Evaluation of Lysis Methods for the Extraction of Bacterial DNA for Analysis of the Vaginal Microbiota

BACKGROUND:Recent studies on the vaginal microbiota have employed molecular techniques such as 16S rRNA gene sequencing to describe the bacterial community as a whole. These techniques require the lysis of bacterial cells to release DNA before purification and PCR amplification of the 16S rRNA gene. Currently, methods for the lysis of bacterial cells are not standardised and there is potential for introducing bias into the results if some bacterial species are lysed less efficiently than others. This study aimed to compare the results of vaginal microbiota profiling using four different pretreatment methods for the lysis of bacterial samples (30 min of lysis with lysozyme, 16 hours of lysis with lysozyme, 60 min of lysis with a mixture of lysozyme, mutanolysin and lysostaphin and 30 min of lysis with lysozyme followed by bead beating) prior to chemical and enzyme-based DNA extraction with a commercial kit. RESULTS:After extraction, DNA yield did not significantly differ between methods with the exception of lysis with lysozyme combined with bead beating which produced significantly lower yields when compared to lysis with the enzyme cocktail or 30 min lysis with lysozyme only. However, this did not result in a statistically significant difference in the observed alpha diversity of samples. The beta diversity (Bray-Curtis dissimilarity) between different lysis methods was statistically significantly different, but this difference was small compared to differences between samples, and did not affect the grouping of samples with similar vaginal bacterial community structure by hierarchical clustering. CONCLUSIONS:An understanding of how laboratory methods affect the results of microbiota studies is vital in order to accurately interpret the results and make valid comparisons between studies. Our results indicate that the choice of lysis method does not prevent the detection of effects relating to the type of vaginal bacterial community one of the main outcome measures of epidemiological studies. However, we recommend that the same method is used on all samples within a particular study

Heat stress on agricultural workers exacerbates crop impacts of climate change

The direct impacts of climate change on crop yields and human health are individually well-studied, but the interaction between the two have received little attention. Here we analyze the consequences of global warming for agricultural workers and the crops they cultivate using a global economic model (GTAP) with explicit treatment of the physiological impacts of heat stress on humans' ability to work. Based on two metrics of heat stress and two labor functions, combined with a meta-analysis of crop yields, we provide an analysis of climate, impacts both on agricultural labor force, as well as on staple crop yields, thereby accounting for the interacting effect of climate change on both land and labor. Here we analyze the two sets of impacts on staple crops, while also expanding the labor impacts to highlight the potential importance on non-staple crops. We find, worldwide, labor and yield impacts within staple grains are equally important at +3 ∘C warming, relative to the 1986–2005 baseline. Furthermore, the widely overlooked labor impacts are dominant in two of the most vulnerable regions: sub-Saharan Africa and Southeast Asia. In those regions, heat stress with 3 ∘C global warming could reduce labor capacity in agriculture by 30%–50%, increasing food prices and requiring much higher levels of employment in the farm sector. The global welfare loss at this level of warming could reach $136 billion, with crop prices rising by 5%, relative to baseline

A melanin-independent interaction between Mc1r and Met signalling pathways is required for HGF-dependent melanoma

Melanocortin 1 receptor (MC1R) signaling stimulates black eumelanin production through a cAMP-dependent pathway. MC1R polymorphisms can impair this process, resulting in a predominance of red phaeomelanin. The red hair, fair skin and UV sensitive phenotype is a well-described melanoma risk factor. MC1R polymorphisms also confer melanoma risk independent of pigment. We investigated the effect of Mc1r deficiency in a mouse model of UV-induced melanoma. C57BL/6-Mc1r+/+-HGF transgenic mice have a characteristic hyperpigmented black phenotype with extra-follicular dermal melanocytes located at the dermal/epidermal junction. UVB induces melanoma, independent of melanin pigmentation, but UVA-induced and spontaneous melanomas are dependent on black eumelanin. We crossed these mice with yellow C57BL/6-Mc1re/e animals which have a non-functional Mc1r and produce predominantly yellow phaeomelanin. Yellow C57BL/6-Mc1re/e-HGF mice produced no melanoma in response to UVR or spontaneously even though the HGF transgene and its receptor Met were expressed. Total melanin was less than in C57BL/6-Mc1r+/+-HGF mice, hyperpigmentation was not observed and there were few extra-follicular melanocytes. Thus, functional Mc1r was required for expression of the transgenic HGF phenotype. Heterozygous C57BL/6-Mc1re/+-HGF mice were black and hyperpigmented and, although extra-follicular melanocytes and skin melanin content were similar to C57BL/6-Mc1r+/+-HGF animals, they developed UV-induced and spontaneous melanomas with significantly less efficiency by all criteria. Thus, heterozygosity for Mc1r was sufficient to restore the transgenic HGF phenotype but insufficient to fully restore melanoma. We conclude that a previously unsuspected melanin-independent interaction between Mc1r and Met signaling pathways is required for HGF-dependent melanoma and postulate that this pathway is involved in human melanoma

Simulating the impact of non-pharmaceutical interventions limiting transmission in COVID-19 epidemics using a membrane computing model

[EN] Epidemics caused by microbial organisms are part of the natural phenomena of increasing biological complexity. The heterogeneity and constant variability of hosts, in terms of age, immunological status, family structure, lifestyle, work activities, social and leisure habits, daily division of time and other demographic characteristics make it extremely difficult to predict the evolution of epidemics. Such prediction is, however, critical for implementing intervention measures in due time and with appropriate intensity. General conclusions should be precluded, given that local parameters dominate the flow of local epidemics. Membrane computing models allows us to reproduce the objects (viruses and hosts) and their interactions (stochastic but also with defined probabilities) with an unprecedented level of detail. Our LOIMOS model helps reproduce the demographics and social aspects of a hypothetical town of 10 320 inhabitants in an average European country where COVID-19 is imported from the outside. The above-mentioned characteristics of hosts and their lifestyle are minutely considered. For the data in the Hospital and the ICU we took advantage of the observations at the Nursery Intensive Care Unit of the Consortium University General Hospital, Valencia, Spain (included as author). The dynamics of the epidemics are reproduced and include the effects on viral transmission of innate and acquired immunity at various ages. The model predicts the consequences of delaying the adoption of non-pharmaceutical interventions (between 15 and 45 days after the first reported cases) and the effect of those interventions on infection and mortality rates (reducing transmission by 20, 50 and 80%) in immunological response groups. The lockdown for the elderly population as a single intervention appears to be effective. This modeling exercise exemplifies the application of membrane computing for designing appropriate multilateral interventions in epidemic situations.MC and FB were sponsored by the Projects COV20 00067 of the Program SARS-COV-2 and COVID-19 infection of the Instituto de Salud Carlos III, Ministerio de Ciencia e Innovacion of Spain, CB06/02/0053 of the Centro de Investigacion Biom edica en Red de Epidemiolog¿a y Salud Publica (CIBERESP), and the Regional Government of Madrid (InGeMICS-B2017/BMD-3691). For JCG, this study was partially founded by the Autonomous Community of Madrid, Spain (COVID-19 Grant, 2020) and the Ramon y Cajal Institute for Health Research (IRYCIS), Madrid, Spain. For AM, this study was supported by grants from the Spanish Ministry of Science and Innovation (PID2019-105969GB-I00), the government of Valencia (project Prometeo/2018/A/133) and cofinanced by the European Regional Development Fund (ERDF).Campos Frances, M.; Sempere Luna, JM.; Galán, JC.; Moya, A.; Llorens, C.; De-Los-Angeles, C.; Baquero-Artigao, F.... (2021). Simulating the impact of non-pharmaceutical interventions limiting transmission in COVID-19 epidemics using a membrane computing model. microLife. 2:1-14. https://doi.org/10.1093/femsml/uqab011S114

“It's Just Presence,” the Contributions of Aboriginal and Torres Strait Islander Health Professionals in Cancer Care in Queensland

Objectives: The aim of this research was to explore health professionals' perspectives on the provision of follow-up cancer care for Aboriginal and Torres Strait Islander patients in Queensland.Methods: Semi-structured interviews were conducted with Indigenous and non-Indigenous health professionals who had experience providing care for Indigenous cancer patients in the primary health care and hospital setting.Results: Participants were recruited from six Aboriginal Community Controlled Health Services (n = 17) and from a tertiary hospital (n = 9) across urban, regional, and remote geographical settings. Culturally safe care, psychological support, determining patient needs, practical assistance, and advocating for Indigenous health were identified as enablers to support the needs of Indigenous patients when accessing cancer care, and Indigenous health professionals were identified as the key enabler.Conclusion: Indigenous health professionals significantly contribute to the provision of culturally competent follow-up cancer care by increasing the accessibility of follow-up cancer care services and by supporting the needs of Indigenous cancer patients. All health professionals need to work together and be sufficiently skilled in the delivery of culturally competent care to improve the Indigenous cancer journey and outcomes for Indigenous people. Effective organizational policies and practices are crucial to enable all health professionals to provide culturally competent and responsive cancer care to Indigenous Australians

An immunoinformatics-derived DNA vaccine encoding human class II T cell epitopes of Ebola virus, Sudan virus, and Venezuelan equine encephalitis virus is immunogenic in HLA transgenic mice

Immunoinformatics tools were used to predict human leukocyte antigen (HLA) class II-restricted T cell epitopes within the envelope glycoproteins and nucleocapsid proteins of Ebola virus (EBOV) and Sudan virus (SUDV) and the structural proteins of Venezuelan equine encephalitis virus (VEEV). Selected epitopes were tested for binding to soluble HLA molecules representing 5 class II alleles (DRB1*0101, DRB1*0301, DRB1*0401, DRB1*0701, and DRB1*1501). All but one of the 25 tested peptides bound to at least one of the DRB1 alleles, and 4 of the peptides bound at least moderately or weakly to all 5 DRB1 alleles. Additional algorithms were used to design a single “string-of-beads” expression construct with 44 selected epitopes arranged to avoid creation of spurious junctional epitopes. Seventeen of these 44 predicted epitopes were conserved between the major histocompatibility complex (MHC) of humans and mice, allowing initial testing in mice. BALB/c mice vaccinated with the multi-epitope construct developed statistically significant cellular immune responses to EBOV, SUDV, and VEEV peptides as measured by interferon (IFN)-γ ELISpot assays. Significant levels of antibodies to VEEV, but not EBOV, were also detected in vaccinated BALB/c mice. To assess immunogenicity in the context of a human MHC, HLA-DR3 transgenic mice were vaccinated with the multi-epitope construct and boosted with a mixture of the 25 peptides used in the binding assays. The vaccinated HLA-DR3 mice developed significant cellular immune responses to 4 of the 25 (16%) tested individual class II peptides as measured by IFN-γ ELISpot assays. In addition, these mice developed antibodies against EBOV and VEEV as measured by ELISA. While a low but significant level of protection was observed in vaccinated transgenic mice after aerosol exposure to VEEV, no protection was observed after intraperitoneal challenge with mouse-adapted EBOV. These studies provide proof of concept for the use of an informatics approach to design a multi-agent, multi-epitope immunogen and provide a basis for further testing aimed at focusing immune responses toward desired protective T cell epitopes

Dicamba Retention in Commercial Sprayers Following Triple Rinse Cleanout Procedures, and Soybean Response to Contamination Concentrations

The commercial launch of dicamba‐tolerant (DT) crops has resulted in increased dicamba usage and a high number of dicamba off‐target movement complaints on sensitive soybeans (Glycine max L.). Dicamba is a synthetic auxin and low dosages as 0.028 g ae ha−1 can induce injury on sensitive soybean. Tank contamination has been identified as one of the sources for unintended sensitive crop exposure. The labels of new dicamba formulations require a triple rinse cleanout procedure following applications. Cleanout efficacy might vary based on the sprayer type and procedure followed. This study was performed to quantify dicamba retention in commercial sprayers and assess the risk for crop injury from remaining contaminants. The results indicate triple rinse with water was comparable to cleanout procedures utilizing ammonium, commercial tank cleaners, and glyphosate in rinses. Dicamba contaminants in final rinsates resulted in \u3c15% visual injury and no yield response when applied to sensitive soybeans at R1 stage. A survey of 25 agricultural sprayers demonstrated a cleanout efficacy of 99.996% by triple rinsing with water following applications of dicamba at 560 g ae ha−1, with concentrations of less than 1 ug mL−1 detected rinsates from the fourth rinse. A dose response experiment predicted dosages causing 5% visual injury and the yield losses were 0.1185 and 2.8525 g ae ha−1. However, symptomology was observed for all tested dosages, including the rate as low as 0.03 g ae ha−1. The results from this study suggest triple rinsing with sufficient amount of water (≥10% of tank volume) is adequate for the removal of dicamba residues from sprayers to avoid sensitive soybean damage. This study can provide producers with confidence in cleanout procedures following dicamba applications, and aid in minimizing risk for off‐target movement through tank contamination