Systematics and biogeography of the Atlantic Forest endemic genus Juliomys (Rodentia: Cricetidae): a test of diversification hypothesis using mitochondrial data

The Atlantic Forest harbors a large species richness and high levels of endemism, but the processes that shaped its biodiversity are poorly studied, especially for mammals. Among them are the endemic mice Juliomys, which comprise forest dwellers distributed in southeastern and southern Brazil, northeastern Argentina, and eastern Paraguay. In this study, we investigate the phylogenetic relationships among species and perform phylogeographic analyses to evaluate the population structure and demographic scenarios through mitochondrial gene cytochrome b sequences. We investigate three hypotheses of diversification (forest refuges, montane isolate, and geomorphological events) to understand the evolution of the Juliomys species. Phylogenetic analyses recovered five clades/lineages, four of which are congruent with species currently recognized. The fifth lineage expands the range of the genus 659 km to the north and may represent a new species. The observed demographic and geographic structure of genetic diversity does not match the forest refuge hypothesis as mechanism to explain the diversification in Juliomys. Our results recovered J. rimofrons and J. ximenezi as sister species, supporting predictions of montane isolate hypothesis. We also detected a shallow genetic structure in J. pictipes and J. ossitenuis. Both phylogeographic breaks were congruent with limits of the São Paulo Basin, an area that has undergone Neogene reactivations of tectonic faults. It is suggested that geomorphological events led to a deformed landscape that influenced the dynamics of sedimentary basins and promoted an incipient population structure in J. pictipes and J. ossitenuis. Our findings demonstrate that the divergences whithin Juliomys species occurred during the Quaternary, too recently to have produced strong geographic structure.

Systematics and biogeography of the atlantic forest endemic genus Juliomys (Rodentia: Cricetidae): A test of diversification hypothesis using mitochondrial data

The Atlantic Forest harbors a large species richness and high levels of endemism, but the processes that shaped its biodiversity are poorly studied, especially for mammals. Among them are the endemic mice Juliomys, which comprise forest dwellers distributed in southeastern and southern Brazil, northeastern Argentina, and eastern Paraguay. In this study, we investigate the phylogenetic relationships among species and perform phylogeographic analyses to evaluate the population structure and demographic scenarios through mitochondrial gene cytochrome b sequences. We investigate three hypotheses of diversification (forest refuges, montane isolate, and geomorphological events) to understand the evolution of the Juliomys species. Phylogenetic analyses recovered five clades/lineages, four of which are congruent with species currently recognized. The fifth lineage expands the range of the genus 659 km to the north and may represent a new species. The observed demographic and geographic structure of genetic diversity does not match the forest refuge hypothesis as mechanism to explain the diversification in Juliomys. Our results recovered J. rimofrons and J. ximenezi as sister species, supporting predictions of montane isolate hypothesis. We also detected a shallow genetic structure in J. pictipes and J. ossitenuis. Both phylogeographic breaks were congruent with limits of the São Paulo Basin, an area that has undergone Neogene reactivations of tectonic faults. It is suggested that geomorphological events led to a deformed landscape that influenced the dynamics of sedimentary basins and promoted an incipient population structure in J. pictipes and J. ossitenuis. Our findings demonstrate that the divergences whithin Juliomys species occurred during the Quaternary, too recently to have produced strong geographic structure.

Reevaluation of Rhipidomys emiliae (J.A. Allen 1916) and description of a new Rhipidomys (Rodentia: Cricetidae) species from Amazonia and Cerrado.

We present the revalidation of the sigmodontinae rodent species R. emiliae, as well as the description of a new species for the genus Rhipidomys. The maximum likelihood analysis recovers R. emiliae as sister species of the clade with Rhipidomys sp. nov. and R. ipukensis, with high bootstrap values. Comparisons between these species based on the external, cranial, and dental morphology identified several unique characters in Rhipidomys sp. nov., including more grayish brown color of the dorsal coat, subsquamosal fenestra wide and long, angular process ends in the same position of the end of condyloid process, conspicuous protostyle and enterostyle. We describe a new karyotype (2n = 44 and FN = 64) for the genus and, based on an integrative analysis together with morphology and molecular phylogeny, assign it to R. emiliae, and assign the karyotype with 2n = 44 and FN = 52 to Rhipidomys sp. nov.. The analysis integrating data indicated that R. emiliae has a geographic distribution restricted to the lowlands of eastern Amazonia, whereas Rhipidomys sp. nov. occurs in the central Amazonia and Cerrado. The data showed that some Rhipidomys species have its distribution currently limited by rivers, as Rhipidomys sp. nov. occurring west of the Araguaia-Tocantins interfluve, R. emiliae east of the Tocantins River, and R. ipukensis between the Tocantins and Araguaia rivers. This work, in addition to revealing a still unknown biodiversity describing a species, brings a new understanding to the genus, and shows how integrating different markers helps in the correct association between the nominal form and the karyotype.

Corrigendum: Somatic DNA damage response and homologous repair gene alterations and its association with tumor variant burden in breast cancer patients with occupational exposure to pesticides.

[This corrects the article DOI: 10.3389/fonc.2022.904813.].

Diverse mutational spectrum in the 13q14 chromosomal region in a Brazilian cohort of retinoblastoma.

Retinoblastoma is a rare childhood tumor caused by the inactivation of both copies of the RB1 gene. Early diagnosis and identification of heritable RB1 mutation carriers can improve the disease outcome and management via genetic counseling. We used the Multiplex Ligation-dependent Probe Amplification (MLPA) method to analyze the RB1 gene and flanking regions in blood samples from 159 retinoblastoma patients previously negative for RB1 point mutations via Sanger sequencing. We detected a wide spectrum of germline chromosomal alterations, ranging from partial loss or duplication of RB1 to large deletions spanning RB1 and adjacent genes. Mutations were validated via karyotyping, fluorescent in situ hybridization (FISH), SNP-arrays (Single Nucleotide Polymorphism-arrays) and/or quantitative relative real-time PCR. Patients with leukocoria as a presenting symptom showed reduced death rate (p = 0.013) and this sign occurred more frequently among carriers of two breakpoints within RB1 (p = 0.05). All unilateral cases presented both breakpoints outside of RB1 (p = 0.0075). Patients with one breakpoint within RB1 were diagnosed at earlier ages (p = 0.017). Our findings characterize the mutational spectrum of a Brazilian cohort of retinoblastoma patients and point to a possible relationship between the mutation breakpoint location and tumor outcome, contributing to a better prospect of the genotype/phenotype correlation and adding to the wide diversity of germline mutations involving RB1 and adjacent regions in retinoblastoma.

An Overview Regarding Pharmacogenomics and Biomarkers Discovery: Focus on Breast Cancer.

Breast cancer represents a health concern worldwide for being the leading cause of cancer- related women's death. The main challenge for breast cancer treatment involves its heterogeneous nature with distinct clinical outcomes. It is clinically categorized into five subtypes: luminal A; luminal B, HER2-positive, luminal-HER, and triple-negative. Despite the significant advances in the past decades, critical issues involving the development of efficient target-specific therapies and overcoming treatment resistance still need to be better addressed. OMICs-based strategies have marked a revolution in cancer biology comprehension in the past two decades. It is a consensus that Next-Generation Sequencing (NGS) is the primary source of this revolution and the development of relevant consortia translating pharmacogenomics into clinical practice. Still, new approaches, such as CRISPR editing and epigenomic sequencing are essential for target and biomarker discoveries. Here, we discuss genomics and epigenomics techniques, how they have been applied in clinical management and to improve therapeutic strategies in breast cancer, as well as the pharmacogenomics translation into the current and upcoming clinical routine.

Somatic DNA Damage Response and Homologous Repair Gene Alterations and Its Association With Tumor Variant Burden in Breast Cancer Patients With Occupational Exposure to Pesticides.

Homologous recombination is a crucial pathway that is specialized in repairing double-strand breaks; thus, alterations in genes of this pathway may lead to loss of genomic stability and cell growth suppression. Pesticide exposure potentially increases cancer risk through several mechanisms, such as the genotoxicity caused by chronic exposure, leading to gene alteration. To analyze this hypothesis, we investigated if breast cancer patients exposed to pesticides present a different mutational pattern in genes related to homologous recombination (BRCA1, BRCA2, PALB2, and RAD51D) and damage-response (TP53) concerning unexposed patients. We performed multiplex PCR-based assays and next-generation sequencing (NGS) of all coding regions and flanking splicing sites of BRCA1, BRCA2, PALB2, TP53, and RAD51D in 158 unpaired tumor samples from breast cancer patients on MiSeq (Illumina) platform. We found that exposed patients had tumors with more pathogenic and likely pathogenic variants than unexposed patients (p = 0.017). In general, tumors that harbored a pathogenic or likely pathogenic variant had a higher mutational burden (p < 0.001). We also observed that breast cancer patients exposed to pesticides had a higher mutational burden when diagnosed before 50 years old (p = 0.00978) and/or when carrying BRCA1 (p = 0.0138), BRCA2 (p = 0.0366), and/or PALB2 (p = 0.00058) variants, a result not found in the unexposed group. Our results show that pesticide exposure impacts the tumor mutational landscape and could be associated with the carcinogenesis process, therapy response, and disease progression. Further studies should increase the observation period in exposed patients to better evaluate the impact of these findings.

Karyotypical Confirmation of Natural Hybridization between Two Manatee Species, Trichechus manatus and Trichechus inunguis.

Two species of manatees are found in Northern Brazil-the Antillean manatee (Trichechus manatus), which is found along the coast from Florida to Northeastern Brazil, and the Amazonian manatee (Trichechus inunguis), endemic to the Amazon drainage basin. These species show a sympatric distribution in the region of the Marajó Archipelago, an estuarine area surrounding the Amazon River mouth. There is evidence of the occurrence of interspecific hybrids in this area, based on mitochondrial DNA analyses, although the use of nuclear markers has not corroborated this proposal. Considering that these species show very distinct karyotypes, despite being closely related (2n = 48 in T. manatus and 2n = 56 in T. inunguis), hybrids would present distinct chromosome numbers. Based on this, we conducted cytogenetic analyses using classic and molecular techniques in three calves found stranded in the Marajó Island and Amapá coast. The results showed that one of them, morphologically classified as T. inunguis, presented the correspondent karyotype, with 2n = 56. However, the other two, which were phenotypically similar to T. manatus, showed 2n = 49. Despite the same diploid number, their G-banding patterns revealed some differences. The results of the distribution of some microsatellite sequences have also confirmed the heterozygosity of some chromosomal pairs in these two individuals. These results are the first indubitable confirmation of the occurrence of natural hybrids between T. manatus and T. inunguis, and also brings about some issues concerning the viability of hybrids, considering that these two individuals do not correspond to an F1 hybrid, but instead, both presented a possible F2 karyotype.

Clinical implications of lipid peroxides levels in plasma and tumor tissue in breast cancer patients.

Oxidative stress can promote the oxidation of lipoproteins and polyunsaturated fatty acids present in cell membranes; an event known as lipid peroxidation (LPO). LPO has been associated with carcinogenesis and cancer progression, however, its meaning concerning the clinicopathological aspects of human breast cancer is not clear. This study investigated LPO profiles in tumor and plasma samples from breast cancer patients (n = 140) considering their clinicopathological features (age at diagnosis, menopausal status, body mass index, tumor histological grade, tumor size, ki-67 proliferation index, presence of metastasis, chemotherapy response, the molecular subtype of cancer and overall survival status). LPO levels were estimated by tert-butyl hydroperoxide-initiated chemiluminescence. High LPO levels were found regarding poor prognosis parameters as young age at diagnosis (p = 0.006 in tissue), premenopausal patients (p = 0.012 in tissue), high-grade tumors (p = 0.010 in tissue and p = 0.002 in plasma), metastatic disease (p = 0.046 in tissue), chemoresistant tumors (p = 0.041 in tissue), disease relapse (p = 0.018 in tissue and p = 0.009 in plasma) and overall survival status (p = 0.001 in plasma). Our findings point out the clinical meaning of LPO and highlight it as an oxidative stress event linked to poor prognosis and disease aggressiveness in breast cancer patients.

Phylogenetic relationships, population demography, and species delimitation of the Alouatta belzebul species complex (Atelidae: Alouattinae).

Howler monkeys (genus Alouatta) exhibit the most extensive distribution among platyrrhines, comprising Mesoamerican and South American species groups, with the South American group including the Brazilian endemic A. belzebul species complex encompassing A. belzebul, A. discolor, and A. ululata. We herein analyzed their phylogenetic relationship, nucleotide and haplotype diversity, and population demography based on the mitochondrial gene cytochrome b. The phylogenetic and median-joining network analyses distinguished A. discolor, distributed in the west bank of the Xingu River, from A. belzebul on the east bank. This river is a zoogeographic barrier for these species. We did not find evidence of phylogenetic structure between the A. belzebul populations of opposite banks of the Tocantins River, likely related to the changes in the position of this river to the northeast in the late Pleistocene. The A. belzebul along this river showed great morphologic and haplotype diversity, and A. belzebul from the Amazon have kept a larger population size than A. discolor. We herein describe the karyotype of A. discolor, which was similar to those described for A. ululata and A. belzebul. Our results showed two well-defined and supported clades for A. discolor and A. belzebul. However, a new assessment of A. ululata across a large distribution of sampling is required due to the lack of a clear phylogenetic structure.

Hidden diversity of the genus Trinomys (Rodentia: Echimyidae): phylogenetic and populational structure analyses uncover putative new lineages

Trinomys, one of the most species-rich spiny rat genera in Brazil, is widely distributed in Caatinga, Cerrado and Atlantic Forest biomes, and currently includes ten recognized species, three of which are polytypic. Although some studies employing molecular data have been conducted to better characterize phylogenetic relationships among species, 19 nominal taxa have been suggested, implying considerable incongruence regarding species boundaries. We addressed this incongruence by intensively sampling all species across the geographic distribution of the genus. In addition to publicly available data, we generated 182 mt-Cytb gene sequences, and employed phylogenetic and computational species delimitation methods to obtain a clearer picture of the genus diversity. Moreover, we evaluated populational diversity within each accepted species, considering their geographical distribution and a timescale for the evolution of the genus. Beyond confirming the general patterns described for the evolution of the group, this new analysis suggests that Trinomys is comprised of at least 16 evolutionary lineages, 13 of them recognized as species or subspecies, and three never before characterized. This study highlights the importance of increased sample sizes and computational species delimitation methods in uncovering hidden diversity in Trinomys.

Correlation between the number of false positive variants and the quality of results using Ion Torrent PGM™ sequencing to screen BRCA genes. / Correlación entre el número de variantes de falsos positivos y la calidad de los resultados en la secuenciación con Ion Torrent PGM™ para seleccionar genes BRCA

INTRODUCTION: Next Generation Sequencing (NGS) is cost-effective and a faster method to study genes, but its protocol is challenging. OBJECTIVE: To analyze different adjustments to the protocol for screening the BRCA genes using Ion Torrent PGM sequencing and correlate the results with the number of false positive (FP) variants. MATERIAL AND METHODS: We conducted a library preparation process and analyzed the number of FP InDels, the library concentration, the number of cycles in the target amplification step, the purity of the nucleic acid, the input, and the number of samples/Ion 314 chips in association with the results obtained by NGS. RESULTS: We carried out 51 reactions and nine adjustments of protocols and observed eight FP InDels in homopolymer regions. No FP Single-Nucleotide Polymorphism variant was observed; 67.5% of protocol variables were jointly associated with the quality of the results obtained (p<0.05). The number of FP InDels decreased when the quality of results increased. CONCLUSION: The Ion AmpliSeq BRCA1/BRCA2 Community Panel had a better performance using four samples per Ion-314 chip instead of eight and the optimum number of cycles in the amplification step, even when using high-quality DNA, was 23. We observed better results with the manual equalization process and not using the Ion Library Equalizer kit. These adjustments provided a higher coverage of the variants and fewer artifacts (6.7-fold). Laboratories must perform internal validation because FP InDel variants can vary according to the quality of results while the NGS assay should be validated with Sanger.

Introducción. La secuenciación de nueva generación es un método rentable y rápido para el estudio de los genes, pero su protocolo entraña desafíos. Objetivo. Investigar diferentes ajustes del protocolo de selección de los genes BRCA mediante secuenciación de Ion Torrent PGM™ y correlacionar los resultados con el número de variantes de falso positivo. Materiales y métodos. El proceso de preparación de la biblioteca, el número de falsos positivos InDels, la concentración de la biblioteca, el número de ciclos en el paso de amplificación de objetivos, la pureza del ácido nucleico, la entrada y el número de muestras por chip del Ion-314 se analizaron en asociación con los resultados obtenidos por secuenciación de nueva generación secuenciación de nueva generación. Resultados. Se hicieron 51 reacciones y nueve ajustes de los protocolos, y se observaron ocho falsos positivos InDels en las regiones de homopolímeros. No se observó ninguna variante de polimorfismo de nucleótido simple falso positivo. En 67,5 % de los casos, las variables de protocolo en su conjunto se asociaron con la calidad de los resultados obtenidos (p<0,05). El número de falsos positivos InDels disminuyó al aumentar la calidad de los resultados. Conclusiones. El panel comunitario Ion AmpliSeq BRCA1/BRCA2 tuvo un mejor rendimiento, con cuatro muestras por chip Ion-314 en lugar de ocho, y el número de ciclos en el paso de amplificación, incluso con ADN de alta calidad, fue mejor con 23. Se observaron mejores resultados con el proceso de ecualización manual y sin el uso del kit Ion Library Equalizer. Estos ajustes proporcionaron una mayor cobertura de las variantes y menos artefactos. Los laboratorios deben realizar la validación interna porque las variantes de falsos positivos InDel pueden variar según la calidad de los resultados. La secuenciación de próxima generación debe validarse con Sanger.

Correlation between the number of false positive variants and the quality of results using Ion Torrent PGM™ sequencing to screen BRCA genes / Correlación entre el número de variantes de falsos positivos y la calidad de los resultados en la secuenciación con Ion Torrent PGM™ para seleccionar genes BRCA

Abstract | Introduction: Next Generation Sequencing (NGS) is cost-effective and a faster method to study genes, but its protocol is challenging. Objective: To analyze different adjustments to the protocol for screening the BRCA genes using Ion Torrent PGM sequencing and correlate the results with the number of false positive (FP) variants. Materials and methods: We conducted a library preparation process and analyzed the number of FP InDels, the library concentration, the number of cycles in the target amplification step, the purity of the nucleic acid, the input, and the number of samples/Ion 314 chips in association with the results obtained by NGS. Results: We carried out 51 reactions and nine adjustments of protocols and observed eight FP InDels in homopolymer regions. No FP Single-Nucleotide Polymorphism variant was observed; 67.5% of protocol variables were jointly associated with the quality of the results obtained (p<0.05). The number of FP InDels decreased when the quality of results increased. Conclusion: The Ion AmpliSeq BRCA1/BRCA2 Community Panel had a better performance using four samples per Ion-314 chip instead of eight and the optimum number of cycles in the amplification step, even when using high-quality DNA, was 23. We observed better results with the manual equalization process and not using the Ion Library Equalizer kit. These adjustments provided a higher coverage of the variants and fewer artifacts (6.7-fold). Laboratories must perform internal validation because FP InDel variants can vary according to the quality of results while the NGS assay should be validated with Sanger.

Resumen | Introducción. La secuenciación de nueva generación es un método rentable y rápido para el estudio de los genes, pero su protocolo entraña desafíos. Objetivo. Investigar diferentes ajustes del protocolo de selección de los genes BRCAmediante secuenciación de Ion Torrent PGM™ y correlacionar los resultados con el número de variantes de falso positivo. Materiales y métodos. El proceso de preparación de la biblioteca, el número de falsos positivos InDels, la concentración de la biblioteca, el número de ciclos en el paso de amplificación de objetivos, la pureza del ácido nucleico, la entrada y el número de muestras por chip del Ion-314 se analizaron en asociación con los resultados obtenidos por secuenciación de nueva generación secuenciación de nueva generación. Resultados. Se hicieron 51 reacciones y nueve ajustes de los protocolos, y se observaron ocho falsos positivos InDels en las regiones de homopolímeros. No se observó ninguna variante de polimorfismo de nucleótido simple falso positivo. En 67,5 % de los casos, las variables de protocolo en su conjunto se asociaron con la calidad de los resultados obtenidos (p<0,05). El número de falsos positivos InDels disminuyó al aumentar la calidad de los resultados. Conclusiones. El panel comunitario Ion AmpliSeq BRCA1/BRCA2 tuvo un mejor rendimiento, con cuatro muestras por chip Ion-314 en lugar de ocho, y el número de ciclos en el paso de amplificación, incluso con ADN de alta calidad, fue mejor con 23. Se observaron mejores resultados con el proceso de ecualización manual y sin el uso del kit Ion Library Equalizer. Estos ajustes proporcionaron una mayor cobertura de las variantes y menos artefactos. Los laboratorios deben realizar la validación interna porque las variantes de falsos positivos InDel pueden variar según la calidad de los resultados. La secuenciación de próxima generación debe validarse con Sanger.

Orthohantavirus Survey in Indigenous Lands in a Savannah-Like Biome, Brazil.

In Brazil, the first confirmed cases of hantavirus cardiopulmonary syndrome in Indigenous populations occurred in 2001. The purpose of this study was to determine the seroprevalence of orthohantavirus infections in the Utiariti Indigenous land located in the southeastern region of the Brazilian Amazon. In December 2014 and 2015, a survey was conducted using an enzyme-linked immunosorbent assay in nine villages belonging to the Haliti-Paresí Indigenous communities. A total of 301 participants were enrolled in the study. Of the two study cohorts, the one from 2014 showed a prevalence of 12.4%, whereas the one from 2015 had a serum prevalence of 13.4%. Analysis of the paired samples of 110 Indigenous people who participated in both stages of the study enabled identification of four individuals who had seroconverted during the study period. Identifying the circulation of orthohantaviruses in the Utiariti Indigenous land highlights a serious public health problem in viral expansion and highlights the need to implement preventive measures appropriate to the sociocultural reality of these communities.

SINE-B1 Distribution and Chromosome Rearrangements in the South American Proechimys gr. goeldii (Echimyidae, Rodentia).

Proechimys species are remarkable for their extensive chromosome rearrangements, representing a good model to understand genome evolution. Herein, we cytogenetically analyzed 3 different cytotypes of Proechimys gr. goeldii to assess their evolutionary relationship. We also mapped the transposable element SINE-B1 on the chromosomes of P. gr. goeldii in order to investigate its distribution among individuals and evaluate its possible contribution to karyotype remodeling in this species. SINE-B1 showed a dispersed distribution along chromosome arms and was also detected at the pericentromeric regions of some chromosomes, including pair 1 and the sex chromosomes, which are involved in chromosome rearrangements. In addition, we describe a new cytotype for P. gr. goeldii, reinforcing the significant role of gross chromosomal rearrangements during the evolution of the genus. The results of FISH with SINE-B1 suggest that this issue should be more deeply investigated for a better understanding of its role in the mechanisms involved in the wide variety of Proechimys karyotypes.

Prevalence of germline variants in consensus moderate-to-high-risk predisposition genes to hereditary breast and ovarian cancer in BRCA1/2-negative Brazilian patients.

PURPOSE: This study aimed to identify and classify genetic variants in consensus moderate-to-high-risk predisposition genes associated with Hereditary Breast and Ovarian Cancer Syndrome (HBOC), in BRCA1/2-negative patients from Brazil. METHODS: The study comprised 126 index patients who met NCCN clinical criteria and tested negative for all coding exons and intronic flanking regions of BRCA1/2 genes. Multiplex PCR-based assays were designed to cover the complete coding regions and flanking splicing sites of six genes implicated in HBOC. Sequencing was performed on HiSeq2500 Genome Analyzer. RESULTS: Overall, we identified 488 unique variants. We identified five patients (3.97%) that harbored pathogenic or likely pathogenic variants in four genes: ATM (1), CHEK2 (2), PALB2 (1), and TP53 (1). One hundred and thirty variants were classified as variants of uncertain significance (VUS), 10 of which were predicted to disrupt mRNA splicing (seven non-coding variants and three coding variants), while other six missense VUS were classified as probably damaging by prediction algorithms. CONCLUSION: A detailed mutational profile of non-BRCA genes is still being described in Brazil. In this study, we contributed to filling this gap, by providing important data on the diversity of genetic variants in a Brazilian high-risk patient cohort. ATM, CHEK2, PALB2 and TP53 are well established as HBOC predisposition genes, and the identification of deleterious variants in such actionable genes contributes to clinical management of probands and relatives.

Checklist of mammals from Goiás, central Brazil / Lista de Mamíferos de Goiás, Brasil central

Abstract: The state of Goiás, in central Brazil, is covered mainly by the Cerrado domain, with the Alto Paraná Atlantic Forest occupying its central-southern portion. Goiás is one of the 20 Brazilian federative units without a mammal checklist. In this study, we provide the first checklist of mammals from Goiás state. We recorded mammal species based primarily on the analysis of specimens housed in scientific collections as well as on literature with associated voucher material. We listed 191 mammalian species belonging to 125 genera, 31 families and 10 orders, which represents 25.2% of the mammal species occurring in Brazil. The most speciose orders were Chiroptera (90 spp.), followed by Rodentia (43 spp.), Carnivora (19 spp.) and Didelphimorphia (17 spp.). The following orders accounted for a smaller portion of the state diversity: Cetartiodactyla (7 spp.), Cingulata (7 spp.), Primates (4 spp.), Pilosa (2 spp.), Lagomorpha (1 sp.), and Perissodactyla (1 sp.). A total of 28 species (14.7%), mainly represented by medium and large-sized mammals, are nationally threatened while 12 (6.3%) are globally threatened. Our results indicate great portions of the state lacking a proper survey of mammals, especially the northwestern portion. We discuss species richness, distribution and conservation status of the mammals of Goiás state in national and regional scenarios. We highlight the need for mammal inventories based on complementary survey techniques with the collection of vouchers in order to provide karyologic, molecular, morphologic, parasitologic, and ecological data. These informations are the basis for integrative studies that lead to the understanding of current mammalian richness and diversity. Indeed, knowledge on species richness distribution in the state will guide conservation strategies, especially in areas undergoing habitat loss and fragmentation, such as the central-southern portion of Goiás.

Resumo: O estado de Goiás, no Brasil central, é coberto principalmente pelo domínio do Cerrado, com a Mata Atlântica do Alto Paraná ocupando sua porção centro-sul. Goiás é uma das 20 unidades federativas brasileiras que ainda não possui uma lista de espécies de mamíferos. Neste estudo, apresentamos a primeira lista de mamíferos para o estado. Registramos as espécies de mamíferos com base principalmente na análise de espécimes depositados em coleções científicas, bem como na literatura apresentando material testemunho associado. Listamos 191 espécies pertencentes a 125 gêneros, 31 famílias e 10 ordens, as quais representam 25,2% das espécies de mamíferos que ocorrem no Brasil. As ordens mais especiosas foram Chiroptera (90 spp.), seguida pelas ordens Rodentia (43 spp.), Carnivora (19 spp.) e Didelphimorphia (17 spp.), com as demais ordens respondendo por uma porção menor da diversidade: Cetartiodactyla (7 spp.), Cingulata (7 spp.), Primates (4 spp.), Pilosa (2 spp.), Lagomorpha (1 sp.) e Perissodactyla (1 sp.). Um total de 28 espécies (14,7%), principalmente representadas por mamíferos de médio e grande porte, estão ameaçadas nacionalmente e 12 (6,3%) encontram-se globalmente ameaçadas. Nossos resultados indicaram grandes porções do estado ainda não devidamente pesquisadas em relação aos seus mamíferos, com informações escassas e fragmentadas, principalmente no que diz respeito à sua porção noroeste. Discutimos a riqueza de espécies, a distribuição e o estado de conservação dos mamíferos do estado de Goiás nos cenários nacional e regional. Ressaltamos a importância da realização de inventários que utilizem técnicas complementares de amostragem, incluindo a coleta de material testemunho, proporcionando a obtenção de dados cariotípicos, moleculares, morfológicos, parasitológicos e ecológicos. Estas informações são a base de estudos integrativos, os quais aumentam nossa compreensão a respeito da riqueza e diversidade atual dos mamíferos. O conhecimento a respeito da distribuição da riqueza de espécies em Goiás é essencial para embasar estratégias de conservação, tão necessárias em áreas que vem sofrendo com a perda e fragmentação de seus hábitats naturais, como a porção centro-sul do estado.

Morphological variation in the genus Juliomys (Rodentia: Cricetidae: Sigmodontinae) and taxonomic status of Juliomys anoblepas (Winge 1887) from the Quaternary of Southeast Brazil.

The region of Lagoa Santa, Minas Gerais, Brazil, is one of the most important karstic areas of the Brazilian Quaternary due to the faunistic diversity of living and extinct forms. Among them, some taxa remain poorly studied, as is the case of Calomys anoblepas Winge 1887. Despite the recent allocation of the taxon within Juliomys, its description and morphological analysis are condensed, based on comparative few specimens and on few informative characters. In this study, we investigate characters proposed to distinguish species of Juliomys, and reevaluate the taxonomic status of the fossil Juliomys anoblepas. We analyzed 80 cranio-dental morphological characters in 233 specimens represented by the four species currently recognized: J. pictipes (Osgood 1933), J. rimofrons Oliveira Bonvicino 2002, J. ossitenuis Costa, Pavan, Leite Fagundes 2007, and J. ximenezi Christoff, Vieira, Oliveira, Gonçalves, Valiati Tomasi 2016. We also performed principal component analysis on eight craniodental measurements available for the J. anoblepas hypodigm. The review of morphological systems and the evaluation of the characters used in the literature revealed that there are no diagnostic characters in the anterior portion of the skull and in the molar series of Juliomys, being difficult to differentiate the fossil from the other living species. Only six qualitative characters were variable and applicable to the hypodigm of J. anoblepas. Characters are polymorphic, invariable, or the fossil is not sufficiently complete to determinate its states. The taxon could not be morphometrically differentiated from J. pictipes and J. ossitenuis. Based on the results presented herein, we consider J. anoblepas as a nomen dubium and restrict its name to the taxon's hypodigm.

Impression Cytology Is a Non-invasive and Effective Method for Ocular Cell Retrieval of Zika Infected Babies: Perspectives in OMIC Studies.

IMPORTANCE: Non-invasive techniques for retrieving ocular surface cells from babies infected by zika virus (ZIKV) during the gestational period remain to be determined. OBJECTIVES: The aim of this study was to describe an optimized impression cytology method for the isolation of viable cells from Zika infected babies with and without Congenital Zika Syndrome (CZS) in satisfactory amount and quality to enable easy adoption in the field and application in the context of genomic and molecular approaches. DESIGN SETTINGS AND PARTICIPANTS: Ocular surface samples were obtained with a hydrophilic nitrocellulose membrane (through optimized impression cytology method) from twelve babies referred to the Pediatric Service of the Antonio Pedro Hospital, Universidade Federal Fluminense (UFF), Niteroi, Rio de Janeiro, Brazil. After an authorized written informed consent from the parents, samples were collected from both eyes of 12 babies (4 babies with maternal ZIKV exposure during gestation and presence of clinical signs which included ocular abnormalities and microcephaly; 4 babies with maternal ZIKV exposure during gestation but no clinical signs; and 4 unaffected control babies with negative PCR for Zika virus and without clinical signs). Cells were used for microscopy analyses and evaluated for their suitability for downstream molecular applications in transcriptomic and proteomic experiments. RESULTS: Our optimized impression cytology protocol enabled the capture of a considerable number of viable cells. The microscopic features of the conjunctival epithelial cells were described by both direct analysis of the membrane-attached cells and analysis of cytospinned captured cells using several staining procedures. Epithelial basal, polyhedral and goblet cells were clearly identified in all groups. All cases of ZIKV infected babies showed potential morphological alterations (cell keratinization, pyknosis, karyolysis, anucleation, and vacuolization). Molecular approaches were also performed in parallel. Genomic DNA and RNA were successfully isolated from all samples to enable the establishment of transcriptomic and proteomic studies. CONCLUSIONS AND RELEVANCE: Our method proved to be a suitable, fast, and non-invasive tool to obtain ocular cell preparations from babies with and without Zika infection. The method yielded sufficient cells for detailed morphological and molecular analyses of samples. We discuss perspectives for the application of impression cytology in the context of ZIKV studies in basic and clinical research.

DNA barcoding of the rodent genus Oligoryzomys (Cricetidae: Sigmodontinae): mitogenomic-anchored database and identification of nuclear mitochondrial translocations (Numts).

DNA barcoding has become a standard method for species identification in taxonomically complex groups. An important step of the barcoding process is the construction of a library of voucher-based material that was properly identified by independent methods, free of inaccurate identification, and paralogs. We provide here a cytochrome oxidase I (mt-Co1) DNA barcode database for species of the genus Oligoryzomys, based on type material and karyotyped specimens, and anchored on the mitochondrial genome of one species of Oligoryzomys, O. stramineus. To evaluate the taxonomic determination of new COI sequences, we assessed species intra/interspecific genetic distances (barcode gap), performed the General Mixed Yule Coalescent method (GMYC) for lineages' delimitation, and identified diagnostic nucleotides for each species of Oligoryzomys. Phylogenetic analyses of Oligoryzomys were performed on 2 datasets including 14 of the 23 recognized species of this genus: a mt-Co1 only matrix, and a concatenated matrix including mt-Co1, cytochrome b (mt-Cytb), and intron 7 of the nuclear fibrinogen beta chain gene (i7Fgb). We recovered nuclear-mitochondrial translocated (Numts) pseudogenes on our samples and identified several published sequences that are cases of Numts. We analyzed the rate of non-synonymous and synonymous substitution, which were higher in Numts in comparison to mtDNA sequences. GMYC delimitations and DNA barcode gap results highlight the need for further work that integrate molecular, karyotypic, and morphological analyses, as well as additional sampling, to tackle persistent problems in the taxonomy of Oligoryzomys.