A novel natural antimicrobial can reduce the in vitro and in vivo pathogenicity of T6SS positive Campylobacter jejuni and campylobacter coli chicken isolates

© 2018 Sima, Stratakos, Ward, Linton, Kelly, Pinkerton, Stef, Gundogdu, Lazar and Corcionivoschi. Human campylobacteriosis is considered one of the most common foodborne diseases worldwide with poultry identified as the main source of infection accounting for 50-80% of human cases. Highly virulent Campylobacter spp., positive for the Type VI secretion system (T6SS), which have an increased ability to adhere to and invade the host gastrointestinal epithelium are highly prevalent in poultry. Multidrug resistant strains of bacteria are rapidly evolving and therefore, new antimicrobials to supplement animal feed that are able to control Campylobacter species, are in great need. The work presented herein indicates that a novel phenolic antimicrobial, Auranta 3001, is able to reduce the adhesion and invasion of human intestinal epithelial cells (HCT-8) by two T6SS positive chicken isolates, C. jejuni RC039 (p < 0.05) and C. coli RC013 (p < 0.001). Exposure of C. jejuni RC039 and C. coli RC013 to Auranta 3001 downregulated the expression of hcp and cetB genes, known to be important in the functionality of T6SS. Furthermore, the reduced adhesion and invasion is associated with a significant decrease in bacterial motility of both isolates (p < 0.05-p < 0.001) in vitro. Most importantly our in vivo results show that Auranta 3001 is able to reduce cecum colonization levels from log 8 CFU/ml to log 2 CFU/ml for C. jejuni RC039 and from log 7 CFU/ml to log 2 CFU/ml for C. coli RC013. In conclusion, this novel antimicrobial is able to reduce the pathogenic properties of T6SS campylobacters in vitro and also to decrease colonization in vivo

The pesticide adjuvant, Toximul™, alters hepatic metabolism through effects on downstream targets of PPARα

AbstractPrevious studies demonstrated that chronic dermal exposure to the pesticide adjuvant (surfactant), Toximul™ (Tox), has significant detrimental effects on hepatic lipid metabolism. This study demonstrated that young mice dermally exposed to Tox for 12 days have significant increases in expression of peroxisomal acyl-CoA oxidase (mRNA and protein), bifunctional enzyme (mRNA) and thiolase (mRNA), as well as the P450 oxidizing enzymes Cyp4A10 and Cyp4A14 (mRNA and protein). Tox produced a similar pattern of increases in wild type adult female mice but did not induce these responses in PPARα-null mice. These data support the hypothesis that Tox, a heterogeneous blend of nonionic and anionic surfactants, modulates hepatic metabolism at least in part through activation of PPARα. Notably, all three groups of Tox-treated mice had increased relative liver weights due to significant accumulation of lipid. This could be endogenous in nature and/or a component(s) of Tox or a metabolite thereof. The ability of Tox and other hydrocarbon pollutants to induce fatty liver despite being PPARα agonists indicates a novel consequence of exposure to this class of chemicals, and may provide a new understanding of fatty liver in populations with industrial exposure

Attenuation of Vibrio parahaemolyticus Virulence Factors by a Mixture of Natural Antimicrobials.

Reducing acute mortality in aquatic crustaceans using natural alternatives to antibiotics has become a necessity, firstly for its positive impact on the aquaculture industry and, secondly, because the extensive use of antibiotics may lead to increased levels of drug resistance in pathogenic microorganisms. This study aimed to investigate the effect of a mixture of natural antimicrobials on the in vitro and in vivo virulence abilities of Type VI secretion system (T6SS)-positive Vibrio parahaemolyticus (A3 and D4), strains known as having potentially harmful health consequences for aquatic crustaceans and consumers. Herein, we report that a natural antimicrobial mixture (A3009) was capable of significantly reducing the virulence of V. parahaemolyticus strains A3 and D4 in an in vitro infection model, using the fish cell line CHSE-214, an effect which correlates with the bacterial downregulation of hcp1 and hcp2 gene expression and with the ability of the antimicrobial to efficiently retain low cytotoxic levels (p < 0.001). We show for the first time that a natural antimicrobial is able to significantly reduce the mortality of shrimps in a challenge experiment and is able to significantly attenuate H2O2 release during infection (p < 0.001), indicating that it could harbor positive intestinal redox balance effects

Attenuation of E. coli O157:H7 virulence by a combination of natural plant extracts and organic acids before and after refrigerated storage

Anti-virulence strategies are an alternative approach to combat zoonotic bacterial pathogens. Although control measures are in place against E. coli O157, it still remains a global health concern with cattle being the most important reservoir. Different feed constituents have varying effects on the virulence of E. coli O157 present in the gut which then are transferred to meat processing surfaces and meat during slaughter. This study explored the anti-virulence properties of a mixture of natural plant extracts and organic acids (citrus, grape and oregano extracts, lactic and citric acid) before and after refrigerated storage as means of reducing the risk from E. coli O157 by reducing its virulence. Assessment of the effects of sub-inhibitory concentrations (0.1 and 0.5 %, v/v) of the antimicrobial mixture before refrigerated storage showed that the pathogen’s motility and adhesion onto HCT-8 cells was significantly reduced (P<0.05) in a dose-dependent manner. Shiga-toxin 2 production was also significantly reduced. Real-time PCR analysis revealed that the mixture of natural antimicrobials repressed expression of adhesion (eae) and shiga-toxin 2 (stx2) genes, which was consistent with the observed reduction in adhesion and toxin production. The same virulence factors were investigated after simulated storage of E. coli O157 in meat simulation medium. Results revealed that after exposure to the antimicrobial mixture, the virulence was significantly lower compared to non-treated control after refrigerated storage. The present work shows the potential of the antimicrobial mixture in reducing virulence and thus risk from E. coli O157 by applying it as a feed additive

The effect of natural antimicrobials on the Campylobacter coli T6SS+/- during in vitro infection assays and on their ability to adhere to chicken skin and carcasses.

Reducing the Campylobacter load on poultry carcasses represents a major tasks for the industry as its ability to reduce their presence is of major interest aiming to increase consumer safety. This study investigated the ability of a mixture of natural antimicrobials (A3001) to reduce the adherence of the T6SS+/-C. coli isolates (NC1hcp-, NC2 hcp- and NC3 hcp+) to chicken neck skin and whole carcasses. Overall, the antimicrobial mixture induced a significant reduction in the capability of our C. coli isolates to colonise the chicken skin (p < 0.05) and carcasses (p < 0.0001) but with a greater effect (≈3 log reduction) on the NC3 isolate. Using the HCT-8 in vitro infection model we also show that at a concentration of 0.5% A3001, the impact on the NC3 isolate is accompanied by the downregulation of the hcp gene (p = 0.0001), and indicator of the T6SS presence. The results described herein also indicated that these isolates are highly resistant to H2O2, up to 20 mM, suggesting a high resilience to environmental stresses. In summary our study shows that natural antimicrobials can reduce the ability of T6SS positive chicken C. coli isolates to adhere to chicken skin or to the whole carcass and to infect epithelial cells in vitro and could be considered a potential intervention at processor level

In vitro and in vivo characterisation of Listeria monocytogenes outbreak isolates

Listeriosis is an important food-borne disease responsible for high rates of morbidity and mortality. L. monocytogenes has been the cause of several foodborne outbreaks and its ability to adapt and survive in a wide range of environmental conditions makes eradication difficult. Many L. monocytogenes strains are avirulent but have the ability to increase their virulence if exposed to environmental stresses. The aim of this study was to explain the observed increase in virulence of outbreak L. monocytogenes isolates by using phenotypic assays and whole genome sequencing. Four L. monocytogenes isolates from sweetcorn and one isolate from a raw milk (control) were sequenced and characterised using a range of phenotypic assays. The four L. monocytogenes sweetcorn isolates displayed a significant increase for in vitro adhesion and invasion of epithelial cells compared to the control isolate. They also showed a higher level of colonisation of the liver and spleen in vivo. In addition, the four L. monocytogenes isolates displayed an increased ability to form biofilms, resist heat stress and resist a combination of antimicrobials. Investigation of the genomes of the four L. monocytogenes sweet corn isolates identified Single Nucleotide Polymorphisms (SNPs) in genes, which may have a role in the observed phenotypes characteristic of these strains, particularly in response to survival properties within the environment or in terms of virulence. We highlight the importance of combining whole genomic sequencing with phenotypic characterisation as a key element in the investigation of outbreaks of foodborne pathogens

The in vitro and ex vivo effect of Auranta 3001 in preventing Cryptosporidium hominis and Cryptosporidium parvum infection

Background: Cryptosporidium is a major cause of diarrhea worldwide in both humans and farm animals with no completely effective treatment available at present. In this study, we assessed the inhibitory effect of different concentrations of Auranta 3001 (0.1, 0.5 and 1%), a novel natural feed supplement, on C. hominis and C. parvum invasion of human ileocecal adenocarcinoma (HCT-8), bovine primary cells and C. parvum invasion of HCT-8, bovine primary cells and bovine intestinal biopsies. The effect of the feed supplement on the production of pro-inflammatory cytokines IL-8 and INF-?, the anti-inflammatory cytokine IL-10, the expression of CpSUB1 protease gene during infection was also assessed by quantitative PCR (q-PCR). Transepithelial electrical resistance (TEER) was employed to measure the integrity of tight junction dynamics of the culture models. Results: Pre-treatment of intestinal cells or oocysts with the Auranta 3001 significantly reduced the invasiveness of C. hominis and C. parvum against HCT-8 and bovine primary cells in a dose dependent manner. The most pronounced reduction in the invasiveness of both parasites was observed when Auranta 3001 was present during infection. Levels of IL-8 were significantly reduced in both HCT-8 and bovine primary cells, while the levels of INF-? and IL-10 showed opposite trends in the two cell lines during infection in the presence of Auranta 3001. CpSUB1 gene protease expression, which mediates infection, was significantly reduced suggesting that this enzyme is a possible target of Auranta 3001. Conclusions: Although, C. hominis and C. parvum use different invasion mechanisms to infect cells, the novel feed additive can significantly attenuate the entry of Cryptosporidium in HCT-8 cells, primary bovine cells and bovine intestinal biopsies and thus provide an alternative method to control cryptosporidiosis

Transcription factor MITF and remodeller BRG1 define chromatin organisation at regulatory elements in melanoma cells

Microphthalmia-associated transcription factor (MITF) is the master regulator of the melanocyte lineage. To understand how MITF regulates transcription, we used tandem affinity purification and mass spectrometry to define a comprehensive MITF interactome identifying novel cofactors involved in transcription, DNA replication and repair, and chromatin organisation. We show that MITF interacts with a PBAF chromatin remodelling complex comprising BRG1 and CHD7. BRG1 is essential for melanoma cell proliferation in vitro and for normal melanocyte development in vivo. MITF and SOX10 actively recruit BRG1 to a set of MITF-associated regulatory elements (MAREs) at active enhancers. Combinations of MITF, SOX10, TFAP2A, and YY1 bind between two BRG1-occupied nucleosomes thus defining both a signature of transcription factors essential for the melanocyte lineage and a specific chromatin organisation of the regulatory elements they occupy. BRG1 also regulates the dynamics of MITF genomic occupancy. MITF-BRG1 interplay thus plays an essential role in transcription regulation in melanoma

Global age-sex-specific mortality, life expectancy, and population estimates in 204 countries and territories and 811 subnational locations, 1950–2021, and the impact of the COVID-19 pandemic: a comprehensive demographic analysis for the Global Burden of Disease Study 2021

Background: Estimates of demographic metrics are crucial to assess levels and trends of population health outcomes. The profound impact of the COVID-19 pandemic on populations worldwide has underscored the need for timely estimates to understand this unprecedented event within the context of long-term population health trends. The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 provides new demographic estimates for 204 countries and territories and 811 additional subnational locations from 1950 to 2021, with a particular emphasis on changes in mortality and life expectancy that occurred during the 2020–21 COVID-19 pandemic period. Methods: 22 223 data sources from vital registration, sample registration, surveys, censuses, and other sources were used to estimate mortality, with a subset of these sources used exclusively to estimate excess mortality due to the COVID-19 pandemic. 2026 data sources were used for population estimation. Additional sources were used to estimate migration; the effects of the HIV epidemic; and demographic discontinuities due to conflicts, famines, natural disasters, and pandemics, which are used as inputs for estimating mortality and population. Spatiotemporal Gaussian process regression (ST-GPR) was used to generate under-5 mortality rates, which synthesised 30 763 location-years of vital registration and sample registration data, 1365 surveys and censuses, and 80 other sources. ST-GPR was also used to estimate adult mortality (between ages 15 and 59 years) based on information from 31 642 location-years of vital registration and sample registration data, 355 surveys and censuses, and 24 other sources. Estimates of child and adult mortality rates were then used to generate life tables with a relational model life table system. For countries with large HIV epidemics, life tables were adjusted using independent estimates of HIV-specific mortality generated via an epidemiological analysis of HIV prevalence surveys, antenatal clinic serosurveillance, and other data sources. Excess mortality due to the COVID-19 pandemic in 2020 and 2021 was determined by subtracting observed all-cause mortality (adjusted for late registration and mortality anomalies) from the mortality expected in the absence of the pandemic. Expected mortality was calculated based on historical trends using an ensemble of models. In location-years where all-cause mortality data were unavailable, we estimated excess mortality rates using a regression model with covariates pertaining to the pandemic. Population size was computed using a Bayesian hierarchical cohort component model. Life expectancy was calculated using age-specific mortality rates and standard demographic methods. Uncertainty intervals (UIs) were calculated for every metric using the 25th and 975th ordered values from a 1000-draw posterior distribution. Findings: Global all-cause mortality followed two distinct patterns over the study period: age-standardised mortality rates declined between 1950 and 2019 (a 62·8% [95% UI 60·5–65·1] decline), and increased during the COVID-19 pandemic period (2020–21; 5·1% [0·9–9·6] increase). In contrast with the overall reverse in mortality trends during the pandemic period, child mortality continued to decline, with 4·66 million (3·98–5·50) global deaths in children younger than 5 years in 2021 compared with 5·21 million (4·50–6·01) in 2019. An estimated 131 million (126–137) people died globally from all causes in 2020 and 2021 combined, of which 15·9 million (14·7–17·2) were due to the COVID-19 pandemic (measured by excess mortality, which includes deaths directly due to SARS-CoV-2 infection and those indirectly due to other social, economic, or behavioural changes associated with the pandemic). Excess mortality rates exceeded 150 deaths per 100 000 population during at least one year of the pandemic in 80 countries and territories, whereas 20 nations had a negative excess mortality rate in 2020 or 2021, indicating that all-cause mortality in these countries was lower during the pandemic than expected based on historical trends. Between 1950 and 2021, global life expectancy at birth increased by 22·7 years (20·8–24·8), from 49·0 years (46·7–51·3) to 71·7 years (70·9–72·5). Global life expectancy at birth declined by 1·6 years (1·0–2·2) between 2019 and 2021, reversing historical trends. An increase in life expectancy was only observed in 32 (15·7%) of 204 countries and territories between 2019 and 2021. The global population reached 7·89 billion (7·67–8·13) people in 2021, by which time 56 of 204 countries and territories had peaked and subsequently populations have declined. The largest proportion of population growth between 2020 and 2021 was in sub-Saharan Africa (39·5% [28·4–52·7]) and south Asia (26·3% [9·0–44·7]). From 2000 to 2021, the ratio of the population aged 65 years and older to the population aged younger than 15 years increased in 188 (92·2%) of 204 nations. Interpretation: Global adult mortality rates markedly increased during the COVID-19 pandemic in 2020 and 2021, reversing past decreasing trends, while child mortality rates continued to decline, albeit more slowly than in earlier years. Although COVID-19 had a substantial impact on many demographic indicators during the first 2 years of the pandemic, overall global health progress over the 72 years evaluated has been profound, with considerable improvements in mortality and life expectancy. Additionally, we observed a deceleration of global population growth since 2017, despite steady or increasing growth in lower-income countries, combined with a continued global shift of population age structures towards older ages. These demographic changes will likely present future challenges to health systems, economies, and societies. The comprehensive demographic estimates reported here will enable researchers, policy makers, health practitioners, and other key stakeholders to better understand and address the profound changes that have occurred in the global health landscape following the first 2 years of the COVID-19 pandemic, and longer-term trends beyond the pandemic

Burden of disease scenarios for 204 countries and territories, 2022–2050: a forecasting analysis for the Global Burden of Disease Study 2021

Background: Future trends in disease burden and drivers of health are of great interest to policy makers and the public at large. This information can be used for policy and long-term health investment, planning, and prioritisation. We have expanded and improved upon previous forecasts produced as part of the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) and provide a reference forecast (the most likely future), and alternative scenarios assessing disease burden trajectories if selected sets of risk factors were eliminated from current levels by 2050. Methods: Using forecasts of major drivers of health such as the Socio-demographic Index (SDI; a composite measure of lag-distributed income per capita, mean years of education, and total fertility under 25 years of age) and the full set of risk factor exposures captured by GBD, we provide cause-specific forecasts of mortality, years of life lost (YLLs), years lived with disability (YLDs), and disability-adjusted life-years (DALYs) by age and sex from 2022 to 2050 for 204 countries and territories, 21 GBD regions, seven super-regions, and the world. All analyses were done at the cause-specific level so that only risk factors deemed causal by the GBD comparative risk assessment influenced future trajectories of mortality for each disease. Cause-specific mortality was modelled using mixed-effects models with SDI and time as the main covariates, and the combined impact of causal risk factors as an offset in the model. At the all-cause mortality level, we captured unexplained variation by modelling residuals with an autoregressive integrated moving average model with drift attenuation. These all-cause forecasts constrained the cause-specific forecasts at successively deeper levels of the GBD cause hierarchy using cascading mortality models, thus ensuring a robust estimate of cause-specific mortality. For non-fatal measures (eg, low back pain), incidence and prevalence were forecasted from mixed-effects models with SDI as the main covariate, and YLDs were computed from the resulting prevalence forecasts and average disability weights from GBD. Alternative future scenarios were constructed by replacing appropriate reference trajectories for risk factors with hypothetical trajectories of gradual elimination of risk factor exposure from current levels to 2050. The scenarios were constructed from various sets of risk factors: environmental risks (Safer Environment scenario), risks associated with communicable, maternal, neonatal, and nutritional diseases (CMNNs; Improved Childhood Nutrition and Vaccination scenario), risks associated with major non-communicable diseases (NCDs; Improved Behavioural and Metabolic Risks scenario), and the combined effects of these three scenarios. Using the Shared Socioeconomic Pathways climate scenarios SSP2-4.5 as reference and SSP1-1.9 as an optimistic alternative in the Safer Environment scenario, we accounted for climate change impact on health by using the most recent Intergovernmental Panel on Climate Change temperature forecasts and published trajectories of ambient air pollution for the same two scenarios. Life expectancy and healthy life expectancy were computed using standard methods. The forecasting framework includes computing the age-sex-specific future population for each location and separately for each scenario. 95% uncertainty intervals (UIs) for each individual future estimate were derived from the 2·5th and 97·5th percentiles of distributions generated from propagating 500 draws through the multistage computational pipeline. Findings: In the reference scenario forecast, global and super-regional life expectancy increased from 2022 to 2050, but improvement was at a slower pace than in the three decades preceding the COVID-19 pandemic (beginning in 2020). Gains in future life expectancy were forecasted to be greatest in super-regions with comparatively low life expectancies (such as sub-Saharan Africa) compared with super-regions with higher life expectancies (such as the high-income super-region), leading to a trend towards convergence in life expectancy across locations between now and 2050. At the super-region level, forecasted healthy life expectancy patterns were similar to those of life expectancies. Forecasts for the reference scenario found that health will improve in the coming decades, with all-cause age-standardised DALY rates decreasing in every GBD super-region. The total DALY burden measured in counts, however, will increase in every super-region, largely a function of population ageing and growth. We also forecasted that both DALY counts and age-standardised DALY rates will continue to shift from CMNNs to NCDs, with the most pronounced shifts occurring in sub-Saharan Africa (60·1% [95% UI 56·8–63·1] of DALYs were from CMNNs in 2022 compared with 35·8% [31·0–45·0] in 2050) and south Asia (31·7% [29·2–34·1] to 15·5% [13·7–17·5]). This shift is reflected in the leading global causes of DALYs, with the top four causes in 2050 being ischaemic heart disease, stroke, diabetes, and chronic obstructive pulmonary disease, compared with 2022, with ischaemic heart disease, neonatal disorders, stroke, and lower respiratory infections at the top. The global proportion of DALYs due to YLDs likewise increased from 33·8% (27·4–40·3) to 41·1% (33·9–48·1) from 2022 to 2050, demonstrating an important shift in overall disease burden towards morbidity and away from premature death. The largest shift of this kind was forecasted for sub-Saharan Africa, from 20·1% (15·6–25·3) of DALYs due to YLDs in 2022 to 35·6% (26·5–43·0) in 2050. In the assessment of alternative future scenarios, the combined effects of the scenarios (Safer Environment, Improved Childhood Nutrition and Vaccination, and Improved Behavioural and Metabolic Risks scenarios) demonstrated an important decrease in the global burden of DALYs in 2050 of 15·4% (13·5–17·5) compared with the reference scenario, with decreases across super-regions ranging from 10·4% (9·7–11·3) in the high-income super-region to 23·9% (20·7–27·3) in north Africa and the Middle East. The Safer Environment scenario had its largest decrease in sub-Saharan Africa (5·2% [3·5–6·8]), the Improved Behavioural and Metabolic Risks scenario in north Africa and the Middle East (23·2% [20·2–26·5]), and the Improved Nutrition and Vaccination scenario in sub-Saharan Africa (2·0% [–0·6 to 3·6]). Interpretation: Globally, life expectancy and age-standardised disease burden were forecasted to improve between 2022 and 2050, with the majority of the burden continuing to shift from CMNNs to NCDs. That said, continued progress on reducing the CMNN disease burden will be dependent on maintaining investment in and policy emphasis on CMNN disease prevention and treatment. Mostly due to growth and ageing of populations, the number of deaths and DALYs due to all causes combined will generally increase. By constructing alternative future scenarios wherein certain risk exposures are eliminated by 2050, we have shown that opportunities exist to substantially improve health outcomes in the future through concerted efforts to prevent exposure to well established risk factors and to expand access to key health interventions