 and -180, and 4.4-DDE were detected in all individuals. This study on toxic elements and organic pollutants in wintering Balearic shearwaters provides baseline data from which deviations can be detected in the future.

Concentration of some toxic elements in Oryctolagus cuniculus and in its intestinal cestode Mosgovoyia ctenoides, in Dunas de Mira (Portugal).

The need for sentinel organisms reflecting small-scale changes in heavy metal pollution of different habitats has been previously stated and the role of terrestrial mammalian parasites has been pointed out as an important field of research aiming at the potential use of parasitic models as bioindicators. The scope of the present study was to assess the concentration of some toxic (Cd, Pb, As and Hg) elements in Dunas de Mira (Natura 2000 PTCON055) while testing the model constituted by the wild rabbit (Oryctolagus cuniculus Linnaeus, 1758) and its intestinal cestode parasite Mosgovoyia ctenoides Railliet, 1890 as potential bioindicators. Wild rabbits were harvested by hunters and analysed for intestinal cestodes. Samples of kidney, liver, intestinal mucosa, muscle of host and also M. ctenoides were taken and deep-frozen for posterior element analysis by ICP-MS. In general, levels of contamination in rabbit tissues were found to be low although some of the elements were detected in higher quantities when compared to other previous reports for wildlife in Europe. The highest quantity of Pb was found in rabbit muscle (3.81 ppm) while highest Cd and Hg values were found in kidney (1.02 and 0.08 ppm). Significant linear relationships were found between Pb concentration in the parasite and Pb concentration in kidney (P=0.0047), muscle (P=0.0002) and intestinal mucosa (P=0.0181) and between As concentration in the parasite and As concentration in liver (P=0.0024), kidney (P=0.0010), muscle (P=0.0003) and intestinal mucosa (P=0.0047). The most significant relationships for Pb and As were detected between element concentration in the cestode and in host muscle. However, maximum Pb and As concentrations in M. ctenoides were only twice as high as those found for rabbit kidney and therefore it is not possible to confirm the role of the model O. cuniculus/M. ctenoides as a promising bioindicator system unlike other mammalian host/cestode models tested in other studies. The contrasting results might be due to differences in the absorption processes in the tegument of different cestode families. Furthermore, the large size of the cestodes used in the present study might also explain the relatively low element concentration values obtained.

Vitamin E (alpha-tocopherol) production by the marine microalgae Dunaliella tertiolecta and Tetraselmis suecica in batch cultivation.

Batch experiments with Dunaliella tertiolecta and Tetraselmis suecica were performed to investigate alpha-tocopherol (alpha-T) production in time, in order to assess the effect of light availability per cell on the production of this antioxidant. In D. tertiolecta alpha-T content increased during growth, in other words, as the cell density increased and the light availability per cell decreased. In T. suecica the pattern was different: alpha-T content was highest during the exponential phase, decreased significantly during the linear phase and increased again towards the end of the cultivation. Chlorophyll (chl-a) content of T. suecica cells decreased after the exponential phase, instead of the expected increase typically observed in shade-adapted cells, suggesting that the culture was nutrient limited. An experiment with extra nutrients showed that chl-a and alpha-T content increased significantly during both the linear and the stationary phase when compared with values in nutrient-deficient conditions. No correlation between alpha-T and chl-a was observed. Our results indicate that diminished light availability does not limit alpha-T production in D. tertiolecta and T. suecica, opening up the possibility of using high cell density, light-limited cultures for the production of this commercially interesting compound.

The helminth parasites of the wild rabbit Oryctolagus cuniculus and their effect on host condition in Dunas de Mira, Portugal.

The present study focuses on the helminth parasite community of the wild rabbit in a sand dune area in Portugal over a 5-year period. The influence of host sex and year on the composition of the helminth community is assessed, along with the potential effect of the detected helminths on host body condition. The basic structure of the helminth community comprises Mosgovoyia ctenoides, Graphidium strigosum, Trichostrongylus retortaeformis, Nematodiroides zembrae and Passalurus ambiguus. Mean intensities of G. strigosum varied between years. General G. strigosum intensities were also found to vary according to both year and host sex, but not according to the interaction of both factors. When assessing the effect of helminths on rabbit body condition (expressed by the kidney fat index), higher burdens of M. ctenoides, a cestode that presents a relatively large body mass, were found to induce a reduction in rabbit condition.

Cadmium and lead concentrations in Gallegoides arfaai (Cestoda: Anoplocephalidae) and Apodemus sylvaticus (Rodentia: Muridae) from Spain.

Information on parasites of vertebrates living in terrestrial ecosystems as sentinels for heavy metal environmental pollution is scarce. The aim of the present study was to assess the concentration of cadmium and lead using the model Apodemus sylvaticus/Gallegoides arfaai in order to test the potential suitability of G. arfaai as a sentinel organism for lead and cadmium under natural field conditions. Samples of 15 A. sylvaticus as well as whole specimens of G. arfaai were analysed for both elements by inductively coupled plasma mass spectrometry. The level of cadmium in G. arfaai was always much lower than in the tissues of A. sylvaticus. Contrarily, values for lead in G. arfaai were found to be 6, 20 and 24-fold higher than in the kidney, liver and muscle of A. sylvaticus. We propose the model A. sylvaticus/G. arfaai as a promising bioindication system to evaluate environmental lead exposure in terrestrial habitats, especially for non-urban areas.