Markers of inflammasome activation in Coronavirus disease 2019 (COVID-19) Egyptian patients: Impact on disease severity.

There are conflicting data regarding the relationship between coronavirus disease 2019 (COVID-19) severity and Caspase-1 (Casp-1), interleukin-1ß (IL-1ß), and IL-18. Our study sought to quantify the levels of IL-18, IL-1ß, and Casp-1 as indicators for inflammasome activation in COVID-19 patients at Assiut University Hospitals and to correlate their levels with parameters of disease severity in COVID-19 patients. Serum levels of Casp-1, IL-1ß and IL-18 were measured in 63 COVID-19 patients and 26 normal controls by an enzyme linked immunosorbent assay (ELISA). Also, arterial blood gas analysis and laboratory parameters including hemoglobin, platelets, lymphocyte count, liver function test, kidney function test, C-reactive protein (CRP), D-dimer, ferritin and LDH were estimated. Serum levels of Casp-1, IL-1ß and IL-18 were significantly higher in the COVID-19 group as compared to controls (p= 0.04, p=0.001 and p=0.03, respectively). Although the three markers were higher in the severe group, yet only IL-1ß showed a significant difference as compared to the non-severe group (p=0.04). IL-18 had significant positive correlations with CRP and ferritin (p = 0.04 and p = 0.02, respectively). IL-1ß was positively correlated with alanine aminotransferase. Casp-1 had significant positive correlations with CRP and lactate dehydrogenase (p=0.045 and p=0.001, respectively). Patients showed weak positive correlations between serum level of Casp-1 and each of IL-1ß and IL-18. Also, a strong positive correlation was found between IL-1ß and IL-18 (p < 0.0001). In conclusion, inflammasome activation was a hallmark in COVID-19 patients. The markers of activation were positively correlated with many parameters of inflammation, may suggest their important roles in the pathophysiology of the disease and its progression. IL-1ß was the only marker to be correlated with disease severity and therefore may be suggested as a potential marker for identifying severe COVID-19 patients.

Impact of Geographical Location on the Gut Microbiota Profile in Egyptian Children with Type 1 Diabetes Mellitus: A Pilot Study.

Purpose: To investigate the compositional and functional characteristics of T1DM-associated gut microbiota in two Egyptian cities and to study the geographical locality effects. Patients and Methods: This case-control study included 32 children with controlled T1DM and 16 controls, selected from two different regions of Egypt. The gut microbiota of both diabetic and control children was analyzed through 16S rRNA gene sequencing; this was done using the Illumina MiSeq platform. Results: Consistent findings among the diabetic children included significantly lower alpha diversity than the control children, as well as a lower mean Firmicutes/Bacteroidetes (F/B) ratio, and reduced proportions of Firmicutes and the genera Prevotella and Ruminococcus. In the diabetic children, there were also significantly enriched representations of Actinobacteria, Bacteroidetes, and Proteobacteria and the genera Lactobacilli, Bacteroides, and Faecalibacterium. When comparing the two diabetic groups, the Ismailia group (IsDM) was found to have a significantly higher F/B ratio and diversity indices, with resultant differences at the functional level. Conclusion: There are a number of consistent changes in the microbiota profile characterizing the diabetic groups irrespective of the geographical location including significantly lower alpha diversity, mean Firmicutes/ Bacteroidetes (F/B) ratio, and reduced proportions of Firmicutes and genera Prevotella and Ruminococcus. There are also significantly enriched representations of Actinobacteria, Bacteroidetes, and Proteobacteria and genera Lactobacilli, Bacteroides, and Faecalibacterium pointing to the greater driving power of the disease.

COVID-19 Infection in Patients with Comorbidities: Clinical and Immunological Insight.

AIM: Our study's objectives were to study the clinical and laboratory characteristics that may serve as biomarkers for predicting disease severity, IL-10 levels, and frequencies of different T cell subsets in comorbid COVID-19 patients. METHODS: Sixty-two hospitalized COVID-19 patients with comorbidities were assessed clinically and radiologically. Blood samples were collected to assess the T lymphocyte subsets by flow cytometry and IL-10 levels by ELISA. RESULTS: The most common comorbidities observed in COVID-19 patients were diabetes mellitus (DM), hypertension, and malignancies. Common symptoms and signs included fever, cough, dyspnea, fatigue, myalgia, and sore throat. CRP, ferritin, D dimer, LDH, urea, creatinine, and direct bilirubin were significantly increased in patients than controls. Lymphocyte count and CD4+ and CD8+ T-cells were significantly decreased in comorbid COVID-19 patients, and CD25 and CD45RA expression were increased. CD4+ and CD8+ regulatory T cells (Tregs) and IL-10 levels were significantly decreased in patients. CONCLUSIONS: Many parameters were found to be predictive of severity in the comorbid patients in our study. Significant reductions in the levels and activation of CD4+ and CD8+ T-cells were found. In addition, CD4+ and CD8+ Tregs were significant decreased in patients, probably pointing to a prominent role of CD8+ Tregs in dampening CD4+ T-cell activation.

Altered Regulatory B Cell Subsets in Children with Type 1 Diabetes Mellitus.

B regulatory cells (Breg) refer to characteristic subsets of B cells that generally exert anti-inflammatory functions and maintain peripheral tolerance mainly through their ability to secrete interleukin-10 (IL10). Dysregulation in the function of Breg cells was reported in several autoimmune diseases. However, the relation between Breg and children with type 1 diabetes (T1D) is poorly understood. Thus, this study is aimed at determining whether Breg cells play a role in T1D in children or not, so we hypothesized that an altered phenotype of B cell subsets is associated with T1D in children. Children with T1D (n = 29) and control children with normal blood glucose levels (n = 14) were recruited. The percentages of different circulating IL10-producing Breg subsets, including B10, immature transitional, and plasmablasts were determined using flow cytometry analysis. Furthermore, the association between different IL10-producing B cells and patient parameters was investigated. The percentage of circulating IL10+CD24hiCD27+ (B10) and IL10+CD24hiCD38hi (immature transitional) subsets of Breg cells was significantly lower in T1D patients than in healthy controls. Moreover, these cells were also negatively correlated with fasting blood glucose and HbA1c levels. Breg cells did not correlate with autoantibody levels in the serum. These findings suggest that certain Breg subsets are numerically deficient in children with T1D. This alteration in frequency is associated with deficient islet function and glycemia. These findings suggest that Breg cells may be involved in the loss of auto-tolerance and consequent destruction of pancreatic cells and could, therefore, be a potential target for immunotherapy.

Autoimmune Hepatitis: Shifts in Gut Microbiota and Metabolic Pathways among Egyptian Patients.

Autoimmune hepatitis (AIH) is a chronic inflammatory disorder with complex immunopathogenesis. Dysbiosis has been linked to many autoimmune diseases, but its detailed role in autoimmune hepatitis (AIH) still needs rigorous evaluation, especially in Egypt. We aimed to identify the shift in the gut microbiota profile and resultant metabolic pathways in AIH Egyptian patients compared to healthy individuals. Stool samples were collected from 15 AIH-naive patients and from 10 healthy individuals. The V3-V4 hyper-variable regions in16S rRNA gene was amplified and sequenced using Illumina MiSeq platform. Significantly lower bacterial diversity in AIH patients was found compared to the controls. A phylum-level analysis showed the overrepresentation of Firmicutes, Bacteroides, and Proteobacteria. At the genus level, AIH-associated enrichment of Faecalibacterium, Blautia, Streptococcus, Haemophilus, Bacteroides, Veillonella, Eubacterium, Lachnospiraceae and Butyricicoccus was reported in contrast to Prevotella, Parabacteroides and Dilaster, which were significantly retracted in such patients. Overall, the predicted metabolic pathways associated with dysbiosis in AIH patients could orchestrate the potential pathogenic roles of gut microbiota in autoimmune disease, though not in a disease-specific manner, calling for future large-scale studies.

Occult Hepatitis B Infection Among Blood Donors in Al Azhar University Hospital, Upper Egypt: The Current Status After 25 years of Vaccine Introduction.

Occult hepatitis B virus infection (OBI) is a challenging topic with much debate regarding its clinical and virological relevance. The reliance on anti HBc seropositivity for diagnosis of OBI is still controversial. We aimed to determine the frequency of OBI among Egyptian blood donors, the role of anti HBc and HBe Ag as predictors of OBI and the possible risk factors. A total of 300 randomly selected hepatitis B surface antigen (HBsAg) negative blood donors at the Blood Bank of Al Azhar University Hospital in Assiut were included. Measurement of liver enzymes and screening for HBV core antibodies (anti HBc) and Hepatitis e antigen (HbeAg) were done. Quantitative HBV DNA measurement was achieved by real time polymerase chain reaction with a detection limit of 20 IU/mL after DNA extraction from the peripheral blood mononuclear cells (PBMC). The prevalence of OBI was 3.7% (11/300) among the donors and the majority had low level viremia (63.6%). Anti HBc was detected in 20.7% (62/300) (group I). OBI was detected in 14.5% (9/62) in group I compared to 0.84% (2/238) in the anti-HBc Ab negative donors (group II). Anti HBc had a sensitivity and specificity of nearly 82% for detection of OBI with a high negative predictive value (99.16%). HBe Ag was detected in only 1.6% (1/62) of group I. There were no statistical significant differences regarding the liver enzymes, demographic data or risk factors between group I and II and even between cases of OBI and those without. We conclude that OBI exists in an alarming percentage among Egyptian blood donors. Anti HBc should be introduced in the routine blood screening. Negative anti HBc results ensures safe blood, while positive results need nucleic acid confirmation especially if given to high risk recipients. More in-depth evaluation of the immunization program is needed.

Health care-associated infections in pre-transplant liver intensive care unit: Perspectives and challenges.

BACKGROUND: Health care-associated infections (HAIs) threaten patient's safety worldwide especially in the intensive care units (ICU). In end-stage liver disease (ESLD), the condition is much more complicated. Data regarding HAIs among ESLD patients is lacking. We aimed to assess the incidence of HAIs, risk factors, causative micro-organisms, antimicrobial susceptibilities and mortality rates among patients with end-stage liver disease (ESLD) admitted to pre-transplant liver intensive care unit (LICU). METHOD: This prospective observational study included 337 ESLD patients admitted to LICU, Al-Rajhi liver center, Assiut University Hospital, Assiut, Egypt between January 2016 and June 2016 and they were followed up for the development of HAI manifestations. The medical history, physical examination and severity of underlying disease were determined. Clinical samples were taken from patients who developed HAIs for microbiological diagnosis and antimicrobial susceptibility testing. RESULTS: A total of 57 (16.9%) ESLD patients developed HAIs with the incidence density of 26.8 per 1000 patient-days. Blood stream infection was the most common (49.1%). Escherichia coli (21.1%) followed by methicillin-resistant Staphylococcus aureus (MRSA) (15.8%) were the commonest bacteria. Multidrug resistant organisms were reported in 52.6% of the isolates. Fungal causes were 15.8% with Candida species dominance. Sphingomonas paucimobilis and Achromobacter dentrificans were reported for the first time as pathogens for HAIs in LICU. Prolonged hospital stay, intravenous line duration, prolonged use of proton pump inhibitors and paracentesis were predictors for HAIs. No significant difference between ESLD patients with and without HAIs regarding mortality (36.8% vs. 48.6%, P=0.2). CONCLUSION: High HAI rate among ESLD patients is a matter of worry. Effective surveillance program, active infection control measures and national antibiotic policies are necessary to reduce the burden of HAIs.

Bloodstream Infections in Febrile Neutropenic Pediatric Cancer Patients: Microbiological and Sepsis Biomarkers Insight.

The diagnosis of blood steam infections (BSIs) in febrile neutropenic pediatric cancer patients (FNPCP) remains a challenge. Although blood culture is the most accurate method; yet the delay in results has urged the need for reliable biomarkers for early diagnosis. The objectives of this study were to identify the bacterial causes of BSIs in FNPCP at SECI and their antimicrobial susceptibility patterns. Also, to assess the value of procalcitonin (PCT), interleukin 6 (IL6), and interleukin 10 (IL 10) for early diagnosis of BSIs. This study included 68 FNPCP with a total of 85 fever episodes. Blood cultures were done at the onset of fever. Identification of the organisms was carried by Vitek 2 system and the antimicrobial susceptibility testing by disc diffusion. The levels of PCT, IL-6 and IL-10 serum levels were measured by ELISA. Blood stream bacterial infection was detected in 29.4% (25/85). Most were Gram positive cocci in 53.6 % (15/28). There were high percentages of multidrug resistant organism (MDRO) (73.3% and 92.3% among Gram positive and negative bacteria, respectively). The least percentage of resistance was to linezolid (0%) and amikacin (15.4%). The levels of the biomarkers were significantly higher in patients with positive bacterial cultures compared to those with negative cultures (P < 0.001). IL -6 had the best sensitivity (96%) (AUC 0.975, cut off 0.925ng/L) with considerable specificity (88.3%). Combined PCT & IL-6 had the highest sensitivity (96%) and specificity (98.3%). We conclude that the percentage of BSIs among FNPCP was considerable. Gram positive bacteria were the commonest causes. High percentages of MDRO were reported. The most efficient antimicrobials were linezolid and amikacin. IL-6 alone had the best sensitivity for early diagnosis of BSIs. The combination of PCT and IL 6 showed the best performance.

Frequency and Implications of Natural Killer and Natural Killer T Cells in Hepatocellular Carcinoma.

A complex role of the immune system has been highlighted in the development and progression of hepatocellular carcinoma (HCC). Natural Killer cells (NK) and natural killer T cells (NKT) cells are among the innate immune lymphocytes that predominate in the liver and can prevent tumor growth and metastasis. The aim of the study was to measure the percentages of NK and NKT cells among a sample of Egyptian patients with HCC and to find the association between their frequencies and disease progression. The study included 2 groups; the HCC patient group (n=40) and the healthy control group (n=20). Blood samples were drawn from all subjects for complete blood picture, liver enzymes and alpha fetoprotein serum level measurement. Flow cytometric analysis was performed for CD3 and CD16/56 for determining the percentages of NK and NKT cells. The frequencies of NK cells and NKT cells were significantly decreased in HCC patients (6.58± 1.76 and 5.26 ± 1.13 respectively) as compared to healthy controls (9.01±1.62 and 6.88 ± 1.88 respectively) (P < 0.001 and 0.0008 respectively) and in HCC patients with metastasis (6.01± 1.11 and 5.07± 1.10 respectively) than HCC patients without metastasis (7.75± 1.98 and 5.89± 0.88 respectively) (P= 0.004 and 0.03 respectively). We concluded that the reduced percentages of NK cells and NKT cells in HCC patients especially in those with metastasis point to their important roles in the occurrence and progression of HCC.

Seroprevalence and risk factors of Toxoplasma gondii infection among pregnant women in south western, Saudi Arabia.

This study aimed to determine the seroprevalence of Toxoplasma gondii in pregnant women in the south western region of Saudi Arabia and to find out the possible risk factors that may lead to infection. This cross sectional hospital based study was carried out at three hospitals in the south western region of Saudi Arabia from January 2008 to August 2010. Blood samples from 487 pregnant women were collected and used to detect anti-T. gondii antibodies IgM and IgG by enzyme linked immunosorbent assay (ELISA). A questionnaire interview was carried out to ask about some risk factors of infection. Among the 487 studied pregnant women, 38.8 % were seropositive for anti T. gondii IgG while 6.2 % were positive for anti T. gondii IgM and 3.3 % were positive for both anti T. gondii IgG & IgM. The only risk factor associated with seropositive anti T. gondii IgM was the history of the intake of immunosuppressive drugs. Regarding anti T. gondii IgG seropositivity, it was found to increase significantly with increased age, number of gravida and parities, and previous history of toxoplasmosis. The seroprevalence of T. gondii IgG & IgM by ELISA among pregnant women in the south western region of Saudi Arabia is considerable with few identifiable significant risk factors reported.

Fungal infection in patients with end-stage liver disease: low frequency or low index of suspicion.

BACKGROUND: End-stage liver disease (ESLD) is associated with dysregulation of the immune system and increased susceptibility to infections. Although invasive fungal infection (IFI) is a growing public health problem, studies of IFI in ESLD are lacking. The aims of this study were to screen for IFI in ESLD and to assess risk factors and serum interleukin 17 (IL-17) as a marker of the cellular immune response. METHODS: Both blood and ascitic fluid samples were collected from 46 patients with ESLD for fungal culture and PCR. Serum IL-17 levels were determined. RESULTS: Seven patients had isolated IFI (four had spontaneous fungal peritonitis, two had fungemia, and one had a disseminated fungal infection) and five cases had combined fungal and bacterial infections. Spontaneous fungal peritonitis was attributed to Candida species, while fungemia was caused by Aspergillus species. Patients with IFI had higher serum IL-17 levels and increased mortality compared to patients without IFI. A history of antibiotic use (p = 0.002), higher model for end-stage liver disease (MELD) scores (p = 0.04), and hepatorenal syndrome (p = 0.006) were risk factors for IFI. CONCLUSIONS: Patients with ESLD had a low frequency of IFI; however, in patients with these infections, delayed diagnosis and treatment may contribute to a high fatality rate. Thus, clinicians should have a high index of suspicion for this unusual but lethal entity, as prompt detection and appropriate treatment can improve the outcome.

Is serum level of methylated RASSF1A valuable in diagnosing hepatocellular carcinoma in patients with chronic viral hepatitis C?

BACKGROUND AND STUDY AIMS: The detection of the promoter hypermethylation of RASSF1A in serum DNA could be a valuable biomarker for early detection of preneoplastic lesions and early cancer development among high-risk populations who are at a high risk of hepatocellular carcinoma (HCC). Therefore, we aimed determining the serum level of methylated RASSF1A sequence in patients with chronic hepatitis C viral infection and to evaluate the predictive value of it as a diagnostic marker for HCC in patients with chronic hepatitis C viral infection. PATIENTS AND METHODS: Serum levels of methylated RASSF1A were detected and measured using real-time PCR after digestion with a methylation-sensitive restriction enzyme in 40 patients with chronic HCV infection, 40 patients with HCC (on top of HCV) and 20 controls. RESULTS: Methylated RASSF1A was detected in 10% of the controls, 62.5% of HCV group and in 90% of HCC group. Chronic HCV patients had insignificantly higher levels than the controls. The levels were significantly higher in patients with HCC compared to the controls (p=0.0001) and chronic HCV patients (p=0.001). By logistic regression analysis, the serum methylated RASSF1A was found to differentiate HCC patients from healthy controls with an AUROC of 0.83nmol/L, and an overall predictive accuracy of 77.5%. It was able to differentiate patients with HCC from those with chronic HCV infection alone with an AUROC of 0.733 and an overall predictive accuracy of 72.5%. CONCLUSION: The mean serum levels of methylated RASSF1A could be of value for early diagnosis of HCC especially in high risk patients with HCV infection.