Early detection of gray mold in grape using conventional and molecular methods

Botrytis cinerea affects grape quality and yield, and can be difficult to manage due in part to non-symptomatic, quiescent infection in berry development. The aim of this study was to develop a dual system for the detection, isolation and quantification of B. cinerea. After three days of samples replication on the modified selective medium (mKERS), the results showed a significant infection effect on the majority of inflorescence samples, especially on the small berries which demonstrated Botrytis infection in all tested plants and appeared to be highly susceptible to Botrytis infection prior to harvest. Moreover, infection variation was determined in almost all inflorescence samples taken from different plants. The real-time PCR assay was used to determine the DNA quantity of B. cinerea in each sample tested. A linear relationship was found in these two systems, conventional and molecular assays, to demonstrate the infection of different samples with B. cinerea. Although, the real-time PCR assay was highly expensive, it appeared to be more rapid and sensitive than the conventional selective medium assay, allowing both detection and quantification of B. cinerea within 3 h. However, conventional assay has an advantage of both detection and isolation of viable cells of B. cinerea, which resulted in making a wide collection of different isolates. Furthermore, this conventional assay is cheaper than molecular test, especially when we carry out a routine work. This dual method proved to be selective and sensitive assays and should be used to monitor Botrytis infection in the field.Key words: Botrytis cinerea, inflorescence infection, latent/quiescent infection, real time polymerase chain reaction/real-time quantitative PCR (PCR/qPCR)

Pseudomonas aeruginosa - Modified Hodge Test (PAE-MHT) and ChromID Carba Agar for Detection of Carbapenemase Producing Pseudomonas Aeruginosa Recovered from Clinical Specimens

AIMS: This study aims to evaluate the ability of ChromID Carba agar, and Pseudomonas aeruginosa modified Hodge test (PAE-MHT) for detection of carbapenemase-producing P. aeruginosa and to determine the associated carbapenemase gene classes by PCR. METHODS: One hundred Carbapenem-resistant P. aeruginosa (CRPA) isolates were tested for: i) carbapenemases production by ChromID carba agar, Modified Hodge test (MHT) and (PAE-MHT) and ii) detection of some carbapenemase genes by PCR. RESULTS: All (100%) of the isolates showed growth on ChromID Carba agar with 100% sensitivity. Using MHT, 54% of isolates were positive, 3% were indeterminate, and 43% were negative, demonstrating 58.9% sensitivity and 80% specificity. On performing PAE-MHT, 91% of the strains were positive, 3% were intermediate, and 6% were negative, demonstrating 97.9% sensitivity and 80% specificity. The most prevalent gene was blaKPC (81%), followed by blaVIM (74%); blaIMP was detected in only one isolate, and blaOXA-48 in 34% of the isolates. CONCLUSIONS: We conclude that PAE-MHT and ChromID Carba are sensitive, specific, simple and cost-effective screening tests for detection of CRPA isolates compared to the traditional MHT

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

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Uterine artery Doppler and subendometrial blood flow in patients with unexplained recurrent miscarriage

Objective: To evaluate uterine artery blood flow and subendometrial blood flow in women with unexplained recurrent miscarriage (RM) compared to normal fertile women. Design: Case control study. Settings: Departments of Obstetrics and Gynaecology, Cairo university and, Beni Souif University, Cairo, Egypt. Patients and methods: Women were divided into two groups: those with history of unexplained recurrent miscarriage (study group, no. = 40), and those with no history of miscarriage and at least one child born at term (control group, no. = 40), transvaginal 3D power Doppler ultrasonography was performed to all patients in the second phase of non-pregnant unstimulated cycle to detect uterine artery pulsatility index (UAPI) and subendometrial area to detect subendometrial blood flow presented by the indices vascularisation index (VI), flow index (FI), and vascular flow index (VFI). The indices between the two groups were compared. Results: UAPI was significantly higher in the study group (p = 0.000), on the other hand VI (p = 0.000), FI (p = 0.000), VFI (p = 0.000) were significantly lower in the study group. A significant positive correlation was found between UAPI and recurrent miscarriage in the study group (r = 0.8, p = 0.00), whereas a significant negative correlation was found between VI (r = −0.56, p = 0.00), and FI (r = −0.79, p = 0.00) and recurrent miscarriage in the study group, but no significant correlation was found between endometrial thickness and endometrial volume between the two groups. However the application of binary logistic regression analysis model found that adding variables has not significantly increased the ability to predict the occurrence of miscarriage. Conclusion: The presence of good uterine and endometrial blood flow is an important prerequisite for successful implantation and continuation of pregnancy as shown by higher uterine artery blood flow resistance and lower endometrial blood flow in recurrent miscarriage cases and those patients with unexplained RPL may have abnormalities in the uterine and endometrial blood flow. Despite these findings we could not find any cut off values that could predict the occurrence of miscarriage which may be attributed to small sample size and short period of follow up so larger prospective studies are needed aiming to confirm such results and reaching values that can accurately predict the occurrence of miscarriage

The catalytic and kinetic characterization of Bacillus subtilis MK775302 milk clotting enzyme: comparison with calf rennet as a coagulant in white soft cheese manufacture

Abstract Background Calf rennet is considered the traditional source of milk clotting enzyme (MCE). However, increasing cheese consumption with decreasing the calf rennet supply had encouraged the quest for new rennet alternatives. The purpose of this study is to acquire more information about the catalytic and kinetic properties of partially purified Bacillus subtilis MK775302 MCE and to assess the role of enzyme in cheese manufacture. Results B. subtilis MK775302 MCE was partially purified by 50% acetone precipitation with 5.6-fold purification. The optimum temperature and pH of the partially purified MCE were 70 °C and 5.0, respectively. The activation energy was calculated as 47.7 kJ/mol. The calculated Km and Vmax values were 36 mg/ml and 833 U/ml, respectively. The enzyme retained full activity at NaCl concentration of 2%. Compared to the commercial calf rennet, the ultra-filtrated white soft cheese produced from the partially purified B. subtilis MK775302 MCE exhibited higher total acidity, higher volatile fatty acids, and improved sensorial properties. Conclusions The partially purified MCE obtained in this study is a promising milk coagulant that can replace calf rennet at a commercial scale to produce better-quality cheese with improved texture and flavor