A novel SDS-stable dimer of a heterogeneous nuclear ribonucleoprotein at presynaptic terminals of squid neurons

Author Posting. © The Author(s), 2015. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Neuroscience 300 (2015): 381-392, doi:10.1016/j.neuroscience.2015.05.040.The presence of mRNAs in synaptic terminals and their regulated translation are important factors in neuronal communication and plasticity. Heterogeneous nuclear ribonucleoprotein (hnRNP) complexes are involved in the translocation, stability, and subcellular localization of mRNA and the regulation of its translation. Defects in these processes and mutations in components of the hnRNP complexes have been related to the formation of cytoplasmic inclusion bodies and neurodegenerative diseases. Despite much data on mRNA localization and evidence for protein synthesis, as well as the presence of translation machinery, in axons and presynaptic terminals, the identity of RNA-binding proteins involved in RNA transport and function in presynaptic regions is lacking. We previously characterized a strongly basic RNA-binding protein (p65), member of the hnRNP A/B subfamily, in squid presynaptic terminals. Intriguingly, in SDS-PAGE, p65 migrated as a 65 kDa protein, whereas members of the hnRNP A/B family typically have molecular masses ranging from 35 to 42 kDa. In this report we present further biochemical and molecular characterization that shows endogenous p65 to be an SDS-stable dimer composed of ~37 kDa hnRNPA/B-like subunits. We cloned and expressed a recombinant protein corresponding to squid hnRNPA/B-like protein and showed its propensity to aggregate and form SDS-stable dimers in vitro. Our data suggest that this unique hnRNPA/B-like protein co-localizes with synaptic vesicle protein 2 and RNA-binding protein ELAV and thus may serve as a link between local mRNA processing and presynaptic function and regulation.Research was supported by grants to REL from the Fundação de Amparo à Pesquisa do Estado de Sao Paulo (FAPESP), the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and the Fundação de Apoio ao Ensino, Pesquisa e Assistência do Hospital das Clínicas da FMRP-USP (FAEPA). JAD received financial support from the RI-INBRE Program Grant #8 P20 GM103430-12 from the National Institute of General Medical Sciences, NIH, Bethesda, MD. DTPL and GSL received research fellowships from FAPESP and CNPq. REL and JCR received the Productivity-in-Research fellowship from CNPq

Worldwide occurrence of feline hemoplasma infections in wild felid species

While hemoplasma infections in domestic cats are well studied, almost no information is available on their occurrence in wild felids. The aims of the present study were to investigate wild felid species as possible reservoirs of feline hemoplasmas and the molecular characterization of the hemoplasma isolates. Blood samples from the following 257 wild felids were analyzed: 35 Iberian lynxes from Spain, 36 Eurasian lynxes from Switzerland, 31 European wildcats from France, 45 lions from Tanzania, and 110 Brazilian wild felids, including 12 wild felid species kept in zoos and one free-ranging ocelot. Using real-time PCR, feline hemoplasmas were detected in samples of the following species: Iberian lynx, Eurasian lynx, European wildcat, lion, puma, oncilla, Geoffroy's cat, margay, and ocelot. "Candidatus Mycoplasma haemominutum" was the most common feline hemoplasma in Iberian lynxes, Eurasian lynxes, Serengeti lions, and Brazilian wild felids, whereas "Candidatus Mycoplasma turicensis" was the most prevalent in European wildcats; hemoplasma coinfections were frequently observed. Hemoplasma infection was associated with species and free-ranging status of the felids in all animals and with feline leukemia virus provirus-positive status in European wildcats. Phylogenetic analyses of the 16S rRNA and the partial RNase P gene revealed that most hemoplasma isolates exhibit high sequence identities to domestic cat-derived isolates, although some isolates form different subclusters within the phylogenetic tree. In conclusion, 9 out of 15 wild felid species from three different continents were found to be infected with feline hemoplasmas. The effect of feline hemoplasma infections on wild felid populations needs to be further investigated

The golden jackal (Canis aureus): A new host for Echinococcus multilocularis and Trichinella britovi in Switzerland.

INTRODUCTION The golden jackal (Canis aureus) is a wild canid new to Switzerland. It is an officially monitored species and all deceased individuals are submitted for post-mortem examination to collect baseline health data. This includes parasitological examinations, with an emphasis on zoonotic, reportable infections, such as those caused by Trichinella spp. or Echinococcus spp. From 2016 to 2021, five golden jackals originating from four Swiss cantons were submitted for full post-mortem examination. In one case only organ samples were available, and therefore parasitological examination was not possible. Parasite stages recovered during necropsy, as well as by routine coproscopical techniques, were morphologically identified. Taeniid eggs and adult tapeworms were processed for molecular species identification. Additionally, tongue and diaphragm were analysed for Trichinella spp. by the artificial digestion technique followed by multiplex-PCR in positive cases. Of the four jackals investigated for parasites, hookworm eggs were detected in one animal, both adult worms and eggs of Echinococcus multilocularis were present in another case, and one animal was free of parasites. Eggs of E. multilocularis as well as eggs of Toxocara canis and sporocysts of Sarcocystis sp. were detected in the intestinal content, and Trichinella britovi larvae were found in the muscle samples of the last case. The health monitoring programme in place for protected carnivores in Switzerland allowed us to add the golden jackal to the list of hosts for the endemic zoonotic parasites E. multilocularis and T. britovi in this country. Hunters, farmers, and other persons who could come in contact with golden jackals should be aware of the associated health risk and handle faeces and carcasses with caution

Estimation of the number of synapses in the hippocampus and brain-wide by volume electron microscopy and genetic labeling

Determining the number of synapses that are present in different brain regions is crucial to understand brain connectivity as a whole. Membrane-associated guanylate kinases (MAGUKs) are a family of scaffolding proteins that are expressed in excitatory glutamatergic synapses. We used genetic labeling of two of these proteins (PSD95 and SAP102), and Spinning Disc confocal Microscopy (SDM), to estimate the number of fluorescent puncta in the CA1 area of the hippocampus. We also used FIB-SEM, a three-dimensional electron microscopy technique, to calculate the actual numbers of synapses in the same area. We then estimated the ratio between the three-dimensional densities obtained with FIB-SEM (synapses/µm) and the bi-dimensional densities obtained with SDM (puncta/100 µm). Given that it is impractical to use FIB-SEM brain-wide, we used previously available SDM data from other brain regions and we applied this ratio as a conversion factor to estimate the minimum density of synapses in those regions. We found the highest densities of synapses in the isocortex, olfactory areas, hippocampal formation and cortical subplate. Low densities were found in the pallidum, hypothalamus, brainstem and cerebellum. Finally, the striatum and thalamus showed a wide range of synapse densities.This work was supported by grants from the following entities: the Spanish “Ministerio de Ciencia, Innovación y Universidades” (Grant PGC2018-094307-B-I00 and the Cajal Blue Brain Project [C080020-09; the Spanish partner of the Blue Brain Project initiative from EPFL, Switzerland]; the European Union’s Horizon 2020 Research and Innovation Programme under grant agreement No. 785907 (Human Brain Project, SGA2); the Wellcome Trust (Technology Development Grant 202932); and the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (695568 SYNNOVATE). L.T.-R. is a recipient of grants from the EMBO Long-term fellowship 2016–2018 and the IBRO-PERC InEurope grants programme

Harmonizing methods for wildlife abundance estimation and pathogen detection in Europe-a questionnaire survey on three selected host-pathogen combinations

__Background:__ The need for wildlife health surveillance as part of disease control in wildlife, domestic animals and humans on the global level is widely recognized. However, the objectives, methods and intensity of existing wildlife health surveillance programs vary greatly among European countries, resulting in a patchwork of data that are difficult to merge and compare. This survey aimed at evaluating the need and potential for data harmonization in wildlife health in Europe. The specific objective was to collect information on methods currently used to estimate host abundance and pathogen prevalence. Questionnaires were designed t

Temporal stability in the genetic structure of Sarcoptes scabiei under the host-taxon law: empirical evidences from wildlife-derived Sarcoptes mite in Asturias, Spain

<p>Abstract</p> <p>Background</p> <p>Implicitly, parasite molecular studies assume temporal genetic stability. In this study we tested, for the first time to our knowledge, the extent of changes in genetic diversity and structure of <it>Sarcoptes </it>mite populations from Pyrenean chamois (<it>Rupicapra pyrenaica</it>) in Asturias (Spain), using one multiplex of 9 microsatellite markers and <it>Sarcoptes </it>samples from sympatric Pyrenean chamois, red deer (<it>Cervus elaphus</it>), roe deer (<it>Capreolus capreolus</it>) and red fox (<it>Vulpes vulpes</it>).</p> <p>Results</p> <p>The analysis of an 11-years interval period found little change in the genetic diversity (allelic diversity, and observed and expected heterozygosity). The temporal stability in the genetic diversity was confirmed by population structure analysis, which was not significantly variable over time. Population structure analysis revealed temporal stability in the genetic diversity of <it>Sarcoptes </it>mite under the host-taxon law (herbivore derived- and carnivore derived-<it>Sarcoptes </it>mite) among the sympatric wild animals from Asturias.</p> <p>Conclusions</p> <p>The confirmation of parasite temporal genetic stability is of vital interest to allow generalizations to be made, which have further implications regarding the genetic structure, epidemiology and monitoring protocols of the ubiquitous <it>Sarcoptes </it>mite. This could eventually be applied to other parasite species.</p

Feline Leukemia Virus and Other Pathogens as Important Threats to the Survival of the Critically Endangered Iberian Lynx (Lynx pardinus)

BACKGROUND: The Iberian lynx (Lynx pardinus) is considered the most endangered felid species in the world. In order to save this species, the Spanish authorities implemented a captive breeding program recruiting lynxes from the wild. In this context, a retrospective survey on prevalence of selected feline pathogens in free-ranging lynxes was initiated. METHODOLOGY/ PRINCIPAL FINDINGS: We systematically analyzed the prevalence and importance of seven viral, one protozoan (Cytauxzoon felis), and several bacterial (e.g., hemotropic mycoplasma) infections in 77 of approximately 200 remaining free-ranging Iberian lynxes of the Doñana and Sierra Morena areas, in Southern Spain, between 2003 and 2007. With the exception of feline immunodeficiency virus (FIV), evidence of infection by all tested feline pathogens was found in Iberian lynxes. Fourteen lynxes were feline leukemia virus (FeLV) provirus-positive; eleven of these were antigenemic (FeLV p27 positive). All 14 animals tested negative for other viral infections. During a six-month period in 2007, six of the provirus-positive antigenemic lynxes died. Infection with FeLV but not with other infectious agents was associated with mortality (p<0.001). Sequencing of the FeLV surface glycoprotein gene revealed a common origin for ten of the eleven samples. The ten sequences were closely related to FeLV-A/61E, originally isolated from cats in the USA. Endogenous FeLV sequences were not detected. CONCLUSIONS/SIGNIFICANCE: It was concluded that the FeLV infection most likely originated from domestic cats invading the lynx's habitats. Data available regarding the time frame, co-infections, and outcome of FeLV-infections suggest that, in contrast to the domestic cat, the FeLV strain affecting the lynxes in 2007 is highly virulent to this species. Our data argue strongly for vaccination of lynxes and domestic cats in and around lynx's habitats in order to prevent further spread of the virus as well as reduction the domestic cat population if the lynx population is to be maintained