Reducing firearm access for youth at risk for suicide in a pediatric emergency department.

Background: Firearm-related suicide is the second leading cause of pediatric firearm death. Lethal means counseling (LMC) can improve firearm safe-storage practices for families with youth at risk of suicide. Objectives: This study aims to evaluate the feasibility of pediatric emergency department (ED) behavioral mental health (BMH) specialists providing LMC to caregivers of youth presenting with BMH complaints and to test for changes in firearm safety practices, pre-post ED LMC intervention, as measures of preliminary efficacy. Methods: Prospective pilot feasibility study of caregivers of youth presenting to a pediatric ED with BMH complaints. Caregivers completed an electronic survey regarding demographics and firearm safe-storage knowledge/practices followed by BMH specialist LMC. Firearm owners were offered a free lockbox and/or trigger lock. One-week follow-up surveys gathered self-reported data on firearm safety practices and intervention acceptability. One-month interviews with randomly sampled firearm owners collected additional firearm safety data. Primary outcomes were feasibility measures, including participant accrual/attrition and LMC intervention acceptability. Secondary outcomes included self-reported firearm safety practice changes. Feasibility benchmarks were manually tabulated, and Likert-scale acceptability responses were dichotomized to strongly agree/agree vs. neutral/disagree/strongly disagree. Descriptive statistics were used for univariate and paired data responses. Results: In total, 81 caregivers were approached; of which, 50 (81%) caregivers enrolled. A total of 44% reported having a firearm at home, 80% completed follow-up at one week. More than 80% affirmed that ED firearm safety education was useful and that the ED is an appropriate place for firearm safety discussions. In total, 58% of participants reported not having prior firearm safety education/counseling. Among firearm owners (n = 22), 18% reported rarely/never previously using a safe-storage device, and 59% of firearm owners requested safe storage devices.At 1-week follow-up (n = 40), a greater proportion of caregivers self-reported asking about firearms before their child visited other homes (+28%). Among firearm owners that completed follow-up (n = 19), 100% reported storing all firearms locked at one week (+23% post-intervention). In total, 10 caregivers reported temporarily/permanently removing firearms from the home. Conclusion: It is feasible to provide LMC in the pediatric ED via BMH specialists to families of high-risk youth. Caregivers were receptive to LMC and reported finding this intervention useful, acceptable, and appropriate. Additionally, LMC and device distribution led to reported changes in safe storage practices.

Presence of pathogen DNA in milk harvested from quarters is associated to changes in cows' milk yield and composition.

BACKGROUND: Intramammary infection is the result of invasion and multiplication of microorganisms in the mammary gland and commonly leads to mastitis in dairy animals. Although much has been done to improve cows' udder health, mastitis remains a significant and costly health issue for dairy farmers, especially if subclinical. In this study, quarter milk samples from clinically healthy cows were harvested to detect pathogens via quantitative PCR (qPCR) and evaluate changes in individual milk traits according to the number of quarters infected and the type of microorganism(s). A commercial qPCR kit was used for detection of Mycoplasma bovis, Mycoplasma spp., Staphylococcus aureus, coagulase-negative staphylococci (CNS), Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, Prototheca spp., Escherichia coli, Klebsiella spp., Enterococcus spp. and Lactococcus lactis ssp. lactis. Quarter and pooled milk information of 383 Holstein, 132 Simmental, 129 Rendena, and 112 Jersey cows in 9 Italian single-breed herds was available. RESULTS: Among the cows with pathogen(s) present in at least 1 quarter, CNS was the most commonly detected DNA, followed by Streptococcus uberis, Mycoplasma bovis, and Streptococcus agalactiae. Cows negative to qPCR were 206 and had the lowest milk somatic cell count. Viceversa, cows with DNA isolated in ≥ 3 quarters were those with the highest somatic cell count. Moreover, when major pathogens were isolated in ≥ 3 quarters, milk had the lowest casein index and lactose content. In animals with pathogen(s) DNA isolated, the extent with whom milk yield and major solids were impaired did not significantly differ between major and minor pathogens. CONCLUSIONS: The effect of the number of affected quarters on the pool milk quality traits was investigated in clinically healthy cows using a commercial kit. Results remark the important negative effect of subclinical udder inflammations on milk yield and quality, but more efforts should be made to investigate the presence of untargeted microorganisms, as they may be potentially dangerous for cows. For a smarter use of antimicrobials, analysis of milk via qPCR is advisable - especially in cows at dry off - to identify quarters at high risk of inflammation and thus apply a targeted/tailored treatment.

Heterologous expression and structure prediction of a xylanase identified from a compost metagenomic library.

Xylanases are key biocatalysts in the degradation of the ß-1,4-glycosidic linkages in the xylan backbone of hemicellulose. These enzymes are potentially applied in a wide range of bioprocessing industries under harsh conditions. Metagenomics has emerged as powerful tools for the bioprospection and discovery of interesting bioactive molecules from extreme ecosystems with unique features, such as high temperatures. In this study, an innovative combination of function-driven screening of a compost metagenomic library and automatic extraction of halo areas with in-house MATLAB functions resulted in the identification of a promising clone with xylanase activity (LP4). The LP4 clone proved to be an effective xylanase producer under submerged fermentation conditions. Sequence and phylogenetic analyses revealed that the xylanase, Xyl4, corresponded to an endo-1,4-ß-xylanase belonging to glycosyl hydrolase family 10 (GH10). When xyl4 was expressed in Escherichia coli BL21(DE3), the enzyme activity increased about 2-fold compared to the LP4 clone. To get insight on the interaction of the enzyme with the substrate and establish possible strategies to improve its activity, the structure of Xyl4 was predicted, refined, and docked with xylohexaose. Our data unveiled, for the first time, the relevance of the amino acids Glu133 and Glu238 for catalysis, and a close inspection of the catalytic site suggested that the replacement of Phe316 by a bulkier Trp may improve Xyl4 activity. Our current findings contribute to enhancing the catalytic performance of Xyl4 towards industrial applications. KEY POINTS: • A GH10 endo-1,4-ß-xylanase (Xyl4) was isolated from a compost metagenomic library • MATLAB's in-house functions were developed to identify the xylanase-producing clones • Computational analysis showed that Glu133 and Glu238 are crucial residues for catalysis.

Unraveling the evolutionary origin of the complex Nuclear Receptor Element (cNRE), a cis-regulatory module required for preferential expression in the atrial chamber.

Cardiac function requires appropriate proteins in each chamber. Atria requires slow myosin to act as reservoirs, while ventricles demand fast myosin for swift pumping. Myosins are thus under chamber-biased cis-regulation, with myosin gene expression imbalances leading to congenital heart dysfunction. To identify regulatory inputs leading to cardiac chamber-biased expression, we computationally and molecularly dissected the quail Slow Myosin Heavy Chain III (SMyHC III) promoter that drives preferential expression to the atria. We show that SMyHC III gene states are orchestrated by a complex Nuclear Receptor Element (cNRE) of 32 base pairs. Using transgenesis in zebrafish and mice, we demonstrate that preferential atrial expression is achieved by a combinatorial regulatory input composed of atrial activation motifs and ventricular repression motifs. Using comparative genomics, we show that the cNRE might have emerged from an endogenous viral element through infection of an ancestral host germline, revealing an evolutionary pathway to cardiac chamber-specific expression.

The link between obesity and the gut microbiota and immune system in early-life.

In early-life, the gut microbiota is highly modifiable, being modulated by external factors such as maternal microbiota, mode of delivery, and feeding strategies. The composition of the child's gut microbiota will deeply impact the development and maturation of its immune system, with consequences for future health. As one of the main sources of microorganisms to the child, the mother represents a crucial factor in the establishment of early-life microbiota, impacting the infant's wellbeing. Recent studies have proposed that dysbiotic maternal gut microbiota could be transmitted to the offspring, influencing the development of its immunity, and leading to the development of diseases such as obesity. This paper aims to review recent findings in gut microbiota and immune system interaction in early-life, highlighting the benefits of a balanced gut microbiota in the regulation of the immune system.

The ability of a handheld near-infrared spectrometer to do a rapid quality assessment of bovine colostrum, including the immunoglobulin G concentration.

Portable infrared-based instruments have made important contributions in different research fields. Within the dairy supply chain, for example, most of portable devices are based on near-infrared spectroscopy (NIRS) and are nowadays an important support for farmers and operators of the dairy sector, allowing fast and real-time decision-making, particularly for feed and milk quality evaluation and animal health and welfare monitoring. The affordability, portability, and ease of use of these instruments have been pivotal factors for their implementation on farm. In fact, pocket-sized devices enable nonexpert users to perform quick, low-cost, and nondestructive analysis on various matrixes without complex preparation. Because bovine colostrum (BC) quality is mostly given by the IgG level, evaluating the ability of portable NIRS tools to measure antibody concentration is advisable. In this study we used the wireless device SCiO manufactured by Consumer Physics Inc. (Tel Aviv, Israel) to collect BC spectra and then attempt to predict IgG concentration and gross and fine composition in individual samples collected immediately after calving (<6 h) in primiparous and pluriparous Holstein cows on 9 Italian farms. Chemometric analyses revealed that SCiO has promising predictive performance for colostral IgG concentration, total Ig concentration, fat, and AA. The coefficient of determination of cross-validation (R2CV) was in fact ≥0.75). Excellent accuracy was observed for dry matter, protein, and S prediction in cross-validation and good prediction ability in external validation (R2CV ≥ 0.93; the coefficient of determination of external validation, R2V, was ≥0.82). Nonetheless, SCiO's ability to discriminate between good- and low-quality samples (IgG ≥ vs. < 50 g/L) was satisfactory. The affordable cost, the accurate predictions, and the user-friendly design, coupled with the increased interest in BC within the dairy sector, may boost the collection of extensive BC data for management and genetic purposes in the near future.

Phylogenetics, Epidemiology and Temporal Patterns of Dengue Virus in Araraquara, São Paulo State.

Dengue virus (DENV) is a prominent arbovirus with global spread, causing approximately 390 million infections each year. In Brazil, yearly epidemics follow a well-documented pattern of serotype replacement every three to four years on average. Araraquara, located in the state of São Paulo, has faced significant impacts from DENV epidemics since the emergence of DENV-1 in 2010. The municipality then transitioned from low to moderate endemicity in less than 10 years. Yet, there remains an insufficient understanding of virus circulation dynamics, particularly concerning DENV-1, in the region, as well as the genetic characteristics of the virus. To address this, we sequenced 37 complete or partial DENV-1 genomes sampled from 2015 to 2022 in Araraquara. Then, using also Brazilian and worldwide DENV-1 sequences we reconstructed the evolutionary history of DENV-1 in Araraquara and estimated the time to the most recent common ancestor (tMRCA) for serotype 1, for genotype V and its main lineages. Within the last ten years, there have been at least three introductions of genotype V in Araraquara, distributed in two main lineages (L Ia and L Ib, and L II). The tMRCA for the first sampled lineage (2015/2016 epidemics) was approximately 15 years ago (in 2008). Crucially, our analysis challenges existing assumptions regarding the emergence time of the DENV-1 genotypes, suggesting that genotype V might have diverged more recently than previously described. The presence of the two lineages of genotype V in the municipality might have contributed to the extended persistence of DENV-1 in the region.

[Eating Habits of People with Type 2 Diabetes Mellitus in Portugal: A Cross-Sectional Study]. / Hábitos Alimentares das Pessoas com Diabetes Mellitus Tipo 2 em Portugal: Um Estudo Transversal.

INTRODUCTION: Nutrition is a cornerstone of diabetes mellitus prevention and management; therefore, it is essential to enable patients to adopt healthy eating habits. Previous studies have not yet documented the main errors in the eating habits of Portuguese people with type 2 diabetes mellitus. This study aims to identify the main errors in the eating habits of people living with type 2 diabetes mellitus in Portugal and to evaluate its associations with sociodemographic variables. METHODS: Cross-sectional multicentric study in a convenience sample of people with type 2 diabetes mellitus in Primary Health Care Units. The UK Diabetes and Diet Questionnaire (UKDDQ) - translated and adapted, was applied from July to October 2022. Descriptive and inferential statistical analyses were conducted. RESULTS: Of the 550 participants, 52.2% were female, 68.3% were 65 years or over, 55.8 % had an education level up to the fourth grade, 24.7% had economic deprivation, and the mean time since diagnosis was 10.60 ± 8.13 years. Only 36.2% of the sample had a healthy UKDDQ score. Less than 50% of the sample had healthy scores for the items "high-fiber rice or pasta", "high-fiber bread", "butter, margarine and vegetable oils" and "vegetables and pulses". Only 8.9% of the sample had a healthy consumption of fiber. About 70.4% reported healthy scores for the consumption of "high-added-sugar foods" and 54.7% for "high-saturated fat". A statistically significant weak positive correlation was found between the UKDDQ score and age (ρ = 0.201, p < 0.001) with a more frequent choice of healthy foods with increasing age. Female respondents reported healthier habits, particularly in the consumption of "high-saturated fat" and "high-fiber foods". CONCLUSION: The majority of our sample did not take advantage of the potential benefits of healthy eating habits. The main food groups whose consumption should be emphasized or discouraged were individualized, particularly the need to encourage the consumption of high-fiber foods. Targeted educational actions must focus especially on younger and/or male patients.

Introdução: A nutrição é uma área de intervenção na prevenção e gestão da diabetes mellitus; por isso, é fulcral promover a capacitação da população para a adoção de hábitos alimentares saudáveis. Ainda que existam alguns estudos nesta área, não se conhecem os principais erros nos hábitos alimentares das pessoas com diabetes em Portugal. Os objetivos deste estudo foram identificar os principais erros nos hábitos alimentares das pessoas com diabetes mellitus tipo 2 em Portugal e avaliar a sua relação com variáveis sociodemográficas. Métodos: Estudo transversal multicêntrico, em amostra de conveniência de pessoas com diabetes mellitus tipo 2 seguidas em Unidades de Cuidados de Saúde Primários. Aplicação do UK Diabetes and Diet Questionnaire (UKDDQ) ­ traduzido e adaptado, de julho a outubro de 2022. Análise estatística descritiva e inferencial. Resultados: Amostra de 550 participantes, 52,2% do sexo feminino, 68,3% com 65 anos ou mais, 55,8% com nível de escolaridade igual ou inferior ao 1.º ciclo do ensino básico, 24,7% com insuficiência económica e tempo desde o diagnóstico médio de 10,60 ± 8,13 anos. Apenas 36,2% da amostra obteve um score UKDDQ considerado saudável. Menos de 50% obteve scores saudáveis para os itens "arroz ou massa ricos em fibras", "pão integral", "manteiga, margarina e óleos vegetais" e "vegetais e leguminosas". Somente 8,9% da amostra obteve score saudável para o consumo de fibras. Cerca de 70,4% obteve score saudável para o consumo de açúcares livres e 54,7% para o consumo de ácidos gordos saturados. Verificou-se a existência de uma correlação com significado estatístico positiva fraca entre o score UKDDQ e a idade (ρ = 0,201, p < 0,001), com escolha mais frequente de alimentos saudáveis com o aumentar da idade. As pessoas do sexo feminino reportaram hábitos alimentares mais saudáveis, particularmente no consumo de fibras e ácidos gordos saturados. Conclusão: A maior parte da nossa amostra não usufruiu do potencial efeito positivo de uma alimentação saudável. Individualizam-se grupos de alimentos cujos consumos devem ser enfatizados ou desencorajados, particularmente, a necessidade de incentivar o consumo de alimentos ricos em fibra. Ações educacionais dirigidas devem ter especial foco em pessoas mais jovens e/ou do sexo masculino.

Teat morphology across five buffalo breeds: a multi-country collaborative study.

The buffalo (Bubalus bubalis) is a species of worldwide importance, raised to produce milk, meat, and hides, and often used as a working animal in rural contexts with low access to hi-tech solutions. In the present study, 100 lactating buffaloes (50 primiparous and 50 pluriparous) of five popular breeds were recruited to characterize and compare teat morphology. In particular, the focus was put on the Nili Ravi, Mediterranean, Egyptian, Bulgarian Murrah, and Azeri buffaloes raised in Pakistan, Italy, Egypt, Bulgaria, and Iran, respectively. In all countries, a longitudinal cross-section ultrasound was obtained before the milking to measure teat parameters at individual level: overall, teat canal length (TCL) averaged 24.13 mm, teat diameter (TD) 30.46 mm, cisternal diameter (CD) 17.80 mm, and teat wall (TW) 7.12 mm. The most variable trait across breeds was TCL which was positively correlated with CD and TD and negatively with TW, regardless of the teat position (front/rear or left/right). A strong negative correlation was found between TW and CD (- 0.43). The analysis of variance revealed that the fixed effect of breed significantly affected all the traits except TD. In fact, Bulgarian Murrah, Azeri, and Egyptian buffaloes presented the greatest estimate of TCL, whereas NR the smallest (14.70 mm). The TW was maximum in Nili Ravi, Egyptian, and Mediterranean buffaloes, with estimates equal to 8.19, 7.59, and 8.74 mm, respectively. Nili Ravi also showed the greatest TL (82.39 mm). In terms of CD, the lowest least square mean was that of Mediterranean buffaloes (12.14 mm). Primiparous and pluriparous buffaloes differed in terms of TD, TW, and TL, with older animals presenting the highest least square mean. In terms of position, instead, significant differences were observed for TD, CD, and TL when comparing front and rear teats, as left and right teats did not differ. Teat anatomy includes a set of heritable morphological features and is therefore breed-dependent. Differences presented in this study could be attributed to the divergent breeding objective and selective pressure across the five breeds; e.g., in some cases such as Mediterranean buffalo, selection for decades was oriented to improve milk production and milkability and achieve optimal conformation for mechanical milking. A better understanding of the mammary gland anatomical descriptors can be informative of the history of a breed and could provide useful insights to guide possible selection.

The effects of dietary iodine content, milking system, and farming practices on milk iodine concentration and quality traits.

Various management practices can influence milk quality traits in dairy cattle. As an example, an increasing investment in automatic milking system to substitute milking parlors has been observed in the last 2 decades in dairy farms which could have affected certain bulk milk quality traits. What is more, milking practices can also affect certain milk parameters; as an example, teat disinfectants containing I are used in commercial farms where pre- or postdipping is performed, leading to presence of some I in the bulk milk. However, this trace mineral is also supplied in cows' diet to fulfill their nutritional requirements, partly contributing to the milk I final concentration. Therefore, the aim of this study was to evaluate the sources of variation of milk I along with other traditional milk quality traits. A total of 91 dairy farms in northeastern Italy were enrolled in the study. In each farm, diet and bulk milk samples were collected on the same day for chemical analysis. Concentration of I, in particular, was determined in both milk and feed with gold standard. Pearson correlations were calculated among the traits available for milk and diet, and a general linear model was used to test significance of fixed effects (feeding system, milking system, farming system, herd size, herd stage of lactation, and sampling month) on milk quality traits including the I concentration. In the case of milk I, diet I and presence of I-based predipping and postdipping teat disinfect application were also tested as fixed effects. Results showed a positive linear correlation between milk and diet I content (correlation coefficient [r] = 0.78). Although milk I was also positively correlated with lactose content (r = 0.25), dietary I was not correlated with other milk traits. Milk I content was significantly affected by dietary I, I-based predipping teat disinfectant application, and herd composition. Compared with conventional farms, organic farms showed lower protein content and greater somatic cell score (SCS) but similar milk I. Milking system significantly affected only lactose content and SCS of milk. Sampling month was only significant for milk urea nitrogen and herd composition, feeding system, herd size, and herd average days in milk did not modify milk gross composition and SCS. In conclusion, dietary supply of I is the main factor affecting milk I concentration and findings suggest that I level in milk can be naturally improved in dairy cows by modulating the I content in the diet administered. However, further research is needed to evaluate the effect of I-based sanitizers on milk I.

Genetic evaluation of gestation length in Italian Holstein breed.

Gestation length (GL) can potentially affect health and performance of both the dam and the newborn calf, and it is controlled by two genetic components, direct and maternal. This means that both the calf (direct effect) and the cow (maternal effect) genotypes contribute to determine GL and its variability. The aims of the present study were to estimate direct and maternal variance components of GL, develop a routine genetic evaluation of GL in Italian Holstein and evaluate potential (un)favourable associations with traits for which selection is undertaken in this population. A multiple-trait repeatability linear animal model was employed for the estimation of variance components considering GL in first and later parities as different traits. The posterior mean (PM) of heritability of the direct effect was 0.43 for first parity and 0.35 for later parities. The PM of heritability of the maternal effect was lower, being 0.08 for primiparae and 0.06 for pluriparae. The posterior standard deviation (PSD) of the heritability estimates was small, ranging from 0.001 to 0.005. The relationship of direct and maternal effects with important traits such as milk yield and fertility indicated that selecting for extreme GL, longer or shorter, may have negative consequences on several traits, suggesting that GL has an intermediate optimum in dairy cattle. In conclusion, this study reveals that selecting an intermediate GL in the Italian Holstein population is advisable. Although scarcely variable compared to other conventional traits for which Italian Holstein is selected, GL is heritable and a deeper knowledge can be useful for decision-making at the farm level.

scFv biofunctionalized nanoparticles to effective and safe targeting of CEA-expressing colorectal cancer cells.

Colorectal cancer (CRC) is one of the deadliest cancers worldwide, with the 5 year survival rate in metastatic cases limited to 12%. The design of targeted and effective therapeutics remains a major unmet clinical need in CRC treatment. Carcinoembryonic antigen (CEA), a glycoprotein overexpressed in most colorectal tumors, may constitute a promising molecule for generating novel CEA-targeted therapeutic strategies for CRC treatment. Here, we developed a smart nanoplatform based on chemical conjugation of an anti-CEA single-chain variable fragment (scFv), MFE-23, with PLGA-PEG polymers to deliver the standard 5-Fluorouracil (5-FU) chemotherapy to CRC cells. We confirmed the specificity of the developed CEA-targeted NPs on the internalization by CEA-expressing CRC cells, with an enhance of threefold in the cell uptake. Additionally, CEA-targeted NPs loaded with 5-FU induced higher cytotoxicity in CEA-expressing cells, after 24 h and 48 h of treatment, reinforcing the specificity of the targeted NPs. Lastly, the safety of CEA-targeted NPs loaded with 5-FU was evaluated in donor-isolated macrophages, with no relevant impact on their metabolic activity nor polarization. Altogether, this proof of concept supports the CEA-mediated internalization of targeted NPs as a promising chemotherapeutic strategy for further investigation in different CEA-associated cancers and respective metastatic sites.Authors: Please confirm if the author names are presented accurately and in the correct sequence (given name, middle name/initial, family name). Author 1 Given name: [Maria José] Last name [Silveira]. Author 7 Given name: [Maria José] Last name [Oliveira]. Also, kindly confirm the details in the metadata are correctokAffiliations: Please check and confirm that the authors and their respective affiliations have been correctly identified and amend if necessary.ok.

Pathogen Detection via Quantitative PCR in Milk of Healthy Cows Collected Using Different Sampling Protocols.

In this study we evaluated the prevalence of pathogens detected via quantitative PCR (qPCR) in milk from apparently healthy cows to identify the most common etiological agents present in Italian dairy farms. Milk samples were collected using a sterile protocol at quarter-level (3239 samples, 822 cows) and a conventional protocol at udder level as composite milk from the functional quarters of each cow (5464 samples, 5464 cows). The qPCR commercial kit detected Mycoplasma bovis, Mycoplasma spp., Staphylococcus aureus, coagulase-negative staphylococci (CNS), Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus uberis, Prototheca spp., Escherichia coli, Klebsiella spp., Enterococcus spp. and Lactococcus lactis ssp. lactis as well as DNA from the penicillin resistance ß-lactamase gene from staphylococci. The prevalence of specific DNA was calculated based on its presence or absence in the samples, factoring in both the sampling protocols and herds. Regardless of the sampling protocol used, the most frequently detected pathogens were CNS (26.6% in sterile and 13.9% in conventional protocol) and Streptococcus uberis (9.6% and 16.5%, respectively). These results underscore the necessity for pathogen-specific interventions at the farm level to enhance the udder health of dairy cows via management recommendations.

Functional and sequence-based metagenomics to uncover carbohydrate-degrading enzymes from composting samples.

The renewable, abundant , and low-cost nature of lignocellulosic biomass can play an important role in the sustainable production of bioenergy and several added-value bioproducts, thus providing alternative solutions to counteract the global energetic and industrial demands. The efficient conversion of lignocellulosic biomass greatly relies on the catalytic activity of carbohydrate-active enzymes (CAZymes). Finding novel and robust biocatalysts, capable of being active under harsh industrial conditions, is thus imperative to achieve an economically feasible process. In this study, thermophilic compost samples from three Portuguese companies were collected, and their metagenomic DNA was extracted and sequenced through shotgun sequencing. A novel multi-step bioinformatic pipeline was developed to find CAZymes and characterize the taxonomic and functional profiles of the microbial communities, using both reads and metagenome-assembled genomes (MAGs) as input. The samples' microbiome was dominated by bacteria, where the classes Gammaproteobacteria, Alphaproteobacteria, and Balneolia stood out for their higher abundance, indicating that the degradation of compost biomass is mainly driven by bacterial enzymatic activity. Furthermore, the functional studies revealed that our samples are a rich reservoir of glycoside hydrolases (GH), particularly of GH5 and GH9 cellulases, and GH3 oligosaccharide-degrading enzymes. We further constructed metagenomic fosmid libraries with the compost DNA and demonstrated that a great number of clones exhibited ß-glucosidase activity. The comparison of our samples with others from the literature showed that, independently of the composition and process conditions, composting is an excellent source of lignocellulose-degrading enzymes. To the best of our knowledge, this is the first comparative study on the CAZyme abundance and taxonomic/functional profiles of Portuguese compost samples. KEY POINTS: • Sequence- and function-based metagenomics were used to find CAZymes in compost samples. • Thermophilic composts proved to be rich in bacterial GH3, GH5, and GH9 enzymes. • Compost-derived fosmid libraries are enriched in clones with ß-glucosidase activity.

Iron metabolism in colorectal cancer: a balancing act.

BACKGROUND: Colorectal cancer (CRC) is the third most commonly diagnosed cancer and the second deadliest malignancy worldwide. Current dietary habits are associated with increased levels of iron and heme, both of which increase the risk of developing CRC. The harmful effects of iron overload are related to the induction of iron-mediated pro-tumorigenic pathways, including carcinogenesis and hyperproliferation. On the other hand, iron deficiency may also promote CRC development and progression by contributing to genome instability, therapy resistance, and diminished immune responses. In addition to the relevance of systemic iron levels, iron-regulatory mechanisms in the tumor microenvironment are also believed to play a significant role in CRC and to influence disease outcome. Furthermore, CRC cells are more prone to escape iron-dependent cell death (ferroptosis) than non-malignant cells due to the constitutive activation of antioxidant genes expression. There is wide evidence that inhibition of ferroptosis may contribute to the resistance of CRC to established chemotherapeutic regimens. As such, ferroptosis inducers represent promising therapeutic drugs for CRC. CONCLUSIONS AND PERSPECTIVES: This review addresses the complex role of iron in CRC, particularly in what concerns the consequences of iron excess or deprivation in tumor development and progression. We also dissect the regulation of cellular iron metabolism in the CRC microenvironment and emphasize the role of hypoxia and of oxidative stress (e.g. ferroptosis) in CRC. Finally, we underline some iron-related players as potential therapeutic targets against CRC malignancy.

Pro-inflammatory polarization and colorectal cancer modulate alternative and intronic polyadenylation in primary human macrophages.

Introduction: Macrophages are essential cells of the immune system that alter their inflammatory profile depending on their microenvironment. Alternative polyadenylation in the 3'UTR (3'UTR-APA) and intronic polyadenylation (IPA) are mechanisms that modulate gene expression, particularly in cancer and activated immune cells. Yet, how polarization and colorectal cancer (CRC) cells affect 3'UTR-APA and IPA in primary human macrophages was unclear. Methods: In this study, we isolated primary human monocytes from healthy donors, differentiated and polarized them into a pro-inflammatory state and performed indirect co-cultures with CRC cells. ChrRNA-Seq and 3'RNA-Seq was performed to quantify gene expression and characterize new 3'UTR-APA and IPA mRNA isoforms. Results: Our results show that polarization of human macrophages from naïve to a pro-inflammatory state causes a marked increase of proximal polyA site selection in the 3'UTR and IPA events in genes relevant to macrophage functions. Additionally, we found a negative correlation between differential gene expression and IPA during pro-inflammatory polarization of primary human macrophages. As macrophages are abundant immune cells in the CRC microenvironment that either promote or abrogate cancer progression, we investigated how indirect exposure to CRC cells affects macrophage gene expression and 3'UTR-APA and IPA events. Co-culture with CRC cells alters the inflammatory phenotype of macrophages, increases the expression of pro-tumoral genes and induces 3'UTR-APA alterations. Notably, some of these gene expression differences were also found in tumor-associated macrophages of CRC patients, indicating that they are physiologically relevant. Upon macrophage pro-inflammatory polarization, SRSF12 is the pre-mRNA processing gene that is most upregulated. After SRSF12 knockdown in M1 macrophages there is a global downregulation of gene expression, in particular in genes involved in gene expression regulation and in immune responses. Discussion: Our results reveal new 3'UTR-APA and IPA mRNA isoforms produced during pro-inflammatory polarization of primary human macrophages and CRC co-culture that may be used in the future as diagnostic or therapeutic tools. Furthermore, our results highlight a function for SRSF12 in pro-inflammatory macrophages, key cells in the tumor response.

Arthrodesis of the Fourth and Fifth Tarsometatarsal Joints in a Sensate Patient with Post-Traumatic Arthritis: A Case Report and Surgical Technique.

Lateral column arthrodesis of the tarsometatarsal joints is a highly controversial topic in foot and ankle surgery, with minimal prospective research and reproducible findings in the current literature. Arthrodesis of the lateral fourth and fifth tarsometatarsal joints, when performed, is most often done secondary to post-traumatic osteoarthritis or Charcot's neuroarthropathy deformity. This case report focuses on arthrodesis of the lateral column in a patient with post-traumatic osteoarthritis from a previously sustained Lisfranc fracture-dislocation. The patient also experienced a cavus foot deformity that was addressed with a lateral displacement calcaneal osteotomy. Arthrodesis of the fourth and fifth tarsometatarsal joints was found to be successful on this patient, with bony union noted to occur radiographically 12 weeks postoperatively. In addition, the patient experienced significant reduction in her preoperative pain and an ability to return to activities of daily living. Regular visits during an 18-month postoperative period occurred, with the patient continuing to have satisfactory results and a significant reduction in preoperative pain levels. One postoperative complication was encountered 15 months postoperatively: painful hardware, which resulted in the removal of both calcaneal screws and one screw from the fourth tarsometatarsal arthrodesis site. This case report proposes that lateral column arthrodesis may be performed successfully in select patients where other joint-preserving procedures may not be applicable. Herein we outline a suggested surgical technique with hardware that can be used to reproduce these findings and assist surgeons who are unfamiliar with performing this procedure.

Stayability in Simmental cattle as affected by muscularity and body condition score between calvings.

The present study aimed to investigate the association between stayability (STAY) traits, muscularity, and body condition score (BCS) in the Italian Simmental dual-purpose cows. Data were collected from 2,656 cows linearly scored in their first lactation from 2002 to 2020 and reared in 324 herds. The binary trait STAY, which is the ability of a cow to stay in the herd, was obtained for each cow-lactation available up to parity 5 (from STAY1-2 to STAY4-5). Analysis of STAY was carried out using logistic regression, considering the fixed effect of energy corrected milk, conception rate, somatic cell score, and muscularity or BCS predicted at different time points. The herd of linear classification and residual error were the random effects. Primiparous cows with a medium BCS and muscularity in early lactation presented a more favorable STAY across life compared to thinner ones (P < 0.05). In fact, cows with an intermediate BCS/muscularity were more likely to stay in the herd after the third lactation (STAY3-4), compared to those presenting a lower BCS/muscularity (P < 0.01). However, cows whose muscularity was high were generally less likely to start the third lactation compared to the others. A potential explanation for this could be the willing to market cows with good conformation for meat purpose. Simmental is in fact a dual-purpose breed known for the good carcass yield and meat quality. This study demonstrates how muscularity and BCS available early in life can be associated with the ability of Simmental cows to stay in the herd.

Total and Differential Somatic Cell Count in Italian Local Cattle Breeds: Phenotypic Variability and Effect on Milk Yield and Composition.

Milk differential somatic cell count (DSCC) represents the percentage of polymorphonuclear neutrophils and lymphocytes out of the total somatic cell count (SCC) and has been proposed in recent years as a proxy for udder health in dairy cows. We investigated phenotypic factors affecting SCC and DSCC using 3978 records of 212 Alpine Grey and 426 Burlina cows farmed in Northern Italy. The linear mixed model accounted for the fixed effects of breed, parity, lactation stage, sampling season, and first-order interactions of breed with the other effects. Cow, herd-test-date nested within breed were random. Subsequently, four udder health status groups (UHS) were created by combining SCC and DSCC to assess the UHS impact on milk yield and quality. DSCC was greater in Alpine Grey (66.2 ± 0.8%) than Burlina cows (63.2 ± 0.6%) and, similarly to SCC, it increased with days in milk and parity regardless of breed. Milk yield and composition were affected by UHS in both breeds. These results suggest that also udder health of local breeds can be monitored on a large scale through SCC and DSCC for reduction in biodiversity loss and increased farm profitability. However, in addition to milk data, the introduction of mastitis recording and monitoring plans is advisable.

Genetic characteristics of colostrum refractive index and its use as a proxy for the concentration of immunoglobulins in Holstein cattle.

BACKGROUND: Colostral concentration of immunoglobulins (Ig) is crucial for the passive transfer of antibodies from the cow to the new-born calf. Direct determination of Ig by the gold standard radial immunodiffusion method is demanding in terms of time and costs. For this reason, a refractometer is commonly used at the farm level for an indirect estimation of colostrum quality, which is given as the Ig concentration. In this study, colostrum samples were collected from 548 Italian Holstein cows within 6 h of calving. The refractive index (BRIX, %) of these samples was assessed using a portable optical refractometer, as well as the concentration of total protein, IgG, IgA, and IgM by radial immunodiffusion. A four-trait animal model was used to estimate genetic parameters for BRIX and the different immunoglobulin isotypes. A receiver operating characteristic analysis was carried out to evaluate the BRIX diagnostic accuracy. RESULTS: Colostral BRIX was moderately heritable (0.26) and its genetic and phenotypic correlations with IgG (0.91, 0.78), IgA (0.57, 0.57), and IgM (0.71, 0.61) were all positive and of similar order, although the genetic correlations were generally higher than the phenotypic correlations. Low-quality colostrum samples, defined as those with an IgG concentration lower than 50 g/L, were accurately identified by the refractive index on the BRIX scale, with an area under the curve of 0.90. CONCLUSIONS: The use of a refractometer is recommended on dairy farms to produce a proxy for colostral Ig concentration. BRIX is a useful phenotyping tool that can be used in cattle to improve the quality of colostrum for first feeding of calves through both traditional genetic and genomic strategies. Improving colostrum quality will reduce the incidence of failure of passive transfer of immunity in young stock.