Enzyme Immunosorbent Assay Research Articles

B-104 Correlation of Anti-Centromere Antibodies (ACA) Detection by Indirect Immunofluorescence (IFA) on HEp-2 Cells and by ELISA Demonstrates High Concordance but Highlights Advantage of IFA

Abstract Background Anti-centromere antibodies (ACA), a type of anti-nuclear antibody (ANA) first identified as a discrete speckled pattern on indirect immunofluorescence assay (IFA), are a hallmark of systemic sclerosis (SSc), especially in its limited cutaneous form (lcSSc) or CREST (calcinosis, Raynaud’s, esophageal dysmotility, sclerodactyly and telangiectasia). These mitosis-related autoantibodies mainly recognize three centromere proteins (CENP), CENP-A, CENP-B, and CENP-C, that comprise the kinetochore important to chromosome segregation, but more recently, at least three other CENP antigens have been identified (CENP-D, CENP-E and CENP-F). Anti-CENP-B is considered the main autoantibody, and many, if not most, ELISA utilize recombinant CENP-B such that antibodies to CENP antigens other than CENP-B are not detected. Here, an ELISA using both CENP-B and CENP-A is compared to the reference method, HEp-2 cell IFA, highlighting its high, but still less than 100%, sensitivity due to multiple centromere subcomponents. Methods Anti-centromere antibodies are determined in patient serum using the reference method, IFA on the HEp-2 cell substrate (Sprinter XL/EUROpattern®, Euroimmun, NJ) which is also the current method for standalone ACA testing as well as screening by selected ANA profiles designed for the rheumatology specialist. Here ACA IFA- positive and negative serum samples were further analyzed by enzyme immunosorbent assay (ELISA) utilizing recombinant CENP-A and CENP-B (QuantaLyzer® 3000, Werfen, CA). Results When compared, samples between ACA pattern by HEp-2 cell IFA and anti- centromere antibodies by ELISA yielded an agreement of 93.2%. McNemar’s Test Chi Square of p>0.999, with a relative sensitivity of 89.7% and a specificity of 100.0%. Conclusions This study demonstrated very high concordance between ELISA and accurate ACA pattern identification on HEp-2 IFA. The ELISA studied here identifies autoantibodies to both CENP-A and CENP-B where many other ELISA identify only anti-CENP-B. Although ACA are relatively specific for SSc, they have also been reported in systemic lupus erythematosus (SLE), primary biliary cholangitis (PBC), rheumatoid arthritis (RA), Sjogren Syndrome, Raynaud's phenomenon and in cancer in individuals with or without evidence of systemic autoimmune rheumatic disease (SARD). Of note, many other commercially available ELISA for anti-centromere antibodies identify only anti-CENP-B, but there at least four other known CENP antigen targets for autoantibodies causing centromere pattern on IFA. Furthermore, almost all laboratories use the ELISA for ACA on ANA panel (or other connective tissue disease) screening. Hence, it should be duly noted that these testing strategies will not detect autoantibodies to all anti-centromere antibodies. Only specialized ANA profiles designed for the difficult-to-diagnose autoimmune patient begin screening using the IFA method for anti-centromere and therefore, are capable of detecting autoantibodies to all CENP antigens. Laboratorians should advise clinicians of these diagnostic limitations especially as our knowledge about centromere subcomponents and their clinical associations increases.

P-100 Prolonged exposure to polyethylene glycol (peg) 6000 severely affects the reproductive system of male Wistar rats

Abstract Study question Studies have shown that microplastics have adverse effects on aquatic reproductive health. Is there any potential effect of Polyethylene glycol (PEG)6000 on rodents reproductive system? Summary answer Despite its wide use in industry, medicine and cosmetic formulations, prolonged exposure to PEG 6000 may pose great risk, especially causing reproductive toxicity. What is known already The usage of Polyethylene glycol (PEG) 6000, a microplastic, is rising in industries, medicine, and cosmetic formulations. They serve as ointment bases, capsules, liquid prescription, suppositories and recently used to alleviate constipation. More so, it is one of the pivotal ingredients in soaps, detergents, and also used as chemical intermediates in textile, leather and plastic industries. Additionally, PEG 6000 is widely used in drug delivery systems and tissue engineering. Although approved for internal consumption by regulatory agencies like FDA, there are evidences of its developmental toxicity. Study design, size, duration Fifteen(15) rats weighing between 200-250g were used in this study. The rats were divided into three groups(A, B and C), of 5 rats in each group. Group A was designated as the control group while groups B and C were the test groups. Group A received distilled water, while groups B and C were administered 50 mg/kg and 200 mg/kg body weight of polyethylene glycol PEG6000 respectively via oral gavage daily for 6 weeks. Participants/materials, setting, methods After 6 weeks, the rats were sacrificed as reproductive organs and serum obtained. Enzyme immunosorbent assay (ELISA) was used for hormonal assays. Reproductive organs were weighed and tissue damage was determined using Haemoxylin /eosin staining. Antioxidant enzymes levels (Catalase, Superoxide dismutase), Glutathione peroxidise) and Lipid peroxidation (Malondialdehyde) were also evaluated. Computer-assisted sperm assay (CASA) was used to evaluate sperm parameters and sperm movement kinetics. Main results and the role of chance PEG 6000 caused a reduction in the weights of the rats in groups B and C when compared with control group. There was no significant change in the relative testicular and seminal vesicle organ/body weight ratio in groups B and C, however a pattern of reduction was seen in the organ/body weight ratio of epididymis and prostate, when compared to control group. Serum hormonal levels of Follicle stimulating hormone and Luteinizing hormone were reduced in the test groups when compared to control, but not significant. Testosterone was significantly reduced in groups B and C, while estradiol levels were significantly elevated in groups B and C when compared to control group. There was significant reduction in the levels of Superoxide dismutase, Catalase and Glutathione peroxidase, and a significant increase in Malondialdehyde levels in groups B and C when compared to the control group. Histopathological changes were also seen in the testes, epididymis, seminal vesicles and prostate glands of rats in the test groups. Semen analyses showed statistically significant reductions (p < 0.05) in sperm concentration, motility, vitality and sperm movement kinetics of rats in the test groups when compared to the control group. Limitations, reasons for caution Although this study was done in rats, it may not be easy to extrapolate the present findings to humans. However, this calls for caution as primates and rodents are both members of the superorder Euarchontoglires. Wider implications of the findings Prolonged ingestion of PEG 6000 affects male reproductive system, alters hormonal levels, reduces sperm parameters and induces oxidative stress. Interestingly, the disruptive effects of PEG 6000 are dose-dependent, and may lead to male infertility. Trial registration number not applicable

THE CONTENT OF PROSTANOIDS AND CYCLOOXYGENASES IN COLON TISSUE IN EXPERIMENTAL ULCERATIVE COLITIS

Introduction. The article examines changes in the content of prostaglandins and cyclooxygenases (COX) in colon tissue in ulcerative colitis induced by 2,4-dinitrobenzene sulfonic acid (DNBS) in a 50% ethanol solution. Based on the obtained results, the authors conclude that changes in the content of the studied parameters, except PGI2, are due to ethanol effect, not DNBS. Both COX isozymes are expressed in normal colon and reduced in ulcerative colitis.
 The aim. To study the prostanoids (PGE2, PGI2, PGF2α, TBX2 and 8-iso-PGF2α) and COX-1 and -2 contents in colon tissue in experimental ulcerative colitis.
 Materials and methods. The determination of prostanoids and cyclooxygenases contents in colon tissue by enzyme immunosorbent assay was carried out on three groups of sexually mature laboratory rats of both sexes of the WAG population (1st control group – intrarectal injection of saline; 2nd control group – injection of 50% ethanol; experimental group – injection of DNBS in 50% ethanol).
 Results. PGE2 and PGI2 contents in colon tissue of experimental group rats were statistically significantly higher compared 1st and 2nd control groups. The content of PGE2 was also increased in 2nd control group versus 1st control one. The increasing PGI2 in 2nd control group versus 1st control was not significant. TBX2 and PGF2α contents in experimental and 2nd control groups were significantly lower compared 1st control. 8-iso-PGF2α (non-enzymatically derived prostanoid) level in experimental group rats was significantly higher compared both controls. 8-iso-PGF2α content in 2nd control group was significantly higher compared 1st one. The content of both COX isoforms in colon tissue in experimental group and 2nd control group rats was significantly lower compared to 1st control group.
 Conclusions. Both isoforms of COX are expressed in control group colon indicating COX-2 involvement in supporting physiological functions of normal colon tissue. All studied indicators changes, except PGI2, are due to ethanol, not DNBS. Both 50% ethanol and DNBS in 50% ethanol stimulate lipid peroxidation, confirmed by significant increase in 8-iso-PGF2α content. PGE2 and PGF2α contents changes against the background of reduced levels of COX-1 and COX-2 in experimental ulcerative colitis are most likely an adaptive response aimed at maintaining colon homeostasis. PGI2 content changes are due to DNBS, and not to ethanol.

Intranasal Insulin Alleviates Traumatic Brain Injury by Inhibiting Autophagy and Endoplasmic Reticulum Stress-mediated Apoptosis Through the PI3K/Akt/mTOR Signaling Pathway.

Intranasal insulin reduces lesion size and enhances memory capacity in traumatic brain injury (TBI) models, but the molecular mechanisms behind this neuroprotective action not yet understood. Here we used Feeney's free-falling method to construct TBI mouse models and administrated intranasal insulin, rapamycin, insulin and rapamycin, or normal saline to assess their effects on neurological functions, cerebral edema, and the expression of Iba1 in microglia through immunofluorescence assay. We also measured concentrations of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in the brain using enzyme immunosorbent assay, investigated apoptosis with TUNEL staining and Western blotting, and evaluated autophagy, endoplasmic reticulum (ER) stress, and PI3K/Akt/mTOR signaling pathway with Western blotting. The autophagosome was assessed through transmission electron microscopy. Our findings demonstrated that intranasal insulin promoted neurological recovery, decreased brain swelling, and reduced injury lesions on days 1, 3, and 7 post TBI. Moreover, intranasal insulin reduced microglia activation and the concentration of IL-1β or TNF-α on the same days. Through Western blotting and transmission electron microscopy, we observed that intranasal insulin suppressed autophagy while activating the PI3K/AKT/mTOR signaling pathway on days 1 and 3 post TBI. TUNEL assay and Western blotting also indicated that intranasal insulin inhibited ER stress-mediated apoptosis. Interestingly, the mTOR inhibitor rapamycin partially blocked the pro-autophagy and anti-apoptosis effects of intranasal insulin both on days 1 and 3 post TBI. Our results suggest that intranasal insulin can ameliorate TBI by regulating autophagy and ER stress-mediated apoptosis through the PI3K/AKT/mTOR signaling pathway, providing a promising therapeutic strategy for TBI.

Preliminary assessment of adjuvant activities of Glycine Max (L.) Merr saponin extract in BALB/c mice immunized with hepatitis B virus vaccine

Background: Vaccine adjuvants are used to increase the immunogenicity of weak antigens. Some saponins have adjuvant effects that are exerted via their immune-stimulatory effects and production of cytokines. Soybean (Glycine max (L.) Merr.) contains saponins that could provide affordable vaccine adjuvants. This study aimed to assess the effect of the saponin extracts of soybean on the immune system in BALB/c mice immunized with the hepatitis B virus (HBV) vaccine and hepatitis B surface antigen (HBsAg). Methods: Saponins were extracted from soybean meal and their presence confirmed by foam generation and Fourier-transform infrared methods. A total of 51 mice were immunized in triplicate with 50 µL of various regimens of concentrations of the extracts and either HBV vaccine or HBsAg. The plasma anti-HBsAg antibody titre was determined using an enzyme immunosorbent assay (ELISA) 14 days post-immunization. Gene expression levels of interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF-α), and haematological profile of the blood samples were determined. Results: When the two immunogens were co-administered with the soybean extract, immune response was slightly higher (0.799 + 0.013 for HBsAg and 0.758 + 0.012 for vaccine) than with the Revac B vaccine used alone. However, this difference was not statistically significant (p-value > 0.467, and 0.416 respectively). Unexpectedly, mice immunized with the vaccine expressed less IL-6 levels than the untreated mice (0.603- and 1.469-fold change in transcription (FCT), respectively). Mice treated with the vaccine expressed higher TNF-α levels than the untreated group (28.84 –and 1.00 FCT respectively) while adding the extract significantly reduced the expression of TNF-α (p < 0.063). Administration of immunogens and extract reduced neutrophil (P-value = 0.027) and platelet (p-value > 0.592) counts. Conclusions: The study demonstrated that soybean extract lacked marked adjuvant activity for HBV vaccine, that HBV vaccine suppressed expression of IL-6 and promoted expression of TNF-α.

Serological and molecular evidence of chikungunya virus infection among febrile outpatients seeking healthcare in Northern Malawi

ABSTRACT Introduction: Despite global evidence of chikungunya fever (CHIKF) in humans that is caused by chikungunya virus (CHIKV), little is known about the occurrence of CHIKF in Malawi. This study was conducted to determine the seroprevalence of CHIKF and to molecularly confirm the presence of CHIKV ribonucleic acid (RNA) among febrile outpatients seeking health care at Mzuzu Central Hospital in the Northern Region of Malawi. Methods: Enzyme-immunosorbent assay (ELISA) was used to detect the presence or absence of specific antibodies against CHIKV. Reverse transcription-polymerase chain reaction (RT-PCR) was conducted on randomly selected anti-CHIKV IgM-positive samples to detect CHIKV RNA. Results: Out of 119 CHIKF suspected samples analyzed, 73 tested positive for anti-CHIKV IgM antibodies, with an overall seroprevalence of 61.3%. Most of the CHIKV infected individuals presented with joint pain, abdominal pain, vomiting and nose bleeding with seroprevalence of 45.2%, 41.1%, 16.4% and 12.3%, respectively. All the randomly selected samples that were positive for CHIKV anti-IgM by ELISA had detectable CHIKV RNA by RT-PCR. Conclusion: The presence of anti-CHIKV IgM antibodies suggests the presence of recent CHIKV infection. We therefore recommend for the inclusion of CHIKF as the differential diagnosis in febrile ill patients in Mzuzu city, Malawi.

Sensitivity and performance of three novel quantitative assays of SARS-CoV-2 nucleoprotein in blood

To assess if SARS-CoV-2 (COVID-19) systemic disease can be determined by available nucleoprotein assays, we compared the performance of three commercial SARS-CoV-2 nucleoprotein (N) assays in plasma. A total of 272 plasma samples collected in the period November–December 2021 were analyzed by the methods Simoa SARS CoV‐2 N Protein Advantage Kit [Quanterix Simoa], Solsten SARS-CoV-2 Antigen enzyme immunosorbent assay (ELISA) [Solsten ELISA], and Elecsys SARS‐CoV‐2 Antigen electrochemiluminescence immunoassay [Elecsys ECLIA]. Additionally, a dilution series of inactivated virus culture was analyzed by the three assays. The SARS CoV-2 PCR-status was not known for the patients. Linear correlation in the pairwise correlation between assays as well as linearity of dilution series of inactivated virus culture was estimated by Spearman score. Sensitivity and specificity were estimated by pairwise comparison. The three assays showed poor agreement on patient samples with regards to concentration. Performance on virus culture was excellent but with different level of detection (LOD). Positive vs negative results show comparable sensitivity and specificity of Quanterix Simoa and Solsten ELISA, with a higher LOD in Elecsys ECLIA and thus lower sensitivity and high specificity. N by all tested assays can be used as a marker for systemic COVID-19 disease.

POTENTIALLY DANGEROUS CAUSES OF BACTERIAL DISEASES OF POTATOES IN UKRAINE

An overview of bacterial causative agents of soft (wet) rot of potatoes is given, the epidemiological and etiological aspects of the diseases caused by them are analyzed. When preparing the article, general scientific methods were used: generalization, comparison, system analysis. Data from the European and Mediterranean Plant Protection Organization (EOPO), as well as data from phytosanitary services of EU countries and Ukraine, scientific literature served as material for the analytical study. Trade in plant materials, including potato seed tubers and ornamental plants, is largely responsible for the widespread distribution of pathogens. Locally, pathogens are also spread through plant debris, soil, waterways, aerosols, alternative hosts, and/or farm machinery. The main causative agents of bacterial wet rot of potatoes are gram-negative bacteria of the genera Pectobacterium and Dickeya and quarantine phytopathogens of the genera Clavibacter and Ralstonia. The main methods of detection and identification in asymptomatic potato tubers on an industrial scale are: phytopathological (visual examination of plantations and registration of symptoms of soft rot), microbiological (cultural-morphological and biochemical method, use of test systems for accelerated identification of microorganisms), immunoenzymatic ( enzyme immunosorbent assay), molecular genetic (PCR with specific primers, BIOLOG, DNA fingerprinting and nucleotide sequencing). To date, there are no completely effective pesticides to control all pathogens, so disease control measures will continue to rely primarily on avoiding infection during plant cultivation, and especially during the production of healthy certified seed. For a crop like potatoes, this is primarily based on obtaining bacteria-free minibulbs, applying strict seed certification schemes and strict phytosanitary restrictions. Knowledge of the sources of pathogens and routes of infection should be the basis for the application of phytosanitary measures, especially during and after harvest. Control of quarantine phytopathogens requires special attention. Soft rot pathogens are the main cause of limiting potato production in many regions of the world, particularly Clavibacter michiganensis subsp. sepedonicum and R. solanacearum are quarantine objects of the A-2 list of the European and Mediterranean Organization for Quarantine and Plant Protection. Upon entering our country, R. solanacearum has a high probability of acclimatization and spread in the country.

Laboratory Diagnosis of Human Adenovirus associated with Respiratory Tract Infection in children

Human Adenovirus (HADV) is classified in Adenoviridae family, it is important viral pathogen of respiratory tract infection (RTIs) among children. This study aimed at detecting HADV infections among hospitalized children under 10 years old with respiratory tract infections in Salahaldin Governorate, in addition to compare different diagnostic methods for the detection of this virus.
 Nasopharyngel Swabs (NPS) and whole blood samples were taken from 400 hospitalized children between end of August 2013, to the end of May 2014, each sample was analyzed for HADV by real time –PCR, IgM and IgG specific to HADV determining from blood specimens in serum using Enzyme Immuno sorbent assay (ELISA) technique.
 Nasopharyngel Swab was more sensitive than the whole blood specimens for the detection of HADV and was identified in 3% by RT-PCR, in 72.2% by IgM – HADV antibodies, 83.1% by IgG-HADV antibodies.
 Autumn – Spring periods were considered as the most common time for HADV infection with peak in September and February.

Comparing the Neuroprotective Effects of Telmisartan, Perindopril, and Nebivolol Against Lipopolysaccharide-Induced Injury in Neuron-Like Cells.

The effect of antihypertensive drugs, especially drugs modulating the renin-angiotensin-aldosterone-system (RAAS), on neurodegenerative diseases still needs to be investigated. This study aimed to compare the effects of three different antihypertensive drugs (telmisartan, perindopril, and nebivolol) on neuroprotection and acetylcholine (ACh) levels against lipopolysaccharide (LPS)-induced injury in a differentiated SH-SY5Y cell line. Cells were treated with retinoic acid for differentiation to a neuronal phenotype. LPS 20 (μg/mL) was applied to the cells for one hour. Then, the cells were treated with 1, 5, and 10 µg/mL concentrations of telmisartan, perindopril, and nebivolol separately for 24 hours, except for the control and LPS alone groups. Cell viability was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. ACh levels were analyzed using an enzyme immunosorbent assay in the culture medium. Tumor necrosis factor-alpha (TNF-α), interleukin 1 beta (IL-1β), and nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) expressions were evaluated using western blot analysis. Telmisartan demonstrated the highest cell viability against LPS-induced injury, whereas the protective effect of perindopril was moderate. Nebivolol showed no neuroprotective effect. The protective effect of 10-µg/mL telmisartan was superior to 10 µg/mL perindopril (p=0.006), 5 µg/mL perindopril (p=0.001), 1 µg/mL perindopril (p=0.001), and 1, 5, and 10 µg/mL nebivolol (p<0.001). Among all the study drugs, only telmisartan provided a statistically significant increase in ACh levels after LPS-induced injury. Additionally, the administration of telmisartan provided a concentration-dependent reduction in TNF-α, IL-1β, and NFκB expression against LPS-induced neuroinflammation. These findings suggest that telmisartan has a superior neuroprotective effect against LPS-induced injury in neuron-like cells compared with both perindopril and nebivolol.

PB2302: HEPARIN-INDUCED THROMBOCYTOPENIA ANTIBODIES CAN MAINTAIN PLATELET-ACTIVATING PROPERTIES AFTER TWO SESSIONS OF THERAPEUTIC PLASMA EXCHANGE

Background: Therapeutic plasma exchange (TPE) has been used as a salvage therapy in a small subset of heparin-induced thrombocytopenia (HIT) patients (severe or refractory HIT, before heparin re-exposure during cardiac surgery, in bleeding patients and in autoimmune HIT). Existing data show that HIT antibodies can lose platelet-activating properties after one or two TPE sessions, while more sessions may be needed to decrease the antibody titer. However, results may be assay dependent. The utility of Heparin-Induced Multiple aggregometry (HIMEA) in this setting has not been evaluated. Aims: To assess the effect of TPE on pathogenic HIT antibodies using HIMEA before an emergency surgery for gynecologic malignancy. Methods: TPE was performed in a 72-year-old patient necessitating emergency surgery for a voluminous uterine sarcoma in the acute phase of HIT (10 days after diagnosis). TPE aimed to reduce post-operative thrombotic risk by decreasing pathogenic HIT antibodies in a patient presenting a high risk of post-operative thrombotic complications. Two consecutive sessions of TPE using Spectra Optia ® Apheresis System were performed immediately before surgery. During each plasma exchange 2 L of plasma were removed and replaced by 5% human albumin. Acid-Citrate-Dextrose (ACD) was used as anticoagulant. Blood samples were taken before and after each TPE session (TPE1 pre, TPE1 after, TPE2 pre, TPE2 after). Removed patient’s plasma collected to the plasmapheresis bag was also tested. IgG anti-PF4/heparin antibodies tested with Enzyme Immunosorbent Assay (EIA) (Hyphen ZYMUTEST™ HIA IgG, Hyphen Biomed) and Heparin-Induced Multiple aggregometry (HIMEA) (Multiplate, Roche) in the absence of heparin (HIMEA saline), in the presence of 1UI/ml unfractionated heparin (UFH) (HIMEA low) and in the presence of 100IU/ml UFH (HIMEA high) were assessed. Results: Patient’s 4T’s score was 7 (2 for platelet count fall, 2 for timing, 2 for catheter associated venous thrombosis and 1 for other causes), strongly positive IgG anti-PF4 antibodies (OD: 2.43, cut-off<0.3 units) and a HIMEA profile favoring HIT (typical sigmoid curve at HIMEA low, >50% decrease of AUC at HIMEA high, absence of platelet aggregation at HIMEA saline). She was treated with fondaparinux at 1.5 mg/ day due to sepsis-associated acute kidney injury requiring hemodialysis. Platelet count (PLT) were 168 G/l before surgery. After two TPE sessions antibody titers remained unchanged (TPE1 pre: 2.63 units, TPE1 after:2.19 units, TPE2 pre:2.51 units, TPE2 after:5.52 units). Moreover, platelet reactivity remained also strong as demonstrated by high area under the curve (AUC) values in HIMEA low (figure 1). Plasma from the collection bag showed strong EIA and platelet activation test reactivity. No thrombosis nor thrombocytopenia were observed in one month follow-up. Image:Summary/Conclusion: Two TPE sessions failed to reduce platelet-activating properties of HIT antibodies as assessed by HIMEA in a cancer patient. TPE could be considered as a therapeutic intervention that can reduce thrombotic risk in patients necessitating an urgent non-cardiac surgery in the acute phase of HIT. However, more data is needed on the clinical benefit and the optimal number of TPE sessions in this clinical setting. Functional assays assessing Fcγ receptor mediated platelet activation may help guide the timing of the surgery.

Seroprevalence of Gestational and Neonatal Toxoplasmosis as well as Risk Factors in Yaoundé, Cameroon.

Background Toxoplasmosis, caused by Toxoplasma gondii in pregnant women, is a significant public health problem due to risk of mother to child transmission. The aim of the study was to determine the seroprevalence of toxoplasmosis in pregnant women and corresponding cord blood among women attending Biyem-Assi and CASS Nkoldongo hospitals in Yaoundé, Cameroon. Methods An institutional based cross-sectional study was conducted between June 2019 and May 2020 on 300 pregnant women from late second trimester to third trimester. A total of 259 cord blood samples were collected at birth from these women. Toxoplasma gondii-specific IgG and IgM antibodies in maternal and cord blood were detected using the Toxoplasma Enzyme Immunosorbent Assay kit, and potential risk factors captured through questionnaire were identified using binary logistic regression model. Statistical significance was measured at P < 0.05. Results The overall seroprevalence of gestational and neonatal toxoplasmosis was 80% and 88%, respectively. IgG seropositivity was 72.7%, IgM only was 1.3% and cooccurrence of IgG/IgM was 6% amongst pregnant women. Out of 259 newborn cord bloods, 72.2% were positive for IgG only, 8.9% for IgM only, and 23.9% for both IgG/IgM. Pregnant women 15-24 years (AOR = 4.6, P = 0.011) and women with primary level of education (AOR = 3.9, P = 0.042) were significantly at risk of infection with Toxoplasma gondii. Conclusion Gestational and neonatal toxoplasmosis appears to be more common with higher risk of infection in younger women and less educated women. Hence, these findings will serve as baseline data for further investigations on mother to child transmission of toxoplasmosis in Yaoundé and the need for reinforcement of pregnant women toxoplasmosis-related health measures.

Seroprevalence and Potential Risk Factors of Hepatitis E Virus among Pregnant Women in Khartoum, Sudan

Although hepatitis E virus (HEV) infection has been widely understood as a public health concern in pregnant females, it is sometimes overlooked because of unusual observations of hepatitis E-induced complications during pregnancy. The objective of this descriptive cross-sectional study was to find out how frequent Hepatitis E virus infection is in pregnant women in Sudan. Blood samples were collected from pregnant women (n = 90) attending Khartoum North Hospital from February to December 2019. The specimens were investigated for Hepatitis E virus IgG and IgM antibodies using the commercial kits based on the enzyme immunosorbent assay (ELISA) technique. Out of ninety pregnant women, 36 (40%) were found positive for IgG antibodies, and 5 (6%) were tested positive for IgM antibodies. In addition, 13 (14.5%) of the positive cases are women in their third trimester. These data showed a significant association between previous Hepatitis E virus infection and miscarriage (p &lt;0.001). This study did not find a significant association between maternal and gestation age, education, water sources, and hepatitis E virus infection. In conclusion, there was a high percentage of HEV infections among pregnant women attending Khartoum North Hospital. This study estimates the Hepatitis E virus burden in Khartoum state, but more extensive studies are required to confirm the burden in Sudan.

Laboratory Diagnosis of Human Adenovirus associated with Respiratory Tract Infection in children

Human Adenovirus (HADV) is classified in Adenoviridae family, it is important viral pathogen of respiratory tract infection (RTIs) among children. This study aimed at detecting HADV infections among hospitalized children under 10 years old with respiratory tract infections in Salahaldin Governorate, in addition to compare different diagnostic methods for the detection of this virus. Nasopharyngel Swabs (NPS) and whole blood samples were taken from 400 hospitalized children between end of August 2013, to the end of May 2014, each sample was analyzed for HADV by real time –PCR, IgM and IgG specific to HADV determining from blood specimens in serum using Enzyme Immuno sorbent assay (ELISA) technique. Nasopharyngel Swab was more sensitive than the whole blood specimens for the detection of HADV and was identified in 3% by RT-PCR, in 72.2% by IgM – HADV antibodies, 83.1% by IgG-HADV antibodies. Autumn – Spring periods were considered as the most common time for HADV infection with peak in September and February

Knowledge and Practice of Hepatitis B and C Prevention among Seropositive and Seronegative Prospective Blood Donors in a Tertiary Care Hospital in Eastern Nigeria

Introduction: Hepatitis B and C infections are endemic in Nigeria and the major causes of chronic liver diseases, including hepatocellular carcinoma. One of the major routes of transmission is by transfusion of infected blood or blood products. Awareness of the disease is important, and this study seeks to compare the knowledge and practice of the disease among the seropositive and seronegative prospective blood donors. Aim: To assess the difference in the level of knowledge and preventive practice of blood borne hepatitis among hepatitis B and hepatitis C seropositive and seronegative prospective blood donors. Materials and Methods: The present study was a descriptive survey carried out between June 2019 to January 2020 in the Blood Donor’s Department of the University of Nigeria Teaching Hospital, Enugu, Nigeria. A pretested self/interviewer’s administered questionnaire was used to assess the knowledge and practice among 100 hepatitis B and hepatitis C seropositive and 305 seronegative prospective blood donors. The hepatitis B and C seropositive subjects were screened using Enzyme Immunosorbent Assay (ELISA) technique. The findings and scores were analysed using IBM Corporation 2019 Statistical Package for the Social Sciences (SPSS) software version 21.0. Results: Total of 305 respondents were seronegative while 82 and 18 were hepatitis B and C seropositive, respectively. The mean age of the study participants was 27.78±7.17 years, whereas it was 28.09±6.36 years and 27.46±7.968 years among seropositive and seronegative respondents, respectively. In the study, 98 (98%) males and 2 (2%) females were in the seropositive group while, 268 (87.9%) males and 37 (12.1%) females were in seronegative group. The overall knowledge score and practice score were 56.3% and 28.1%, respectively. In the seropositive group, the average knowledge score was 27.3% and the average practice score was 17%. While in the seronegative group, the average knowledge score was 65.7% and the average score on practice score was 31.8%. There was a significant association between seropositive and being a male (p-value=0.001), less educated (p-value=0.002), currently unmarried (p-value=0.029), knowledge above average (p-value=0.001), and practice below average (p=0.002). None of the respondents had hepatitis B vaccination. Conclusion: Based on the results obtained from the present study, it is evident that the level of knowledge and practice among the seropositive respondent is significantly low when compared with the seronegative groups. There is a need for a serious awareness campaign on the transmission and prevention of hepatitis B and C, and the possibility of extending hepatitis B immunisation coverage in the country owing to the increasing burden of the disease.

Faecal Triiodothyronine Hormone Levels in Female Iguana during Weekday and Weekend in In Situ Conservation

Iguanas are exotic animals that are widely kept as pets in our society,their performance and color being their main attraction. Jogja Exotarium is anin situ conservation park that carries the back to nature concept. Animals thatare kept at the Jogja Exotarium are allowed direct contact with visitors. Thisstudy aims to determine the hormone triiodothyronine in female iguanas that are raised and conserved in situ. This study used feces obtained from iguanas kept in Jogja Exotarium, sampling was carried out on Wednesdays and Sundays for 2 weeks. The collected faecal samples were processed by usingfreeze-drying method and then extracted using 80% methanol. The extracted samples were analyzed using the enzyme immunosorbent assay (EIA) method. The results showed during the week day was 107.1 ± 7.01 ng/gr dry feces, while the week end was 119.95 ± 4.3 ng/gr dry feces. Based on the results of the study, it can be concluded that there is no difference in the hormone triiodothyronine in female iguanas during weekday and weekend

Determination of glyphosate in breast milk of lactating women in a rural area from Paraná state, Brazil.

The aim of this study was to verify the presence of glyphosate in breast milk and to characterize maternal environmental exposure. Sixty-seven milk samples were collected from lactating women in the city of Francisco Beltrão, Paraná, living in urban (n=26) and rural (n=41) areas, at the peak of glyphosate application in corn and soy crops in the region (April and May 2018). To characterize the study population, socio-epidemiological data of the women were collected. To determine glyphosate levels, a commercial enzyme immunosorbent assay kit was used. Glyphosate was detected in all breast milk samples analyzed with a mean value of 1.45 µg/L. Despite some descriptive differences, there were no statistically significant differences (P<0.05) between the categories of the variables tested. Also, glyphosate was detected in drinking water samples from the urban area and in artesian well water from the rural area of the region where the studied population lived. The estimation of the total amount of glyphosate ingested by breastfeeding babies in a period of 6 months was significant. These results suggest that the studied lactating population was contaminated with glyphosate, possibly through continued environmental exposure.

Soluble Complement Component 1q Receptor 1 (sCD93) Is Associated with Graft Function in Kidney Transplant Recipients.

Cluster of differentiation 93 (CD93), also known as complement component 1q receptor 1 is a transmembrane glycoprotein expressed in endothelial and hematopoietic cells and associated with phagocytosis, cell adhesion, angiogenesis and inflammation. The extracellular part, soluble CD93 (sCD93), is released to body fluids in inflammation. Data on sCD93 in kidney diseases are limited. Our aim was to evaluate serum sCD93 in long-term kidney transplant recipients as a marker of inflammation and endothelial dysfunction that may be potentially useful in early recognition of graft dysfunction. Seventy-eight adult patients with functioning kidney graft and stable clinical state were examined at least one year after kidney transplantation. Serum sCD93 was measured by enzyme immunosorbent assay. Estimated glomerular filtration rate (eGFR) and albuminuria or proteinuria were assessed at baseline and over one-year follow-up. Increased sCD93 was associated with lower baseline eGFR independently of the confounders. Moreover, sCD93 was negatively associated with eGFR during one-year follow-up in simple analysis; however, this was not confirmed after adjustment for confounders. Baseline sCD93 was positively associated with baseline albuminuria and with increased proteinuria during the follow-up. Serum sCD93 was not correlated with other studied inflammatory markers (interleukin 6, C-reactive protein, procalcitonin and C3 and C4 complement components). To the best of our knowledge, this is the first report regarding the concentrations of sCD93 in kidney transplant recipients and one of the first reports showing the inverse association between sCD93 and renal function. Serum sCD93 should be further evaluated as a diagnostic and prognostic marker in renal transplantation.

Diurnal relationship between core clock gene BMAL1, antioxidant SOD1 and oxidative RNA/DNA damage in young and older healthy women.

BMAL1 is a core clock gene that positively regulates circadian rhythms. In animals, BMAL1 further acts as transcription factor for the SOD1 gene which encodes the major antioxidant enzyme superoxide dismutase. SOD1 protects against oxidative damage that is a major factor for human aging. Mice lacking mBmal1 exhibit premature aging phenotypes which might be related to a reduced expression of SOD1. The purpose of this study was to explore the circadian relationship between BMAL1, SOD1, oxidative RNA/DNA damage and aging in healthy humans. In a cross-sectional study design, buccal mucosa cells and saliva samples were obtained from 21 young (23.7±2.3yrs) and 21 older women (66.8±5.7yrs) within 24h at 4-h intervals (08:00, 12:00, 16:00, 20:00, 24:00 and 04:00h). Transcript levels of BMAL1 and SOD1 were measured by real-time quantitative PCR. Protein levels of SOD1 were determined by immunoblotting and densitometry. Levels of oxidative RNA/DNA damage and melatonin were quantified by enzyme immunosorbent assays. Transcript levels of BMAL1 and SOD1 mRNAs as well as protein levels of SOD1 and melatonin exhibited significant 24-h variation in each age group (P<0.010, Friedman tests). The mRNA expression patterns of BMAL1 and SOD1 showed similar 24-h rhythmicity and positive relationships were found. Strongest relationships occurred in young women at 12:00h (rs=0.81, P=0.005) and in older women at 08:00h (r=0.84, P<0.001). Maximum levels of SOD1 mRNA appeared within 24h in both age groups at 24:00h. In both age groups, the timing of maximum level for SOD1 protein was delayed relative to the timing of maximum level for SOD1 mRNA. This delay was larger in older women (8h) compared to young women (4h). Older women showed higher oxidative RNA/DNA damage at all time-points compared to young women (P<0.020). The oxidative RNA/DNA damage decreased continuously from 08:00 to 20:00h (P<0.001) in both age groups. Although oxidative damage and SOD1 protein levels declined simultaneously, only weak and non-significant relationships were noted. This study demonstrates that transcript levels of SOD1 and protein level of SOD1 follow a circadian pattern of expression in healthy young and older women. The time-shift found between the 24-h maximum levels of SOD1 mRNA and SOD1 protein could be explained by the time needed for translation of SOD1 protein. The positive relationship found between expression levels of SOD1 mRNA and BMAL1 mRNA is in line with animal studies showing that BMAL1 acts as transcription factor and regulates the circadian synthesis of SOD1 mRNA. Differences between young and older women found in the timing of 24-h maxima as well as differences observed in 24-h relationships between characters might be due to age-related alterations in the circadian system. Absence of relationships between levels of SOD1 and oxidative RNA/DNA damage argue for a more complex interaction between the antioxidant system and the circadian system.

GP88/PGRN Serum Levels Are Associated with Prognosis for Oral Squamous Cell Carcinoma Patients.

Progranulin (PGRN)/GP88 is a growth factor that is expressed in a wide range of tumor tissues. The secreted form is involved in various biological processes including proliferation and inflammation. In several tumor types, the serum GP88 level is associated with a patient's prognosis; however, data for oral squamous cell carcinomas (OSCCs) have not yet been reported. We measured the serum GP88 levels in 96 OSCC patients by an enzyme immunosorbent assay (EIA) and correlated these data with clinicopathological parameters and patient outcomes. The GP88 levels in the serum of OSCC patients and healthy volunteers were comparable. In OSCC patients, the levels did not correlate with age, sex, or TNM status. In a Kaplan-Meier survival analysis, a serum GP88 level < 68 ng/mL was significantly associated with worsened survival (p = 0.0005, log-rank-test) as well as in uni- and multivariate Cox regression analyses (RR = 4.6 [1.6-12.9], p = 0.004 and RR = 4.2 [1.2-12.0], p = 0.008). This effect was predominant in OSCC patients older than 60.5 years (p = 0.027), while in younger patients no significant association between serum GP88 levels and prognosis could be observed. Altogether, lower serum GP88 levels are significantly associated with a worsened outcome for an OSCC and may be an interesting candidate for risk stratification during OSCC therapy.