Biological properties of TW01 cells expressing latent membrane protein-1 gene of EBV-derived from nasopharyngeal carcinoma cells at different stages of malignancy

Background and Aim: Epstein—Barr virus (EBV), a human gammaherpesvirus is intimately associated with nasopharyngeal carcinoma (NPC), with the incidence of the virus detected in malignant tissues being close to 100% in NPC endemic areas. The viral latent gene, latent membrane protein 1 (LMP1), has all the typical characteristics of an oncogene and extensive studies have shown beyond doubt its abilities in cellular transformation giving rise to malignant phenotypes. The present study compares the gene sequence and biological properties of LMP1 gene derived from two patients with different stages of NPC – one presented with dysplastic, pre-malignant lesion and the other with malignant lesion. Methods: The sequences of the LMP1 genes derived from pre-malignant (NORLMP1) and malignant (NPCLMP1) tissues were compared. NORLMP1 and NPCLMP1 were expressed in TW01 cells and their biological properties conferred were assessed in terms of in vitro invasion capability, ability to resist apoptosis and regulation of E-cadherin (CDH1) expression. Results: NORLMP1 and NPCLMP1, possessed numerous amino acid changes with respect to the wild-type B95.8 LMP1. Both LMP1 variants demonstrated distinct biological properties when expressed in TW01 cells. NORLMP1-expressing TW01 cells exhibited more aggressive phenotypes than the NPCLMP1-expressing counterpart in terms of invasive ability, resistance to stimuli-induces apoptosis and regulation of CDH1 expression. Conclusion: It could be inferred from the present study that LMP1 genes derived from two patients with different stages of NPC differed in their gene sequences that manifested in distinct biological properties.Обоснование и цель: вирус Эпштейна—Барр (ВЭБ), герпесвирус человека, ассоциирован с развитием рака носоглотки (РНГ): вирус выявляют в малигнизированной ткани практически в 100% случаев.  Ген, кодирующий латентный мембранный белок 1 (LMP , обладает типичными характеристиками онкогена; в ряде исследований продемонстрирована его способность вызывать трансформацию клеток и развитие злокачественного фенотипа. В данной работе проведено сравнение нуклеотидной последовательности генов LMP , выделенных из ткани РНГ двух больных (с дисплазией/предраком и РНГ ІІ стадии соответственно) и биологических свойств клеток, трансфецированных этими генами. Методы: сравнивали нуклеотидные последовательности генов LMP из образцов предраковой (NORLMP и опухолевой (NPCLMP тканей и соответственно аминокислотные последовательности кодируемых ими белков. Гены NORLMP и NPCLMP встраивали в клетки ВЭБ-отрицательной линии TW01 и исслеовали иологические свойства трансфецированных клеток, в частности их способность к инвазивному росту in, устойчивость к апоптозу и экспрессии Е-кадгерина (CDH1). Результаты: оба белка — NORLMP1 и NPCLMP1 — значительно отличались по аминокислотной послеовательности от елка LMP1 дикого типа (B95,8), и клетки линии TW01, в которых они кспрессировались, различались по иологическим свойствам. Клетки TW01, экспрессирующие NORLMP1, обладали более злокачественным фенотипом, чем таковые, экспрессирующие NPCLMP1, по показателям инвазивной спосоности, устойчивости к апоптозу и регуляции кспрессии CDH1. Выводы: полученные данные свидетельствуют о том, что гены LMP , выделенные из образцов биопсии двух больных с различными стадиями РНГ, отличаются по нуклеотидной последовательности, что проявляется в различиях биологических свойств клеток, трансфецированных соотвествующими генами

Health outcomes, pathogenesis and epidemiology of severe acute malnutrition (HOPE-SAM): rationale and methods of a longitudinal observational study

Introduction: Mortality among children hospitalised for complicated severe acute malnutrition (SAM) remains high despite the implementation of WHO guidelines, particularly in settings of high HIV prevalence. Children continue to be at high risk of morbidity, mortality and relapse after discharge from hospital although long-term outcomes are not well documented. Better understanding the pathogenesis of SAM and the factors associated with poor outcomes may inform new therapeutic interventions. Methods and analysis: The Health Outcomes, Pathogenesis and Epidemiology of Severe Acute Malnutrition (HOPE-SAM) study is a longitudinal observational cohort that aims to evaluate the short-term and long-term clinical outcomes of HIV-positive and HIV-negative children with complicated SAM, and to identify the risk factors at admission and discharge from hospital that independently predict poor outcomes. Children aged 0–59 months hospitalised for SAM are being enrolled at three tertiary hospitals in Harare, Zimbabwe and Lusaka, Zambia. Longitudinal mortality, morbidity and nutritional data are being collected at admission, discharge and for 48 weeks post discharge. Nested laboratory substudies are exploring the role of enteropathy, gut microbiota, metabolomics and cellular immune function in the pathogenesis of SAM using stool, urine and blood collected from participants and from well-nourished controls

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

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

Clarithromycin resistance and point mutations in the 23S rRNA gene in Helicobacter pylori isolates from Malaysia

OBJECTIVE: To determine the prevalence of primary clarithromycin resistance amongst Helicobacter pylori (H. pylori) strains in Malaysian patients with gastroduodenal diseases, by using restriction fragment length polymorphism (RFLP) in domain V of 23S rRNA. METHODS: Gastric biopsies were obtained from H. pylori positive patients undergoing gastroscopy. DNA extraction was followed by PCR amplification using the primers Hp23-1 and Hp23-2 flanking a region of 425bp within the bacterial 23S rRNA peptidyltranferase (Hp23S fragment). Analysis of the 23S rRNA gene mutations is based on the generation of restriction sites for two restriction enzymes: BbsI and BsaI, which correspond to the base substitutions characteristic of clarithromycin resistance from A to G at positions 2142 and 2143, respectively. RESULTS: Gastric biopsy samples were obtained from 107 patients. A fragment of size 425bp corresponding to that expected from amplification of domain V of 23S rRNA was PCR-amplified from only 105 samples. The amplicon was subsequently subjected to restriction by BbsI and BsaI. Only 1 sample (0.95%) had the BbsI mutation (base substitution at A2142G) and 2 samples (1.90%) the BsaI mutation (base substitution at A2143G). Thus 3 of 105 (2.9%) samples harbored clarithromycin resistant strains. CONCLUSION: In our experience, PCR-RFLP is a rapid and precise method to detect the resistance of H. pylori to clarithromycin. Using this method, a low prevalence of clarithromycin resistance was detected in our local Malaysian strains. This augurs well for the continued use of clarithromycin as a first line drug in the treatment and eradication of H. pylori infection

Sensitization profiles of Malaysian and Singaporean subjects to allergens from Dermatophagoides pteronyssinus and Blomia tropicalis

10.1159/000074302International Archives of Allergy and Immunology1323215-220IAAI

Detection of Epstein-Barr virus DNA in nasopharyngeal carcinoma using a non-radioactive digoxigenin-labelled probe

The presence of Epstein Barr virus (EBV) DNA in biopsies from the post-nasal space (PNS) of patients suspected of nasopharyngeal carcinoma (NPC) was detected by in situ cytohybridization with an EBV DNA probe labelled with the novel labelling compound digoxigenin. The digoxigenin probe was hybridised to cryostat sections of NPC biopsies and subsequently detected by an enzyme immunoassay procedure. It was found that in situ cytohybridization using the digoxigenin probe was much more rapid and sensitive (96 h compared to five weeks) than the current method of using 3H-labelled probe. Using the digoxigenin EBV probe, it was found that in all the eighteen NPC biopsies tested, EBV DNA was detected in malignant epithelial cells and infiltrating lymphocytes. EBV DNA was also detected in some normal epithelial cells in these NPC biopsies. EBV DNA was not detected in epithelial cells of non-malignant biopsies

Relationship between serial serum neutrophil-lymphocyte ratio, cardiovascular mortality, and all-cause mortality in Chinese peritoneal dialysis patients

Background It is believed that the excessive cardiovascular (CV) burden of patients on peritoneal dialysis (PD) is closely associated with chronic inflammation. Neutrophil-lymphocyte ratio (NLR) is an inflammatory marker that was shown to correlate with CV outcomes. However, little is known about the significance of serial monitoring of serum NLR. We aimed to determine the prognostic value of serial NLR on all-cause mortality and CV mortality in PD patients. Methods Serial measurement of NLR was obtained from 225 incident PD patients in a single center, with each measurement one year apart. Patients were divided into two groups (‘high’ versus ‘low’) by the median value of NLR. The primary and secondary outcome measure was all-cause and CV mortality respectively. Results After a median of follow up for 43.9 months, patients with lower baseline NLR demonstrated a higher survival rate (p=0.01). Patients with persistently high NLR values on serial measurement had the lowest survival rate (p=0.03). Multivariate Cox regression showed that this group of patients had significantly higher all-cause mortality (HR 1.74, 95%CI 1.09 to 2.79, p=0.02). However, the NLR failed to demonstrate a statistically significant relationship with CV mortality. Conclusions While baseline NLR was an independent predictor of all-cause mortality in PD patients, persistent elevation in NLR appeared to further amplify the risk. Regular monitoring of serial serum NLR may enable early identification of patients who are at risk of adverse outcome