Characterization of vitamin D3 biotransformation by the cell lysate of Actinomyces hyovaginalis CCASU-A11-2.

A former work conducted in our Lab, lead to in a effective scale up of vitamin D3 bioconversion into calcitriol by Actinomyces (A.) hyovaginalis isolate CCASU-A11-2 in Lab fermenter (14 L) resulting in 32.8 µg/100 mL of calcitriol. However, the time needed for such a bioconversion process was up to 5 days. Therefore, the objective of this study was to shorten the bioconversion time by using cell-free lysate and studying different factors influencing bioconversion. The crude cell lysate was prepared, freeze-dried, and primarily fractionated into nine fractions, of which, only three fractions, 50, 100, and 150 mM NaCl elution buffers showed 22, 12, and 2 µg/10 mL, calcitriol production, respectively. Ammonium sulfate was used for protein precipitation, and it did not affect the bioconversion process except at a concentration of 10%w/v. Secondary fractionation was carried out using 80 mL of the 50 mM NaCl elution buffer and the results showed the 80 mL eluent volume was enough for the complete elution of the active protein. The pH 7.8, temperature 28 °C, and 6 h reaction time were optimum for maximum calcitriol production (31 µg/10 mL). In conclusion, the transformation of vitamin D3 into calcitriol was successfully carried out within 6 h and at pH 7.8 and 28 °C using fractionated cell lysate. This process resulted in a 10-fold increase in calcitriol as compared to that produced in our previous study using a 14 L fermenter (32.8 µg/100 mL). Therefore, cell-free lysate should be considered for industrial and scaling up vitamin D3 bioconversion into calcitriol.

Bioconversion of vitamin D3 into calcitriol by Actinomyces hyovaginalis isolate CCASU- A11-2.

Vitamin D3 is a fat-soluble prohormone that is activated inside the liver to produce 25-hydroxyvitamin D3 (calcidiol), and in the kidney to produce the fully active 1α, 25-dihydroxy vitamin D3 (calcitriol). A previous work piloted in our laboratory, resulted in a successful recovery of a local soil-promising Actinomyces hyovaginalis isolate CCASU-A11-2 capable of converting vitamin D3 into calcitriol. Despite the rising amount of research on vitamin D3 bioconversion into calcitriol, further deliberate studies on this topic can significantly contribute to the improvement of such a bioconversion process. Therefore, this work aimed to improve the bioconversion process, using the study isolate, in a 14 L laboratory fermenter (4 L fermentation medium composed of fructose (15 g/L), defatted soybean (15 g/L), NaCl (5 g/L), CaCO3 2 g/L); K2HPO4, (1 g/L) NaF (0.5 g/L) and initial of pH 7.8) where different experiments were undertaken to investigate the effect of different culture conditions on the bioconversion process. Using the 14 L laboratory fermenter, the calcitriol production was increased by about 2.5-fold (32.8 µg/100 mL) to that obtained in the shake flask (12.4 µg/100 mL). The optimal bioconversion conditions were inoculum size of 2% v/v, agitation rate of 200 rpm, aeration rate of 1 vvm, initial pH of 7.8 (uncontrolled); addition of vitamin D3 (substrate) 48 h after the start of the main culture. In conclusion, the bioconversion of vitamin D3 into calcitriol in a laboratory fermenter showed a 2.5-fold increase as compared to the shake flask level where, the important factors influencing the bioconversion process were the aeration rate, inoculum size, the timing of substrate addition, and the fixed pH of the fermentation medium. So, those factors should be critically considered for the scaling-up of the biotransformation process.

Phytoestrogen ß-Sitosterol Exhibits Potent In Vitro Antiviral Activity against Influenza A Viruses.

Influenza is a contagious infection in humans that is caused frequently by low pathogenic seasonal influenza viruses and occasionally by pathogenic avian influenza viruses (AIV) of H5, H7, and H9 subtypes. Recently, the clinical sector in poultry and humans has been confronted with many challenges, including the limited number of antiviral drugs and the rapid evolution of drug-resistant variants. Herein, the anti-influenza activities of various plant-derived phytochemicals were investigated against highly pathogenic avian influenza A/H5N1 virus (HPAIV H5N1) and seasonal low pathogenic human influenza A/H1N1 virus (LPHIV H1N1). Out of the 22 tested phytochemicals, the steroid compounds ß-sitosterol and ß-sitosterol-O-glucoside have very potent activity against the predefined influenza A viruses (IAV). Both steroids could induce such activity by affecting multiple stages during IAV replication cycles, including viral adsorption and replication with a major and significant impact on the virus directly in a cell-free status "viricidal effect". On a molecular level, several molecular docking studies suggested that ß-sitosterol and ß-sitosterol-O-glucoside exhibited viricidal effects through blocking active binding sites of the hemagglutinin surface protein, as well as showing inhibitory effects against replication through the binding with influenza neuraminidase activity and blocking the active sites of the M2 proton channel activity. The phytoestrogen ß-sitosterol has structural similarity with the active form of the female sex hormone estradiol, and this similarity is likely one of the molecular determinants that enables the phytoestrogen ß-sitosterol and its derivative to control IAV infection in vitro. This promising anti-influenza activity of ß-sitosterol and its O-glycoside derivative, according to both in vitro and cheminformatics studies, recommend both phytochemicals for further studies going through preclinical and clinical phases as efficient anti-influenza drug candidates.

Characterization and anti-biofilm activity of bacteriophages against urinary tract Enterococcus faecalis isolates.

Strong biofilm-forming Enterococcus feacalis urinary tract pathogens (n = 35) were used to determine the lytic spectrum of six bacteriophages isolated from sewage samples. Only 17 Enterococcus feacalis isolates gave lytic zones with the tested bacteriophages from which five isolates were susceptible to all of them. The isolated enterococcal phages are characterized by wide range of thermal (30-90 °C) and pH (3-10) stability. They belong to order Caudovirales, from which four bacteriophages (EPA, EPB, EPD, EPF) belong to family Myoviridae and two (EPC, EPE) belong to family Siphoviridae. In addition, they have promising antibiofilm activity against the tested strong-forming biofilm E. faecalis isolates. The enterococcal phages reduced the formed and preformed biofilms to a range of 38.02-45.7% and 71.0-80.0%, respectively, as compared to the control. The same promising activities were obtained on studying the anti-adherent effect of the tested bacteriophages on the adherence of bacterial cells to the surface of urinary catheter segments. They reduced the number of adherent cells to a range of 30.8-43.8% and eradicated the pre-adherent cells to a range of 48.2-71.1%, as compared to the control. Overall, the obtained promising antibiofilm activity makes these phages good candidates for application in preventing and treating biofilm associated Enterococcus faecalis infections.

A new grading system for female sexual dysfunction based on the female sexual function index in Egyptian women: a cross-sectional study.

OBJECTIVE: To provide a grading system that accurately reflects the grades of female sexual dysfunction (FSD) severity. PATIENTS AND METHODS: A cross-sectional study was conducted in Assiut University Hospital. It included 500 women who answered the Arabic version of the Female Sexual Function Index (FSFI) after getting their consent. A gradient of FSD severity was created, classifying FSD into five grades: severe, moderate, mild to moderate, mild, and no FSD. RESULTS: According to our grading system, FSD was detected in 339 women (67.8 %); Mild FSD in 20.4%, mild to moderate in 41.6%, moderate in 15.3%, and severe in 22.7%. Mean scores of desire show a linear trend of reduction from 3.8 in mild to 3.36 in mild to moderate to 2.25 in moderate and markedly reduced to 2.1 in severe grade. This difference was highly statistically significant (p= 0.002). The same was reported in arousal, orgasm, and satisfaction domains, while in lubrication and pain domains, the difference was not statistically significant. CONCLUSION: In this study, our grading system was complementary to the FSFI. Moreover, it seems to be more practical and useful in grading the severity of FSD.

Serotyping and Antimicrobial Resistance Profile of Enteric Nontyphoidal Salmonella Recovered from Febrile Neutropenic Patients and Poultry in Egypt.

A total of 300 human fecal samples were collected from febrile neutropenic patients suffering from severe gastroenteritis, followed by identification and serological characterization of recovered isolates. Fifty nontyphoidal Salmonella (NTS) serovars were recovered. A total of serologically identified 50 NTS serovars recovered from poultry of the same geographical area and during the same period as well as one standard strain S. Poona were supplied by the Bacterial Bank of Animal Health Research Institute of Egypt. Antibiogram analysis revealed that the human and poultry serovars exhibited similar antimicrobial resistance patterns against 28 different antimicrobial agents, particularly against ampicillin, cefotaxime, oxytetracycline, and erythromycin. Plasmids harboring blaCTX-m, blaSHV, blaTEM, and aac(6')-Ib were detected in 11 (22%) and 8 (16%) of human and poultry serovars, respectively. Molecular detection of the most clinically relevant virulence genes and analysis of the associated virulence genotypes proved that the human (n = 11) and poultry serovars (n = 12) shared 11 genotypes. Enterobacterial repetitive intergenic consensus PCR analysis revealed that human and poultry serovars were clustered together in 3 out of the 4 clusters with a similarity index ranged from 0.15 to 1. Since poultry are usually consumed by humans, the presence of resistant bacteria harboring transmissible genetic elements is of great health concern.

Better outcome of COVID-19 positive kidney transplant recipients during the unremitting stage with optimized anticoagulation and immunosuppression.

INTRODUCTION: COVID-19 is an ongoing pandemic with high morbidity and mortality and with a reported high risk of severe disease in kidney transplant recipients (KTR). AIM: We aimed to report the largest number of COVID-19-positive cases in KTR in a single center and to discuss their demographics, management, and evolution. METHODS: We enrolled all the two thousand KTR followed up in our center in Kuwait and collected the data of all COVID-19-positive KTR (104) from the start of the outbreak till the end of July 2020 and have reported the clinical features, management details, and both patient and graft outcomes. RESULTS: Out of the one hundred and four cases reported, most of them were males aged 49.3 ± 14.7 years. Eighty-two of them needed hospitalization, of which thirty-one were managed in the intensive care unit (ICU). Main comorbidities among these patients were hypertension in 64.4%, diabetes in 51%, and ischemic heart disease in 20.2%. Management strategies included anticoagulation in 56.7%, withdrawal of antimetabolites in 54.8%, calcineurin inhibitor (CNI) withdrawal in 33.7%, the addition of antibiotics in 57.7%, Tocilizumab in 8.7%, and antivirals in 16.3%. During a follow-up of 30 days, the reported number of acute kidney injury (AKI) was 28.7%, respiratory failure requiring oxygen therapy 46.2%, and overall mortality rate was 10.6% with hospital mortality of 13.4% including an ICU mortality rate of 35.5%. CONCLUSION: Better outcome of COVID-19-positive KTR in our cohort during this unremitting stage could be due to the younger age of patients and early optimized management of anticoagulation, modification of immunosuppression, and prompt treatment of secondary bacterial infections. Mild cases can successfully be managed at home without any change in immunosuppression.

Effect of female genital mutilation/cutting; types I and II on sexual function: case-controlled study.

BACKGROUND: The existing literature is contradictory regarding effects of female genital mutilation/cutting (FGM/C) on sexual functions. The aim of this study was to explore the impact of type I and II FGM/C on sexual function of Egyptian women. METHODS: We recruited 197 cut women and 197 control women from those visiting Assiut University hospitals for different reasons. We asked each woman to fill the Arabic female sexual function index (FSFI) (a self reported 19-item questionnaire assessing the main domains of female sexual function). Genital Examination was done to confirm the type of FGM. RESULTS: Female sexual dysfunction (FSD) was found in 83.8% of FGM/C cases in contrast to 64.5% of the control. The total FSFI score in the FGM/C group (19.82 ± 7.1) was significantly lower than in the control group (23.34 ± 8.1). Concerning the types of FGM/C, type 73.6% of cases had type I and 26.4% had type II. Type I FGM/C was performed mainly by physicians (62.1%) while type II was performed mainly by midwives (44.4%). FSD was found in 83.4% of FGM/C I cases and in 84.6% of FGM/C II cases. There was no statistically significant difference between the two types of FGM/C as regards total and individual domain scores except for the pain domain. There were significantly lower total and individual domain scores in both FGM/C types except for the desire domain compared to control. CONCLUSION: In this study, FGM/C was associated with reduced scores of FSFI on all domains scores, and among both types I and II, both were associated with sexual dysfunction.