Weekly Nowcasting of New COVID-19 Cases Using Past Viral Load Measurements.

The rapid spread of the coronavirus disease COVID-19 has imposed clinical and financial burdens on hospitals and governments attempting to provide patients with medical care and implement disease-controlling policies. The transmissibility of the disease was shown to be correlated with the patient's viral load, which can be measured during testing using the cycle threshold (Ct). Previous models have utilized Ct to forecast the trajectory of the spread, which can provide valuable information to better allocate resources and change policies. However, these models combined other variables specific to medical institutions or came in the form of compartmental models that rely on epidemiological assumptions, all of which could impose prediction uncertainties. In this study, we overcome these limitations using data-driven modeling that utilizes Ct and previous number of cases, two institution-independent variables. We collected three groups of patients (n = 6296, n = 3228, and n = 12,096) from different time periods to train, validate, and independently validate the models. We used three machine learning algorithms and three deep learning algorithms that can model the temporal dynamic behavior of the number of cases. The endpoint was 7-week forward number of cases, and the prediction was evaluated using mean square error (MSE). The sequence-to-sequence model showed the best prediction during validation (MSE = 0.025), while polynomial regression (OLS) and support vector machine regression (SVR) had better performance during independent validation (MSE = 0.1596, and MSE = 0.16754, respectively), which exhibited better generalizability of the latter. The OLS and SVR models were used on a dataset from an external institution and showed promise in predicting COVID-19 incidences across institutions. These models may support clinical and logistic decision-making after prospective validation.

Evaluation of Knowledge, Attitudes, and Practices Related to Osteoporosis and Correlates of Perceived High Risk among People Living in Two Main Districts of Lebanon.

Background: The prevalence of osteoporosis is increasing in Lebanon. Aim: We evaluated the knowledge, attitudes, and practices related to osteoporosis and correlates of its perceived high risk among people living in Beirut and Mount Lebanon districts of Lebanon. Methods: This study is a cross-sectional study which consisted of 376 participants that were selected from the two districts within two or three households after two geographical areas were randomly selected from each stratum classified by education and altitude. They were then asked to fill a KAP survey on osteoporosis and provide information on factors likely related to its perceived high risk. Results: The majority of participants had a low (20.2%) and moderate (65.4%) knowledge of osteoporosis, with a higher knowledge in females than in males. A higher percentage of young people perceived it as a serious health risk than that of older people. In contrast, 85.9% participants reported drinking caffeinated beverages and 51.6% participants reported that they do not exercise. Glucose intolerance due to epigenetic and genetic factors, female sex, and older age were risk factors of a perceived high risk of osteoporosis, while any physical exercise, abstention from caffeine for 48 to 72 hours, and higher education were protective factors. Conclusion: A nationwide KAP study should be conducted; likewise, awareness campaigns should be adopted.

Non-nutritive sweeteners and type 2 diabetes: Should we ring the bell?

Non-nutritive sweeteners (NNS) were thought to be healthy sugar substitutes used instead of sugar for caloric and glycemic control but evidences blaming them for contributing to type 2 diabetes (T2D) are rising. We aim to investigate whether NNS consumption prevents or causes T2D. Articles of all designs conducted on humans were retrieved from three databases in addition to manually reviewed articles. The literature is highly heterogeneous, and conclusions vary with different studies' types and designs. While some studies highlighted the neutral effect of NNS on T2D or reported inconclusive evidences to make their conclusion, others either found NNS culprit for increasing the risk for T2D or reported their protective effect against it. Those results were changing after adjustment for confounders. Due to the inconsistency in the literature, well-designed studies that take into consideration all types of confounders are needed. RESEARCH IN CONTEXT: Evidence before this study: The epidemic of obesity is the result of many factors and causes several chronic diseases where its association with type 2 diabetes is well established. The first line of treatment for obesity is lifestyle changes including physical activity and dietary intervention where non-nutritive sweeteners have received a high attention; those were thought to be healthy sugar substitutes used instead of sugar for caloric and glycemic control but several evidences have blamed them for having a role in the development of type 2 diabetes. In our paper, we aim to investigate whether non-nutritive sweeteners consumption prevents or causes type 2 diabetes. To respond to this question, an extensive search of the literature was conducted between October and December 2018 using the key terms "non-nutritive sweeteners", "artificial sweeteners", "high-intensity sweeteners", "type 2 diabetes" and "prediabetes" on three databases including Pubmed, Science direct and Scopus. Additional search for relevant articles was carried out manually from the reference list of selected articles. Animal studies, studies involving sugar alcohols or those conducted on children, adolescents, pregnant women, or on participants with diabetes were excluded. Human studies conducted from January 2004 to October 2018 were included and divided into observational, interventional, and systematic review and meta-analysis for discussion and analysis Added value of this study: In the literature, the term high-intensity sweeteners has been used interchangeably with non-nutritive sweeteners, artificial sweeteners or low-calorie sweeteners but few articles, if any, clarified the difference between them. In our review, we gathered the different definitions and classifications and summarized them to help the reader understand the difference. Since artificial sweeteners are nowadays widely used and prescribed for caloric and glycemic control, and are unintentionally consumed because they enter in the manufacturing process of thousands of products and due to their potential side effects reported in several studies, we found it interesting to gather, summarize and discuss the available results assessing the role of non-nutritive sweeteners in the development of type 2 diabetes. Those results showed the heterogeneity of the literature and the difficulty in having a firm conclusion; this helps researchers to profit from our study and to conduct well-designed studies leading to firm conclusions and recommendations. Implications of all the available evidence: Despite the absence of strong conclusion that confirms the fact that non-nutritive sweeteners consumption increases the risk for diabetes, no firm conclusion rejects this statement. In result, the existing evidences in addition to our study should ring the bell for clinicians and practitioners who are prescribing those sugar-alternatives as "healthy substitute" to white sugar. Moreover, this encourages the manufacturers to search for a healthy natural alternative to artificial sweeteners to be used in the manufacturing process.

Shisha microbiota: the good, the bad and the not so ugly.

OBJECTIVE: Over the last decade, there has been a rapid expansion of the trendy water pipe smoking around the world especially among younger adults. The initial objective of this study was to identify the microbiota of the shisha, which may either be of no harm for the smoker or enhance the threat on his well-being. The total DNA for the metagenomics study was conducted on three different shishas from three different delivery shops in Jounieh, Lebanon. The microbiota in two solid parts of the shisha, shaft and hose, were analysed including the fresh tobacco and the water in the bowl. All samples were analysed using high-throughput sequencing of 16S rRNA gene amplicons. RESULTS: Overall, more than 40 bacterial genera were found in the three investigated shishas, some are commensal others are pathogenic. All three shishas showed similar microbial content regarding the bacteria inhabiting in water, shaft, or hose. From the results of this study it appears that a very large quantity of bacteria was found in the water pipes, some are harmful and others beneficial. We assume that the presence of gut dependent microbiota is related to the loose hygienic conditions in which the shisha is prepared.

[Quantitative PCR (qPCR) and the guide to good practices MIQE: adapting and relevance in the clinical biology context]. / La PCR quantitative (qPCR) et le guide de bonnes pratiques MIQE : adaptation et pertinence dans le contexte de la biologie clinique.

The qPCR has been introduced in clinical and biomedical research for over 10 years from now. Its use in trials and diagnostics is continuously increasing. Due to this heavy use, the question of relyability and relevance of qPCR results has to be asked. This review proposes a documented and evidence based answer to this question, thanks to the MIQE (minimum information for publication of quantitative real-time PCR experiments) guideline. The whole analysis process is addressed, from nucleic acids extraction to data management. Simple answers are given, taking into account the technical constraints from clinical research in order to allow a realistic application of this guideline.

Passaging impact of H9N2 avian influenza virus in hamsters on its pathogenicity and genetic variability.

INTRODUCTION: Avian influenza viruses of the H9N2 subtype have been reported to cause human infections. This study demonstrates the impact of nasal viral passaging of avian H9N2 in hamsters on its cross species-pathogenic adaptability and variability of amino acid sequences of the hemagglutinin (HA) and neuraminidase (NA) stalk. METHODOLOGY: Three intranasal passagings of avian H9N2 in hamsters P1, P2, and P3 were accomplished. Morbidity signs and lesions were observed three days post viral inoculation. The HA test was used for presumptive detection of H9N2 virus in the trachea and lungs of the hamsters challenged with the differently passaged viruses. Different primers were used for PCR amplification of the HA1 and NA stalk regions of the differently passaged H9N2 viruses, followed by sequence alignment. RESULTS: The morbidity signs indicated low pathogenicity of the differently passaged H9N2 viruses in hamsters. The frequency of gross and microscopic lesions in the tracheas and lungs were insignificantly different among hamsters challenged with the differently passaged H9N2 viruses (p > 0.05). There was 100% similarity in the amino acid sequence of the HA gene of most passaged viruses. The amino acid sequence of the neuraminidase in the third passaged H9N2 virus recovered from lungs showed a R46P mutation that might have a role in the pathogenic adaptability of P3 viruses in hamsters' lungs. CONCLUSIONS: The apparent adaptation of avian H9N2 virus to mammalian cells is in agreement with the World Health Organization's alertness for a possible public health threat by this adaptable virus.

Five years MIQE guidelines: the case of the Arabian countries.

The quantitative real time polymerase chain reaction (qPCR) has become a key molecular enabling technology with an immense range of research, clinical, forensic as well as diagnostic applications. Its relatively moderate instrumentation and reagent requirements have led to its adoption by numerous laboratories, including those located in the Arabian world, where qPCR, which targets DNA, and reverse transcription qPCR (RT-qPCR), which targets RNA, are widely used for region-specific biotechnology, agricultural and human genetic studies. However, it has become increasingly apparent that there are significant problems with both the quality of qPCR-based data as well as the transparency of reporting. This realisation led to the publication of the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines in 2009 and their more widespread adoption in the last couple of years. An analysis of the performance of biomedical research in the Arabian world between 2001-2005 suggests that the Arabian world is producing fewer biomedical publications of lower quality than other Middle Eastern countries. Hence we have analysed specifically the quality of RT-qPCR-based peer-reviewed papers published since 2009 from Arabian researchers using a bespoke iOS/Android app developed by one of the authors. Our results show that compliance with 15 essential MIQE criteria was low (median of 40%, range 0-93%) and few details on RNA quality controls (22% compliance), assays design (12%), RT strategies (32%), amplification efficiencies (30%) and the normalisation process (3%). These data indicate that one of the reasons for the poor performance of Arabian world biomedical research may be the low standard of any supporting qPCR experiments and identify which aspects of qPCR experiments require significant improvements.

Impact of embryonic passaging of H9N2 virus on pathogenicity and stability of HA1- amino acid sequence cleavage site.

BACKGROUND: Highly virulent Avian Influenza viruses might arise from avirulent strains following viral passaging. This work aims at studying the impact of embryonic passaging of H9N2 on the stability of the HA1 amino acid sequence and its relatedness to pathogenicity. MATERIAL/METHODS: The original H9N2 virus was propagated for 3 consecutive passages in embryonated chicken eggs. Pathogenicity and amino acids sequences at the HA1 gene level of the original (P0), and the once (P1), twice (P2), and three times (P3) passaged viruses were compared. RESULTS: The percent mortality significantly increased in embryos inoculated with P2 (86.7%) and P3 of H9N2 (100%) in comparison to P0 (0.0%) and P1 of H9N2 (46.1%) (P<0.05), while the density of propagated H9N2 declined with passaging. The R-S-S-R motif was stable at the HA1 cleavage site of P0, P1, P2, and P3 viruses. The similarity in the HA1 sequences among the differently passaged viruses ranged between 93.2 to 100%. CONCLUSIONS: The pathogenicity increased significantly upon passaging in chicken embryos in spite of the presence of the same motif at the HA1 cleavage site. Further investigations will target the study of changes in the whole HA protein and of Neuraminidases that could be responsible for a higher pathogenicity.

Simultaneous determination of genomic DNA methylation and uracil misincorporation.

OBJECTIVE: To develop a method for the simultaneous measurement of 5-methylcytosine (5-metC) and 2'-deoxyuridine monophosphate (dU). MATERIALS AND METHODS: Genomic DNA was extracted from the HepG2 cell line grown in experimental complete medium or in folate-depleted medium. Samples were treated with RNAse A and RNAse T1 to avoid any RNA contamination. High-performance liquid chromatography (HPLC)/electrospray ionization mass spectrometric (ESI-MS) method was used to separate nucleotides after enzymatic hydrolysis of DNA with nuclease P1, phosphodiesterase I and alkaline phosphatase. RESULTS: Using this sensitive new methodology, we were able to quantify simultaneously the concentration of DNA-5-metC and DNA-uracil in DNA. The linear correlation coefficient (R(2)) between the MS signal and the concentration of 5-metC in a range of 0.5-5 microM or dU in a range of 10-100 microM was 0.9954 and 0.9999, respectively. The coefficient of variation was 16.94 and 14.77%, respectively. The applicability of this assay is demonstrated by detection of a decrease in 5-metC% and elevation of dU/thymidylate (dT) into genomic DNA extracted from the HepG2 cell line grown in a folate-depleted medium. CONCLUSION: Our results confirm that the HPLC/ESI-MS method reported earlier for measuring 5-metC allows measurement of uracil misincorporation into DNA.

Standardisation of a new model of H9N2/Escherichia coli challenge in broilers in the Lebanon.

Primary infection by low pathogenic avian influenza (LPAI) predisposes for secondary infection by Escherichia coli in poultry, leading to significant economic losses. Future research in control of this ailment requires the establishment of a successful controlled challenge by avian influenza virus (AIV)/E. coli. Six groups of broilers (6 birds/group) were included for the standardisation of the controlled challenge by AIV/E. coli. Birds in groups 1, 2, 3, 4 and 5 received an intra-tracheal challenge of 0.5 ml of two haemagglutinating units of H9N2 virus at 20 days of age. At the age of 23 days, birds in group 1 received an intra-thoracic (right air sac)-E. coli challenge equivalent to 1.6 x 10 colony-forming units (cfu)/0.5 ml/bird, while birds in groups 2, 3, 4 and 5 received E. coli by the same route and in the following respective decreasing order of viable cells: 1.6 x 10(6), 1.6 x 10(5), 1.6 x 10(4) and 1.6 x 10(3); cfu. Birds in control group 6 were deprived of H9N2 and E. coli challenge. Results showed significant early mortality in group 1 that was challenged with the highest number of E. coli, in comparison to groups 2-6 (p<0.05); however, the average weight at 28 days of age was similar in surviving birds of groups 2-6 (p>0.05). The frequencies of four signs at 2 days and at 5 days post E. coli challenge (conjunctivitis, diarrhoea, ocular exudates and rales) in the surviving birds of groups 2-5 were most often higher than those observed in control group 6 (p<0.05). These four signs and five gross lesions (abdominal airsacculitis, left thoracic airsacculitis, pericarditis, right thoracic airsacculitis and tracheitis) had a decreasing pattern of frequency related to a decrease in the E. coli count used in the challenge.

Comparison of the level of organic contaminants in bile and muscle of Mugil spp. following a major oil spill in the eastern Mediterranean Sea.

The total polyaromatic hydrocarbons (TPAH) and the total polychlorinated biphenyls (TPCB) that originate from oil spills in sea and ocean waters are toxic to fish and their offspring. The authors compare the levels of organic contaminants (TPAH and TPCB) recovered from the bile versus the dorsal muscles of 120 individual Mugil spp. that were harvested from six sites in the eastern Mediterranean following a significant heavy oil spill. Results showed an insignificant difference between the mean of means of TPAH and TPCB (six means of individual Mugil spp. from six respective sites) in bile versus dorsal muscle. In addition, the correlation equation between the level of bile TPAH and the level of bile carcinogenic polyaromatic hydrocarbons (cPAH) was established. This data suggests the possibility of substituting the analysis of organic contaminants in muscles by using the liquid bile of Mugil spp., thus eliminating the time-consuming steps of lyophilisation and homogenisation of muscle. In addition, the bile cPAH could be predicted from the bile TPAH by a regression relationship.

Folate receptor and human reduced folate carrier expression in HepG2 cell line exposed to fumonisin B1 and folate deficiency.

Fumonisin B1 (FB1) induces apoptosis and decreases the cellular uptake of 5-methyltetrahydrofolate. Two folate transporters (folate receptor, FR, and Reduced Folate Carrier, hRFC1) are involved in the cell uptake of folate. We aimed to study whether FB1 modifies the expression of the FR and the hRFC1 and whether its apoptotic effect is influenced by folate. Incubation of HepG2 cells with FB1 induced apoptosis in concentration and time-dependent manner in complete medium (experimental control medium, ECM), as well as in folate-depleted medium (FDM). FDM increased the toxicity of FB1 as the cells developed apoptosis within 24 h at 1 microM of FB1 instead of 100 microM in ECM. Whereas FR protein expression in cells grown in ECM was significantly inhibited after apoptosis event, protein expression of the hRFC1 was rather increased. The hrfc1 transcription was decreased in the treated cells. Under folate-deficient conditions, dramatic changes were observed on both transcriptional and post-transcriptional expression of the two transporters. FDM alone reduced FR protein expression by 12 +/- 2% and 43 +/- 1% at 48 and 72 h, respectively. The 5-methytetrahydrofolate attenuates apoptosis in a greater extent than the folic acid. However, its effects in preventing decrease of both folate transporters have not been observed. In conclusion, this study shows that the changes in the expression of FR after FB1 addition are probably a consequence of the FB1 toxicity. The response to FB1 by HepG2 cell lines is influenced by folate status and by folate form. 5-methyltetrahydrofolate appears to be more effective in preventing apoptosis than folic acid.

Holistic efficacy of specific nutrient synergy against avian flu virus: pathology and immunomodulation.

The authors evaluated the holistic efficacy of nine specific nutrient synergy (NS) against avian influenza virus (AIV) strain Lebanon 1 (H9N2). The study included two segments; the first was designed to determine the minimum dose among four doses (1X, 2X, 3X and 4X in which X = 24.4 mg/ml/bird) of NS, administered intraoesophageally, once per day between 7 and 14 days of age, resulting in an improvement of chicken performance without any toxic side-effects; the second aimed at reducing pathological effects and inducing immunomodulation by the determined safe dose of NS in chickens exposed to AIV. The first segment showed that the daily oral administration of the NS to birds between 7 to 14 days of age at the 2X dose-level (320 mg/kg body weight or 48.8 mg/ml/bird) resulted in a consistent and significant improvement in the feed conversion (P<0.05) at 10 and 14 days of age, associated with a significant (P<0.05) increase in the liver weight index. In addition, the administration of NS at 2X level resulted in complete absence of toxicity signs (swollen infraorbital sinuses, ocular exudate, nasal discharge, thick oral saliva, diarrhoea, lameness and huddling) and complete absence of toxicity lesions (airsacculitis, hydropericardium signs, pericarditis, perihepatitis, splenomegaly and tracheitis). The four groups of birds that received levels 1X to 4X levels had significantly higher frequency of birds with gaseous caeca compared to the control group deprived of NS (P<0.05), a sign of higher fermentation activity in this organ. Data from the second segment of this research showed that the daily administration of NS at a level of 48.8 mg/ml/bird, between 7 to 14 days of age, to H9N2-challenged birds reduced specific pathological effects at 14 days of age namely: absence of rales at 3 days post H9N2 challenge and gross lesions (absence of tracheitis and enteritis at 7 days post challenge). Such reductions in signs and gross lesions were associated with a 63.4% reduction in immune responses to the hemagglutinin protein of the AIV, an indication that NS has a reducing effect on the viral infectivity in chickens.