Shepherding the past: High-resolution data on Neolithic Southern Iberian livestock management at Cueva de El Toro (Antequera, Málaga).

The feeding strategies of the first domesticated herds had to manage the risks arising from the novelty of livestock practices in territories often distant from the animals' primary habitats. The Iberian Peninsula is characterised by a great diversity of environments, which undoubtedly influenced these dynamics. At the beginning of the Neolithic period these led the possibility to combine diverse livestock farming practices based on different animal feeding habits. This variability is also consistent with the rythms of adoption of domesticated animals, being later on the northern area. In order to address this issue, this work focuses on the dietary regimes of early sheep herds from southern Iberia, an area for which information is currently scarce. This study utilises high-resolution radiocarbon dating and stable isotope data on teeth to investigate sheep husbandry management strategies in Cueva de El Toro (Antequera, Málaga). The radiocarbon dates on the analysed remains evidenced they were deposited at the site over a short period, supporting the recurrent use of the cave. The sequential analysis of oxygen and carbon isotopes in tooth enamel reveals distinct livestock management strategies, reproduction patterns, feeding habits, and mobility during this short period. This variability demonstrates that livestock management practices in the western Mediterranean are more diverse than previously considered. Furthermore, these findings support the hypothesis that early Neolithic communities in the southern Iberian Peninsula were able to adopt different feeding strategies within the same herd, depending on their ecological and productive needs.

Ancient chicken remains reveal the origins of virulence in Marek's disease virus.

The pronounced growth in livestock populations since the 1950s has altered the epidemiological and evolutionary trajectory of their associated pathogens. For example, Marek's disease virus (MDV), which causes lymphoid tumors in chickens, has experienced a marked increase in virulence over the past century. Today, MDV infections kill >90% of unvaccinated birds, and controlling it costs more than US$1 billion annually. By sequencing MDV genomes derived from archeological chickens, we demonstrate that it has been circulating for at least 1000 years. We functionally tested the Meq oncogene, one of 49 viral genes positively selected in modern strains, demonstrating that ancient MDV was likely incapable of driving tumor formation. Our results demonstrate the power of ancient DNA approaches to trace the molecular basis of virulence in economically relevant pathogens.

Antimicrobial Dispensing Practices during COVID-19 and the Implications for Pakistan.

Antibiotics are one of the most frequently dispensed classes of medicines. However, excessive misuse and abuse enhances antimicrobial resistance (AMR). Previous studies in Pakistan have documented extensive dispensing of 'Watch' and 'Reserve' antibiotics, which is a concern. In view of this, there is a need to assess current dispensing patterns following COVID-19 in Pakistan. A cross-sectional study was undertaken, collecting dispensing data from 39 pharmacies and 53 drug stores from November 2022 to February 2023. Outlets were principally in urban areas (60.9%), with pharmacists/pharmacy technicians present in 32.6% of outlets. In total, 11,092 prescriptions were analyzed; 67.1% of patients were supplied at least one antimicrobial, 74.3% antibiotics, 10.2% antifungals and 7.9% anthelmintics. A total of 33.2% of antimicrobials were supplied without a prescription. Common indications for dispensed antibiotics were respiratory (34.3%) and gastrointestinal (16.8%) infections, which can be self-limiting. In addition, 12% of antibiotics were dispensed for the prevention or treatment of COVID-19. The most frequent antibiotics dispensed were ceftriaxone (18.4%) and amoxicillin (15.4%). Overall, 59.2% antibiotics were 'Watch' antibiotics, followed by 'Access' (40.3%) and 'Reserve' (0.5%) antibiotics. Of the total antibiotics dispensed for treating COVID-19, 68.3% were 'Watch' and 31.7% 'Access'. Overall, there appeared to be an appreciable number of antibiotics dispensed during the recent pandemic, including for patients with COVID-19, alongside generally extensive dispensing of 'Watch' antibiotics. This needs to be urgently addressed with appropriate programs among pharmacists/pharmacy technicians to reduce AMR.

Socio-ecological impact of monogenetic volcanism in the La Garrotxa Volcanic Field (NE Iberia).

Volcanism can cause major impacts, including climate change and mass extinctions. However, the impact of monogenetic volcanism is often considered as limited in volcanological research. This work provides for the first time an interdisciplinary approach to the socio-ecological impact of monogenetic volcanism in a key region, the La Garrotxa Volcanic Field (GVF, Girona, NE Iberia), where intense monogenetic volcanic activity occurred in the past. The analyses of a sedimentary sequence from the GVF enabled identifying previously unknown volcanic eruptions in the time interval 14-8.4 ka cal BP, constrain their volcanic stratigraphy and age, and unfold the effects of environmental change on geomorphology, vegetation, aquatic organisms and humans. Moreover, we reconstruct the major palaeoenvironmental changes caused by the eruptions in terms of fire episodes and subsequent disturbance on vegetation, hydrology and limnological conditions. When put in context with the archaeological record, it appears that the last hunter-gatherer communities were resilient at an extra-local scale, facing episodes of vulnerability due to volcanic activity, suggesting that their flexible nomadic patterns and foraging economies were an efficient source of risk management against the volcanic eruptions and their ecological impacts.

Bones geometric morphometrics illustrate 10th millennium cal. BP domestication of autochthonous Cypriot wild boar (Sus scrofa circeus nov. ssp).

Epipaleolithic hunter-gatherers from the Near East introduced wild boars (Sus scrofa) to Cyprus, with the Early Pre-Pottery Neolithic (PPN) settlers hunting the wild descendants of these boars. However, the geographic origin of the Cypriot boar and how they were integrated into the earliest forms of pig husbandry remain unsolved. Here, we present data on 11,000 to 9000 cal. BP Sus scrofa from the PPN sites of Klimonas and Shillourokambos. We compared them to contemporaneous populations from the Near East and to Neolithic and modern populations in Corsica, exploring their origin and evolution using biosystematic signals from molar teeth and heel bones (calcanei), using 2D and 3D geometric morphometrics. We found that the Cypriot PPN lineage of Sus scrofa originates from the Northern Levant. Yet, their phenotypic idiosyncrasy suggest that they evolved into an insular sub-species that we named Sus scrofa circeus, referring to Circe, the metamorphosis goddess that changed Ulysses companions into pigs. The phenotypic homogeneity among PPNA Klimonas wild boars and managed populations of PPNB Shillourokambos suggests that local domestication has been undertaken on the endemic S. s. circeus, strengthening the idea that Cyprus was integrated into the core region of animal domestication.

Effects of Cage Enrichment on Behavior, Welfare and Outcome Variability in Female Mice.

The manner in which laboratory rodents are housed is driven by economics (minimal use of space and resources), ergonomics (ease of handling and visibility of animals), hygiene, and standardization (reduction of variation). This has resulted in housing conditions that lack sensory and motor stimulation and restrict the expression of species-typical behavior. In mice, such housing conditions have been associated with indicators of impaired welfare, including abnormal repetitive behavior (stereotypies, compulsive behavior), enhanced anxiety and stress reactivity, and thermal stress. However, due to concerns that more complex environmental conditions might increase variation in experimental results, there has been considerable resistance to the implementation of environmental enrichment beyond the provision of nesting material. Here, using 96 C57BL/6 and SWISS female mice, respectively, we systematically varied environmental enrichment across four levels spanning the range of common enrichment strategies: (1) bedding alone; (2) bedding + nesting material; (3) deeper bedding + nesting material + shelter + increased vertical space; and (4) semi-naturalistic conditions, including weekly changes of enrichment items. We studied how these different forms of environmental enrichment affected measures of animal welfare, including home-cage behavior (time-budget and stereotypic behavior), anxiety (open field behavior, elevated plus-maze behavior), growth (food and water intake, body mass), stress physiology (glucocorticoid metabolites in fecal boluses and adrenal mass), brain function (recurrent perseveration in a two-choice guessing task) and emotional valence (judgment bias). Our results highlight the difficulty in making general recommendations across common strains of mice and for selecting enrichment strategies within specific strains. Overall, the greatest benefit was observed in animals housed with the greatest degree of enrichment. Thus, in the super-enriched housing condition, stereotypic behavior, behavioral measures of anxiety, growth and stress physiology varied in a manner consistent with improved animal welfare compared to the other housing conditions with less enrichment. Similar to other studies, we found no evidence, in the measures assessed here, that environmental enrichment increased variation in experimental results.

A cross-species judgement bias task: integrating active trial initiation into a spatial Go/No-go task.

Judgement bias tasks are promising tools to assess emotional valence in animals, however current designs are often time-consuming and lack aspects of validity. This study aimed to establish an improved design that addresses these issues and can be used across species. Horses, rats, and mice were trained on a spatial Go/No-go task where animals could initiate each trial. The location of an open goal-box, at either end of a row of five goal-boxes, signalled either reward (positive trial) or non-reward (negative trial). Animals first learned to approach the goal-box in positive trials (Go) and to re-initiate/not approach in negative trials (No-go). Animals were then tested for responses to ambiguous trials where goal-boxes at intermediate locations were opened. The Go:No-go response ratio was used as a measure of judgement bias. Most animals quickly learned the Go/No-go discrimination and performed trials at a high rate compared to previous studies. Subjects of all species reliably discriminated between reference cues and ambiguous cues, demonstrating a monotonic graded response across the different cue locations, with no evidence of learning about the outcome of ambiguous trials. This novel test protocol is an important step towards a practical task for comparative studies on judgement biases in animals.

First evidence of Smith-Magenis syndrome in mother and daughter due to a novel RAI mutation.

Smith-Magenis syndrome (SMS) is a complex genetic disorder caused by interstitial 17p11.2 deletions encompassing multiple genes, including the retinoic acid induced 1 gene-RAI1-or mutations in RAI1 itself. The clinical spectrum includes developmental delay, cognitive impairment, and behavioral abnormalities, with distinctive physical features that become more evident with age. No patients have been reported to have had offspring. We here describe a girl with developmental delay, mainly compromising the speech area, and her mother with mild intellectual disabilities and minor dysmorphic features. Both had sleep disturbance and attention deficit disorder, but no other atypical behaviors have been reported. In both, CGH-array analysis detected a 15q13.3 interstitial duplication, encompassing CHRNA7. However, the same duplication has been observed in several, apparently healthy, maternal relatives. We, thus, performed a whole exome sequencing analysis, which detected a frameshift mutation in RAI1, de novo in the mother, and transmitted to her daughter. No other family members carried this mutation. This is the first report of an SMS patient having offspring. Our experience confirms the importance of searching for alternative causative genetic mechanisms in case of confounding/inconclusive findings such as a CGH-array result of uncertain significance. © 2016 Wiley Periodicals, Inc.

A Novel HRAS Mutation Independently Contributes to Left Ventricular Hypertrophy in a Family with a Known MYH7 Mutation.

Several genetic conditions can lead to left ventricular hypertrophy (LVH). Among them, hypertrophic cardiomyopathy (HCM), caused by mutations in sarcomere genes, is the most common inherited cardiac disease. Instead, RASopathies, a rare class of disorders characterized by neuro-cardio-facial-cutaneous abnormalities and sometimes presenting with LVH, are caused by mutations in the RAS-MAPK pathway. We report on a 62-years-old male who presented isolated severe obstructive LVH but did not carry the sarcomere mutation previously identified in his affected relatives. By exome sequencing, we detected a novel mutation in HRAS gene (NM_005343.2:p.Arg68Trp), present also in the proband's daughter, who showed mild LVH and severe intellectual disability. The cardiac phenotype was indistinguishable between family members carrying either mutation. In silico studies suggested that the mutated HRAS protein is constitutionally activated. Consistently, functional characterization in vitro confirmed elevated HRAS-GTP accumulation and downstream RAS-MAPK pathway activation that are known to drive cell proliferation in LVH. Our study emphasizes the role of RAS signaling in cardiac hypertrophy and highlights the complexity in differential diagnosis of RASopathies. In fact, the mild features of RASopathy and the recurrence of sarcomeric HCM in this family delayed the correct diagnosis until comprehensive genetic testing was performed.

Caracterização e organização da enfermagem do hospital alemão Oswaldo Cruz no contexto da II guerra / Characterization and organization of hospital alemão Oswaldo Cruz nursing in II World War context

Trata-se de um estudo documental com abordagem histórico-social com objetivo de analisar a composição e a estruturação da Enfermagem do Hospital Alemão Oswaldo Cruz (HAOC) entre 1941 e 1945. Foram utilizadas duas fontes documentais: "Livro de Registro de Empregados" e "Fichas Profissionais de Registro de Empregados". Antes de serem correlacionados, os dados de cada fonte foram estudados separadamente. Os dados dos livros foram copiados eletrostaticamente, certificados, transcritos, organizados em planilha Excel®, para então serem analisados quantitativamente e qualitativamente sob o contexto histórico, social, econômico e político. A pesquisa revelou que, desde a década de 1940, essa instituição utilizava de ferramentas de gestão de recursos humanos para controlar headcount e folha de pagamento. Existia divisão formal e hierárquica do trabalho. A força de trabalho de enfermagem era composta por 16 diferentes cargos subdivididos em três segmentos (líderes, graduados, não graduados) atuantes em nível estratégico, tático e operacional. Concluiu-se, que, já da década de 1940, a organização dos profissionais da Enfermagem, sustentada pelo modelo assistencial e de gestão, possibilitou, ao HAOC, ser considerado uma instituição modelo-referência.

This article is a documentary study involving a social and historical approach with the objective of analyze the composition and the structure of the nursing workforce in the Hospital Alemão Oswaldo Cruz, between the year of 1941 and 1945. It was appreciated two documentary sources: "Book of Employee's Register" and "Professional Sheets" from HAOC. Beforebeing correlated, the data from each source were studied separately. The data from each book was copied electrostatically, certificated, transcript, organized into Excel® spreadsheet, and then analyzed quantitatively and qualitatively in the historical, social, economic and political context. The research revealed that since the 1940s, the institution was using human resource management tools to control headcount and payroll. There was formal and hierarchical division of labor. The nursing workforce was composed by 16 different designations of positions that was divided into three segments (leaders, graduates, non-graduates) acting in strategic, tactical and operational levels. It was concluded the organization of nursing professionals, supported by the care and management model, enabled HAOC to be considered a model institution-reference.

Microcephaly, ectodermal dysplasia, multiple skeletal anomalies and distinctive facial appearance: delineation of cerebro-dermato-osseous-dysplasia.

In 1980, a novel multiple malformation syndrome has been described in a 17-year-old woman with micro- and turricephaly, intellectual disability, distinctive facial appearance, congenital atrichia, and multiple skeletal anomalies mainly affecting the limbs. Four further sporadic patients and a couple of affected sibs are also reported with a broad clinical variability. Here, we describe a 4-year-old girl strikingly resembling the original report. Phenotype comparison identified a recurrent pattern of multisystem features involving the central nervous system, and skin and bones in five sporadic patients (including ours), while the two sibs and a further sporadic case show significant phenotypic divergence. Marked clinical variability within the same entity versus syndrome splitting is discussed and the term "cerebro-dermato-osseous dysplasia" is introduced to define this condition.

A large scale expression study associates uc.283-plus lncRNA with pluripotent stem cells and human glioma.

BACKGROUND: There are 481 ultra-conserved regions (UCRs) longer than 200 bases in the genomes of human, mouse and rat. These DNA sequences are absolutely conserved and show 100% identity with no insertions or deletions. About half of these UCRs are reported as transcribed and many correspond to long non-coding RNAs (lncRNAs). METHODS: We used custom microarrays with 962 probes representing sense and antisense sequences for the 481 UCRs to examine their expression across 374 normal samples from 46 different tissues and 510 samples representing 10 different types of cancer. The expression in embryonic stem cells of selected UCRs was validated by real time PCR. RESULTS: We identified tissue selective UCRs and studied UCRs in embryonic and induced pluripotent stem cells. Among the normal tissues, the uc.283 lncRNA was highly specific for pluripotent stem cells. Intriguingly, the uc.283-plus lncRNA was highly expressed in some solid cancers, particularly in one of the most untreatable types, glioma. CONCLUSION: Our results suggest that uc.283-plus lncRNA might have a role in pluripotency of stem cells and in the biology of glioma.

Pluripotent stem cell miRNAs and metastasis in invasive breast cancer.

BACKGROUND: The purpose of this study is to determine whether microRNA for pluripotent stem cells are also expressed in breast cancer and are associated with metastasis and outcome. METHODS: We studied global microRNA profiles during differentiation of human embryonic stem cells (n =26) and in breast cancer patients (n = 33) and human cell lines (n = 35). Using in situ hybridization, we then investigated MIR302 expression in 318 untreated breast cancer patients (test cohort, n = 22 and validation cohort, n = 296). In parallel, using next-generation sequencing data from breast cancer patients (n = 684), we assessed microRNA association with stem cell markers. All statistical tests were two-sided. RESULTS: In healthy tissues, the MIR302 (high)/MIR203 (low) asymmetry was exclusive for pluripotent stem cells. MIR302 was expressed in a small population of cancer cells within invasive ductal carcinoma, but not in normal breast (P < .001). Furthermore, MIR302 was expressed in the tumor cells together with stem cell markers, such as CD44 and BMI1. Conversely, MIR203 expression in 684 breast tumors negatively correlated with CD44 (Spearman correlation, Rho = -0.08, P = .04) and BMI1 (Rho = -0.11, P = .004), but positively correlated with differentiation marker CD24 (Rho = 0.15, P < .001). Primary tumors with lymph node metastasis had cancer cells showing scattered expression of MIR302 and widespread repression of MIR203. Finally, overall survival was statistically significantly shorter in patients with MIR302-positive cancer cells (P = .03). CONCLUSIONS: In healthy tissues the MIR302(high)/MIR203(low) asymmetry was characteristic of embryonic and induced pluripotency. In invasive ductal carcinoma, the MIR302/MIR203 asymmetry was associated with stem cell markers, metastasis, and shorter survival.

Novel α-actinin 2 variant associated with familial hypertrophic cardiomyopathy and juvenile atrial arrhythmias: a massively parallel sequencing study.

BACKGROUND: Next-generation sequencing might be particularly advantageous in genetically heterogeneous conditions, such as hypertrophic cardiomyopathy (HCM), in which a considerable proportion of patients remain undiagnosed after Sanger. In this study, we present an Italian family with atypical HCM in which a novel disease-causing variant in α-actinin 2 (ACTN2) was identified by next-generation sequencing. METHODS AND RESULTS: A large family spanning 4 generations was examined, exhibiting an autosomal dominant cardiomyopathic trait comprising a variable spectrum of (1) midapical HCM with restrictive evolution with marked biatrial dilatation, (2) early-onset atrial fibrillation and atrioventricular block, and (3) left ventricular noncompaction. In the proband, 48 disease genes for HCM, selected on the basis of published reports, were analyzed by targeted resequencing with a customized enrichment system. After bioinformatics analysis, 4 likely pathogenic variants were identified: TTN c.21977G>A (p.Arg7326Gln); TTN c.8749A>C (p.Thr2917Pro); ACTN2 c.683T>C (p.Met228Thr); and OBSCN c.13475T>G (p.Leu4492Arg). The novel variant ACTN2 c.683T>C (p.Met228Thr), located in the actin-binding domain, proved to be the only mutation fully cosegregating with the cardiomyopathic trait in 18 additional family members (of whom 11 clinically affected). ACTN2 c.683T>C (p.Met228Thr) was absent in 570 alleles of healthy controls and in 1000 Genomes Project and was labeled as Damaging by in silico analysis using polymorphism phenotyping v2, as Deleterious by sorts intolerant from tolerant, and as Disease-Causing by Mutation Taster. CONCLUSIONS: A targeted next-generation sequencing approach allowed the identification of a novel ACTN2 variant associated with midapical HCM and juvenile onset of atrial fibrillation, emphasizing the potential of such approach in HCM diagnostic screening.

Identification of a novel de novo deletion in RAF1 associated with biventricular hypertrophy in Noonan syndrome.

Biventricular hypertrophy (BVH) is a disease state characterized by the thickening of the ventricle walls. The differential diagnosis of BVH with other congenital and familial diseases in which increased ventricle wall thickness is a prominent clinical feature is fundamental due to its therapeutic and prognostic value, mainly during infancy. We describe a 2-month-old infant presenting BVH. Using exome sequencing, we identified a novel de novo 3-bp deletion in the RAF1 gene that is located in the binding active site for the 14-3-3 peptide. Based on docking calculations, we demonstrate that this novel mutation impairs protein/target binding, thus constitutively activating Ras signaling, which is a dysregulation associated with Noonan syndrome. Finally, our study underlines the importance of molecular modeling to understand the roles of novel mutations in pathogenesis.

A new mutational mechanism for hypertrophic cardiomyopathy.

We describe a male patient affected by hypertrophic cardiomyopathy (HCM) with no point mutations in the eight sarcomeric genes most commonly involved in the disease. By multiple ligation-dependent probe amplification (MLPA) we have identified a multi-exons C-terminus deletion in the cardiac myosin binding protein C (MYBPC3) gene. The rearrangement has been confirmed by long PCR and breakpoints have been defined by sequencing. The 3.5 kb terminal deletion is mediated by Alu-repeat elements and is predicted to result in haploinsufficiency of MYBPC3. To exclude the presence of other rare pathogenic variants in additional HCM genes, we performed targeted next-generation sequencing (NGS) of 88 cardiomyopathy-associated genes but we did not identify any further mutation. Interestingly, the MYBPC3 multi-exons deletion was detectable by NGS. This finding broadens the range of mutational spectrum observed in HCM, contributing to understanding the genetic basis of the most common inherited cardiovascular disease. Moreover, our data suggest that NGS may represent a new tool to achieve a deeper insight into molecular basis of complex diseases, allowing to detect in a single experiment both point mutations and gene rearrangements.

Breast cancer signatures for invasiveness and prognosis defined by deep sequencing of microRNA.

The transition from ductal carcinoma in situ to invasive ductal carcinoma is a key event in breast cancer progression that is still not well understood. To discover the microRNAs regulating this critical transition, we used 80 biopsies from invasive ductal carcinoma, 8 from ductal carcinoma in situ, and 6 from normal breast. We selected them from a recently published deep-sequencing dataset [Farazi TA, et al. (2011) Cancer Res 71:4443-4453]. The microRNA profile established for the normal breast to ductal carcinoma in situ transition was largely maintained in the in situ to invasive ductal carcinoma transition. Nevertheless, a nine-microRNA signature was identified that differentiated invasive from in situ carcinoma. Specifically, let-7d, miR-210, and -221 were down-regulated in the in situ and up-regulated in the invasive transition, thus featuring an expression reversal along the cancer progression path. Additionally, we identified microRNAs for overall survival and time to metastasis. Five noncoding genes were associated with both prognostic signatures--miR-210, -21, -106b*, -197, and let-7i, with miR-210 the only one also involved in the invasive transition. To pinpoint critical cellular functions affected in the invasive transition, we identified the protein coding genes with inversely related profiles to miR-210: BRCA1, FANCD, FANCF, PARP1, E-cadherin, and Rb1 were all activated in the in situ and down-regulated in the invasive carcinoma. Additionally, we detected differential splicing isoforms with special features, including a truncated EGFR lacking the kinase domain and overexpressed only in ductal carcinoma in situ.