Bacterial septicemia and herpesvirus infection in Antarctic fur seals (Arctocephalus gazella) stranded in the São Paulo coast, Brazil.

In August 2021, two juvenile male Antarctic fur seals (Arctocephalus gazella) stranded in the southeastern Brazilian coast and were referred to rehabilitation centers. The animals presented increased body temperature, prostration, respiratory distress and despite treatment died. A necropsy following a standardized protocol was performed, and formalin-fixed tissues were processed for microscopic examination. Samples were screened for morbillivirus, herpesvirus, and Brucella spp. by molecular analyses (PCR, RT-PCR). Bacteriological culture was performed in samples collected from the lungs, trachea, and lymph nodes of both cases. The main histopathologic findings were of infectious nature, including multifocal necrotizing and fibrinous mixed interstitial pneumonia, bronchiolitis, and bronchitis, with intralesional myriad bacteria associated with vascular fibrinoid necrosis. Pseudomonas aeruginosa was isolated from tracheal and lung swabs of Case 1, and Klebsiella oxytoca was found in nostril swabs, tracheobronchial lymph nodes, and lung of Case 2. Gammaherpesvirus infection was detected in both cases, and the sequences retrieved were classified into the genus Percavirus. All tested samples were PCR-negative for Brucella spp. and morbillivirus. We hypothesize that the deficient immunological status in association with starvation predisposed the reactivation of herpesvirus and secondary bacterial co-infections. To the authors' knowledge, this is the first molecular detection of herpesvirus in an Antarctic pinniped. These findings reinforce that Otariid gammaherpesvirus circulating in the Southern Hemisphere are likely endemic in the Arctocephalus genus. This report contributes to the current knowledge of health aspects affecting wild pinnipeds, especially in the poorly studied Antarctic species.

Facial Squamous Cell Carcinoma and Abdominal Peripheral Nerve Sheath Tumour with Rhabdomyoblastic Differentiation in a Rough-toothed Dolphin (Steno bredanensis).

We report the pathological features of a facial squamous cell carcinoma (SCC) and an abdominal peripheral nerve sheath tumour (PNST) with rhabdomyoblastic differentiation in an aged free-ranging rough-toothed dolphin (Steno bredanensis). The animal was found stranded dead in poor body condition. On external examination, there was a 25 × 7 × 3 cm extensively ulcerated area on the right maxillary region of the rostrum, involving the oral mucocutaneous junction with prominent nodular edges, severe soft tissue loss and extensive maxillary and premaxillary bone lysis. On abdominal dissection, a 5 × 4 × 3.5 cm pale tan to red, raised mass expanded the inner aspect of the right transverse abdominis muscle. Microscopically, the aggressive facial lesion was an acantholytic SCC with extensive osteolysis; there was no evidence of metastasis in the tissues examined. The abdominal mass had cytohistomorphological features compatible with a localized PNST, including whorling, Antoni A and Antoni B areas and Verocay bodies intermixed with rhabdomyoblastic components, as suggested by phosphotungstic acid haematoxylin stain. This neoplasm was locally infiltrative, yet no metastases were observed in the tissues examined. No immunohistochemical investigations could be performed due to lack of tissue availability. Total DNA from the formalin-fixed and paraffin wax-embedded SCC was extracted and tested by polymerase chain reaction for herpesvirus and papillomavirus genetic material. There was no amplification for either of these genera. Other pathological findings observed in this animal were related to the 'live-stranding stress response'. The severity and extent of the facial SCC likely related to anorexia and poor body condition and might have played a role in the stranding and death of this dolphin. These two tumour subtypes add to the relatively uncommon reports of neoplasia in cetaceans. Specifically, these appear to be the first neoplasia records for rough-toothed dolphins, including the first documentation of a PNST with features compatible with rhabdomyoblastic differentiation in a marine mammal.

The Negro River (Ancash-Peru): A unique case of water pollution, three environmental scenarios and an unresolved issue.

This paper is focused on the hydrogeochemical characterization of the Negro River along its course, as well as in the proposal of a functioning model for the contamination processes in order to establish potential cause-effect relationships between water quality, geology (ARD), mining activities (AMD) and the tectonic framework as transmission vector of acidity, metals and sulphates. The scenario shows a heavily-contaminated river compared to the unaffected regional background. By graphical and statistical treatments of physico-chemical data of Negro River and the unaffected values of regional background and other AMD/ARD representative rivers' it is possible to conclude that Antamina Mine, is not the cause of the Negro River contamination, without the need of isotopic tracers, but just through the inexistent concentrations of Cu, Bi and Mo found in the waters. In the proposed contamination model, climatic factors (glacial retreat) activate geological (ARD) processes. The tectonic scenario (faults) intervenes as a transport medium of the contamination flux from the sulphide oxidation surface in upper altitudes until the spring in lower altitudes. At the end, it is concluded that this contamination comes from the recent glacial retreat in areas near the Cordillera Blanca that has left massive amounts of sulphide materials exposed to weathering conditions, oxidizing naturally (ARD processes) and finally contributing to the contamination of the Negro River through faults. In this case, we would face an ARD process in the strict sense, which is the direct oxidation of sulphides outcropping in the upper part of the mountain with the generation of sulphates, the release of hydrogen ions and the consequent generation of acid and the dissolution of the metals. This ARD process would come from the glacial retreat, which, through the faults, transports contaminated water until the spring.

Application of fuzzy logic tools for the biogeochemical characterisation of (un)contaminated waters from Aljustrel mining area (South Portugal).

Aljustrel mining area (South Portugal) belongs to the Iberian Pyrite Belt (IPB). It is classified of high environmental risk due to its large tailings and to the Acid Mine Drainage (AMD) affected waters, generated by sulphides' oxidation. Integrating biological parameters (for the first time) in the input data matrix of the software PreFuRGe, allowed a better discrimination of the diatoms' responses to the stimuli caused by the hydrochemical changes imposed by the processes affecting water quality. Each hydrochemical scenario, was modeled by imposing maximum and minimum limits for each antecedent, according to the conditions imposed by the consequent, which in this case were the number of diatom species and pH. Thus, PreFuRGe evidenced some qualitative aspects that could not be achieved by classic statistics. pH appeared as the main discriminator of diversity and diatom species composition, nevertheless and due to the complex environment under study other chemical interactions must be considered: (a) AMD waters, with extremely low pH values, but also with extremely high hydrogeochemical complexity, represented by a mixture of metals, do not allow to associate, unequivocally, the reduction in diatom diversity to pH, but also to high metal (loid)s concentrations; (b) in the most alkaline waters, with higher abundance of diatom species, average to high concentrations of Na and Cl (due to Cenozoic sediments) do not seem to affect diatom diversity. This methodology proved to be an efficient tool to establish, for the first time, cause-effect relationships, improving the comprehension between biological (diatoms) and hydrochemical parameters.

Hydrogeochemical characteristics of the Tinto and Odiel Rivers (SW Spain). Factors controlling metal contents.

The Tinto and Odiel Rivers are strongly affected by acid mine drainage (AMD) due to the intense sulphide mining developed in their basins over the past 5000 years. In this study the results obtained from a weekly sampling in both rivers, before their mouth in the Ría of Huelva, over three and a half years of control are analysed. In the Tinto River, the concentrations of sulphates, Al, Cd, Co, Li and Zn are double to those of the Odiel as a consequence of lower dilution. However, the concentration of Fe in the Odiel River is 20 times lower, since the precipitation of Fe oxyhydroxysulphates caused by neutralisation processes is more intense. Lower As, Cr, Cu and Pb concentrations are also found in the Odiel River as, to a greater or lesser extent, they are sorbed and/or coprecipitated with Fe. Other elements such as Be, Mn, Ni and Mg show similar values in both systems, which is ascribed to lithological factors. The seasonal evolution of contaminants is typical of rivers affected by AMD, reaching a maximum in autumn due to the dissolution of evaporitic salts precipitated during the summer. Nevertheless, in the Tinto River, Ca, Na and Sr show a strong increase during the summer, probably due to a greater water interaction with marly materials, through which the last reach of the river flows. Barium has a different behaviour from the rest of the metals and its concentration seems to be controlled by the solubility of barite. Iron, As and Pb show different behaviours in both rivers, those for Fe and As possibly linked to the prevalence of different dissolved species of Fe. The different Pb pattern is probably due to the control of Pb solubility by anglesite or other minerals rich in Pb in the Tinto River.

Rare earth elements mobility processes in an AMD-affected estuary: Huelva Estuary (SW Spain).

Huelva Estuary is a transition zone where REE-rich acidic waters interact with saline-alkaline seawater. This mixing process influences the geochemical and mineralogical characteristics of particulate and dissolved fractions. The Tinto River has >11,000µgL-1 dissolved REE (pH=1.66), whereas seawater only reaches 8.75·10-2µgL-1 dissolved REE (pH=7.87). REE-normalized patterns in "pH<6 solutions" are parallel and show similarities, diminishing their concentration as pH increases. Sequential extraction performed on the generated precipitates of mixed solutions indicates that most REE are associated to the residual phase. In a second order, REE are associated with soluble salts at pH3 and 3.5 whereas in sediments generated at pH4 and 5, they are distributed in salts (1° extraction), poorly crystallized Fe-bearing minerals (schwertmannite, 3° extraction) and well crystallized Fe-bearing minerals (goethite - hematite, 4° extraction). Finally, precipitated REE are highest at pH6 newly formed minerals with a release to solution in higher pH.

Geochemical behavior of an acid drainage system: the case of the Amarillo River, Famatina (La Rioja, Argentina).

The Amarillo River (Famatina range, Argentina, ~29° S and ~67° W) is unusual because acid mine drainage (AMD) is superimposed on the previously existing acid rock drainage (ARD) scenario, as a Holocene paleolake sedimentary sequence shows. In a markedly oxidizing environment, its water is currently ferrous and of the sulfate-magnesium type with high electrical conductivity (>10 mS cm-1 in uppermost catchments). At the time of sampling, the interaction of the mineralized zone with the remnants of mining labors determined an increase in some elements (e.g., Cu ~3 to ~45 mg L-1; As ~0.2 to ~0.5 mg L-1). Dissolved concentrations were controlled by pH, decreasing significantly by precipitation of neoformed minerals (jarosite and schwertmannite) and subsequent metal sorption (~700 mg kg-1 As, 320 mg kg-1 Zn). Dilution also played a significant role (i.e., by the mixing with circumneutral waters which reduces the dissolved concentration and also enhances mineral precipitation). Downstream, most metals exhibited a significant attenuation (As 100 %, Fe 100 %, Zn 99 %). PHREEQC-calculated saturation indices (SI) indicated that Fe-bearing minerals, especially schwertmannite, were supersaturated throughout the basin. All positive SI increased through the input of circumneutral water. PHREEQC inverse geochemical models showed throughout the upper and middle basin, that about 1.5 mmol L-1 of Fe-bearing minerals were precipitated. The modeling exercise of mixing different waters yielded results with a >99 % of correlation between observed and modeled data.

Assessment of metal contamination, bioavailability, toxicity and bioaccumulation in extreme metallic environments (Iberian Pyrite Belt) using Corbicula fluminea.

The Iberian Pyrite Belt (SW Iberian Peninsula) has intense mining activity. Currently, its fluvial networks receive extremely acid lixiviate residue discharges that are rich in sulphates and metals in solution (acid mine drainage, AMD) from abandoned mines. In the current study, the sediment and water quality were analysed in three different areas of the Odiel River to assess the risk associated with the metal content and its speciation and bioavailability. Furthermore, sediment contact bioassays were performed using the freshwater clam Corbicula fluminea to determine its adequacy as a biomonitoring tool in relation to theoretical risk indexes and regulatory thresholds. Reburial activity and mortality were used as the toxic responses of clams when exposed to contaminated sediment. The results showed coherence between the water and sediment chemical contamination for most of the metals. The reburial activity was correlated with the metal toxicity, but no clam mortality was registered. The bioaccumulation of the studied metals in the clam did not have a significant correlation with the bioavailable fraction of the metal content in the environment, which could be related to a potential different speciation in this singular environment. The bioaccumulation responses were negative for As, Cd and Zn in highly contaminated environments and were characterized as severe, considerable and low potential environmental risks, respectively. The results show that C. fluminea is a good biomonitor of Cu and Pb.

Seasonal water quality variations in a river affected by acid mine drainage: the Odiel River (South West Spain).

This paper intends to analyse seasonal variations of the quality of the water of the Odiel River. This river, together with the Tinto River, drains the Iberian Pyrite Belt (IPB), a region containing an abundance of massive sulphide deposits. Because of mining activity dating back to prehistoric times, these two rivers are heavily contaminated. The Odiel and Tinto Rivers drain into a shared estuary known as the Ría of Huelva. This work studies dissolved contaminant data in water of the Odiel River collected by various organisations, between October 1980 and October 2002, close to the rivers entry into the estuary. Flow data for this location were also obtained. The most abundant metals in the water, in order of abundance, are zinc (Zn), iron (Fe), manganese (Mn) and copper (Cu). Arsenic (As), cadmium (Cd) and lead (Pb) are also present but in much lower quantities. The quality of the river water is linked to precipitation; the maximum sulphate, Fe, Zn, Mn, Cd and Pb concentrations occur during the autumn rains, which dissolve the Fe hydroxysulphates that were precipitated during the summer months. In winter, the intense rains cause an increase in the river flow, producing a dilution of the contaminants and a slight increase in the pH. During spring and summer, the sulphate and metal concentration (except Fe) recover and once again increase. The Fe concentration pattern displays a low value during summer due to increased precipitation of ferric oxyhydroxides. The arsenic concentration displays a different evolution, with maximum values in winter, and minimum in spring and summer as they are strongly adsorbed and/or coprecipitated by the ferric oxyhydroxides. Mn and sulphates are the most conservative species in the water. Relative to sulphate, Mn, Zn and Cd, copper displays greater values in winter and lower ones in summer, probably due to its coprecipitation with hydroxysulphates during the spring and summer months. Cd and Zn also appear to be affected by the same process, although to a lower degree than Cu, experiencing a slight reduction in summer with respect to Mn and sulphates.

[Fulminating malignant hyperthermia in swine--an alternative therapy concept]. / Die fulminante maligne Hyperthermie beim Schwein--Ein alternatives Therapiekonzept.

An inexpensive therapeutic concept compared to the dantrolene-therapy to counter the malignant hyperthermia (MH) is to be discussed, using a case-study from swine-anaesthesia. Hyperventilating the animals (with O2), administering metamizol, beta-blocker, bicarbonate and sufficiently cooling the patient can, if the symptoms are recognized early enough, arrest the hypermetabolic cascade in its track. All the animals that were treated according to this scheme survived the (MH) crisis without any lasting post operative damage.

Metal mobility and toxicity to microalgae associated with acidification of sediments: CO2 and acid comparison.

The injection and storage of CO2 into marine geological formations has been suggested as a mitigation measure to prevent global warming. However, storage leaks are possible resulting in several effects in the ecosystem. Laboratory-scale experiments were performed to evaluate the effects of CO2 leakage on the fate of metals and on the growth of the microalgae Phaeodactylum tricornutum. Metal contaminated sediments were collected and submitted to acidification by means of CO2 injection or by adding HCl. Sediments elutriate were prepared to perform toxicity tests. The results showed that sediment acidification enhanced the release of metals to elutriates. Iron and zinc were the metals most influenced by this process and their concentration increased greatly with pH decreases. Diatom growth was inhibited by both processes: acidification and the presence of metals. Data obtained is this study is useful to calculate the potential risk of CCS activities to the marine environment.

Metal cycling during sediment early diagenesis in a water reservoir affected by acid mine drainage.

The discharge of acid mine drainage (AMD) into a reservoir may seriously affect the water quality. To investigate the metal transfer between the water and the sediment, three cores were collected from the Sancho Reservoir (Iberian Pyrite Belt, SW Spain) during different seasons: turnover event; oxic, stratified period; anoxic and under shallow perennially oxic conditions. The cores were sliced in an oxygen-free atmosphere, after which pore water was extracted by centrifugation and analyzed. A sequential extraction was then applied to the sediments to extract the water-soluble, monosulfide, low crystallinity Fe(III)-oxyhydroxide, crystalline Fe(III)-oxide, organic, pyrite and residual phases. The results showed that, despite the acidic chemistry of the water column (pH<4), the reservoir accumulated a high amount of autochthonous organic matter (up to 12 wt.%). Oxygen was consumed in 1mm of sediment due to organic matter and sulfide oxidation. Below the oxic layer, Fe(III) and sulfate reduction peaks developed concomitantly and the resulting Fe(II) and S(II) were removed as sulfides and probably as S linked to organic matter. During the oxic season, schwertmannite precipitated in the water column and was redissolved in the organic-rich sediment, after which iron and arsenic diffused upwards again to the water column. The flux of precipitates was found to be two orders of magnitude higher than the aqueous one, and therefore the sediment acted as a sink for As and Fe. Trace metals (Cu, Zn, Cd, Pb, Ni, Co) and Al always diffused from the reservoir water and were incorporated into the sediments as sulfides and oxyhydroxides, respectively. In spite of the fact that the benthic fluxes estimated for trace metal and Al were much higher than those reported for lake and marine sediments, they only accounted for less than 10% of their total inventory dissolved in the column water.

Source and impact of lead contamination on δ-aminolevulinic acid dehydratase activity in several marine bivalve species along the Gulf of Cadiz.

Coastal areas and estuaries are particularly sensitive to metal contamination from anthropogenic sources and in the last few decades the study of space-time distribution and variation of metals has been extensively researched. The Gulf of Cadiz is no exception, with several rivers draining one of the largest concentrations of sulphide deposits in the world, the Iberian Pyrite Belt (IPB). Of these rivers, the Guadiana, one of the most important in the Iberian Peninsula, together with smaller rivers like the Tinto and Odiel, delivers a very high metal load to the adjacent coastal areas. The purpose of this work was to study the source and impact of lead (Pb) drained from historical or active mining areas in the IPB on the activity of a Pb inhibited enzyme (δ-aminolevulinic acid dehydratase, ALAD) in several bivalve species along the Gulf of Cadiz. Seven marine species (Chamelea gallina, Mactra corallina, Donax trunculus, Cerastoderma edule, Mytilus galloprovincialis, Scrobicularia plana and Crassostrea angulata) were collected at 12 sites from Mazagón, near the mouth of the rivers Tinto and Odiel (Spain), to Cacela Velha (Ria Formosa lagoon system, Portugal). Lead concentrations, ALAD activity and lead isotope ratios ((206)Pb/(204)Pb, (207)Pb/(204)Pb and (208)Pb/(204)Pb) were determined in the whole soft tissues. The highest Pb concentrations were determined in S. plana (3.50±1.09 µg g(-1) Pb d.w.) and D. trunculus (1.95±0.10 µg g(-1) Pb d.w.), while M. galloprovincialis and C. angulata showed the lowest Pb levels (<0.38 µg g(-1) Pb d.w.). In general, ALAD activity is negatively correlated with total Pb concentration. However this relationship is species dependent (e.g. linear for C. gallina ALAD=-0.36[Pb]+0.79; r=0.837; or exponential for M. galloprovincialis ALAD=2.48e(-8.3[Pb]); r=0.911). This indicates that ALAD activity has considerable potential as a biomarker of Pb and moreover, in marine bivalve species with different feeding habits. Lead isotope data showed significant seasonal and spatial changes in bivalve isotopic composition reflecting seasonal and geographic differences in bioaccumulation. Within the study area, Pb can be modelled as a mixing between geogenic Pb and mine-related, discharges of Pb from the IPB. For some sites at the mouth of the Guadiana River, the bivalves show contamination from other anthropogenic sources, such as leaded boat/aviation fuel and/or leaded paint. Finally, the study demonstrates convincingly the need to consider species-specific variation when using bivalve ALAD activity as a biomarker for Pb.

Lacaziosis-like disease in Tursiops truncatus from Brazil: a histopathological and immunohistochemical approach

ABSTRACT: Cetacean lacaziosis-like disease or lobomycosis-like disease (LLD) is a chronic skin condition caused by a non-cultivable yeast of the order Onygenales, which also includes Lacazia loboi, as well as Paracoccidioides brasiliensis and P. lutzii, respectively responsible for lacaziosis and paracoccidioidomycosis in humans. Complete identification and phylogenetic classification of the LLD etiological agent still needs to be elucidated, but preliminary phylogenetic analyses have shown a closer relationship of the LLD agent to Paracoccidioides spp. than to L. loboi. Cases of LLD in South American cetaceans based on photographic identification have been reported; however, to date, only 3 histologically confirmed cases of LLD have been described. We evaluated multiple tissue samples from 4 Tursiops truncatus stranded in the states of Santa Catarina (n = 3) and Rio Grande do Sul (n = 1), southern Brazil. Macroscopically, all animals presented lesions consistent with LLD. Hematoxylin-eosin, periodic acid-Schiff, Grocott's methenamine silver, and Mayer's mucicarmin stains were used for histological evaluation. Microscopically, numerous refractile yeasts (4-9 µm in diameter) were observed in skin samples (4/4), and for the first time in dolphins, also in a skeletal muscle abscess (1/4). Immunohistochemistry using anti-P. brasiliensis glycoprotein gp43 as a primary antibody, which is known to cross-react with L. loboi and the LLD agent, was performed and results were positive in all 4 cases. We describe 3 new cases of LLD in cetaceans based on histopathology and immunohistochemistry. This is the first report of LLD in the muscle of cetaceans.

Inorganic arsenic speciation at river basin scales: the Tinto and Odiel rivers in the Iberian Pyrite Belt, SW Spain.

The Tinto and Odiel rivers are heavily affected by acid mine drainage from mining areas in the Iberian Pyrite Belt. In this work we have conducted a study along these rivers where surface water samples have been collected. Field measurements, total dissolved metals and Fe and inorganic As speciation analysis were performed. The average total concentration of As in the Tinto river (1975 microg L(-1)) is larger than in the Odiel river (441 microg L(-1)); however, the mean concentration of As(III) is almost four times higher in the Odiel. In wet seasons the mean pH levels of both rivers (2.4 and 3.2 for the Tinto and Odiel, respectively) increase slightly and the amount of dissolved total arsenic tend to decrease, while the As(III)/(V) ratio strongly increase. Besides, the concentration of the reduced As species increase along the water course. As a result, As(III)/(V) ratio can be up to 100 times higher in the lower part of the basins. An estimation of the As(III) load transported by both rivers into the Atlantic Ocean has been performed, resulting in about 60 kg yr(-1) and 2.7t yr(-1) by the Tinto and Odiel rivers, respectively.

New preservation method for inorganic arsenic speciation in acid mine drainage samples.

A new preservation method has been proposed for the speciation of As(III) and As(V) in acid mine drainage (AMD) samples, characterised by low pH and high metallic content. Samples were taken from a polymetallic sulphides mining area in the province of Huelva (SW Spain), under exploitation until the 1960s for its Cu, Pb and Zn sulphides. The abandoned mine works and the numerous waste rocks heaps produce AMD with high As content, an aqueous pollution source for the nearby streams. Short-term (from few hours to 1 week) preservation of the two inorganic arsenic species was studied, trying different containers (polyethylene, glass), presence or absence of light, temperatures (ambient, refrigerated, frozen), preserving agents and procedures (EDTA, HCl or AcH acids, cation-exchange resin). The speciation results obtained by liquid chromatography-hydride generation-atomic fluorescence spectrometry (HPLC-HG-AFS) indicated a rapid conversion of the samples with most of the preservation procedures reported in the literature after 3h after sample collection. A promising method for arsenic preservation has been developed in this work, which maintains the arsenic species distribution in the original samples for a longer time. It consists in the use of opaque glass containers, acidification of the samples with HCl and in situ cleanup with cationic exchange resin, which allowed to preserve the samples for As speciation for at least 48 h.