Search results for: viral polymerase

Authors: Christi Jackson, Chuka Onaga

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Introduction: We report a case of delayed diagnosis of extra-pulmonary INH-mono-resistant Tuberculosis (TB) in a South African patient with drug-resistant HIV. Case Presentation: A 36-year old male was initiated on 1st line (NNRTI-based) anti-retroviral therapy (ART) in September 2009 and switched to 2nd line (PI-based) ART in 2011, according to local guidelines. He was following up at the outpatient wellness unit of a public hospital, where he was diagnosed with Protease Inhibitor resistant HIV in March 2016. He had an HIV viral load (HIVVL) of 737000 copies/mL, CD4-count of 10 cells/µL and presented with complaints of productive cough, weight loss, chronic diarrhoea and a septic buttock wound. Several investigations were done on sputum, stool and pus samples but all were negative for TB. The patient was treated with antibiotics and the cough and the buttock wound improved. He was subsequently started on a 3rd-line ART regimen of Darunavir, Ritonavir, Etravirine, Raltegravir, Tenofovir and Emtricitabine in May 2016. He continued losing weight, became too weak to stand unsupported and started complaining of abdominal pain. Further investigations were done in September 2016, including a urine specimen for Line Probe Assay (LPA), which showed M. tuberculosis sensitive to Rifampicin but resistant to INH. A lymph node biopsy also showed histological confirmation of TB. Management and outcome: He was started on Rifabutin, Pyrazinamide and Ethambutol in September 2016, and Etravirine was discontinued. After 6 months on ART and 2 months on TB treatment, his HIVVL had dropped to 286 copies/mL, CD4 improved to 179 cells/µL and he showed clinical improvement. Pharmacy supply of his individualised drugs was unreliable and presented some challenges to continuity of treatment. He successfully completed his treatment in June 2017 while still maintaining virological suppression. Discussion: Several laboratory-related factors delayed the diagnosis of TB, including the unavailability of urine-lipoarabinomannan (LAM) and urine-GeneXpert (GXP) tests at this facility. Once the diagnosis was made, it presented a treatment dilemma due to the expected drug-drug interactions between his 3rd-line ART regimen and his INH-resistant TB regimen, and specialist input was required. Conclusion: TB is more difficult to diagnose in patients with severe immunosuppression, therefore additional tests like urine-LAM and urine-GXP can be helpful in expediting the diagnosis in these cases. Patients with non-standard drug regimens should always be discussed with a specialist in order to avoid potentially harmful drug-drug interactions.

Keywords: drug-resistance, HIV, line probe assay, tuberculosis

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Authors: Z. Yousefniayejahr, S. Bolognini, A. Bonini, C. Campobasso, N. Poma, F. Vivaldi, M. Di Luca, A. Tavanti, F. Di Francesco

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Pathogenic bacteria represent a threat to healthcare systems and the food industry because their rapid detection remains challenging. Electrochemical biosensors are gaining prominence as a novel technology for the detection of pathogens due to intrinsic features such as low cost, rapid response time, and portability, which make them a valuable alternative to traditional methodologies. These sensors use biorecognition elements that are crucial for the identification of specific bacteria. In this context, bacteriophages are promising tools for their inherent high selectivity towards bacterial hosts, which is of fundamental importance when detecting bacterial pathogens in complex biological samples. In this study, we present the development of a low-cost and portable sensor based on the Zeno phage for the rapid detection of Staphylococcus aureus. Screen-printed gold electrodes functionalized with the Zeno phage were used, and electrochemical impedance spectroscopy was applied to evaluate the change of the charge transfer resistance (Rct) as a result of the interaction with S. aureus MRSA ATCC 43300. The phage-based biosensor showed a linear range from 101 to 104 CFU/mL with a 20-minute response time and a limit of detection (LOD) of 1.2 CFU/mL under physiological conditions. The biosensor’s ability to recognize various strains of staphylococci was also successfully demonstrated in the presence of clinical isolates collected from different geographic areas. Assays using S. epidermidis were also carried out to verify the species-specificity of the phage sensor. We only observed a remarkable change of the Rct in the presence of the target S. aureus bacteria, while no substantial binding to S. epidermidis occurred. This confirmed that the Zeno phage sensor only targets S. aureus species within the genus Staphylococcus. In addition, the biosensor's specificity with respect to other bacterial species, including gram-positive bacteria like Enterococcus faecium and the gram-negative bacterium Pseudomonas aeruginosa, was evaluated, and a non-significant impedimetric signal was observed. Notably, the biosensor successfully identified S. aureus bacterial cells in a complex matrix such as a nasal swab, opening the possibility of its use in a real-case scenario. We diluted different concentrations of S. aureus from 108 to 100 CFU/mL with a ratio of 1:10 in the nasal swap matrices collected from healthy donors. Three different sensors were applied to measure various concentrations of bacteria. Our sensor indicated high selectivity to detect S. aureus in biological matrices compared to time-consuming traditional methods, such as enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR), and radioimmunoassay (RIA), etc. With the aim to study the possibility to use this biosensor to address the challenge associated to pathogen detection, ongoing research is focused on the assessment of the biosensor’s analytical performances in different biological samples and the discovery of new phage bioreceptors.

Keywords: electrochemical impedance spectroscopy, bacteriophage, biosensor, Staphylococcus aureus

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Authors: David Tennison

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In the wake of the Covid-19 pandemic, a new chronic illness emerged onto the global stage: Long Covid. Long Covid presents with several symptoms commonly seen in other poorly-understood illnesses, such as fibromyalgia (FM) and myalgic encephalomyelitis/ chronic fatigue syndrome (ME/CFS). However, while Long Covid has swiftly become a recognised illness, FM and ME/CFS are still seen as contested, which impacts patient care and healthcare experiences. This study aims to examine what the differences are between Long Covid and FM; and if the Long Covid case can provide guidance for how to address the healthcare challenge of contested illnesses. To address this question, this study performed comprehensive research into the history of FM; our current biomedical understanding of it; and available healthcare interventions (within the context of the UK NHS). Analysis was undertaken of the stigma and stereotypes around FM, and a comparison made between FM and the emerging Long Covid literature, along with the healthcare response to Long Covid. This study finds that healthcare for chronic contested illnesses in the UK is vastly insufficient - in terms of pharmaceutical and holistic interventions, and the provision of secondary care options. Interestingly, for Long Covid, many of the treatment suggestions are pulled directly from those used for contested illnesses. The key difference is in terms of funding and momentum – Long Covid has generated exponentially more interest and research in a short time than there has been in the last few decades of contested illness research. This stands to help people with FM and ME/CFS – for example, research has recently been funded into “brain fog”, a previously elusive and misunderstood symptom. FM is culturally regarded as a “women’s disease” and FM stigma stems from notions of “hysteria”. A key finding is that the idea of FM affecting women disproportionally is not reflected in modern population studies. Emerging data on Long Covid also suggests a slight leaning towards more female patients, however it is less feminised, potentially due to it emerging in the global historical moment of the pandemic. Another key difference is that FM is rated as an extremely low-prestige illness by healthcare professionals, while it was in large part due to the advocacy of affected healthcare professionals that Long Covid was so quickly recognised by science and medicine. In conclusion, Long Covid (and the risk of future pandemics and post-viral illnesses) highlight a crucial need for implementing new, and reinforcing existing, care networks for chronic illnesses. The difference in how contested illnesses like FM, and new ones like Long Covid are treated have a lot to do with the historical moment in which they emerge – but cultural stereotypes, from within and without medicine, need updating. Particularly as they contribute to disease stigma that causes genuine harm to patients. However, widespread understanding and acceptance of Long Covid could help fight contested illness stigma, and the attention, funding and research into Long Covid may actually help raise the profile of contested illnesses and uncover answers about their symptomatology.

Keywords: long COVID, fibromyalgia, myalgic encephalomyelitis, chronic fatigue syndrome, NHS, healthcare, contested illnesses, chronic illnesses, COVID-19 pandemic

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Authors: Sophio Kobauri, Temur Kantaria, Nina Kulikova, David Tugushi, Ramaz Katsarava

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The elaboration of effective drug delivery vehicles is still topical nowadays since targeted drug delivery is one of the most important challenges of the modern nanomedicine. The last decade has witnessed enormous research focused on synthetic cationic polymers (CPs) due to their flexible properties, in particular as non-viral gene delivery systems, facile synthesis, robustness, not oncogenic and proven gene delivery efficiency. However, the toxicity is still an obstacle to the application in pharmacotherapy. For overcoming the problem, creation of new cationic compounds including the polymeric nano-size particles – nano-containers (NCs) loading with different pharmaceuticals and biologicals is still relevant. In this regard, a variety of NCs-based drug delivery systems have been developed. We have found that amino acid-based biodegradable polymers called as pseudo-proteins (PPs), which can be cleared from the body after the fulfillment of their function are highly suitable for designing pharmaceutical NCs. Among them, one of the most promising are NCs made of biodegradable Cationic PPs (CPPs). For preparing new cationic NCs (CNCs), we used CPPs composed of positively charged amino acid L-arginine (R). The CNCs were fabricated by two approaches using: (1) R-based homo-CPPs; (2) Blends of R-based CPPs with regular (neutral) PPs. According to the first approach NCs we prepared from CPPs 8R3 (composed of R, sebacic acid and 1,3-propanediol) and 8R6 (composed of R, sebacic acid and 1,6-hexanediol). The NCs prepared from these CPPs were 72-101 nm in size with zeta potential within +30 ÷ +35 mV at a concentration 6 mg/mL. According to the second approach, CPPs 8R6 was blended in organic phase with neutral PPs 8L6 (composed of leucine, sebacic acid and 1,6-hexanediol). The NCs prepared from the blends were 130-140 nm in size with zeta potential within +20 ÷ +28 mV depending on 8R6/8L6 ratio. The stability studies of fabricated NCs showed that no substantial change of the particle size and distribution and no big particles’ formation is observed after three months storage. In vitro biocompatibility study of the obtained NPs with four different stable cell lines: A549 (human), U-937 (human), RAW264.7 (murine), Hepa 1-6 (murine) showed both type cathionic NCs are biocompatible. The obtained data allow concluding that the obtained CNCs are promising for the application as biodegradable drug delivery vehicles. This work was supported by the joint grant from the Science and Technology Center in Ukraine and Shota Rustaveli National Science Foundation of Georgia #6298 'New biodegradable cationic polymers composed of arginine and spermine-versatile biomaterials for various biomedical applications'.

Keywords: biodegradable polymers, cationic pseudo-proteins, nano-containers, drug delivery vehicles

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Authors: Mohammad Hossein Pazandeh, Monir Doudi, Sona Rostampour Yasouri

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Irregular consumption of current antibiotic makes increases of antibiotic resistance between urin pathogens on all worlds. This study selected based on this great community problem. The aim of this study was the biosynthesis of silver nanoparticles from Zataria multiflora extract and then to investigate its antibacterial effect on gram-negative bacilli common in Urinary Tract Infections (UTI) and MDR. The plant used in the present research was Zataria multiflora whose extract was prepared through Soxhlet extraction method. Green synthesis condition of silver nanoparticles was investigated in terms of three parameters including the extract amount, concentration of silver nitrate salt, and temperature. The seizes of nanoparticles were determined by Zetasizer. In order to identify synthesized silver nanoparticles Transmission Electron Microscopy (TEM) and X-ray Diffraction (XRD) methods were used. For evaluating the antibacterial effects of nanoparticles synthesized through biological method different concentrations of silver nanoparticles were studied on 140 cases of Muliple Drug Resistance (MDR) bacteria strains Escherichia coli, Klebsiella pneumoniae, Enterobacter aerogenes, Proteus vulgaris,Citrobacter freundii, Acinetobacter bumanii and Pseudomonas aeruginosa, (each genus of bacteria, 20 samples), which all were MDR and cause urinary tract infections , for identification of bacteria were used of Polymerase Chain Reaction (PCR) test and laboratory methods (Agar well diffusion and Microdilution methods) to assess their sensitivity to Nanoparticles. The data were analyzed using SPSS software by nonparametric Kruskal-Wallis and Mann-Whitney tests. Significant results were found about the effects of silver nitrate concentration, different amounts of Zataria multiflora extract, and temperature on nanoparticles; that is, by increasing the concentration of silver nitrate, extract amount, and temperature, the sizes of synthesized nanoparticles declined. However, the effect of above mentioned factors on particles diffusion index was not significant. Based on the TEM results, particles were mainly spherical shape with a diameter range of 25 to 50 nm. The results of XRD Analysis indicated the formation of Nanostructures and Nanocrystals of silver.. The obtained results of antibacterial effects of different concentrations of silver nanoparticles on according to agar well diffusion and microdilution method, biologically synthesized nanoparticles showed 1000 mg /ml highest and lowest mean inhibition zone diameter in E.coli , Acinetobacter bumanii 23 and 15mm, respectively. MIC was observed for all of bacteria 125mg/ml and for Acinetobacter bumanii 250mg/ml.Comparing the growth inhibitory effect of chemically synthesized Nanoparticles and biologically synthesized Nanoparticles showed that in the chemical method the highest growth inhibition belonged to the concentration of 62.5 mg /ml. The inhibitory effect on the growth all of bacteria causes of urine infection and MDR was observed and by increasing silver ion concentration in Nanoparticles, antibacterial activity increased. Generally, the biological synthesis can be considered an efficient way not only in making Nanoparticles but also for having anti-bacterial properties. It is more biocompatible and may be possess less toxicity than the Nanoparticles synthesized chemically.

Keywords: biosynthesis, MDR bacteria, silver nanoparticles, UTI

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Authors: Muhammad Tariq, Muhammad Waseem, Muhammad Hidayat Rasool

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Isolation and characterization of chromium tolerant Staphylococcus aureus from industrial wastewater and their potential use to bioremediate environmental chromium. Objectives: Chromium with its great economic importance in industrial use is major metal pollutant of the environment. Chromium are used in different industries for various applications such as textile, dyeing and pigmentation, wood preservation, manufacturing pulp and paper, chrome plating, steel and tanning. The release of untreated chromium in industrial effluents causes serious threat to environment and human health, therefore, the current study designed to isolate chromium tolerant Staphylococcus aureus for removal of chromium prior to their final discharge into the environment due to its cost effective and beneficial advantage over physical and chemical methods. Methods: Wastewater samples were collected from discharge point of different industries. Heavy metal analysis by atomic absorption spectrophotometer and microbiological analysis such as total viable count, total coliform, fecal coliform and Escherichia coli were conducted. Staphylococcus aureus was identified through gram’s staining, biomeriux vitek 2 microbial identification system and 16S rRNA gene amplification by polymerase chain reaction. Optimum growth conditions with respect to temperature, pH, salt concentrations and effect of chromium on the growth of bacteria, resistance to other heavy metal ions, minimum inhibitory concentration and chromium uptake ability of Staphylococcus aureus strain K1 was determined by spectrophotometer. Antibiotic sensitivity pattern was also determined by disc diffusion method. Furthermore, chromium uptake ability was confirmed by Fourier transform infrared spectroscopy (FTIR) and scanning electron microscope equipped with Oxford Energy Dipersive X-ray (EDX) micro analysis system. Results: The results presented that optimum temperature was 35ᵒC, pH was 8.0 and salt concentration was 0.5% for growth of Staphylococcus aureus K1. The maximum uptake ability of chromium by bacteria was 20mM than other heavy metal ions. The antibiotic sensitivity pattern revealed that Staphylococcus aureus was vancomycin and methicillin sensitive. Non hemolytic activity on blood agar and negative coagulase reaction showed that it was non-pathogenic. Furthermore, the growth of bacteria decreases in the presence of chromium and maximum chromium uptake by bacteria observed at optimum growth conditions. Fourier transform infrared spectroscopy (FTIR), scanning electron microscope (SEM) and Energy dispersive X-ray (EDX) analysis confirmed the presence of chromium uptake by Staphylococcus aureus K1. Conclusion: The study revealed that Staphylococcus aureus K1 have the potential to bio-remediate chromium toxicity from wastewater. Gradually, this biological treatment becomes more important due to its advantage over physical and chemical methods to protect environment and human health.

Keywords: wastewater, staphylococcus, chromium, bioremediation

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Authors: Eleonora Di Salvo, Antonio Panebianco, Graziella Ziino

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Background: Vibrio parahaemolyticus is a human pathogen that is widely distributed in marine environments. It is frequently isolated from raw seafood, particularly shellfish. Consumption of raw or undercooked seafood contaminated with V. parahaemolyticus may lead to acute gastroenteritis. Vibrio spp. has excellent resistance to low temperatures so it can be found in frozen products for a long time. Recently, the viable but non-culturable state (VBNC) of bacteria has attracted great attention, and more than 85 species of bacteria have been demonstrated to be capable of entering this state. VBNC cells cannot grow in conventional culture medium but are viable and maintain metabolic activity, which may constitute an unrecognized source of food contamination and infection. Also V. parahaemolyticus could exist in VBNC state under nutrient starvation or low-temperature conditions. Aim: The aim of the present study was to optimize methods and investigate V. parahaemolyticus VBNC cells and their presence in frozen bivalve molluscs, regularly marketed. Materials and Methods: propidium monoazide (PMA) was integrated with real-time polymerase chain reaction (qPCR) targeting the tl gene to detect and quantify V. parahaemolyticus in the VBNC state. PMA-qPCR resulted highly specific to V. parahaemolyticus with a limit of detection (LOD) of 10-1 log CFU/mL in pure bacterial culture. A standard curve for V. parahaemolyticus cell concentrations was established with the correlation coefficient of 0.9999 at the linear range of 1.0 to 8.0 log CFU/mL. A total of 77 samples of frozen bivalve molluscs (35 mussels; 42 clams) were subsequently subjected to the qualitative (on alkaline phosphate buffer solution) and quantitative research of V. parahaemolyticus on thiosulfate-citrate-bile salts-sucrose (TCBS) agar (DIFCO) NaCl 2.5%, and incubation at 30°C for 24-48 hours. Real-time PCR was conducted on homogenate samples, in duplicate, with and without propidium monoazide (PMA) dye, and exposed for 45 min under halogen lights (650 W). Total DNA was extracted from cell suspension in homogenate samples according to bolliture protocol. The Real-time PCR was conducted with species-specific primers for V. parahaemolitycus. The RT-PCR was performed in a final volume of 20 µL, containing 10 µL of SYBR Green Mixture (Applied Biosystems), 2 µL of template DNA, 2 µL of each primer (final concentration 0.6 mM), and H2O 4 µL. The qPCR was carried out on CFX96 TouchTM (Bio-Rad, USA). Results: All samples were negative both to the quantitative and qualitative detection of V. parahaemolyticus by the classical culturing technique. The PMA-qPCR let us individuating VBNC V. parahaemolyticus in the 20,78% of the samples evaluated with a value between the Log 10-1 and Log 10-3 CFU/g. Only clams samples were positive for PMA-qPCR detection. Conclusion: The present research is the first evaluating PMA-qPCR assay for detection of VBNC V. parahaemolyticus in bivalve molluscs samples, and the used method was applicable to the rapid control of marketed bivalve molluscs. We strongly recommend to use of PMA-qPCR in order to identify VBNC forms, undetectable by the classic microbiological methods. A precise knowledge of the V.parahaemolyticus in a VBNC form is fundamental for the correct risk assessment not only in bivalve molluscs but also in other seafood.

Keywords: food safety, frozen bivalve molluscs, PMA dye, Real-time PCR, VBNC state, Vibrio parahaemolyticus

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Authors: Tatyana Savchuk, Yelena Grinvald, Mohamed Ali, Ramune Sepetiene, Dinara Sadvakassova, Saniya Saussakova, Kuralay Zhangazieva, Dulat Imashpayev

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Introduction. The problem of viral hepatitis B (HBV) takes a vital place in the global health system. The existing risk of HBV transmission through blood transfusions is associated with transfusion of blood taken from infected individuals during the “serological window” period or from patients with latent HBV infection, the marker of which is anti-HBcore. In the absence of information about other markers of hepatitis B, the presence of anti-HBcore suggests that a person may be actively infected or has suffered hepatitis B in the past and has immunity. Aim. To study the risk factors influencing the positive anti-HBcore indicators among the donor population. Materials and Methods. The study was conducted in 2021 in the Scientific and Production Center of Transfusiology of the Ministry of Healthcare in Kazakhstan. The samples taken from blood donors were tested for anti-HBcore, by CLIA on the Architect i2000SR (ABBOTT). A special questionnaire was developed for the blood donors’ socio-demographic characteristics. Statistical analysis was conducted by the R software (version 4.1.1, USA, 2021). Results.5709 people aged 18 to 66 years were included in the study, the proportion of men and women was 68.17% and 31.83%, respectively. The average age of the participants was 35.7 years. A weighted multivariable mixed effects logistic regression analysis showed that age (p<0.001), ethnicity (p<0.05), and marital status (p<0.05) were statistically associated with anti-HBcore positivity. In particular, analysis adjusting for gender, nationality, education, marital status, family history of hepatitis, blood transfusion, injections, and surgical interventions, with a one-year increase in age (adjOR=1.06, 95%CI:1.05-1.07), showed an 6% growth in odds of having anti-HBcore positive results. Those who were russian ethnicity (adjOR=0.65, 95%CI:0.46-0.93) and representatives of other nationality groups (adjOR=0.56, 95%CI:0.37-0.85) had lower odds of having anti-HBcore when compared to Kazakhs when controlling for other covariant variables. Among singles, the odds of having a positive anti-HBcore were lower by 29% (adjOR = 0.71, 95%CI:0.57-0.89) compared to married participants when adjusting for other variables. Conclusions.Kazakhstan is one of the countries with medium endemicity of HBV prevalence (2%-7%). Results of the study demonstrated the possibility to form a profile of risk factors (age, nationality, marital status). Taking into account the data, it is recommended to increase attention to donor questionnaires by adding leading questions and to improve preventive measures to prevent HBV. Funding. This research was supported by a grant from Abbott Laboratories.

Keywords: anti-HBcore, blood donor, donation, hepatitis B virus, occult hepatitis

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Authors: Khaled Alabduljabbar

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Background: Benign, isolated, recurrent sixth nerve palsy is very rare in children. Here we report a case of recurrent abducens nerve palsy with no obvious etiology. It is a diagnosis of exclusion. A recurrent benign form of 6th nerve palsy, a rarer still palsy, has been described in the literature, and it is of most likely secondary to inflammatory causes, e.g, following viral and bacterial infections. Purpose: To present a case of 14 months old girl with recurrent attacks of isolated left sixth cranial nerve palsy following upper respiratory tract infection. Observation: The patient presented to opthalmology clinic with sudden onset of inward deviation (esotropia) of the left eye with a compensatory left face turn one week following signs of upper respiratory tract infection. Ophthalmological examination revealed large angle esotropia of the left eye in primary position, with complete limitation of abduction of the left eye, no palpebral fissure changes, and abnormal position of the head (left face turn). Visual acuity was normal, and no significant refractive error on cycloplegic refraction for her age. Fundus examination was normal with no evidence of papilledema. There was no relative afferent pupillary defect (RAPD) and no anisocoria. Past medical history and family history were unremarkable, with no history of convulsion attacks or head trauma. Additional workout include CBC. Erythrocyte sedimentation rate, Urgent magnetic resonance imaging (MRI), and angiography of the brain were performed and demonstrated the absence of intracranial and orbital lesions. Referral to pediatric neurologist was also done and concluded no significant finding. The patient showed improvement of the left sixth cranial nerve palsy and left face turn over a period of two months. Seven months since the first attack, she experienced a recurrent attack of left eye esotropia with left face turn concurrent with URTI. The rest of eye examination was again unremarkable. CT scan and MRI scan of brain and orbit were performed and showed only signs of sinusitis with no intracranial pathology. The palsy resolved spontaneously within two months. A third episode of left 6th nerve palsy occurred 6 months later, whichrecovered over one month. Examination and neuroimagingwere unremarkable. A diagnosis of benign recurrent left 6th cranial nerve palsy was made. Conclusion: Benign sixth cranial nerve palsy is always a diagnosis of exclusion given the more serious and life-threatening alternative causes. It seems to have a good prognosis with only supportive measures. The likelihood of benign 6th cranial nerve palsy to resolve completely and spontaneously is high. Observation for at least 6 months without intervention is advisable.

Keywords: 6th nerve pasy, abducens nerve pasy, recurrent nerve palsy, cranial nerve palsy

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Authors: Okolie Pius Ifeanyi, Emerenini Emilymary Chima

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Aims: The study investigated the diversity and identities of Lactic Acid Bacteria (LAB) isolated from different fresh fruits using Molecular Nested PCR analysis and the efficacy of cell free supernatants from Lactic Acid Bacteria (LAB) isolated from fresh fruits for in vitro control of some tomato pathogens. Study Design: Nested PCR approach was used in this study employing universal 16S rRNA gene primers in the first round PCR and LAB specific Primers in the second round PCR with the view of generating specific Nested PCR products for the LAB diversity present in the samples. The inhibitory potentials of supernatant obtained from LAB isolates of fruits origin that were molecularly characterized were investigated against some tomato phytopathogens using agar-well method with the view to develop biological agents for some tomato disease causing organisms. Methodology: Gram positive, catalase negative strains of LAB were isolated from fresh fruits on Man Rogosa and Sharpe agar (Lab M) using streaking method. Isolates obtained were molecularly characterized by means of genomic DNA extraction kit (Norgen Biotek, Canada) method. Standard methods were used for Nested Polymerase Chain Reaction (PCR) amplification targeting the 16S rRNA gene using universal 16S rRNA gene and LAB specific primers, agarose gel electrophoresis, purification and sequencing of generated Nested PCR products (Macrogen Inc., USA). The partial sequences obtained were identified by blasting in the non-redundant nucleotide database of National Center for Biotechnology Information (NCBI). The antimicrobial activities of characterized LAB against some tomato phytopathogenic bacteria which include (Xanthomonas campestries, Erwinia caratovora, and Pseudomonas syringae) were obtained by using the agar well diffusion method. Results: The partial sequences obtained were deposited in the database of National Centre for Biotechnology Information (NCBI). Isolates were identified based upon the sequences as Weissella cibaria (4, 18.18%), Weissella confusa (3, 13.64%), Leuconostoc paramensenteroides (1, 4.55%), Lactobacillus plantarum (8, 36.36%), Lactobacillus paraplantarum (1, 4.55%) and Lactobacillus pentosus (1, 4.55%). The cell free supernatants of LAB from fresh fruits origin (Weissella cibaria, Weissella confusa, Leuconostoc paramensenteroides, Lactobacillus plantarum, Lactobacillus paraplantarum and Lactobacillus pentosus) can inhibits these bacteria by creating clear zones of inhibition around the wells containing cell free supernatants of the above mentioned strains of lactic acid bacteria. Conclusion: This study shows that potentially LAB can be quickly characterized by molecular methods to specie level by nested PCR analysis of the bacteria isolate genomic DNA using universal 16S rRNA primers and LAB specific primer. Tomato disease causing organisms can be most likely biologically controlled by using extracts from LAB. This finding will reduce the potential hazard from the use of chemical herbicides on plant.

Keywords: nested pcr, molecular characterization, 16s rRNA gene, lactic acid bacteria

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Authors: Tatsiana Ihnatovich, Darya Nizheharodava, Mikalai Halabarodzka, Tatsiana Savitskaya, Marina Zafranskaya

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Liver fibrosis is a complex of histological changes resulting from chronic liver disease accompanied by an excessive production and deposition of extracellular matrix components in the hepatic parenchyma. Liver fibrosis is a serious medical and social problem. Hepatic stellate cells (HSCs) make a significant contribution to the extracellular matrix deposition due to liver injury. Mesenchymal stem cells (MSCs) have a pronounced anti-inflammatory, regenerative and immunomodulatory effect; they are able to differentiate into hepatocytes and induce apoptosis of activated HSCs that opens the prospect of their use for preventing the excessive fibro-formation and the development of liver cirrhosis. The aim of the study is to evaluate the effect of MSCs therapy on the expression of fibrogenesis markers genes in liver tissue and HSCs cultures of rats with experimental liver cirrhosis (ELC). Materials and methods: ELC was induced by the common bile duct ligation (CBDL) in female Wistar rats (n = 19) with an average body weight of 250 (220 ÷ 270) g. Animals from the control group (n = 10) were sham-operated. On the 56th day after the CBDL, the rats of the experimental (n = 12) and the control (n = 5) groups received intraportal MSCs in concentration of 1×106 cells/animal (previously obtained from rat’s bone marrow) or saline, respectively. The animals were taken out of the experiment on the 21st day. HSCs were isolated by sequential liver perfusion in situ with following disaggregation, enzymatic treatment and centrifugation of cell suspension on a two-stage density gradient. The expression of collagen type I (Col1a1) and type III (Col3a1), matrix metalloproteinase type 2 (MMP2) and type 9 (MMP9), tissue inhibitor of matrix metalloproteinases type 1 (TIMP1), transforming growth factor β type 1 (TGFβ1) and type 3 (TGFβ3) was determined by real-time polymerase chain reaction. Statistical analysis was performed using Statistica 10.0. Results: In ELC rats compared to sham-operated animals, a significant increase of all studied markers expression was observed. The administration of MSCs led to a significant decrease of all detectable markers in the experimental group compared to rats without cell therapy. In ELC rats, an increased MMP9/TIMP1 ratio after cell therapy was also detected. The infusion of MSCs in the sham-operated animals did not lead to any changes. In the HSCs from ELC animals, the expression of Col1a1 and Col3a1 exceeded the similar parameters of the control group (p <0.05) and statistically decreased after the MSCs administration. The correlation between Col3a1 (Rs = 0.51, p <0.05), TGFβ1 (Rs = 0.6, p <0.01), and TGFβ3 (Rs = 0.75, p <0.001) expression in HSCs cultures and liver tissue has been found. Conclusion: Intraportal administration of MSCs to rats with ELC leads to a decreased Col1a1 and Col3a1, MMP2 and MMP9, TIMP1, TGFβ1 and TGFβ3 expression. The correlation between the expression of Col3a1, TGFβ1 and TGFβ3 in liver tissue and in HSCs cultures indicates the involvement of activated HSCs in the fibrogenesis that allows considering HSCs to be the main cell therapy target in ELC.

Keywords: cell therapy, experimental liver cirrhosis, hepatic stellate cells, mesenchymal stem cells

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Authors: Iman Farasat, Howard M. Salis

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Engineered genetic circuits reprogram cellular behavior to act as living computers with applications in detecting cancer, creating self-controlling artificial tissues, and dynamically regulating metabolic pathways. Phenemenological models are often used to simulate and design genetic circuit behavior towards a desired behavior. While such models assume that each circuit component’s function is modular and independent, even small changes in a circuit (e.g. a new promoter, a change in transcription factor expression level, or even a new media) can have significant effects on the circuit’s function. Here, we use statistical thermodynamics to account for the several factors that control transcriptional regulation in bacteria, and experimentally demonstrate the model’s accuracy across 825 measurements in several genetic contexts and hosts. We then employ our first principles model to design, experimentally construct, and characterize a family of signal amplifying genetic circuits (genetic OpAmps) that expand the dynamic range of cell sensors. To develop these models, we needed a new approach to measuring the in vivo binding free energies of transcription factors (TFs), a key ingredient of statistical thermodynamic models of gene regulation. We developed a new high-throughput assay to measure RNA polymerase and TF binding free energies, requiring the construction and characterization of only a few constructs and data analysis (Figure 1A). We experimentally verified the assay on 6 TetR-homolog repressors and a CRISPR/dCas9 guide RNA. We found that our binding free energy measurements quantitatively explains why changing TF expression levels alters circuit function. Altogether, by combining these measurements with our biophysical model of translation (the RBS Calculator) as well as other measurements (Figure 1B), our model can account for changes in TF binding sites, TF expression levels, circuit copy number, host genome size, and host growth rate (Figure 1C). Model predictions correctly accounted for how these 8 factors control a promoter’s transcription rate (Figure 1D). Using the model, we developed a design framework for engineering multi-promoter genetic circuits that greatly reduces the number of degrees of freedom (8 factors per promoter) to a single dimensionless unit. We propose the Ptashne (Pt) number to encapsulate the 8 co-dependent factors that control transcriptional regulation into a single number. Therefore, a single number controls a promoter’s output rather than these 8 co-dependent factors, and designing a genetic circuit with N promoters requires specification of only N Pt numbers. We demonstrate how to design genetic circuits in Pt number space by constructing and characterizing 15 2-repressor OpAmp circuits that act as signal amplifiers when within an optimal Pt region. We experimentally show that OpAmp circuits using different TFs and TF expression levels will only amplify the dynamic range of input signals when their corresponding Pt numbers are within the optimal region. Thus, the use of the Pt number greatly simplifies the genetic circuit design, particularly important as circuits employ more TFs to perform increasingly complex functions.

Keywords: transcription factor, synthetic biology, genetic circuit, biophysical model, binding energy measurement

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Authors: Nouha Bouayed Abdelmoula, Rim Louati, Nawel Abdellaoui, Balkiss Abdelmoula, Oldez Kaabi, Walid Smaoui, Samir Aloulou

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Background and Aims: Cardiofaciocutaneous syndrome (CFC) is an autosomal dominant disorder with the vast majority of cases arising by a new mutation of BRAF, MEK1, MEK2, or rarely, KRAS genes. Here, we report a rare Tunisian case of CFC syndrome for whom we identify SOS1 mutation. Methods: Genomic DNA was obtained from peripheral blood collected in an EDTA tube and extracted from leukocytes using the phenol/chloroform method according to standard protocols. High resolution melting (HRM) analysis for screening of mutations in the entire coding sequence of PTPN11 was conducted first. Then, HRM assays to look for hot spot mutations coding regions of the other genes of the RAS-MAPK pathway (RAt Sarcoma viral oncogene homolog Mitogen-Activated Protein Kinases Pathway): SOS1, SHOC2, KRAS, RAF1, KRAS, NRAS, CBL, BRAF, MEK1, MEK2, HRAS, and RIT1, were applied. Results: Heterozygous SOS1 point mutation clustered in exon 10, which encodes for the PH domain of SOS1, was identified: c.1655 G > A. The patient was a 9-year-old female born from a consanguineous couple. She exhibited pulmonic valvular stenosis as congenital heart disease. She had facial features and other malformations of Noonan syndrome, including macrocephaly, hypertelorism, ptosis, downslanting palpebral fissures, sparse eyebrows, a short and broad nose with upturned tip, low-set ears, high forehead commonly associated with bitemporal narrowing and prominent supraorbital ridges, short and/or webbed neck and short stature. However, the phenotype is also suggestive of CFC syndrome with the presence of more severe ectodermal abnormalities, including curly hair, keloid scars, hyperkeratotic skin, deep plantar creases, and delayed permanent dentition with agenesis of the right maxillary first molar. Moreover, the familial history of the patient revealed recurrent brain malignancies in the paternal family and epileptic disease in the maternal family. Conclusions: This case report of an overlapping RASopathy associated with SOS1 mutation and familial history of brain tumorigenesis is exceptional. The evidence suggests that RASopathies are truly cancer-prone syndromes, but the magnitude of the cancer risk and the types of cancer partially overlap.

Keywords: cardiofaciocutaneous syndrome, CFC, SOS1, brain cancer, germline mutation

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Authors: Ewelina Wisnik, Zsolt Regdon, Kinga Chmielewska, Laszlo Virag, Agnieszka Robaszkiewicz

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Apart from protecting genome, PARP1 has been documented to regulate many intracellular processes inter alia gene transcription by physically interacting with chromatin bound proteins and by their ADP-ribosylation. Our recent findings indicate that expression of PARP1 decreases during the differentiation of human CD34+ hematopoietic stem cells to monocytes as a consequence of differentiation-associated cell growth arrest and formation of E2F4-RBL2-HDAC1-SWI/SNF repressive complex at the promoter of this gene. Since the RBL2 complexes repress genes in a E2F-dependent manner and are widespread in the genome in G0 arrested cells, we asked (a) if RBL2 directly contributes to defining monocyte phenotype and function by targeting gene promoters and (b) if RBL2 controls gene transcription indirectly by repressing PARP1. For identification of genes controlled by RBL2 and/or PARP1,we used primer libraries for surface receptors and TLR signaling mediators, genes were silenced by siRNA or shRNA, analysis of gene promoter occupation by selected proteins was carried out by ChIP-qPCR, while statistical analysis in GraphPad Prism 5 and STATISTICA, ChIP-Seq data were analysed in Galaxy 2.5.0.0. On the list of 28 genes regulated by RBL2, we identified only four solely repressed by RBL2-E2F4-HDAC1-BRM complex. Surprisingly, 24 out of 28 emerged genes controlled by RBL2 were co-regulated by PARP1 in six different manners. In one mode of RBL2/PARP1 co-operation, represented by MAP2K6 and MAPK3, PARP1 was found to associate with gene promoters upon RBL2 silencing, which was previously shown to restore PARP1 expression in monocytes. PARP1 effect on gene transcription was observed only in the presence of active EP300, which acetylated gene promoters and activated transcription. Further analysis revealed that PARP1 binding to MA2K6 and MAPK3 promoters enabled recruitment of EP300 in monocytes, while in proliferating cancer cell lines, which actively transcribe PARP1, this protein maintained EP300 at the promoters of MA2K6 and MAPK3. Genome-wide analysis revealed a similar distribution of PARP1 and EP300 around transcription start sites and the co-occupancy of some gene promoters by PARP1 and EP300 in cancer cells. Here, we described a new RBL2/PARP1/EP300 axis which controls gene transcription regardless of the cell type. In this model cell, cycle-dependent transcription of PARP1 regulates expression of some genes repressed by RBL2 upon cell cycle limitation. Thus, RBL2 may indirectly regulate transcription of some genes by controlling the expression of EP300-recruiting PARP1. Acknowledgement: This work was financed by Polish National Science Centre grants nr DEC-2013/11/D/NZ2/00033 and DEC-2015/19/N/NZ2/01735. L.V. is funded by the National Research, Development and Innovation Office grants GINOP-2.3.2-15-2016-00020 TUMORDNS, GINOP-2.3.2-15-2016-00048-STAYALIVE and OTKA K112336. AR is supported by Polish Ministry of Science and Higher Education 776/STYP/11/2016.

Keywords: retinoblastoma transcriptional co-repressor like 2 (RBL2), poly(ADP-ribose) polymerase 1 (PARP1), E1A binding protein p300 (EP300), monocytes

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Authors: Neda Jebellie

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New information and communication technologies (ICT) not only has revolutionized the world of communication but has also strongly impacted the state of international terrorism. Using the potential of social media, the new wave of terrorism easily can recruit new jihadi members, spread their violent ideology and garner financial support. IS (Islamic State) as the most dangerous terrorist group has already conquered a great deal of social media space and has deployed sophisticated web-based strategies to promote its extremist doctrine. In this respect the vastly popular social media are the perfect tools for IS to establish its virtual Caliphate (e-caliphate) and e-Ommah (e-citizen).Using social media to release violent videos of beheading journalists, burning their hostages alive and mass killing of prisoners are IS strategies to terrorize and subjugate its enemies. Several Twitter and Facebook accounts which are IS affiliations have targeted young generation of Muslims all around the world. In fact IS terrorists use modern resources of communication not only to share information and conduct operations but also justify their violent acts. The strict Wahhabi doctrine of ISIS is based on a fundamental interpretation of Islam in which religious war against non Muslims (Jihad) and killing infidels (Qatal) have been praised and recommended. Via social media IS disseminates its propaganda to inspire sympathizers across the globe. Combating this new wave of terrorism which is exploiting new communication technologies is the most significant challenge for authorities. Before the rise of internet and social media governments had to control only mosques and religious gathering such as Friday sermons(Jamaah Pray) to prevent spreading extremism among Muslims community in their country. ICT and new communication technologies have heighten the challenge of dealing with Islamic radicalism and have amplified its threat .According to the official reports even some of the governments such as UK have created a special force of Facebook warriors to engage in unconventional warfare in digital age. In compare with other terrorist groups, IS has effectively grasped social media potential. Their horrifying released videos on YouTube easily got viral and were re-twitted and shared by thousands of social media users. While some of the social media such as Twitter and Facebook have shut down many accounts alleged to IS but new ones create immediately so only blocking their websites and suspending their accounts cannot solve the problem as terrorists recreate new accounts. To combat cyber terrorism focusing on disseminating counter narrative strategies can be a solution. Creating websites and providing online materials to propagate peaceful and moderate interpretation of Islam can provide a cogent alternative to extremist views.

Keywords: IS-islamic state, cyber terrorism, social media, terrorism, information, communication technologies

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Authors: Mariam Khaled, Noha Farag, Ghada Mohamed Abdel Salam, Khaled Abu-Aisha, Mohamed El-Azizi

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Gastric and colorectal cancers are among the most frequent causes of cancer-associated mortalities in Africa. They have been considered as a global public health concern, as nearly one million new cases are reported per year. IL-1β is a pro-inflammatory cytokine-produced by activated macrophages and monocytes- and a member of the IL-1 family. The inactive IL-1β precursor is cleaved and activated by caspase-1 enzyme, which itself is activated by the assembly of intracellular structures defined as NLRP3 (Nod Like receptor P3) inflammasomes. Activated IL-1β stimulates the Interleukin-1 receptor type-1 (IL-1R1), which is responsible for the initiation of a signal transduction pathway leading to cell proliferation. It has been proven that the IL-1β gene is a highly polymorphic gene in which single nucleotide polymorphisms (SNPs) may affect its expression. It has been previously reported that SNPs including base transitions between C and T at positions, -511 (C-T; dbSNP: rs16944) and -31 (C-T; dbSNP: rs1143627), from the transcriptional start site, contribute to the pathogenesis of gastric and colorectal cancers by affecting IL-1β levels. Altered production of IL-1β due to such polymorphisms is suspected to stimulate an amplified inflammatory response and promote Epithelial Mesenchymal Transition leading to malignancy. Allele frequency distribution of the IL-1β-31 and -511 SNPs, in different populations, and their correlation to the incidence of gastric and colorectal cancers, has been intriguing to researchers worldwide. The current study aims to investigate allele distributions of the IL-1β SNPs among gastric and colorectal cancers Egyptian patients. In order to achieve to that, 89 Biopsy and surgical specimens from the antrum and corpus mucosa of chronic gastritis subjects and gastric and colorectal carcinoma patients was collected for DNA extraction followed by restriction fragment length polymorphism polymerase chain reaction (RFLP-PCR). The amplified PCR products of IL-1β-31C > T and IL-1β-511T > C were digested by incubation with the restriction endonuclease enzymes ALu1 and Ava1. Statistical analysis was carried out to determine the allele frequency distribution in the three studied groups. Also, the effect of the IL-1β -31 and -511 SNPs on nuclear factor binding was analyzed using Fluorescence Electrophoretic Mobility Shift Assay (EMSA), preceded by nuclear factor extraction from gastric and colorectal tissue samples and LPS stimulated monocytes. The results of this study showed that a significantly higher percentage of Egyptian gastric cancer patients have a homozygous CC genotype at the IL-1β-31 position and a heterozygous TC genotype at the IL-1β-511 position. Moreover, a significantly higher percentage of the colorectal cancer patients have a homozygous CC genotype at the IL-1β-31 and -511 positions as compared to the control group. In addition, the EMSA results showed that IL-1β-31C/T and IL-1β-511T/C SNPs do not affect nuclear factor binding. Results of this study suggest that the IL-1β-31 C/T and IL-1β-511 T/C may be correlated to the incidence of gastric cancer in Egyptian patients; however, similar findings couldn’t be proven in the colorectal cancer patients group for the IL-1β-511 T/C SNP. This is the first study to investigate IL-1β -31 and -511 SNPs in the Egyptian population.

Keywords: colorectal cancer, Egyptian patients, gastric cancer, interleukin-1β, single nucleotide polymorphisms

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Authors: Spira A., Marabelle A., Kientop D., Moser E., Mumm J.

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Background: Both interleukin-2 (IL-2) and interleukin-10 (IL-10) have been extensively studied for their stimulatory function on T cells and their potential to obtain sustainable tumor control in RCC, melanoma, lung, and pancreatic cancer as monotherapy, as well as combination with PD-1 blockers, radiation, and chemotherapy. While approved, IL-2 retains significant toxicity, preventing its widespread use. The significant efforts undertaken to uncouple IL-2 toxicity from its anti-tumor function have been unsuccessful, and early phase clinical safety observed with PEGylated IL-10 was not met in a blinded Phase 3 trial. Deka Biosciences has engineered a novel molecule coupling wild-type IL-2 to a high affinity variant of Epstein Barr Viral (EBV) IL-10 via a scaffold (scFv) that binds to epidermal growth factor receptors (EGFR). This patented molecule, termed DK210 (EGFR), is retained at high levels within the tumor microenvironment for days after dosing. In addition to overlapping and non-redundant anti-tumor function, IL-10 reduces IL-2 mediated cytokine release syndrome risks and inhibits IL-2 mediated T regulatory cell proliferation. Methods: DK210 (EGFR) is being evaluated in an open-label, dose-escalation (Phase 1) study with 5 (0.025-0.3 mg/kg) monotherapy dose levels and (expansion cohorts) in combination with PD-1 blockers, or radiation or chemotherapy in patients with advanced solid tumors overexpressing EGFR. Key eligibility criteria include 1) confirmed progressive disease on at least one line of systemic treatment, 2) EGFR overexpression or amplification documented in histology reports, 3) at least a 4 week or 5 half-lives window since last treatment, and 4) excluding subjects with long QT syndrome, multiple myeloma, multiple sclerosis, myasthenia gravis or uncontrolled infectious, psychiatric, neurologic, or cancer disease. Plasma and tissue samples will be investigated for pharmacodynamic and predictive biomarkers and genetic signatures associated with IFN-gamma secretion, aiming to select subjects for treatment in Phase 2. Conclusion: Through successful coupling of wild-type IL-2 with a high affinity IL-10 and targeting directly to the tumor microenvironment, DK210 (EGFR) has the potential to harness IL-2 and IL-10’s known anti-cancer promise while reducing immunogenicity and toxicity risks enabling safe concomitant cytokine treatment with other anti-cancer modalities.

Keywords: cytokine, EGFR over expression, interleukine-2, interleukine-10, clinical trial

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Authors: Yu-Mei Hsueh, Wei-Jen Chen, Yung-Kai Huang, Cheng-Shiuan Tsai, Kuo-Cheng Yeh

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Prostate cancer occurs in men over the age of 50, and rank sixth of the top ten cancers in Taiwan, and the incidence increased gradually over the past decade in Taiwan. Arsenic is confirmed as a carcinogen by International Agency for Research on (IARC). Arsenic induces oxidative stress may be a risk factor for prostate cancer, but the mechanism is not clear. Selenium is an important antioxidant element. Whether the association between plasma selenium levels and risk of prostate cancer are modified by different genotype of selenoprotein is still unknown. Glutathione peroxidase, selenoprotein P (SEPP1) and 15 kDa selenoprotein (SEP 15) are selenoprotein and regulates selenium transport and the oxidation and reduction reaction. However, the association between gene polymorphisms of selenoprotein and prostate cancer is not yet clear. The aim of this study is to determine the relationship between plasma selenium, polymorphism of selenoprotein, urinary total arsenic concentration and prostate cancer. This study is a hospital-based case-control study. Three hundred twenty-two cases of prostate cancer and age (±5 years) 1:1 matched 322 control group were recruited from National Taiwan University Hospital, Taipei Medical University Hospital, and Wan Fang Hospital. Well-trained personnel carried out standardized personal interviews based on a structured questionnaire. Information collected included demographic and socioeconomic characteristics, lifestyle and disease history. Blood and urine samples were also collected at the same time. The Research Ethics Committee of National Taiwan University Hospital, Taipei, Taiwan, approved the study. All patients provided informed consent forms before sample and data collection. Buffy coat was to extract DNA, and the polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) was used to measure the genotypes of SEPP1 rs3797310, SEP15 rs5859, GPX1 rs1050450, GPX2 rs4902346, GPX3 rs4958872, and GPX4 rs2075710. Plasma concentrations of selenium were determined by inductively coupled plasma mass spectrometry (ICP-MS).Urinary arsenic species concentrations were measured by high-performance liquid chromatography links hydride generator and atomic absorption spectrometer (HPLC-HG-AAS). Subject with high education level compared to those with low educational level had a lower prostate cancer odds ratio (OR) Mainland Chinese and aboriginal people had a lower OR of prostate cancer compared to Fukien Taiwanese. After adjustment for age, educational level, subjects with GPX1 rs1050450 CT and TT genotype compared to the CC genotype have lower, OR of prostate cancer, the OR and 95% confidence interval (Cl) was 0.53 (0.31-0.90). SEPP1 rs3797310 CT+TT genotype compared to those with CC genotype had a marginally significantly lower OR of PC. The low levels of plasma selenium and the high urinary total arsenic concentrations had the high OR of prostate cancer in a significant dose-response manner, and SEPP1 rs3797310 genotype modified this joint association.

Keywords: prostate cancer, plasma selenium concentration, urinary total arsenic concentrations, glutathione peroxidase, selenoprotein P, selenoprotein 15, gene polymorphism

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Authors: Mohamad F. Jamiluddin, Emeline Sarry, Ana Bejanariu, Cécile Bauche

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Introduction: Over 360 million people are chronically infected with hepatitis B virus (HBV), of whom 1 million die each year from HBV-associated liver cirrhosis or hepatocellular carcinoma. Current treatment options for chronic hepatitis B depend on interferon-α (IFNα) or nucleos(t)ide analogs, which control virus replication but rarely eliminate the virus. Treatment with PEG-IFNα leads to a sustained antiviral response in only one third of patients. After withdrawal of the drugs, the rebound of viremia is observed in the majority of patients. Furthermore, the long-term treatment is subsequently associated with the appearance of drug resistant HBV strains that is often the cause of the therapy failure. Among the new therapeutic avenues being developed, therapeutic vaccine aimed at inducing immune responses similar to those found in resolvers is of growing interest. The high prevalence of chronic hepatitis B necessitates the design of better vaccination strategies capable of eliciting broad-spectrum of cell-mediated immunity(CMI) and humoral immune response that can control chronic hepatitis B. Induction of HBV-specific T cells and B cells by therapeutic vaccination may be an innovative strategy to overcome virus persistence. Lentiviral vectors developed and optimized by THERAVECTYS, due to their ability to transduce non-dividing cells, including dendritic cells, and induce CMI response, have demonstrated their effectiveness as vaccination tools. Method: To develop a HBV therapeutic vaccine that can induce a broad but specific immune response, we generated recombinant lentiviral vector carrying IRES(Internal Ribosome Entry Site)-containing bicistronic constructs which allow the coexpression of two vaccine products, namely HBV T- cell epitope vaccine and HBV virus like particle (VLP) vaccine. HBV T-cell epitope vaccine consists of immunodominant cluster of CD4 and CD8 epitopes with spacer in between them and epitopes are derived from HBV surface protein, HBV core, HBV X and polymerase. While HBV VLP vaccine is a HBV core protein based chimeric VLP with surface protein B-cell epitopes displayed. In order to evaluate the immunogenicity, mice were immunized with lentiviral constructs by intramuscular injection. The T cell and antibody immune responses of the two vaccine products were analyzed using IFN-γ ELISpot assay and ELISA respectively to quantify the adaptive response to HBV antigens. Results: Following a single administration in mice, lentiviral construct elicited robust antigen-specific IFN-γ responses to the encoded antigens. The HBV T- cell epitope vaccine demonstrated significantly higher T cell immunogenicity than HBV VLP vaccine. Importantly, we demonstrated by ELISA that antibodies are induced against both HBV surface protein and HBV core protein when mice injected with vaccine construct (p < 0.05). Conclusion: Our results highlight that THERAVECTYS lentiviral vectors may represent a powerful platform for immunization strategy against chronic hepatitis B. Our data suggests the likely importance of Lentiviral vector based novel bicistronic construct for further study, in combination with drugs or as standalone antigens, as a therapeutic lentiviral based HBV vaccines. THERAVECTYS bicistronic HBV vaccine will be further evaluated in animal efficacy studies.

Keywords: chronic hepatitis B, lentiviral vectors, therapeutic vaccine, virus-like particle

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Authors: Francesca Nardi, Kashish Aneja, Katherine Ginsbach

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The COVID-19 pandemic has demonstrated both a need for evidence-based and rights-based public health policy and how challenging it can be to make effective decisions with limited information, evidence, and data. The O’Neill Institute, in conjunction with several partners, has been working since the beginning of the pandemic to collect, analyze, and distribute critical data on public health policies enacted in response to COVID-19 around the world in the COVID-19 Law Lab. Well-designed laws and policies can help build strong health systems, implement necessary measures to combat viral transmission, enforce actions that promote public health and safety for everyone, and on the individual level have a direct impact on health outcomes. Poorly designed laws and policies, on the other hand, can fail to achieve the intended results and/or obstruct the realization of fundamental human rights, further disease spread, or cause unintended collateral harms. When done properly, laws can provide the foundation that brings clarity to complexity, embrace nuance, and identifies gaps of uncertainty. However, laws can also shape the societal factors that make disease possible. Law is inseparable from the rest of society, and COVID-19 has exposed just how much laws and policies intersects all facets of society. In the COVID-19 context, evidence-based and well-informed law and policy decisions—made at the right time and in the right place—can and have meant the difference between life or death for many. Having a solid evidentiary base of legal information can promote the understanding of what works well and where, and it can drive resources and action to where they are needed most. We know that legal mechanisms can enable nations to reduce inequities and prepare for emerging threats, like novel pathogens that result in deadly disease outbreaks or antibiotic resistance. The collection and analysis of data on these legal mechanisms is a critical step towards ensuring that legal interventions and legal landscapes are effectively incorporated into more traditional kinds of health science data analyses. The COVID-19 Law Labs see a unique opportunity to collect and analyze this kind of non-traditional data to inform policy using laws and policies from across the globe and across diseases. This global view is critical to assessing the efficacy of policies in a wide range of cultural, economic, and demographic circumstances. The COVID-19 Law Lab is not just a collection of legal texts relating to COVID-19; it is a dataset of concise and actionable legal information that can be used by health researchers, social scientists, academics, human rights advocates, law and policymakers, government decision-makers, and others for cross-disciplinary quantitative and qualitative analysis to identify best practices from this outbreak, and previous ones, to be better prepared for potential future public health events.

Keywords: public health law, surveillance, policy, legal, data

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Authors: Tiera Tanksley, Amanda M. McLeroy

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The advent of technology and social media has ushered in a new era of communication, providing platforms for news dissemination and cause advocacy. However, this digital landscape has also exposed a distressing phenomenon termed "Black death," or trauma porn. This paper delves into the profound effects of repeated exposure to traumatic content on Black youth via social media, exploring the psychological impacts and potential reinforcing of stereotypes. Employing Critical Race Technology Theory (CRTT), the study sheds light on algorithmic anti-blackness and its influence on Black youth's lives and educational experiences. Through ethnographic content analysis, the research investigates common manifestations of Black death encountered online by Black adolescents. Findings unveil distressing viral videos, traumatic images, racial slurs, and hate speech, perpetuating stereotypes. However, amidst the distress, the study identifies narratives of activism and social justice on social media platforms, empowering Black youth to engage in positive change. Coping mechanisms and community support emerge as significant factors in navigating the digital landscape. The study underscores the need for comprehensive interventions and policies informed by evidence-based research. By addressing algorithmic anti-blackness and promoting digital resilience, the paper advocates for a more empathetic and inclusive online environment. Understanding coping mechanisms and community support becomes imperative for fostering mental well-being among Black adolescents navigating social media. In education, the implications are substantial. Acknowledging the impact of Black death content, educators play a pivotal role in promoting media literacy and digital resilience. Creating inclusive and safe online spaces, educators can mitigate negative effects and encourage open discussions about traumatic content. The application of CRTT in educational technology emphasizes dismantling systemic biases and promoting equity. In conclusion, this study calls for educators to be cognizant of the impact of Black death content on social media. By prioritizing media literacy, fostering digital resilience, and advocating for unbiased technologies, educators contribute to an inclusive and just educational environment for all students, irrespective of their race or background. Addressing challenges related to Black death content proactively ensures the well-being and mental health of Black adolescents, fostering an empathetic and inclusive digital space.

Keywords: algorithmic anti-Blackness, digital resilience, media literacy, traumatic content

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Authors: Nahla Elsherbiny, Ahmed Ahmed, Hamada Mohammed, Mohamed Ali

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Background: The enterococci may be considered as opportunistic agents particularly in immunocompromised patients. It is one of the top three pathogens causing many healthcare associated infections (HAIs). Resistance to several commonly used antimicrobial agents is a remarkable characteristic of most species which may carry various genes contributing to virulence. Objectives: to determine the prevalence of enterococci species in different intensive care units (ICUs) causing health care-associated infections (HAIs), intestinal carriage and environmental contamination. Also, to study the antimicrobial susceptibility pattern of the isolates with special reference to vancomycin resistance. In addition to phenotypic and genotypic detection of gelatinase, cytolysin and biofilm formation among isolates. Patients and Methods: This study was carried out in the infection control laboratory at Assiut University Hospitals over a period of one year. Clinical samples were collected from 285 patients with various (HAIs) acquired after admission to different ICUs. Rectal swabs were taken from 14 cases for detection of enterococci carriage. In addition, 1377 environmental samples were collected from the surroundings of the patients. Identification was done by conventional bacteriological methods and confirmed by analytical profile index (API). Antimicrobial sensitivity testing was performed by Kirby Bauer disc diffusion method and detection of vancomycin resistance was done by agar screen method. For the isolates, phenotypic detection of cytolysin, gelatinase production and detection of biofilm by tube method, Congo red method and microtiter plate. We performed polymerase chain reaction (PCR) for detection of some virulence genes (gelE, cylA, vanA, vanB and esp). Results: Enterococci caused 10.5% of the HAIs. Respiratory tract infection was the predominant type (86.7%). The commonest species were E.gallinarum (36.7%), E.casseliflavus (30%), E.faecalis (30%), and E.durans (3.4 %). Vancomycin resistance was detected in a total of 40% (12/30) of those isolates. The risk factors associated with acquiring vancomycin resistant enterococci (VRE) were immune suppression (P= 0.031) and artificial feeding (P= 0.008). For the rectal swabs, enterococci species were detected in 71.4% of samples with the predominance of E. casseliflavus (50%). Most of the isolates were vancomycin resistant (70%). Out of a total 1377 environmental samples, 577 (42%) samples were contaminated with different microorganisms. Enterococci were detected in 1.7% (10/577) of total contaminated samples, 50% of which were vancomycin resistant. All isolates were resistant to penicillin, ampicillin, oxacillin, ciprofloxacin, amikacin, erythromycin, clindamycin and trimethoprim-sulfamethaxazole. For the remaining antibiotics, variable percentages of resistance were reported. Cytolysin and gelatinase were detected phenotypically in 16% and 48 % of the isolates respectively. The microtiter plate method showed the highest percentages of detection of biofilm among all isolated species (100%). The studied virulence genes gelE, esp, vanA and vanB were detected in 62%, 12%, 2% and 12% respectively, while cylA gene was not detected in any isolates. Conclusions: A significant percentage of enterococci was isolated from patients and environments in the ICUs. Many virulence factors were detected phenotypically and genotypically among isolates. The high percentage of resistance, coupled with the risk of cross transmission to other patients make enterococci infections a significant infection control issue in hospitals.

Keywords: antimicrobial resistance, enterococci, ICUs, virulence factors

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Authors: Michelle S. Pereira, Analice C. Azevedo, Julliane D. Medeiros, Ana Claudia S. Martins, Didier S. Castellano-Filho, Claudio G. Diniz, Vania L. Silva

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Vaginal microbiota is a complex ecosystem, composed by aerobic and anaerobic bacteria, living in a dynamic equilibrium. Lactobacillus spp. are predominant in vaginal ecosystem, and factors such as immunity and hormonal variations may lead to disruptions, resulting in proliferation of opportunistic pathogens. Bacterial vaginosis (BV) is a polymicrobial syndrome, caused by an increasing of anaerobic bacteria replacing Lactobacillus spp. Microorganisms such as Gardnerella vaginalis, Mycoplasma hominis, Mobiluncus spp., and Atopobium vaginae can be found in BV, which may also be associated to other infections such as by Human Papillomavirus (HPV). HPV is highly prevalent in sexually active women, and is considered a risk factor for development of cervical cancer. As long as few data is available on vaginal microbiota of women with HPV-associated cervical lesions, our objectives were to evaluate the diversity in vaginal ecosystem in these women. To all patients, clinical and socio-demographic data were collected after gynecological examination. This study was approved by the Ethics Committee from Federal University of Juiz de Fora, Minas Gerais, Brazil. Vaginal secretion and cervical scraping were collected. Gram-stained smears were evaluated to establish Nugent score for BV determination. Viral and bacterial DNA obtained was used as template for HPV genotyping (PCR) and bacterial fingerprint (REP-PCR). In total 31 patients were included (mean age 35 and 93.6% sexually active). The Nugent score showed that 38.7% were BV. From the medical records, Pap smear tests showed that 32.3% had low grade squamous epithelial lesion (LSIL), 29% had high grade squamous epithelial lesion (HSIL), 25.8% had atypical squamous cells of undetermined significance (ASC-US) and 12.9% with atypical squamous cells that would not exclude high-grade lesion (ASC-H). All participants were HPV+. HPV-16 was the most frequent (87.1%), followed by HPV-18 (61.3%). HPV-31, HPV-52 and HPV-58 were also detected. Coinfection HPV-16/HPV-18 was observed in 75%. In the 18-30 age group, HPV-16 was detected in 40%, and HPV-16/HPV-18 coinfection in 35%. HPV-16 was associated to 30% of ASC-H and 20% of HSIL patients. BV was observed in 50% of HPV-16+ participants and in 45% of HPV-16/HPV-18+. Fingerprints of bacterial communities showed clusters with low similarity suggesting high heterogeneity in vaginal microbiota within the sampled group. Overall, the data is worrisome once cervical-cancer highly risk-associated HPV-types were identified. The high microbial diversity observed may be related to the different levels of cellular lesions, and different physiological conditions of the participants (age, social behavior, education). Further prospective studies are needed to better address correlations and BV and microbial imbalance in vaginal ecosystems which would be related to the different cellular lesions in women with HPV infections. Supported by FAPEMIG, CNPq, CAPES, PPGCBIO/UFJF.

Keywords: human papillomavirus, bacterial vaginosis, bacterial diversity, cervical cancer

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Authors: Denise De Fátima Fernandes Barbosa, Tyane Mayara Ferreira Oliveira, Diego Jorge Maia Lima, Paula Renata Amorim Lessa, Ana Karina Bezerra Pinheiro, Cintia Gondim Pereira Calou, Glauberto Da Silva Quirino, Hellen Lívia Oliveira Catunda, Tatiana Gomes Guedes, Nicolau Da Costa

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The aim of this study is to evaluate the use of Digital Cervicography (DC) in the diagnosis of precancerous lesions related to Human Papillomavirus (HPV). Cross-sectional study with a quantitative approach, of evaluative type, held in a health unit linked to the Pro Dean of Extension of the Federal University of Ceará, in the period of July to August 2015 with a sample of 33 women. Data collecting was conducted through interviews with enforcement tool. Franco (2005) standardized the technique used for DC. Polymerase Chain Reaction (PCR) was performed to identify high-risk HPV genotypes. DC were evaluated and classified by 3 judges. The results of DC and PCR were classified as positive, negative or inconclusive. The data of the collecting instruments were compiled and analyzed by the software Statistical Package for Social Sciences (SPSS) with descriptive statistics and cross-references. Sociodemographic, sexual and reproductive variables were analyzed through absolute frequencies (N) and their respective percentage (%). Kappa coefficient (κ) was applied to determine the existence of agreement between the DC of reports among evaluators with PCR and also among the judges about the DC results. The Pearson's chi-square test was used for analysis of sociodemographic, sexual and reproductive variables with the PCR reports. It was considered statistically significant (p<0.05). Ethical aspects of research involving human beings were respected, according to 466/2012 Resolution. Regarding the socio-demographic profile, the most prevalent ages and equally were those belonging to the groups 21-30 and 41-50 years old (24.2%). The brown color was reported in excess (84.8%) and 96.9% out of them had completed primary and secondary school or studying. 51.5% were married, 72.7% Catholic, 54.5% employed and 48.5% with income between one and two minimum wages. As for the sexual and reproductive characteristics, prevailed heterosexual (93.9%) who did not use condoms during sexual intercourse (72.7%). 51.5% had a previous history of Sexually Transmitted Infection (STI), and HPV the most prevalent STI (76.5%). 57.6% did not use contraception, 78.8% underwent examination Cancer Prevention Uterus (PCCU) with shorter time interval or equal to one year, 72.7% had no cases of Cervical Cancer in the family, 63.6% were multiparous and 97% were not vaccinated against HPV. DC identified good level of agreement between raters (κ=0.542), had a specificity of 77.8% and sensitivity of 25% when compared their results with PCR. Only the variable race showed a statistically significant association with CRP (p=0.042). DC had 100% acceptance amongst women in the sample, revealing the possibility of other experiments in using this method so that it proves as a viable technique. The DC positivity criteria were developed by nurses and these professionals also perform PCCU in Brazil, which means that DC can be an important complementary diagnostic method for the appreciation of these professional’s quality of examinations.

Keywords: gynecological examination, human papillomavirus, nursing, papillomavirus infections, uterine lasmsneop

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Authors: Reem Alajmi, Hind Al Harbi, Tahany Ayaad, Zainab Al Musawi

Abstract:

Hard ticks Hyalomma spp. (family: Ixodidae) are obligate ectoparasite in their all life stages on some domestic animals mainly camels and cattle. Ticks may lead to many economic and public health problems because of their blood feeding behavior. Also, they act as vectors for many bacterial, viral and protozoan agents which may cause serious diseases such as tick-born encephalitis, Rocky-mountain spotted fever, Q-fever and Lyme disease which can affect human and/or animals. In the present study, molecular identification of ticks that attack camels in Riyadh region, Saudi Arabia based on the partial sequence of mitochondrial 16s rRNA gene was applied. Also, the present study aims to detect natural infections of collected camel ticks with Rickessia spp. and Borelia spp. using PCR/hybridization of Citrate synthase encoding gene present in bacterial cells. Hard ticks infesting camels were collected from different camels located in a farm in Riyadh region, Saudi Arabia. Results of the present study showed that the collected specimens belong to two species: Hyalomma dromedari represent 99% of the identified specimens and Hyalomma marginatum which account for 1 % of identified ticks. The molecular identification was made through blasting the obtained sequence of this study with sequences already present and identified in GeneBank. All obtained sequences of H. dromedarii specimens showed 97-100% identity with the same gene sequence of the same species (Accession # L34306.1) which was used as a reference. Meanwhile, no intraspecific variations of H. marginatum mesured because only one specimen was collected. Results also had shown that the intraspecific variability between individuals of H. dromedarii obtained in 92 % of samples ranging from 0.2- 6.6%, while the remaining 7 % of the total samples of H. dromedarii showed about 10.3 % individual differences. However, the interspecific variability between H. dromedarii and H. marginatum was approximately 18.3 %. On the other hand, by using the technique of PCR/hybridization, we could detect natural infection of camel ticks with Rickettsia spp. and Borrelia spp. Results revealed the natural presence of both bacteria in collected ticks. Rickettsial spp. infection present in 29% of collected ticks, while 35% of collected specimen were infected with Borrelia spp. The valuable results obtained from the present study are a new record for the molecular identification of camel ticks in Riyadh, Saudi Arabia and their natural infection with both Rickettsia spp. and Borrelia spp. These results may help scientists to provide a good and direct control strategy of ticks in order to protect one of the most important economic animals which are camels. Also results of this project spotlight on the disease that might be transmitted by ticks to put out a direct protective plan to prevent spreading of these dangerous agents. Further molecular studies are needed to confirm the results of the present study by using other mitochondrial and nuclear genes for tick identification.

Keywords: Camel ticks, Rickessia spp. , Borelia spp. , mitochondrial 16s rRNA gene

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Authors: Suman Chaudhary, Rupinder K. Kanwar, Jagat R. Kanwar

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Azadirachta indica, also known as Neem belonging to the mahogany family is actively gaining interest in the era of modern day medicine due to its extensive applications in homeopathic medicine such as Ayurveda and Unani. More than 140 phytochemicals have been extracted from neem leaves, seed, bark and flowers for agro-medicinal applications. Among the various components, neem leaf protein (NLP) is currently the most investigated active ingredient, due to its immunomodulatory activities against tumor growth. However, these therapeutic ingredients of neem are susceptible to degradation and cannot withstand the drastic pH changes under physiological environment, and therefore, there is an urgent need of an alternative strategy such as a nano-delivery system to exploit its medicinal benefits. This study hypothesizes that guar gum (GG) derived biodegradable nano-carrier based encapsulation of NLP will improve its stability, specificity and sensitivity, thus facilitating targeted anti-cancer therapeutics. GG is a galactomannan derived from the endosperm of the guar beans seeds. Synthesis of guar nanocapsules (NCs) was performed using nanoprecipitation technique where the GG was encapsulated with NLP. Preliminary experiments conducted to characterize the NCs confirmed spherical morphology with a narrow size distribution of 30-40 nm. Differential scanning colorimetric analysis (DSC) validated the stability of these NCs even at a temperature range of 50-60°C which was well within the physiological and storage conditions. Thermogravimetric (TGA) analysis indicated high decomposition temperature of these NCs ranging upto 350°C. Additionally, Fourier Transform Infrared spectroscopy (FTIR) and the SDS-PAGE data acquired confirmed the successful encapsulation of NLP in the NCs. The anti-cancerous therapeutic property of this NC was tested on colon cancer cells (caco-2) as they are one of the most prevalent form of cancer. These NCs (both NLP loaded and void) were also tested on human intestinal epithelial cells (FHs 74) cells to evaluate their effect on normal cells. Cytotoxicity evaluation of the NCs in the cell lines confirmed that the IC50 for NLP in FHs 74 cells was ~2 fold higher than in caco-2 cells, indicating that this nanoformulation system possessed biocompatible anti-cancerous properties Immunoconfocal microscopy analysis confirmed the time dependent internalization of the NCs within 6h. Recent findings performed using Annexin V and PI staining indicated a significant increase (p ≤ 0.001) in the early and late apoptotic cell population when treated with the NCs signifying the role of NLP in inducing apoptosis in caco-2 cells. This was further validated using Western blot, Polymerase chain reaction (PCR) and Fluorescence activated cell sorter (FACS) aided protein expressional analysis which presented a downregulation of survivin, an anti-apoptotic cell marker and upregulation of Bax/Bcl-2 ratio (pro-apoptotic indicator). Further, both the NLP NC and unencapsulated NLP treatment destabilized the mitochondrial membrane potential subsequently facilitating the release of the pro-apoptotic caspase cascade initiator, cytochrome-c. Future studies will be focused towards granting specificity to these NCs towards cancer cells, along with a comprehensive analysis of the anti-cancer potential of this naturally occurring compound in different cancer and in vivo animal models, will validate the clinical application of this unprecedented protein therapeutic.

Keywords: anti-tumor, guar gum, nanocapsules, neem leaf protein

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Authors: Pedro M. Rodrigues, Claudia Raposo, Denise Schrama, Marco Cerqueira

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Farmed fish welfare is a very recent concept, widely discussed among the scientific community. Consumers’ interest regarding farmed animal welfare standards has significantly increased in the last years posing a huge challenge to producers in order to maintain an equilibrium between good welfare principles and productivity, while simultaneously achieve public acceptance. The major bottleneck of standard aquaculture is to impair considerably fish welfare throughout the production cycle and with this, the quality of fish protein. Welfare assessment in farmed fish is undertaken through the evaluation of fish stress responses. Primary and secondary stress responses include release of cortisol and glucose and lactate to the blood stream, respectively, which are currently the most commonly used indicators of stress exposure. However, the reliability of these indicators is highly dubious, due to a high variability of fish responses to an acute stress and the adaptation of the animal to a repetitive chronic stress. Our objective is to use comparative proteomics to identify and validate a fingerprint of proteins that can present an more reliable alternative to the already established welfare indicators. In this way, the culture conditions will improve and there will be a higher perception of mechanisms and metabolic pathway involved in the produced organism’s welfare. Due to its high economical importance in Portuguese aquaculture Gilthead seabream will be the elected species for this study. Protein extracts from Gilthead Seabream fish muscle, liver and plasma, reared for a 3 month period under optimized culture conditions (control) and induced stress conditions (Handling, high densities, and Hipoxia) are collected and used to identify a putative fish welfare protein markers fingerprint using a proteomics approach. Three tanks per condition and 3 biological replicates per tank are used for each analisys. Briefly, proteins from target tissue/fluid are extracted using standard established protocols. Protein extracts are then separated using 2D-DIGE (Difference gel electrophoresis). Proteins differentially expressed between control and induced stress conditions will be identified by mass spectrometry (LC-Ms/Ms) using NCBInr (taxonomic level - Actinopterygii) databank and Mascot search engine. The statistical analysis is performed using the R software environment, having used a one-tailed Mann-Whitney U-test (p < 0.05) to assess which proteins were differentially expressed in a statistically significant way. Validation of these proteins will be done by comparison of the RT-qPCR (Quantitative reverse transcription polymerase chain reaction) expressed genes pattern with the proteomic profile. Cortisol, glucose, and lactate are also measured in order to confirm or refute the reliability of these indicators. The identified liver proteins under handling and high densities induced stress conditions are responsible and involved in several metabolic pathways like primary metabolism (i.e. glycolysis, gluconeogenesis), ammonia metabolism, cytoskeleton proteins, signalizing proteins, lipid transport. Validition of these proteins as well as identical analysis in muscle and plasma are underway. Proteomics is a promising high-throughput technique that can be successfully applied to identify putative welfare protein biomarkers in farmed fish.

Keywords: aquaculture, fish welfare, proteomics, welfare biomarkers

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Authors: Tristan R. Fraser, Christopher D. Steadman, Christopher J. Boos

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Myopericarditis is an inflammation syndrome characterised by clinical diagnostic criteria for pericarditis, such as chest pain, combined with evidence of myocardial involvement, such as elevation of biomarkers of myocardial damage, e.g., troponins. It can rarely be a complication of therapeutics used for dysregulated immune-mediated diseases such as inflammatory bowel disease (IBD), for example, mesalazine. The infrequency of mesalazine-induced myopericarditis adds to the challenge in its recognition. Rapid diagnosis and the early introduction of treatment are crucial. This case report follows a 24-year-old professional footballer with a past medical history of ulcerative colitis, recently started on mesalazine for disease control. Three weeks after mesalazine was initiated, he was admitted with fever, shortness of breath, and chest pain worse whilst supine and on deep inspiration, as well as elevated venous blood cardiac troponin T level (cTnT, 288ng/L; normal: <13ng/L). Myocarditis was confirmed on initial inpatient cardiac MRI, revealing the presence of florid myocarditis with preserved left ventricular systolic function and an ejection fraction of 67%. This was a longitudinal case study following the progress of a single individual with myopericarditis over four acute hospital admissions over nine weeks, with admissions ranging from two to five days. Parameters examined included clinical signs and symptoms, serum troponin, transthoracic echocardiogram, and cardiac MRI. Serial measurements of cardiac function, including cardiac MRI and transthoracic echocardiogram, showed progressive deterioration of cardiac function whilst mesalazine was continued. Prior to cessation of mesalazine, transthoracic echocardiography revealed a small global pericardial effusion of < 1cm and worsening left ventricular systolic function with an ejection fraction of 45%. After recognition of mesalazine as a potential cause and consequent cessation of the drug, symptoms resolved, with cardiac MRI performed as an outpatient showing resolution of myocardial oedema. The patient plans to make a return to competitive sport. Patients suffering from myopericarditis are advised to refrain from competitive sport for at least six months in order to reduce the risk of cardiac remodelling and sudden cardiac death. Additional considerations must be taken in individuals for whom competitive sport is an essential component of their livelihood, such as professional athletes. Myopericarditis is an uncommon, however potentially serious medical condition with a wide variety of aetiologies, including viral, autoimmune, and drug-related causes. Management is mainly supportive and relies on prompt recognition and removal of the aetiological process. Mesalazine-induced myopericarditis is a rare condition; as such increasing awareness of mesalazine as a precipitant of myopericarditis is vital for optimising the management of these patients.

Keywords: myopericarditis, mesalazine, inflammatory bowel disease, professional athlete

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Authors: Maryam Nazari

Abstract:

COVID-19 is a respiratory disease triggered by the novel coronavirus, SARS-CoV-2, that has reached pandemic status today. Acute inflammation and immune cells infiltration into lung injuries result in multi-organ failure. The presence of other non-communicable diseases (NCDs) with systemic inflammation derived from COVID-19 may exacerbate the patient's situation and increase the risk for adverse effects and mortality. This pandemic is a novel situation and the scientific community at this time is looking for vaccines or drugs to treat the pathology. One of the biggest challenges is focused on reducing inflammation without compromising the correct immune response of the patient. In this regard, addressing the nutritional factors should not be overlooked not only as a matter of avoiding the presence of NCDs with severe infections but also as an adjunctive way to modulate the inflammatory status of the patients. Despite the pivotal role of nutrition in modifying immune response, due to the novelty of the COVID-19 disease, information about the effects of specific dietary agents is limited in this area. From the macronutrients point of view, protein deficiency (quantity or quality) has negative effects on the number of functional immunoglobulins and gut-associated lymphoid tissue (GALT). High biological value proteins or some amino acids like arginine and glutamine are well known for their ability to augment the immune system. Among lipids, fish oil has the ability to inactivate enveloped viruses, suppress pro-inflammatory prostaglandin production and block platelet-activating factors and their receptors. In addition, protectin D1, which is an Omega-3 PUFAs derivation, is a novel antiviral drug. So it seems that these fatty acids can reduce the severity and/or improve recovery of patients with COVID-19. Carbohydrates with lower glycemic index and fibers are associated with lower levels of inflammatory cytokines (CRP, TNF-α, and IL-6). Short-Chain Fatty acids not only exert a direct anti-inflammatory effect but also provide appropriate gut microbial, which is important in gastrointestinal issues related to COVID-19. From the micronutrients point of view, Vitamins A, C, D, E, iron, magnesium, zinc, selenium and copper play a vital role in the maintenance of immune function. Inadequate status in these nutrients may result in decreased resistance against COVID-19 infection. There are specific bioactive compounds in the diet that interact with the ACE2 receptor, which is the gateway for SARS and SARS-CoV-2, and thus controls the viral infection. Regarding this, the potential benefits of probiotics, resveratrol (a polyphenol found in grape), oleoylethanolamide (derived from oleic acid), and natural peroxisome proliferator-activated receptor γ agonists in foodstuffs (like curcumin, pomegranate, hot pepper) are suggested. Yet, it should be pointed out that most of these results have been reported in animal models and further human studies are needed to be verified.

Keywords: Covid-19, inflammation, nutrition, dietary agents

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Authors: M. D. Nirmani, K. L. N. Perera, G. H. Galhena

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Dengue has become one of the most important arbo-viral disease in all tropical and subtropical regions of the world. Aedes aegypti, is the principal vector of the virus, vary in both epidemiological and behavioral characteristics, which could be finely measured through DNA sequence comparison at their population level. Such knowledge in the population differences can assist in implementation of effective vector control strategies allowing to make estimates of the gene flow and adaptive genomic changes, which are important predictors of the spread of Wolbachia infection or insecticide resistance. As such, this study was undertaken to investigate the phylogenetic relationships of Ae. aegypti from Galle and Colombo, Sri Lanka, based on the ribosomal protein region which spans between two exons, in order to understand the geographical distribution of genetically distinct mosquito clades and its impact on mosquito control measures. A 320bp DNA region spanning from 681-930 bp, corresponding to the ribosomal protein, was sequenced in 62 Ae. aegypti larvae collected from Galle (N=30) and Colombo (N=32), Sri Lanka. The sequences were aligned using ClustalW and the haplotypes were determined with DnaSP 5.10. Phylogenetic relationships among haplotypes were constructed using the maximum likelihood method under Tamura 3 parameter model in MEGA 7.0.14 including three previously reported sequences of Australian (N=2) and Brazilian (N=1) Ae. aegypti. The bootstrap support was calculated using 1000 replicates and the tree was rooted using Aedes notoscriptus (GenBank accession No. KJ194101). Among all sequences, nineteen different haplotypes were found among which five haplotypes were shared between 80% of mosquitoes in the two populations. Seven haplotypes were unique to each of the population. Phylogenetic tree revealed two basal clades and a single derived clade. All observed haplotypes of the two Ae. aegypti populations were distributed in all the three clades, indicating a lack of genetic differentiation between populations. The Brazilian Ae. aegypti haplotype and one of the Australian haplotypes were grouped together with the Sri Lankan basal haplotype in the same basal clade, whereas the other Australian haplotype was found in the derived clade. Phylogram showed that Galle and Colombo Ae. aegypti populations are highly related to each other despite the large geographic distance (129 Km) indicating a substantial genetic similarity between them. This may have probably arisen from passive migration assisted by human travelling and trade through both land and water as the two areas are bordered by the sea. In addition, studied Sri Lankan mosquito populations were closely related to Australian and Brazilian samples. Probably this might have caused by shipping industry between the three countries as all of them are fully or partially enclosed by sea. For example, illegal fishing boats migrating to Australia by sea is perhaps a good mean of transportation of all life stages of mosquitoes from Sri Lanka. These findings indicate that extensive mosquito migrations occur between populations not only within the country, but also among other countries in the world which might be a main barrier to the successful vector control measures.

Keywords: Aedes aegypti, dengue control, extensive mosquito migration, haplotypes, phylogeny, ribosomal protein

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