Enzyme-linked Immunoassay Test Research Articles

Design and Development of an Antigen Test for SARS-CoV-2 Nucleocapsid Protein to Validate the Viral Quality Assurance Panels.

The continuing mutability of the SARS-CoV-2 virus can result in failures of diagnostic assays. To address this, we describe a generalizable bioinformatics-to-biology pipeline developed for the calibration and quality assurance of inactivated SARS-CoV-2 variant panels provided to Radical Acceleration of Diagnostics programs (RADx)-radical program awardees. A heuristic genetic analysis based on variant-defining mutations demonstrated the lowest genetic variance in the Nucleocapsid protein (Np)-C-terminal domain (CTD) across all SARS-CoV-2 variants. We then employed the Shannon entropy method on (Np) sequences collected from the major variants, verifying the CTD with lower entropy (less prone to mutations) than other Np regions. Polyclonal and monoclonal antibodies were raised against this target CTD antigen and used to develop an Enzyme-linked immunoassay (ELISA) test for SARS-CoV-2. Blinded Viral Quality Assurance (VQA) panels comprised of UV-inactivated SARS-CoV-2 variants (XBB.1.5, BF.7, BA.1, B.1.617.2, and WA1) and distractor respiratory viruses (CoV 229E, CoV OC43, RSV A2, RSV B, IAV H1N1, and IBV) were assembled by the RADx-rad Diagnostics core and tested using the ELISA described here. The assay tested positive for all variants with high sensitivity (limit of detection: 1.72-8.78 ng/mL) and negative for the distractor virus panel. Epitope mapping for the monoclonal antibodies identified a 20 amino acid antigenic peptide on the Np-CTD that an in-silico program also predicted for the highest antigenicity. This work provides a template for a bioinformatics pipeline to select genetic regions with a low propensity for mutation (low Shannon entropy) to develop robust 'pan-variant' antigen-based assays for viruses prone to high mutational rates.

Novel MHC BLB2 gene polymorphism and its association with IgY concentration and Newcastle disease antibody titer in IPB-D2 chickens.

This study aimed to identify the polymorphism of the B Locus Beta 2 (BLB2) gene and its association with immunoglobulin Y (IgY) concentration and Newcastle disease (ND) antibody titer; we analyzed BLB2 gene expression in different categories of ND antibody titers in IPB-D2 chickens. The total sample used was 100 IPB-D2 chickens. Blood samples were collected at 21 weeks old for an ELISA (enzyme-linked immunoassay) test, an HI (hemagglutination inhibition) test, and genotyping. The method for BLB2 polymorphism was Sanger sequencing. Analysis of BLB2 gene expression was performed using the cecal tonsil tissue of IPB-D2 chickens. Polymorphism data were analyzed using SNPstats and DNAsp (DNA Sequence Polymorphism) software. The association of the single-nucleotide polymorphisms (SNPs) with IgY concentration and ND antibody titer was analyzed using SAS software (version 9.2). The genotype mean values were compared by means of a T test. The relative mRNA expression analysis was performed using a quantitative real-time polymerase chain reaction (qRT-PCR). The results showed that 13 SNPs were found in exon 2 and exon 3 in the BLB2 gene. As many as 4 out of the 13 SNPs were associated with IgY concentration. As many as 9 out the 13 SNPs may have changed amino acids. The Ct value showed that the expression of the BLB2 gene in IPB-D2 chickens with high ND antibody titers is higher than IPB-D2 chickens with low ND antibody titers. In conclusion, the AA genotype of g.458 T A was associated with high IgY concentrations, and the BLB2 gene presented with a high expression in IPB-D2 chickens with high ND antibody titers.

Designing and developing a sensitive and specific SARS-CoV-2 RBD IgG detection kit for identifying positive human samples

More than 3 y into the coronavirus 2019 (COVID-19) pandemic, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to undergo mutations. In this context, the Receptor Binding Domain (RBD) is the most antigenic region among the SARS-CoV-2 Spike protein and has emerged as a promising candidate for immunological development. We designed an IgG-based indirect enzyme-linked immunoassay (ELISA) kit based on recombinant RBD, which was produced from the laboratory to 10L industry scales in Pichia pastoris. A recombinant-RBD comprising 283 residues (31kDa) was constructed after epitope analyses. The target gene was initially cloned into an Escherichia coli TOP10 genotype and transformed into Pichia pastoris CBS7435 muts for protein production. Production was scaled up in a 10L fermenter after a 1L shake-flask cultivation. The product was ultrafiltered and purified using ion-exchange chromatography. IgG-positive human sera for SARS-CoV-2 were employed by an ELISA test to evaluate the antigenicity and specific binding of the produced protein. Bioreactor cultivation yielded 4g/l of the target protein after 160h of fermentation, and ion-exchange chromatography indicated a purity>95%. A human serum ELISA test was performed in 4 parts, and the ROC area under the curve (AUC) was>0.96 for each part. The mean specificity and sensitivity of each part was 100% and 91.5%, respectively. A highly specific and sensitive IgG-based serologic kit was developed for improved diagnostic purposes in patients with COVID-19 after generating an RBD antigen in Pichia pastoris at laboratory and 10L fermentation scales.

Prevalence of Human Adenoviruses in Respiratory Tract-infected Patients in Anbar Governorate (West of Iraq): A Descriptive Cross-sectional Study

Background: Human adenoviruses (HAdVs) are a significant worldwide problem that causes viral respiratory tract infections affecting millions worldwide each year, especially in children and immunocompromised adults. Objective: To know the prevalence of HAdVs strains 3, 4, 7 among respiratory tract infected patients aged between 15–42 years in Anbar province (west of Iraq) using enzyme-linked immunoassay (ELISA) test with polymerase chain reaction (PCR) technique. Material and methods: A descriptive cross-sectional study was done between 22th of January 2021 - October 11, 2021 to notice the frequency of HAdVs in Anbar governorate children and adults with respiratory infections from the various general hospitals and private clinics. Depending on our questionnaires, blood samples were taken from all those patients for hematological and serological parameters. The ELIZA test with PCR has been done depending on the manufacturer’s instructions. Results: A total of 104 respiratory tract Infected patients, 11(10.6%) patients were ELISA IgM positive, 9(8.7%) of ELISA IgM positive patients were positive using PCR technology. The 6(5.8%) patients were ELISA IgG positive for HAdV. Out of 32 mild pneumonia patients, 1(3.1%) was positive for HAdV using IgM ELIZA. Out of 46 moderate pneumonia patients, 6(13.0%) were positive for HAdV using IgM ELIZA. Out of 26 sever pneumonia patients, 4(15.4%) were positive for HAdV using IgM ELIZA. Out of 32 mild pneumonia patients, 1(3.1%) was positive for HAdV using IgM ELIZA. Out of 46 moderate pneumonia patients, 6(13.0%) were positive for HAdV using IgM ELIZA. Out of 26 sever pneumonia patients, 4(15.4%) were positive for HAdV using IgM ELIZA. Conclusion: The prevalence of HAdVs strain 7 in children with respiratory infection was 4.8%, whereas its prevalence in adults in age groups 18–50 and 51+ years was 2.9% for each group using the IgM ELIZA test.

Protocol of Detection of West Nile Virus in Clinical Samples.

West Nile virus (WNV) is one of the leading causes of arboviral encephalitis in the United States but is often underdiagnosed. Despite the wide breadth of WNV-induced clinical disease syndromes, many of the symptoms associated with WNV are nonspecific at the time of presentation; thus, choosing the right diagnostic tool is essential to not only understand the true burden of disease but also provide pathogen-directed interventions for WNV-infected patients. In this chapter, we briefly discuss the three most common types of diagnostic methods for WNV in human clinical samples: nucleic acid detection, enzyme-linked immunoassay (ELISA), and plaque reduction neutralization test (PRNT) and present the method for PRNT.

Anti-inflammatory potential of berberine-rich extract via modulation of inflammation biomarkers.

Berberine-rich extract (BRE) prepared from Berberis lycium root bark using green extraction approach and its marker compound berberine has a broad spectrum of clinical applications. Berberine's potential pharmacological effects include anticancer, antidiarrheal, antidiabetic, antimicrobial and anti-inflammatory activities. In current work, BRE and berberine were evaluated for their therapeutic prospects in inflammation models. The comparative effect of BRE and berberine against inflammation was determined through in vitro chemiluminescence technique. The in vivo anti-inflammatory evaluation of BRE and berberine (25, 75, and 125 mg/kg) compared to diclofenac (10mg/kg) was performed in carrageenan and formaldehyde-induced inflammation in Wistar rats. Histopathological and biochemical studies were conducted to find the comparative anti-inflammatory potential of BRE and berberine on pathological hallmarks induced by formaldehyde. Moreover, the modulatory effects on inflammatory biomarkers were also investigated through qPCR. ELISA (enzyme-linked immunoassay test assay) was performed to investigate the expression of pathological protein biomarkers like TNF-α and IL-6 and levels of antioxidant enzymes were estimated in liver homogenates. Both BRE and berberine markedly (p < .001) reduced paw diameter and inflammation in carrageenan and formaldehyde-induced inflammation. The levels of antioxidant enzymes were recovered (p < .001) by BRE and berberine treatments, and compared to the formaldehyde-treated inflammation model. Both BRE and berberine remarkably downregulated the mRNA and protein expression of inflammatory biomarkers. BRE similar to berberine mitigated the level of antioxidant enzymes in liver homogenate. The undertaken study suggests that BRE, a natural, green, and therapeutically bioequivalent to berberine could be used as an economical phytomedicine in the treatment of inflammatory disorders. PRACTICAL APPLICATIONS: Anti-inflammatory drugs like NSAIDS are associated with serious adverse effects like gastrointestinal ulcer, worsening of preexisting cardiovascular disorders, and renal failure. Therefore, there is a constant demand to develop novel, inexpensive therapeutic strategies to treat the inflammatory disorder with the least harmful effects. Pure phytochemicals with anti-inflammatory potential are costly and hard to isolate, therefore green microwave-assisted extraction technique is developed to get the rich bioequivalent extract. Berberis lycium a medicinal plant with berberine as a major bioactive constituent, has wide acceptance in traditionally used medicine and as food. Pharmacological studies revealed its hepatoprotective, anticancer, antidiabetic, and antihypertensive activities. BRE was prepared by green microwave-assisted extraction and enrichment by resin column to get a higher yield of berberine. The comparative anti-inflammatory effect of BRE and berberine was determined by in vitro and in vivo studies. Results obtained from this experimental work contribute beneficial guidance that reinforces the use of the BREto treat inflammatory disorders.

Detection of EhCRT Gene Expression in Entamoeba histolytica-Infected Children and its Correlation with Interleukin 25 and Tumor Necrosis Factor Alpha

Objectives: Entamoeba histolytica is a human enteric protozoan, which is the causative agent of amebiasis. The host activates a series of immunological responses to protect against the parasite after contact with the ameba and further invasion of the gut epithelium layer. As a result, the ameba has developed a variety of evasion mechanisms to hold out the immune response and continue to survive and cause disease. The calreticulin (EhCRT) is one of the immunogenic molecules of E. histolytica that induces an immune response in the human host. Increase in the expression of the EhCRT gene could provide control mechanism that allows the parasite to adapt and survive in host tissues. Aim of the Study: This study was designed to detect the EhCRT gene of E. histolytica by real-time polymerase chain reaction (PCR) in stool samples of children with amebiasis and its roles in host–parasite relationship via measuring the concentration of tumor necrosis factor alpha (TNFα) and interleukin 25 (IL25) by enzyme-linked immunoassay (ELISA) technique in their serum. Materials and Methods: A total of 86 diarrheal fecal samples were collected from children in age &lt;1 year to 13 years suspected to be infected with E. histolytica during the period from December 30, 2020, to September 1, 2021. Microscopically positive samples were the subject to conventional PCR and real-time PCR for the detection of E. histolytica HM1:IMSS strain using (Psp) gene sequences and detection of calreticulin (EhCRT) expression. Blood was withdrawn from each child included in the study for ELISA test to measure the level of IL25 and TNFα. Results: Fecal samples for microscopic examination revealed that 71 (82.6%) children had amebic colitis, E. histolytica gene was detected in 44 samples (71%) using conventional PCR, and the immunogene EhCRT was expressed in 36 stool samples using real-time PCR. The results of the recent study showed highly significant elevation in the level of TNFα and IL25 in the amebic group (Eh+ve PCR). The majority of amebic children were in the age group of 1–4 years, had mucoid, acute, and with primary episodes of diarrhea. Conclusion: E. histolytica is a protozoan parasite highly prevalent among diarrheal children and is responsible for gastrointestinal amebiasis in the human host. The PCR is a useful tool in the diagnosis of E. histolytica infection. It is clear that the expression of the calreticulin gene (EhCRT) concedes with the duration of diarrhea a virulence factor that plays a role in host pathogenic pathways. The findings of this study showed that the level of TNFα in the serum of children infected with amebic colitis (Eh gene + ve) is significantly increased during the course of infection and the cytokine IL25 exhibits a significant drops in the same children.

West Nile virus in the Republic of Serbia-Diagnostic performance of five serological tests in dog and horse sera.

West Nile virus (WNV) is a zoonotic mosquito-borne virus classified as family Flaviviridae and genus Flavivirus. The first WNV outbreak in humans in the Republic of Serbia was recorded in 2012. Equids and dogs can show clinical symptoms after WNV infection and are often used as sentinels. This study aimed to (i) give insight into seropositivity for WNV in clinically healthy dog and horse sera in different regions of Serbia and (ii) compare diagnostic value of 'in-house' and commercially available indirect immunofluorescence (IFA) and enzyme-linked immunoassay (ELISA) tests to 'gold standard' virus neutralization test (VNT). Due to cross-reactivity, sera were tested for Usutu virus and tick-borne encephalitis virus in VNT based on the epidemiological data of field presence. Blood sera of dogs (n=184) and horses (n=232) were collected from 2011 to 2013. The seropositivity was confirmed by VNT in 36.9 % tested dog sera and 34.9% tested horse sera with highest positivity in regions near two big rivers, while in four dog and seven horse sera, positivity resulted from Usutu virus infection. Comparative results of diagnostic tests in dogs ranged from 18.7 % seropositivity by 'in-house' ELISA to 31.9% by commercially available ELISA. In horses, seropositivity ranged from 36.2% by 'in-house' IFA to 32.5% by commercially available IFA and from 26.3% by 'in-house' IgG ELISA to 20.9% by commercially available ELISA. There were no statistically significant differences according to the McNemar test between 'in-house' and commercially available IFA and ELISA test in horse sera, while the same was not true for two ELISAs used in dog sera (χ2 =8.647, p=.003). Established seropositivity in dogs and horses was in accordance with the epidemiological situation and WNV spread in the Republic of Serbia and proven Usutu virus co-circulation. 'In-house' tests remain a valuable tool in early diagnostics of WNV.

Correlation between CD14 and CD163 in duodenal ulcer and gastric cancer patients infected with Helicobacter Pylori

Most people around the world are infected with Helicobacter pylori (H.pylori); it can lead to duodenal ulcer (DU) and gastric cancer (GC). The main objective of this study is to determine the correlation between CD14 and CD163 in individuals with DU and GC infected with H.pylori. Sixty nine individuals were included in this work; 27 patients infected with H.pylori only (H.Pylori+), 22 patients infected with H.pylori with DU and 20 patients infected with H.pylori with GC. CD14 and CD163 were measured in all individuals' serum using the Enzyme-Linked Immunoassay (ELISA) test. The results proved there was negative correlation between CD14 and CD163 in H.pylori-positive patients (y= -0.0263x+1.4179). While, there was positive correlation in H.pylori-positive patients with DU (y= 0.0932x+1.6647) and with GC (y= 0.0607x + 1.9824). In conclusion: CD14 and CD163 have a synergistic protected effect in H.pylori-positive patients with DU and GC.

Helicobacter Pylori-oncogenic protein cytotoxin-associated gene A and assessment of CD14 and CD163 in duodenal ulcer and gastric cancer patients

Nearly half of the world's population is infected with Helicobacter pylori (H.pylori). A duodenal ulcer or stomach cancer can be caused by the infection by this bacterium. The aim of this work is to assess the levels of CD14 and CD163 in H.Pylori-positive patients infected with duodenal ulcer (DU) and gastric cancer (GC) and determine the prevalence of Helicobacter Pylori-oncogenic protein cytotoxin-associated gene A strains (H.pylori-CagA). This study included 89 individuals distributed as follows: 20 healthy individuals as controls and 69 patients infected with H. Pylori have been divided as follows: 27 patients infected with H.pylori only (H.Pylori+), 22 H.pylori+DU and 20 H.pylori+GC. H. Pylori-oncogenic protein cytotoxin-associated gene A strains (H-pylori-CagA) were diagnosed based on a qualitative reverse-phase Enzyme Immunoassay Technique. CD163 and CD14 were measured in all individuals' serum using the Enzyme-Linked Immunoassay (ELISA) test. Out of 69 patients infected with H.pylori, there was one CagA strain in H.pylori+; two and five strains were recorded in H.pylori+DU and H.pylori+GC, respectively. CD14 and CD163 serum concentrations were significantly higher (P≤0.05) in H. pylori+, H. pylori+DU and H. pylori+GC than in controls. Conclusions: Patients with CagA strains infection are at risk of developing a duodenal ulcer and stomach cancer.

Free Papers Compiled

INTRODUCTION:Blood borne viral infections (hepatitis C, hepatitis B, human immune deficiency virus), constitute a significant public health concern among patients with opioid dependence, especially among intravenous drug users (IDU).AIM:To measure the prevalence of hepatitis C among patients with intravenous opioid dependence, visiting de-addiction clinic, in a tertiary care hospital in the state of Himachal Pradesh.MATERIAL AND METHOD:A cross-sectional study was conducted between January 2021 and June 2021 at de-addiction clinic, department of Psychiatry, IGMC, Shimla. Socio-demographic and clinical profiles of patients with diagnosis of opioid dependence was entered in a pre-structured performa. All these patients were investigated for HCV antibody using ELISA (enzyme linked immunoassay) test.RESULTS:The study sample comprised of 200 male patients, and majority were between age 21-40 years (96.5%), from nuclear families (57%), urban background (56.5%). One fourth (25.5%) reported suicidal ideation/attempt. Intravenous mode of drug use (IDU) was reported by 121 (60.5%) patients, and majority were from nuclear families (64.46%), urban background (49.59%), and 30.58% reported suicidal ideation/attempt. 29 patients (23.97%) tested positive for hepatitis C infection.CONCLUSION:There is high rate of hepatitis C infection among patients with opioid (intravenous) dependence, at our de-addiction clinic. Future studies with large sample size exploring the high risk behaviour practices and studies with longitudinal design to assess treatment retention, are needed.

Humoral Immunity and Adverse Events after Vaccination Against COVID-19 By a Vector Based Vaccine Sputnik V in Patients with Chronic Myeloid Leukemia

Humoral Immunity and Adverse Events after Vaccination Against COVID-19 By a Vector Based Vaccine Sputnik V in Patients with Chronic Myeloid Leukemia

A longitudinal follow-up of COVID-19 patients in the convalescent phase showed recovery in radiological results, the dynamics of lymphocytes, and a decrease in the level of IgG antibody: a single-centre, observational study.

BackgroundGiven the high prevalence of coronavirus disease 2019 (COVID-19) globally, and the increased number of patients being discharged, it is necessary to understand the health consequences of COVID-19 to formulate and manage public policy concerning convalescent patients.MethodsA longitudinal follow-up investigation of 25 patients from a tertiary hospital in Henan provincial was conducted 8 weeks after discharge. Of these patients, 15 attended a second follow-up appointment 8 weeks after that. A throat swab reverse transcription-polymerase chain reaction (RT-PCR) analysis for SARS-CoV-2 and chest computerized tomography (CT) scans were implemented at the first follow-up appointment, and a total of 40 blood samples (25 from the first and 15 from the second follow-up appointment) were collected. Patients’ levels of Immunoglobulin G (IgG) antibody against S-Receptor binding domain (S-RBD) and Nucleocapsid Protein (NP) of SARS-CoV-2 and the subpopulation of lymphocytes were evaluated using an enzyme-linked immunoassay (ELISA) test and flow cytometry, respectively.ResultsAt the first follow-up appointment, 10 of the 25 patients (40.0%) showed complete radiological resolution. Of these patients, 80.0% were classified as moderate, and 80.0% were younger (those whose age was ≤ the median age of all the patients). The predominant patterns of abnormalities included an irregular line (12/25, 48.0%), ground-glass opacity (GGO) (44.0%), and multiple GGOs (28.0%). At the first follow-up appointment, 40.0% (10/25) of patients still had lymphopenia. Of the 15 patients who were followed-up with twice, the ratio of lymphopenia was 80% (12/15), 60.0% (9/15), and 46.7% (7/15) at 0, 8, and 16 weeks after discharge, respectively. This was mainly due to a decrease in the cluster of differentiation (CD) 4+ T lymphocyte, which was observed in 60% (9/15), 60% (9/15), and 46.7% (7/15) of total patients at 0, 8, and 16 weeks after discharge, respectively. All of the patients were S-RBD and NP IgG antibody positive at the first follow-up appointment. 40.0% (6/15) and 66.7% (10/15) of patients showed a decrease over 50.0% in the level of NP and S-RBD IgG antibodies, respectively, at the second follow-up appointment. The NP and S-RBD IgG antibodies’ levels declined to 44.6% (P=0.044) and 28.1% (P=0.18), respectively. 0 and 26.7% (4/15) of patients turned from NP and S-RBD IgG antibody positive to negative, respectively.ConclusionsAbout half of the patients still showed at least 1 abnormality in chest CT scans 8 weeks after discharge and lymphopenia 16 weeks after discharge. The level of the IgG antibody had declined by the follow-up appointment. Notably, the S-RBD IgG antibody declined more dramatically than that of NP. These results may have implications in the making of policies related to disease prevention, the long-term management of discharged patients, and vaccines’ development.

The "false-positive" conundrum: IgA reference level overestimates the seroprevalence of antibodies to SARS-CoV-2.

BackgroundOn 12 June 2020, Brazil reached the second position worldwide in the number of COVID-19 cases. Authorities increased the number of tests performed, including the identification of antibodies to SARS-CoV-2 (IgG, IgA, and IgM). There was an overflooding of the market with several tests, and the presence of possible false-positive results became a challenge. The purpose of this study was to describe the seroprevalence and immunoglobulin blood levels in a group of asymptomatic individuals using the reference levels provided by the manufacturer.MethodsLevels of IgG and IgA antibodies to SARS-CoV-2 were determined in blood serum by the same ELISA (enzyme-linked immunoassay) test. Patients must be free of symptoms.ResultsFrom 20 to 22 May 2020, 938 individuals were tested. There were 441 (47%) men, age 53 years (interquartile range (IQR) = 39-63.2). The sample included 335 (35.7%) subjects aged ≥60 years old. Subjects with a positive test were 54 (5.8%) for IgG and 96 (10.2%) for IgA and 42 (4.5%) for both IgG and IgA. The prevalence of IgG and IgA positive test was not different in men and women and not different in individuals under 60 and over 60 years of age. Conversely, analysing only individuals with positive tests, the levels of IgG in positive subjects were significantly higher than those with an IgA positive test, 3.00 (IQR = 1.68-5.65), and 1.95 (IQR = 1.40-3.38), respectively; P = 0.017. Additionally, individuals with isolated IgA positive tests had significantly lower levels of IgA than those with both IgA and IgG positive tests: 1.95 (IQR = 1.60-2.40) and 3.15 (IQR = 2.20-3.90), respectively, P = 0.005. These latter data suggest that IgA shows a deviation of the distribution to the left in comparison to IgG distribution data. Indeed, many subjects reported as IgA positive had immunoglobulin levels slightly elevated.ConclusionsIn conclusion, we strongly suggest caution in the interpretation of IgA test results. This recommendation is more important for those with positive IgA just above the reference level.

Concentration of dioxin and screening level ecotoxicity of pore water from bottom sediments in relation to organic carbon contents

The information about concentrations of dioxin in pore water, ecotoxicity and DOC and TOC content can be key factor for the prediction of the fate of dioxins in the aquatic environment as well as an ecological risk assessment. The aims of the study were to assess the concentration of PCDDs/PCDFs and ecotoxicity of pore water and to compare above results in relation to the dissolved organic carbon (DOC) and total organic carbon (TOC) content. The concentration of dioxins was assessed using an enzyme-linked immunoassay test, while the ecotoxicity of pore water was determined using a crustacean Daphnia magna and bacteria Aliivibrio fischeri. The studies were conducted on two different dammed reservoirs Rożnów (catchment basin of an agricultural character) and Rybnik (catchment basin of an industrial character) located in southern Poland. The concentration of dioxins in pore water was between 8.56 to 90.92 ng EQ/L, with a significantly higher concentration in the pore water from the Rożnów Reservoir than the Rybnik Reservoir. The DOC content in pore water was from 30.29 to 63.02 mg/L (Rożnów Reservoir) and from 35.46 to 60.53 mg/L (Rybnik Reservoir). Higher toxic responses were recorded for A. fischeri than for D. magna. Moreover a significantly higher toxicity for both tested organisms was indicated in pore water from the Rożnów Reservoir. Besides of TOC and DOC, the fine fractions of the sediments were particularly important in the concentration of dioxin in pore water. The other pore water parameters, such as pH and EC can influence the toxicity of water for organisms. The result indicate complex relationships between the PCDD/F, ecotoxicity and DOC, TOC concentration in pore water and confirms that these parameters are important in terms of water environmental contamination.

Degradation of aflatoxin in maize using Ferulic acid (phydroxy-3-methyl cinnamic acid) catalyzed by Hydrogen peroxide

Purpose: The study aimed to determine the rate of degradation of aflatoxin in contaminated maize using ferulic acid catalyzed by hydrogen peroxideMethodology: 100 g of dried maize grain was grounded using a laboratory hammer mill and divided into 2 portions of 50 g each. 20 g sample was taken per portion and treated with 100 mL solution of methanol and deionized water in the ration of 8:1, 50 mL of Acetonitrile, 1 g NaCl and 4 g of anhydrous magnesium sulphate, then blended at 120 RPM for 30 min. Aflatoxin content in each extract was analysed using enzyme-linked immunoassay test kits and confirmed using high performance liquid chromatography (HPLC) coupled with fluorescence detector. Further experiments tested the effect of coating, size, concentration, catalyst and reaction time on degradation of aflatoxin in maize. Data analysis was conducted using SPSS and Microsoft excel.Findings: Four-hour treatment of contaminated maize with 0.5 mM ferulic acid reduced aflatoxin by 91.0% for whole maize, 90.5% for dehulled maize and 90.9% for ground maize. Addition of 20 mL of 0.5% hydrogen peroxide to the reaction mixtures increased degradation of aflatoxin load to 99.0% for whole maize, 99.1% for dehulled maize and 99.1% for ground maize within 4-hour reaction time. The rate of decontamination followed first order kinetics with R2 values of 0.919, 0.916 and 0.930 for the whole maize, dehulled maize, ground maize, respectively and achieved degradation half-lives of 43.59, 41.26 and 39.84 minutes in the same order.Unique contribution to theory, practice and policy: Ferulic acid combined with hydrogen peroxide is an effective degrader of aflatoxin in maize. The rate degradation is dependent on the nature of maize pre-treatment, the concentration of ferulic acid, and the catalyst. Ferulic acid and hydrogen peroxide reacted with the lactone ring of the coumarin moity of aflatoxin. Recommendations; Further studies on degradation of aflatoxin in maize should elucidate the pathways and metabolites formed in the ferulic acid degradation process and determine their toxicities

Screening for SARS-CoV-2: Health Policy Implications in Low- and Middle-Income Countries

Screening for SARS-CoV-2: Health Policy Implications in Low- and Middle-Income Countries

Occurrence and Distribution of Aflatoxin in Maize from Selected Counties, Eastern Region, Kenya

Purpose: The study aimed to assess the occurrence and distribution of aflatoxin contamination on dry maize in different types of stores in Meru, Embu, Isiolo, Makueni and Machakos Counties of Eastern region of Kenya.Methodology: Automatic spear sampler was used to collect maize samples from each bag at even intervals. 280 maize samples were collected from 29 stores in five Counties. 100 g of each maize sample was ground, resampled into 50g, blended, extracted, centrifuged, filtered and a quantified for Aflatoxin B1, B2, G1 and G2. Samples were prepared and extracted with methanol/water. The bulky of the samples were analyzed with enzyme-linked immunoassay test kits. Confirmation of positive samples was done with high performance liquid chromatography (HPLC) coupled with fluorescence detector. Data analysis was done with SPSS and Microsoft excel.Findings: Maize samples from Counties in eastern region of Kenya had significantly high levels of (93.10%) aflatoxin contamination. The mean values for aflatoxin B1, B2, G1 and G2 were: 50.08± 4.42, 17.26±1.08, 30.17±2.06 and 10.54± 1.52 (ng/g) in that order. Only nine samples had total aflatoxin within the accepted limit for human consumption of 15 ng/g. The highest total aflatoxin contamination recorded was 198.45ng/g in Makueni county and the lowest recorded was 8.76ng/g in Embu county. Makueni and Embu had mean values for aflatoxin B1, B2, G1 and G2 being (83.07±7.53, 22.15± 1.36, 49.38±3.11, 20.52± 0.70 ng/g) and (18.71 ±2.63, 8.07 ±0.64, 17.02 ±1.38, 8.86 ±1.62 ng/g). Makueni NCPB depot had the highest mean contamination with aflatoxin B1 of 92.67± 5.78 ng/g and Embu had the lowest with 6.26 ± 4.14 ng/g. All the county markets recorded high aflatoxin B1 contamination with exception of Embu county which had a mean of 4.0 ±0.84, Makueni (83.67± 10.42 ng/g), Isiolo (51.27± 32.29 ng/g), Meru (46.02± 23.88 ng/g) and Machakos (36.34± 26.27 ng/g). The stores had aflatoxin load varied from on store to the other and county to county.Unique contribution to theory, policy and practice: The counties in the region had high occurrence and distribution of aflatoxin B1, B2, G1 and G2 in maize in all stores where samples were picked. Location for maize stores should be in areas with low levels of carbon dioxide because mycotoxins are produced under aerobic conditions. The design for maize threshing machines should not course shocks, breakage and cracks on maize grains to decrease chances of mycotoxins infestation during their storage.

Perspectives on meeting the COVID‐19 testing challenge: A dental school collaborative

Perspectives on meeting the COVID‐19 testing challenge: A dental school collaborative

Accuracy of serological tests for diagnosis of chronic pulmonary aspergillosis: A systematic review and meta-analysis.

Chronic pulmonary aspergillosis (CPA) is a slow and progressive disease that develops in preexisting lung cavities of patients with tuberculosis sequelae, and it is associated with a high mortality rate. Serological tests such as double agar gel immunodiffusion test (DID) or counterimmunoelectrophoresis (CIE) test have been routinely used for CPA diagnosis in the absence of positive cultures. However, these tests have been replaced with enzyme-linked immunoassay (ELISA) and, a variety of methods. This systematic review compares ELISA accuracy to reference test (DID and/or CIE) accuracy in CPA diagnosis. It was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). The study was registered in PROSPERO under the registration number CRD42016046057. We searched the electronic databases MEDLINE (PubMed), EMBASE (Elsevier), LILACS (VHL), Cochrane library, and ISI Web of Science. Gray literature was researched using Google Scholar and conference abstracts. We included articles with patients or serum samples from patients with CPA who underwent two serological tests: ELISA (index test) and IDD and/or CIE (reference test). We used the test accuracy as a result. Original articles were considered without a restriction of date or language. The pooled sensitivity, specificity, and summary receiver operating characteristic curves were estimated. We included 14 studies in the review, but only four were included in the meta-analysis. The pooled sensitivities and specificities were 0.93 and 0.97 for the ELISA test. These values were 0.64 and 0.99 for the reference test (DID and/or CIE). Analyses of summary receiver operating characteristic curves yielded 0.99 for ELISA and 0.99 for the reference test (DID and/or CIE). Our meta-analysis suggests that the diagnostic accuracy of ELISA is greater than the reference tests (DID and/or CIE) for early CPA detection.