Association of Cytokine IL-17, IL-4, IL-6, and IL-12 Gene Polymorphisms in Rheumatoid Arthritis Patients in a Tertiary Care Hospital in Bangladesh.

Rheumatoid arthritis (RA) is a chronic autoimmune inflammatory disease that involves cytokines in its pathogenesis. This study is aimed at investigating if gene polymorphisms in cytokines like IL-17, IL-4, IL-6, and IL-12 affect RA susceptibility and severity in the Bangladeshi population. This was a cross-sectional comparative study that included 40 diagnosed RA patients according to the American College of Rheumatology (ACR) criteria 2010, who were free from other rheumatological diseases, and 40 healthy subjects for comparison. The study used PCR-RFLP to determine the IL-17, IL-4, IL-6, and IL-12 cytokine gene polymorphisms. Patients had a mean age of 37.22 ± 6.70 years. Among the patients, 31 were female and 9 were male. The mean disease duration was 18.11 ± 7.39 months. The study found that rheumatoid arthritis patients with the IL-17F (7488 A/G) polymorphism with GG genotype (P = 0.006, OR = 8.56, 95% CI = 1.77 - 41.33) and IL-12B (1188 A/C) polymorphism with AC (P = 0.012, OR = 3.69, 95% CI = 1.43 - 9.53) and CC (P = 0.013, OR = 7.58, 95% CI = 1.56 - 36.88) genotypes were significantly associated with disease risk. Furthermore, patients with the IL-17F (7488) GG genotype and IL-12B (1188) AC and CC genotypes had higher rheumatoid arthritis disease severity and activity parameters. The study found no significant association between polymorphisms involving IL-4 (590 C/T) and IL-6 (174 G/C) genes and rheumatoid arthritis disease risk in the Bangladeshi population. Gene polymorphisms in cytokines IL-17F (7488 A/G) and IL-12B (1188 A/C) can predict disease susceptibility and severity in Bangladeshi rheumatoid arthritis patients.

Hypothalamic-Pituitary-Adrenal Axis Activity in SARS-CoV-2 Infected Noncritically Ill Hospitalized Patients.

Objectives: This study determined the baseline hypothalamic-pituitary-adrenal axis hormonal levels and their associated factors in noncritically ill hospitalized patients with coronavirus disease 2019 (COVID-19). Methodology: This cross-sectional study was carried out in 91 noncritical RT-PCR-confirmed COVID-19 patients (aged 18 to 65 years) recruited consecutively from the COVID unit of two tertiary care hospitals over a period of six months. After the screening, relevant history and physical examinations were done, and blood was drawn between 07:00 am to 09:00 am in a fasting state to measure serum cortisol and plasma adrenocorticotropic hormone (ACTH) by chemiluminescent microparticle immunoassay. Results: Of 91 patients, 54, 26, and 11 had mild, moderate, and severe COVID-19, respectively. Median values of serum cortisol (p = 0.057) and plasma ACTH (p = 0.910) were statistically similar among the severity groups. Considering a cortisol cut-off of 276 nmol/L (<10 µg/dL), the highest percent of adrenal insufficiency was present in severe (27.3%), followed by mild (25.9%) and least in the moderate (3.8%) COVID-19 cases. Using the cortisol/ACTH ratio >15, only 6.6% had enough reserve. Conclusions: The adrenocortical response was compromised in a significant percentage of noncritically ill hospitalized patients with COVID-19, with the highest percentage of adrenal insufficiency present in severely infected cases. The HPA axis parameters of serum cortisol, plasma ACTH and cortisol/ACTH were similar across the severity of noncritical patients with COVID-19.

Sex-Specific Total Testosterone and Dehydroepiandrosterone Sulfate Status in Noncritically Ill Hospitalized Patients with Coronavirus Disease 2019: A Cross-Sectional Study.

BACKGROUND: In individuals with coronavirus disease 2019 (COVID-19), male subjects have consistently been linked to poor severity and prognosis. Data on sex hormones in non-critical COVID-19-infected patients are scarce. The aim of this study was to assess the status of total testosterone (TT) and dehydroepiandrosterone sulfate (DHEAS) among noncritical patients with COVID-19 according to sex and their associations with clinical and biochemical features. MATERIALS AND METHODS: This cross-sectional observational study was done in the COVID-19 unit of a University hospital during the period of September 2021 to February 2022 among 91 adults (18-65 years) with reverse transcriptase- polymerase chain reaction confirmed noncritical COVID-19 patients. Blood was drawn by venipuncture before receiving steroids between 07:00 to 09:00 a.m. in a fasting state to measure serum TT and DHEAS by chemiluminescent microparticle immunoassay. Diagnosis and classification of COVID-19 were done according to World Health Organization's interim guidance. Age- and sex-specific laboratory reference values were used to classify the TT and DHEAS status of the patients. RESULTS: Only three males (8.1%) had low TT and the rest had normal TT. On the other hand, 15 (27.8%) of the females had high TT with normal levels in the rest. Similarly, 11 (29.7%) males had low DHEAS. Females had low, normal, and high DHEAS in four (7.4%), 48 (88.9%), and two (3.7%) cases respectively. Males with moderate severity of COVID-19 had significantly lower DHEAS (post hoc P=0.038) than the mild group. Both TT (P=0.008) and DHEAS (P=0.023) significantly correlated with neutrophils/lymphocytes ratio and only DHEAS with platelets/lymphocytes ratio (P=0.044) in males. In females, TT significantly correlated with serum sodium (P=0.034). CONCLUSION: In noncritical COVID-19 patients, substantial gender variations in TT and DHEAS were detected and correlated with severity markers in males.

Measuring the Effectiveness of COVID-19 Vaccines Used during a Surge of the Delta Variant of SARS-CoV-2 in Bangladesh: A Test-Negative Design Evaluation.

BACKGROUND: From May to December 2021, Bangladesh experienced a major surge in the Delta variant of SARS-CoV-2. The earlier rollout of several vaccines offered the opportunity to evaluate vaccine effectiveness against this variant. METHODS: A prospective, test-negative case-control study was conducted in five large hospitals in Dhaka between September and December 2021. The subjects were patients of at least 18 years of age who presented themselves for care, suffering COVID-like symptoms of less than 10 days' duration. The cases had PCR-confirmed infections with SARS-CoV-2, and up to 4 PCR test-negative controls were matched to each case, according to hospital, date of presentation, and age. Vaccine protection was assessed as being the association between the receipt of a complete course of vaccine and the occurrence of SARS-CoV-2 disease, with symptoms beginning at least 14 days after the final vaccine dose. RESULTS: In total, 313 cases were matched to 1196 controls. The genotyping of case isolates revealed 99.6% to be the Delta variant. Receipt of any vaccine was associated with 12% (95% CI: -21 to 37, p = 0.423) protection against all episodes of SARS-CoV-2. Among the three vaccines for which protection was evaluable (Moderna (mRNA-1273); Sinopharm (Vero Cell-Inactivated); Serum Institute of India (ChAdOx1 nCoV-19)), only the Moderna vaccine was associated with significant protection (64%; 95% CI: 10 to 86, p = 0.029). Protection by the receipt of any vaccine against severe disease was 85% (95% CI: 27 to 97, p = 0.019), with protection estimates of 75% to 100% for the three vaccines. CONCLUSIONS: Vaccine protection against COVID-19 disease of any severity caused by the Delta variant was modest in magnitude and significant for only one of the three evaluable vaccines. In contrast, protection against severe disease was high in magnitude and consistent for all three vaccines. Because our findings are not in complete accord with evaluations of the same vaccines in more affluent settings, our study underscores the need for country-level COVID-19 vaccine evaluations in developing countries.

Association of interferon gamma inducible protein-10, monocyte chemoattractant protein-1, macrophage inflammatory protein-1 alpha, interleukin-6, and rs12252 single nucleotide polymorphism of interferon-induced transmembrane protein-3 gene with the severity of COVID-19 infection.

Background: Evaluating the serum levels of IP-10, MCP-1, MIP-1α, and IL-6 and genotyping of rs12252 SNP of IFITM3 gene among different categories of COVID-19 patients might aid in understanding the pathogenesis of COVID-19 and contribute to developing disease-specific biomarkers and therapeutic strategies. Methods: This is a cross-sectional study involving a total of 84 COVID-19 patients confirmed by positive RT-PCR and 28 healthy controls. COVID-19 patients were recruited from the intensive care unit (ICU) and COVID unit of Bangabandhu Sheikh Mujib Medical University, Shahbag, Dhaka. COVID-19 patients were categorized into moderate, severe, and critically ill groups according to the World Health Organization classification. The serum IP-10, MCP-1, and MIP-1α levels were measured by cytometric bead array assay by flow cytometry, and serum IL-6 level was detected by the chemiluminescence method. rs12252 SNP of the IFITM3 gene was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR RFLP). Results: The serum IP-10, MCP-1, MIP-1α, and IL-6 levels among critically ill COVID-19 patients were significantly higher than that in patients with moderate disease and healthy controls (p < 0.001). Genotype distribution for rs12252 (42 T/C) SNP of the IFITM3 gene between the different groups of COVID-19 patients and healthy controls showed that CC genotype was statistically associated with disease severity (p < 0.001). Conclusions: IP-10 and MCP-1, MIP-α, IL-6, and CC genotype of rs12252 (42 T/C) SNP of IFITM3 gene are associated with COVID-19 severity.

Isolation and characterisation of carbapenem-resistant Pseudomonas aeruginosa from hospital environments in tertiary care hospitals in Dhaka, Bangladesh.

OBJECTIVES: Increasing evidence of carbapenem-resistant Pseudomonas aeruginosa (CRPA) infection in healthcare facilities poses an alarming threat to public health. There is little evidence on the occurrence of this organism in Bangladeshi hospitals. METHODS: We collected 117 environmental swab samples from two tertiary care hospitals in Dhaka, Bangladesh and tested for Pseudomonas species by nonselective enrichment of swabs followed by plating on Cetrimide agar. We confirmed the isolates as P. aeruginosa by API 20NE test and polymerase chain reaction Polymerase Chain Reaction (PCR) for 16S rRNA gene. We analysed P. aeruginosa isolates for susceptibility against 15 clinically important antibiotics and tested the carbapenem-resistant isolates for metallo ß-lactamase (MBL). All CRPA isolates were characterised for carbapenem-resistant genes, virulence genes and biofilm formation genes. RESULTS: Of 117 swab samples, 82 (70%) were tested positive for P. aeruginosa. All P. aeruginosa isolates were multidrug-resistant, and 39% (n = 32) of isolates were CRPA. Around 56% (n = 18) of CRPA were MBL-producing; 22% (n = 7) of isolates were positive for carbapenemase gene blaNDM followed by 16% (n = 5) for blaVIM and 13% (n = 4) for blaIMP. Sequencing identified these genes as blaNDM-1, blaIMP-13, blaVIM-2 variants. Based on optical density values, 94% (n = 30) of CRPA isolates were capable of producing biofilms. All CRPA isolates (n = 32) were positive for at least 1 of 6 biofilm-associated genes and 4 of 12 virulence genes tested in the study. CONCLUSION: Hospital environments in Bangladesh are contaminated with highly virulent CRPA, which might be a potential source of hospital-acquired infections, accentuating the need for strengthening hospital infection control programs.

Endometrial cytokine expression from clinically suspected genital tuberculosis patients at tertiary care hospitals in Dhaka.

OBJECTIVE: The objective of this study was to measure gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α) expression in endometrial tissue and/or aspirate from suspected genital tuberculosis patients with ectopic pregnancy and infertility in Bangladesh. METHODOLOGY: A total 78 women of clinically suspected genital tuberculosis patients were enrolled as study population. These patients underwent manual vaccum aspiration (MVA) procedure, and endometrial tissues and/or aspirates were collected. Ziehl -Neelsen staining (Z-N staining) and Lowen-Stein Jensen (L-J) culture were done to detect Mycobacterium. The study participants were categorized as genital tuberculosis positive cases, genital tuberculosis negative cases and presumptive for tuberculosis cases based on the case definition used in this study. TNF-α and IFN-γ were measured by ELISA. Statistical analysis was done using SPSS (version-22). RESULTS: Out of 78 participants, pro-inflammatory cytokines IFN-γ and TNF-α were significantly increased in TB positive patients than TB negative patients (p < 0.05). IFN-γ value of TB positive patients (41.26 ± 41.05) was higher than TB negative (22.94 ± 44.51) patients. TNF-α value (44.31 ± 64.22) of TB positive patients was higher than TB negative (15.86 ± 41.45) patients. IFN-γ and TNF-α value of presumptive for tuberculosis cases were not statistically significant. According to ROC analysis, cut off value for IFN-γ was 23.5 and for TNF-α was 10 with highest sensitivity and specificity of 66.7%, 89.3%, and 66.7% and 73.1% respectively. CONCLUSION: IFN-γ and TNF-α were significantly higher in TB positive patients and it may act as a potential biomarker for diagnosis of genital tuberculosis.

Molecular epidemiology of toxigenic Clostridioides difficile isolates from hospitalized patients and the hospital environment in Dhaka, Bangladesh.

Epidemiology of Clostridioides difficile (syn. Clostridium difficile) infection (CDI) in Bangladesh is poorly understood. This study assessed the epidemiology of CDI in hospitalized patients and hospital environmental contamination of toxigenic C. difficile at two large urban Bangladesh hospitals. This 12-month prospective observational cohort study collected stool samples from adults with diarrhea and recent antimicrobial exposure during 2017. Environmental samples were collected by swabbing surfaces of hospital common areas. Samples underwent toxigenic culture. C. difficile isolates were tested for toxins A and B and PCR-ribotyped. Of 208 stool samples, 18 (8.7%) were positive for toxigenic C. difficile. Of 400 environmental samples, 45 (11%) were positive for toxigenic C. difficile. Ribotypes present in ≥10% of stool isolates were 017 (38%), 053-163 (13%), and a novel ribotype (FP435 [13%]). Common ribotypes in environmental isolates were 017 (22%), 053-163 (11%), 106 (24%). This is the first report describing current epidemiology of CDI in at risk hospitalized adult patients in Bangladesh.

Association of single nucleotide polymorphism at PNPLA3 with fatty liver, steatohepatitis, and cirrhosis of liver.

BACKGROUND/PURPOSE: The aim of this study was to determine the association of single nucleotide polymorphism (SNP) in patatin-like phospholipase domain-containing 3 (PNPLA3) at I148 with histological severity of non-alcoholic fatty liver disease (NAFLD). METHODS: Patients were selected for the study if they had histological evidence of NAFLD and clinical evidence of non-alcoholic steatohepatits (NASH) cirrhosis. We included 50 NASH cirrhosis, 99 patients of NAFLD including 36 non-NASH fatty liver (NNFL) along with 63 NASH and 75 healthy controls. PNPLA3 genotyping was done by real-time PCR using a Taqman assay for rs738409. RESULTS: CC, CG, and GG frequencies were 45 (60.0%)/27 (36.0%)/3 (4.0%) in healthy control, 19 (52.8%)/14 (38.9%)/ 3 (8.3%) in NNFL, 18 (28.6%)/29 (46.0%)/16 (25.4%) in NASH, and 7 (14.6%), 25 (52.1%), 16 (33.3%) in cirrhosis. The frequency of G allele was significantly higher (62.6%) in NAFLD than in healthy control. The GG genotype had 20.25 times odds of NAFLD. The GG genotype had 6.53 times odds of having NASH. HOMA-IR > 1.6 had 3.81 times odds of having NASH. Regression analysis revealed that G allele odds of having cirrhosis was 3.9 times compared to C. The G allele was also significantly associated with steatosis, lobular inflammation, NAFLD activity score, and fibrosis. CONCLUSION: PNPLA3 genotype showed an association with NAFLD, NASH, fibrosis, and cirrhosis.

Abdominal lump with a rare pathology.

Actinomycosis is one of the rarest pathologies causing intra-abdominal lump in the normal population. Diagnosis is often difficult because of the least suspicion and failure in the pre-operative diagnosis even by taking the aid of modern imaging techniques like Ultrasonogram, CT scan or MRI. Even FNAC may be misleading because of the technical problems and proper interpretation of cytology. Diagnosis is usually made during laparotomy by an experienced Surgeon and finally by the histopathological examination of the resected specimen. We have come across three patients with actinomycosis presenting with intra-abdominal lumps, two as appendicular lumps and 3rd one as a lump of left colonic origin in our institute over the last two years. Ultrasonogram examination revealed lump of gut origin in all the three cases. FNAC in one case revealed the lump as a low grade sarcoma. Laparotomy, excision and or resection of the lumps along with the segment of involved gut with subsequent histopathology revealed these cases as actinomycosis. We report these cases for its extreme rarity and a puzzling pre-operative diagnostic dilemma.

Inhibitory effect of arsenic on aerobic gut flora in rat.

An in vivo study was carried on rats to see the influence of arsenic on aerobic gut flora. A significant inhibition of gut flora was observed after 2 weeks of administration of arsenic (1 mg/L) ad libitum with a decrease in stool arsenic level and increase in liver arsenic level. However, this inhibitory effect of arsenic on gut flora was not observed in presence of vitamin E (1 mg/day) or selenium (0.4 microg/day). Pretreatment with streptomycin (500 mg twice daily) showed similar results. Rats that received folic acid (200 microg/day) showed inhibition of gut floral count but there were decreased liver arsenic level.

Ethanol production from xylose by recombinant Saccharomyces cerevisiae expressing protein engineered NADP+-dependent xylitol dehydrogenase.

Effects of reversal coenzyme specificity toward NADP+ and thermostabilization of xylitol dehydrogenase (XDH) from Pichia stipitis on fermentation of xylose to ethanol were estimated using a recombinant Saccharomyces cerevisiae expressing together with a native xylose reductase from P. stipitis. The mutated XDHs performed the similar enzyme properties in S. cerevisiae cells, compared with those in vitro. The significant enhancement(s) was found in Y-ARSdR strain, in which NADP+-dependent XDH was expressed; 86% decrease of unfavorable xylitol excretion with 41% increased ethanol production, when compared with the reference strain expressing the wild-type XDH.

The positive effect of the decreased NADPH-preferring activity of xylose reductase from Pichia stipitis on ethanol production using xylose-fermenting recombinant Saccharomyces cerevisiae.

We focused on the effects of a mutation of xylose reductase from Pichia stipitis (PsXR) on xylose-to-ethanol fermentation using recombinant Saccharomyces cerevisiae transformed with PsXR and PsXDH (xylitol dehydrogenase from P. stipitis) genes. Based on inherent NADH-preferring XR and several site-directed mutagenetic studies using other aldo-keto reductase enzymes, we designed several single PsXR mutants. K270R showing decreased NADPH-preferring activity without a change in NADH-preferring activity was found to be a potent mutant. Strain Y-K270R transformed with K270R PsXR and wild-type PsXDH showed a 31% decrease in unfavorable xylitol excretion with 5.1% increased ethanol production as compared to the control in the fermentation of 15 g l(-1) xylose and 5 g l(-1) glucose.

Thermostabilization of Pichia stipitis xylitol dehydrogenase by mutation of structural zinc-binding loop.

Xylitol dehydrogenase from Pichia stipitis (PsXDH) is one of the key enzymes for the bio-ethanol fermentation system from xylose. Previously, we constructed the C4 mutant (S96C/S99C/Y102C) with enhanced thermostability by introduction of structural zinc. In this study, for further improvement of PsXDH thermostability, we constructed the appropriate structural zinc-binding loop by comparison with other polyol dehydrogenase family members. A high thermostability of PsXDH was obtained by subsequent site-directed mutagenesis of the structural zinc-binding loop. The best mutant in this study (C4/F98R/E101F) showed a 10.8 degrees C higher thermal transition temperature (T(CD)) and 20.8 degrees C higher half denaturation temperature (T(1/2)) compared with wild-type.

Construction of various mutants of xylose metabolizing enzymes for efficient conversion of biomass to ethanol.

We applied protein engineering to construct an efficient biomass-ethanol conversion system using Saccharomyces cerevisiae. Intercellular redox imbalance caused by the different coenzyme specificity of xylose reductase (XR) and xylitol dehydrogenase (XDH) has been thought to be one of the main factors of xylitol excretion. Introduction of NADH-dependant XR generated in this study reduced the xylitol excretion probably because of maintaining the intercellular redox balance. Ethanol fermentation was measured in batch culture under anaerobic conditions. The best strain R276H produced a maximum of 5.94 g/l ethanol with yield of 0.43 g/g from 5 g glucose/l plus 15 g xylose/l.

Site-directed mutagenesis of a yeast gene for improvement of enzyme thermostability.

Enzyme stability is one of the critical factors to construct an efficient biological conversion system. Xylitol dehydrogenase (XDH) from Pichia stipitis is one of the key enzymes for bio-ethanol fermentation system from xylose. Previously, we tried to improve thermostability of XDH by introduction of structural zinc into the enzyme and successfully obtained a mutant, named C4 mutant, with an increased unfolding temperature (J. Biol. Chem., 280:10340-10349, 2005). We focused on further improvement of the thermostability of XDH in this study and employed subsequent site directed mutagenesis in structural zinc binding region for stabilizing the structural zinc binding loop. Two variants (C4/F98R and C4/E101F) showed higher thermostability than C4 mutant judged by thermal inactivation of enzyme activity and thermal transition temperature.