Gene expression changes with tumor disease and leech parasitism in the juvenile green sea turtle skin transcriptome.

Emerging infectious diseases are a major threat to biodiversity in the 21st century. Fibropapillomatosis (FP) is an epithelial tumor disease that affects immature and adult marine turtles worldwide, particularly green turtles (Chelonia mydas). We know little about the host factors contributing to FP susceptibility, in part because transcriptomic studies that compare transcript expression in turtles with and without FP are lacking. Here, we performed RNA-Seq on healthy skin tissue from immature C. mydas in the Indian River Lagoon, Florida, USA, comparing turtles (1) with and without FP and (2) with and without leech parasites, a putative vector of FP. We assembled a de novo C. mydas skin transcriptome to identify transcripts with significant differential expression (DE) across FP and leech categories. Significant DE transcripts were found across FP and leech comparisons, including 10 of the same transcripts with DE across both comparisons. Leech-positive individuals significantly upregulated different immune and viral interaction transcripts than did leech-negative individuals, including viral interaction transcripts associated with herpesvirus interactions. This finding strengthens the role of marine leeches as mechanical vectors of Chelonid herpesvirus 5 (ChHV5) which has been implicated as a causative agent of FP. FP-positive turtles upregulated several tumor progression and suppression transcripts relative to FP-negative turtles, which had no significant DE tumor progression transcripts. FP-positive turtles also upregulated significantly more protein interaction transcripts than FP-negative turtles. DE transcripts across leech comparisons showed no functional enrichment, whereas DE transcripts across FP comparisons showed some GO terms were enriched in FP-positive and FP negative turtles. Notably, only FP-negative turtles were enriched for GO terms involved in acquired and inflammatory immune gene regulation. Overall, our DE transcripts included several candidate genes that may play important roles in C. mydas resistance to or recovery from FP, highlighting that transcriptomics provides a promising venue to understand this impactful disease. Continued investigation of C. mydas responses to FP and leech affliction is imperative for species persistence and the conservation of marine ecosystems worldwide due to the essential role of sea turtles in ecosystem function and stability.

Developmental assays using invasive cane toads, Rhinella marina, reveal safety concerns of a common formulation of the rice herbicide, butachlor.

Identifying the adverse impacts of pesticide exposure is essential to guide regulations that are protective of wildlife and human health. Within rice ecosystems, amphibians are valuable indicators because pesticide applications coincide with sensitive reproductive and developmental life stages. We conducted two experiments using wild cane toads (Rhinella marina) to test 1) whether environmentally relevant exposure to a commercial formulation of butachlor, an acetanilide herbicide used extensively in rice, affects amphibian development and 2) whether cane toad tadpoles are capable of acclimatizing to sub-lethal exposure. First, we exposed wild cane toads to 0.002, 0.02, or 0.2 mg/L of butachlor (Machete EC), during distinct development stages (as eggs and hatchlings, as tadpoles, or continuously) for 12 days. Next, we exposed a subset of animals from the first experiment to a second, lethal concentration and examined survivorship. We found that cane toads exposed to butachlor developed slower and weighed less than controls, and that development of the thyroid gland was affected: exposed individuals had smaller thyroid glands and thyrocyte cells, and more individual follicles. Analyses of the transcriptome revealed that butachlor exposure resulted in downregulation of transcripts related to metabolic processes, anatomic structure development, immune system function, and response to stress. Last, we observed evidence of acclimatization, where animals exposed to butachlor early in life performed better than naïve animals during a second exposure. Our findings indicate that the commercial formulation of butachlor, Machete EC, causes thyroid endocrine disruption in vertebrates, and suggest that exposure in lowland irrigated rice fields presents a concern for wildlife and human health. Furthermore, we establish that developmental assays with cane toads can be used to screen for adverse effects of pesticides in rice fields.

Genetic delimitation of Pristimantisorestes (Lynch, 1979) and P.saturninoi Brito et al., 2017 and description of two new terrestrial frogs from the Pristimantisorestes species group (Anura, Strabomantidae).

In the genus Pristimantis, species are often combined into taxonomic units called species groups. The taxonomy of these groups is frequently inaccurate due to the absence of genetic data from type series and repeated misidentifications generated by high morphological resemblance between taxa. Here, we focus on the P.orestes species group, providing the first genetic assessment of P.orestes sensu stricto from individuals collected from the type locality, with a reviewed diagnosis and description of advertisement calls. We find that two lineages previously named P.orestes are genetically distinct and should be separated into two different species. Based on genetic and morphological data, we name one of these species P.cajanuma sp. nov. This new species is morphologically distinct from other members of the group by having shagreen dorsal skin, evident dorsolateral folds, broader discs on toes and fingers and pale gray ventral coloration. Additionally, P.saturninoi is placed within the P.orestes species group based on genetic data from its type series. However, we find that one of its paratypes is genetically distinct and belongs to a clade containing a new species we name P.quintanai sp. nov. This new species is morphologically distinguished by lacking a tympanic membrane and vocal sacs in males, and by having expanded discs on toes and fingers, finely tuberculated dorsal skin and irregular white or cream spots in the groin and concealed surfaces of thighs. Our findings highlight the importance of providing genetic characterization and placement from the type series in taxonomic challenging groups, such as Pristimantis. We also suggest that the diversity of species within the P.orestes group will increase as more sampling is achieved in the southern Andes of Ecuador.

ResumenLas especies pertenecientes al género Pristimantis usualmente están agrupadas en unidades taxonómicas llamadas grupos de especies. A menudo la taxonomía de estos grupos es problemática debido a la ausencia de información genética de las series tipo de las especies o debido a identificaciones erróneas generadas por la elevada similitud morfológica entre especies. Aquí nos enfocamos en el grupo de especies P.orestes y proveemos la primera evaluación genética de P.orestes sensu stricto en base a individuos colectados en la localidad tipo de la especie con una diagnosis revisada y descripción de vocalizaciones. Encontramos que dos linajes previamente nombrados como P.orestes son genéticamente distintos y deberían ser considerados como dos distintas especies. En base a evidencia genética y morfológica nombramos a una de estas especies P.cajanuma sp. nov. La nueva especie es distinta de otras especies del grupo por presentar piel dorsal con textura finamente granular, pliegues dorsolaterales evidentes, discos amplios en dedos de pie y manos y una coloración ventral gris pálido. Adicionalmente, P.saturninoi es colocada dentro del grupo de especies P.orestes en base a información genética de especímenes tipo. Sin embargo, encontramos que uno de los paratipos es genéticamente distinto y está dentro de un clado que incluye a una nueva especie morfológicamente similar que nombramos como P.quintanai sp. nov. Esta nueva especie se distingue de otros Pristimantis del grupo por carecer de una membrana timpánica diferenciada, machos sin sacos vocales y por presentar discos expandidos en los dedos de pies y manos, una piel dorsal con textura finamente tubercular y manchas irregulares blancas o crema-blanquecinas en las ingles y superficies ocultas de los muslos. Nuestros resultados resaltan la importancia de proveer caracterizaciones genéticas de especímenes tipos en grupos taxonómicamente complejos como los Pristimantis. Sugerimos que la diversidad de especies dentro del grupo de especies P.orestes incrementara a medida que más expediciones de campo se realicen en el sur de los Andes de Ecuador.

Gene expression plasticity across hosts of an invasive scale insect species.

For plant-eating insects, we still have only a nascent understanding of the genetic basis of host-use promiscuity. Here, to improve that situation, we investigated host-induced gene expression plasticity in the invasive lobate lac scale insect, Paratachardina pseudolobata (Hemiptera: Keriidae). We were particularly interested in the differential expression of detoxification and effector genes, which are thought to be critical for overcoming a plant's chemical defenses. We collected RNA samples from P. pseudolobata on three different host plant species, assembled transcriptomes de novo, and identified transcripts with significant host-induced gene expression changes. Gene expression plasticity was pervasive, but the expression of most detoxification and effector genes was insensitive to the host environment. Nevertheless, some types of detoxification genes were more differentially expressed than expected by chance. Moreover, we found evidence of a trade-off between expression of genes involved in primary and secondary metabolism; hosts that induced lower expression of genes for detoxification induced higher expression of genes for growth. Our findings are largely consonant with those of several recently published studies of other plant-eating insect species. Thus, across plant-eating insect species, there may be a common set of gene expression changes that enable host-use promiscuity.

Discrete nuclear structures in actively growing neuroblastoma cells are revealed by antibodies raised against phosphorylated neurofilament proteins.

BACKGROUND: Nuclear objects that have in common the property of being recognized by monoclonal antibodies specific for phosphoprotein epitopes and cytoplasmic intermediate filaments (in particular, SMI-31 and RT-97) have been reported in glial and neuronal cells, in situ and in vitro. Since neurofilament and glial filaments are generally considered to be restricted to the cytoplasm, we were interested in exploring the identity of the structures labeled in the nucleus as well as the conditions under which they could be found there. RESULTS: Using confocal microscopy and western analysis techniques, we determined 1) the immunolabeled structures are truly within the nucleus; 2) the phosphoepitope labeled by SMI-31 and RT-97 is not specific to neurofilaments (NFs) and it can be identified on other intermediate filament proteins (IFs) in other cell types; and 3) there is a close relationship between DNA synthesis and the amount of nuclear staining by these antibodies thought to be specific for cytoplasmic proteins. Searches of protein data bases for putative phosphorylation motifs revealed that lamins, NF-H, and GFAP each contain a single tyrosine phosphorylation motif with nearly identical amino acid sequence. CONCLUSION: We therefore suggest that this sequence may be the epitope recognized by SMI-31 and RT-97 mABs, and that the nuclear structures previously reported and shown here are likely phosphorylated lamin intermediate filaments, while the cytoplasmic labeling revealed by the same mABs indicates phosphorylated NFs in neurons or GFAP in glia.