Search results for: enzyme-linked immunosorbent assay (ELISA)

Authors: A. M. Gadanya, B. A. Ahmad, U. Maigatari

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Effect of oral administration of Gadagi tea on superoxide dismutase activity was assessed on twenty (20) male subjects (aged 21-40years). Ten (10) male non Gadagi tea consumers (aged 20-26 years), were used as control. Blood samples were collected from the subjects and analysed for serum superoxide dismutase activity using R&D Enzyme Linked Immunosorbent Assay method (ELISA). The subjects were grouped into four based on age i.e group I (21-25 years), group II (26-30 years), and also based on duration of the tea consumption, i.e group A (5-9 years) , group B (10-14 years), group C (15-19 years) and group D (20-24 years). The subjects in group I (0.12 U mg-l +0.05), group II (0.11 U mg-l +0.01), group III (0.14 U mg-l +0.08) and group IV (0.17 U mg-l +0.11) showed increased activity of serum superoxide dismutase when compared with the control subjects (0.88 U mg-l +0.02) (P<0.05). There was no statistical significant difference in superoxide dismutase activity within the case groups (P<0.05), based on age and duration of consumption of the tea. Thus, Gadagi tea consumption could increase serum superoxide dismutase activity in humans.

Keywords: “Gadagi” tea, Serum, Superoxide dismutase, Humans.

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Authors: Ezeugwunne Ifeoma Priscilla, Charles Chinedum Onyenekwe, Joseph Eberendu Ahaneku, Rosemary Adanma Analike, Adesuwa Peace Eidangbe

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This study was designed to assess the effect of malaria parasitaemia on serum reproductive hormone levels of asymptomatic HIV adult subjects. A total of 271 participants aged between 17 and 58 ears were conveniently recruited. 135 asymptomatic HIV-infected subjects participated in the study; 67 of them had malaria parasitaemia. 136 HIV seropositive control subjects, 68 of them had malaria parasitaemia. Blood samples were collected from the participants for the determination of HIV status by immunoassay and immunochromatography. Enzyme-linked immunosorbent assay (ELISA) was used to assay for serum LH, FSH, Estrogen, testosterone, progesterone, prolactin, and PSA levels, CD4+T cell counts by Cyflow method, thick and thin films determination of malaria parasitaemia count and density by WHO. Student's t-tests and ANOVA were used to compare means. P<0.05 was considered statistically significant. The results showed significant differences in serum levels of LH, FSH, PSA, estrogen, progesterone, and testosterone amongst the groups at P<0.05, respectively. The serum levels of LH, FSH, and PSA were significantly higher in malaria-infected asymptomatic HIV subjects than in asymptomatic HIV subjects with malaria parasitaemia (P<0.05 in each case). Also, the serum levels of LH, FSH, PSA, estrogen, and progesterone were significantly higher in malaria-infected asymptomatic HIV subjects compared with malaria-infected HIV seronegative subjects (P<0.05, respectively). The mean MP counts and MP density were significantly higher in asymptomatic HIV subjects compared to HIV seronegative subjects (P<0.05, in each case). The mean serum levels of testosterone were significantly lower in both malaria-infected and malaria uninfected HIV seronegative subjects (P<0.05, in each case). In conclusion, Malaria and HIV co-infection might increase the burden of hypogonadism as well as primary testicular failure, hyperprogesteronaemia, elevated levels of estrogen, and PSA in adult males asymptomatic HIV subjects.

Keywords: malaria parasitaemia, HIV, CD4, reproductive hormones

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Authors: Minh-Phuong Ngoc Bui, Abdennour Abbas

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Microbial spoilage of food/drug has been a constant nuisance and an unavoidable problem throughout history that affects food/drug quality and safety in a variety of ways. A simple and rapid test of fungi and bacteria in food/drugs and environmental clinical samples is essential for proper management of contamination. A number of different techniques have been developed for detection and enumeration of foodborne microorganism including plate counting, enzyme-linked immunosorbent assay (ELISA), polymer chain reaction (PCR), nucleic acid sensor, electrical and microscopy methods. However, the significant drawbacks of these techniques are highly demand of operation skills and the time and cost involved. In this report, we introduce a rapid method for detection of bacteria and fungi in food/drug products using a specific interaction between a reducing agent (tris(2-carboxylethyl)phosphine (TCEP)) and the microbial surface proteins. The chemical reaction was transferred to a transduction system using gold nanoplates-enhanced chemiluminescence. We have optimized our nanoplates synthetic conditions, characterized the chemiluminescence parameters and optimized conditions for the microbial assay. The new detection method was applied for rapid detection of bacteria (E.coli sp. and Lactobacillus sp.) and fungi (Mucor sp.), with limit of detection as low as single digit cells per mL within 10 min using a portable luminometer. We expect our simple and rapid detection method to be a powerful alternative to the conventional plate counting and immunoassay methods for rapid screening of microorganisms in food/drug products.

Keywords: microorganism testing, gold nanoplates, chemiluminescence, reducing agents, luminol

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Authors: Mohammadjavad Mehrabanpour, Maryam Ranjbar Bushehri, Dorsa Mehrabanpour

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Respiratory diseases are the most important problems in the country’s poultry industry, particularly when it comes to broiler flocks. Ornithobacterium rhinotracheale (ORT) is a species that causes poor performance in growth rate, egg production, and mortality. This pathogen causes a respiratory infection including pulmonary alveolar inflammation, and pneumonia of birds throughout the world. Newcastle disease (ND) is a highly contagious disease in poultry, and also, it causes considerable losses to the poultry industry. The aim of this study was to evaluate the simultaneous occurrence of ORT and ND and NDV isolation by inoculation in embryonated eggs and confirmed by RT-PCR in broiler chicken flocks in Fars province. In this study, 318 blood and 85 tissue samples (brain, trachea, liver, and cecal tonsils) were collected from 15 broiler chicken farms. Survey serum antibody titers against ORT by using a commercial enzyme-linked immunosorbent assay (ELISA) kit performed. Evaluation of antibody titer against ND virus is performed by hemagglutination inhibition test. Virus isolation with chick embryo eggs 9-11 and RT-PCR method were carried out. A total of 318 serum samples, 135 samples (42.5%) were positive for antibodies to ORT and titer of HI antibodies against NDV in 122 serum samples (38/4%) were 7-10 (log2) and 61 serum samples (19/2%) had occurrence antibody titer against Newcastle virus and ORT. Results of the present study indicated that 20 tissue samples were positive in embryonated egg and in rapid hemagglutination (HA) test. HI test with specific ND positive serum confirmed that 6 of 20 samples. PCR confirmed that all six samples were positive and PCR products of samples indicated 535-base pair fragments in electrophrosis. Due to the great economic importance of these two diseases in the poultry industry, it is necessary to design and implement a comprehensive plan for prevention and control of these diseases.

Keywords: ELISA, Ornithobacterium rhinotracheale, newcastle disease, seroprevalence

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Authors: Tauseef Ahmad, Muhammad Ishaq, Mathew Eapen, Ahyoung Park, Sam Karpiniec, Vanni Caruso, Rajaraman Eri

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Fucoidans are complex, fucose-rich sulfated polymers discovered in brown seaweeds. Fucoidans are popular around the world, particularly in the nutraceutical and pharmaceutical industries, due to their promising medicinal properties. Fucoidans have been shown to have a variety of biological activities, including anti-inflammatory effects. They are known to inhibit inflammatory processes through a variety of mechanisms, including enzyme inhibition and selectin blockade. Inflammation is a part of the complicated biological response of living systems to damaging stimuli, and it plays a role in the pathogenesis of a variety of disorders, including arthritis, inflammatory bowel disease, cancer, and allergies. In the current investigation, various fucoidan extracts from Undaria pinnatifida, Fucus vesiculosus, Macrocystis pyrifera, Ascophyllum nodosum, and Laminaria japonica were assessed for inhibition of pro-inflammatory cytokine production (TNF-α, IL-1β, and IL-6) in LPS induced human macrophage cell line (THP-1) and human peripheral blood mononuclear cells (PBMCs). Furthermore, we also sought to catalogue these extracts based on their anti-inflammatory effects in the current in-vitro cell model. Materials and Methods: To assess the cytotoxicity of fucoidan extracts, MTT (3-[4,5-dimethylthiazol-2-yl]-2,5, -diphenyltetrazolium bromide) cell viability assay was performed. Furthermore, a dose-response for fucoidan extracts was performed in LPS induced THP-1 cells and PBMCs after pre-treatment for 24 hours, and levels of TNF-α, IL-1β, and IL-6 cytokines were measured using Enzyme-Linked Immunosorbent Assay (ELISA). Results: The MTT cell viability assay demonstrated that fucoidan extracts exhibited no evidence of cytotoxicity in THP-1 cells or PBMCs after 48 hours of incubation. The results of the sandwich ELISA revealed that all fucoidan extracts suppressed cytokine production in LPS-stimulated PBMCs and human THP-1 cells in a dose-dependent manner. Notably, at lower concentrations, the lower molecular fucoidan (5-30 kDa) extract from Macrocystis pyrifera was a highly efficient inhibitor of pro-inflammatory cytokines. Fucoidan extracts from all species including Undaria pinnatifida, Fucus vesiculosus, Macrocystis pyrifera, Ascophyllum nodosum, and Laminaria japonica exhibited significant anti-inflammatory effects. These findings on several fucoidan extracts provide insight into strategies for improving their efficacy against inflammation-related diseases. Conclusion: In the current research, we have successfully catalogued several fucoidan extracts based on their efficiency in LPS-induced macrophages and PBMCs in downregulating the key pro-inflammatory cytokines (TNF-, IL-1 and IL-6), which are prospective targets in human inflammatory illnesses. Further research would provide more information on the mechanism of action, allowing it to be tested for therapeutic purposes as an anti-inflammatory medication.

Keywords: fucoidan, PBMCs, THP-1, TNF-α, IL-1β, IL-6, inflammation

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Authors: Manal M. Kame, Hanan G. El-Baz, Zeinab A.Demerdash, Engy M. Abd El-Moneem, Mohamed A. Hendawy, Ibrahim R. Bayoumi

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There is a constant need to improve the performance of current diagnostic assays of schistosomiasis as well as develop innovative testing strategies to meet new testing challenges. This study aims at increasing the diagnostic efficiency of monoclonal antibody (MAb)-based antigen detection assays through gold nanoparticles conjugated with specific anti-Schistosoma mansoni monoclonal antibodies. In this study, several hybidoma cell lines secreting MAbs against adult worm tegumental Schistosoma antigen (AWTA) were produced at Immunology Department of Theodor Bilharz Research Institute and preserved in liquid nitrogen. One MAb (6D/6F) was chosen for this study due to its high reactivity to schistosome antigens with highest optical density (OD) values. Gold nanoparticles (AuNPs) were functionalized and conjugated with MAb (6D/6F). The study was conducted on serum samples of 116 subjects: 71 patients with S. mansoni eggs in their stool samples group (gp 1), 25 with other parasites (gp2) and 20 negative healthy controls (gp3). Patients in gp1 were further subdivided according to egg count in their stool samples into Light infection {≤ 50 egg per gram(epg) (n= 17)}, moderate {51-100 epg (n= 33)} and severe infection {>100 epg(n= 21)}. Sandwich ELISA was performed using (AuNPs -MAb) for detection of circulating schistosomal antigen (CSA) levels in serum samples of all groups and the results were compared with that after using MAb/ sandwich ELISA system. Results Gold- MAb/ ELISA system reached a lower detection limit of 10 ng/ml compared to 85 ng/ml on using MAb/ ELISA and the optimal concentrations of AuNPs -MAb were found to be 12 folds less than that of MAb/ ELISA system for detection of CSA. The sensitivity and specificity of sandwich ELISA for detection of CSA levels using AuNPs -MAb were 100% & 97.8 % respectively compared to 87.3% &93.38% respectively on using MAb/ ELISA system. It was found that CSA was detected in 9 out of 71 S.mansoni infected patients on using AuNPs - MAb/ ELISA system and was not detected by MAb/ ELISA system. All those patients (9) was found to have an egg count below 50 epg feces (patients with light infections). ROC curve analyses revealed that sandwich ELISA using gold-MAb was an excellent diagnostic investigator that could differentiate Schistosoma patients from healthy controls, on the other hand it revealed that sandwich ELISA using MAb was not accurate enough as it could not recognize nine out of 71 patients with light infections. Conclusion Our data demonstrated that: Loading gold nanoparticles with MAb (6D/6F) increases the sensitivity and specificity of sandwich ELISA for detection of CSA, thus active (early) and light infections could be easily detected. Moreover this binding will decrease the amount of MAb consumed in the assay and lower the coast. The significant positive correlation that was detected between ova count (intensity of infection) and OD reading in sandwich ELISA using gold- MAb enables its use to detect the severity of infections and follow up patients after treatment for monitoring of cure.

Keywords: Schistosomiasis, nanoparticles, gold, monoclonal antibodies, ELISA

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Authors: D. Jablonskytė-Raščė, A. Mankevičienė, S. Supronienė, I. Kerienė, S. Maikštėnienė, S. Bliznikas, R. Česnulevičienė

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Organic farming does not allow the use of conventional mineral fertilizers and crop protection products. As a result, in our experiments we chose to grow different species of cereals and to see how cereal species affects mycotoxin accumulation. From the phytopathological and entomological viewpoint, the glumes of spelt grain perform a positive role since they protect grain from the infection of pathogenic microorganisms. On the background of the above-mentioned infection, there were more Fusarium–affected grains of spelt than of common wheat. It can be assumed that spelt is more susceptible to the Fusarium fungi infection than common wheat. This study describes the occurrence of DON, ZEA and T2/HT2 toxin in a survey of spelt and common wheat and their bran as well as flour. The analysis was conducted using the enzyme-linked immunosorbent assay (ELISA) method. The concentrations of DON, ZEA, and T2/HT2 in Triticum spelta and Triticum aestivum are influenced by species, cereal type and year interaction. The highest concentration of mycotoxin was found in spelt grain with glumes. The obtained results indicate the significantly higher concentrations of Fusarium toxins in glumes than in dehulled grain which implicate the possible protective effect of spelt wheat glumes. The lowest DON, ZEA, and T2/HT2 concentration was determined in spelt grain without glumes.

Keywords: Fusarium mycotoxins, organic farming, spelt

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Authors: Hatem A. El-Mezayen, Hossam Darwesh

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Background/Aim: Hepatocellular carcinoma (HCC) is often diagnosed at advanced stage where effective therapies are lacking. Identification of new scoring system is needed to discriminate HCC patients from those with chronic liver disease. Based on the link between vascular endothelial growth factor (VEGF) and HCC progression, we aimed to develop a novel score based on combination of VEGF and routine laboratory tests for early prediction of HCC. Methods: VEGF was assayed for HCC group (123), liver cirrhosis group (210) and control group (50) by Enzyme Linked Immunosorbent Assay (ELISA). Data from all groups were retrospectively analyzed including α feto protein (AFP), international normalized ratio (INR), albumin and platelet count, transaminases, and age. Areas under ROC curve were used to develop the score. Results: A novel index named hepatocellular carcinoma-vascular endothelial growth factor score (HCC-VEGF score)=1.26 (numerical constant) + 0.05 ×AFP (U L-1)+0.038 × VEGF(ng ml-1)+0.004× INR –1.02 × Albumin (g l-1)–0.002 × Platelet count × 109 l-1 was developed. HCC-VEGF score produce area under ROC curve of 0.98 for discriminating HCC patients from liver cirrhosis with sensitivity of 91% and specificity of 82% at cut-off 4.4 (ie less than 4.4 considered cirrhosis and greater than 4.4 considered HCC). Conclusion: Hepatocellular carcinoma-VEGF score could replace AFP in HCC screening and follow up of cirrhotic patients.

Keywords: Hepatocellular carcinoma, cirrhosis, HCV, diagnosis, tumor markers

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Authors: Aml M. El-Sharkawy, Hossam M. Darwesh

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Abstract— Background/Aim: Hepatocellular carcinoma (HCC) is often diagnosed at advanced stage where effective therapies are lacking. Identification of new scoring system is needed to discriminate HCC patients from those with chronic liver disease. Based on the link between tumor associated trypsin inhibitor (TATI) and HCC progression, we aimed to develop a novel score based on combination of TATI and routine laboratory tests for early prediction of HCC. Methods: TATI was assayed for HCC group (123), liver cirrhosis group (210) and control group (50) by Enzyme Linked Immunosorbent Assay (ELISA). Data from all groups were retrospectively analyzed including α feto protein (AFP), international normalized ratio (INR), albumin and platelet count, transaminases, and age. Areas under ROC curve were used to develop the score. Results: A novel index named hepatocellular carcinoma-vascular endothelial growth factor score (HCC-TATI score) = 3.1 (numerical constant) + 0.09 ×AFP (U L-1) + 0.067 × TATI (ng ml-1) + 0.16 × INR – 1.17 × Albumin (g l-1) – 0.032 × Platelet count × 109 l-1 was developed. HCC-TATI score produce area under ROC curve of 0.98 for discriminating HCC patients from liver cirrhosis with sensitivity of 91% and specificity of 82% at cut-off 6.5 (ie less than 6.5 considered cirrhosis and greater than 4.4 considered HCC). Conclusion: Hepatocellular carcinoma-TATI score could replace AFP in HCC screening and follow up of cirrhotic patients.

Keywords: Hepatocellular carcinoma, cirrhosis, HCV, diagnosis, TATI

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Authors: Lorena GóMez Escorcia, Gustavo Aroca MartíNez, Jose Luiz Villarreal, Elkin Navarro Quiroz

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Lupus nephritis (LN) is a high-cost disease, occurring in about half of patients with Systemic Lupus Erythematosus (SLE). Renal biopsy constitutes the only protocol that, to date, allows a correct diagnosis of the level of renal involvement in these patients. However, this procedure can have various adverse effects such as kidney bleeding, muscle bleeding, infection, pain, among others. Therefore, the development of new diagnostic alternatives is required. The neutrophil gelatinase-associated lipocalin (NGAL) has been emerging as a novel biomarker of acute kidney injury. The aim of this study was to assess urinary NGAL levels as a marker for disease activity in patients with lupus nephritis. For this work included 50 systemic lupus erythematosus (SLE) patients, 50 with active lupus nephritis (LN), and 50 without autoimmune and renal disease as controls. TNGAL in urine samples was measured by enzyme-linked immunosorbent assay (ELISA). The results revealed that patients with kidney damage had an elevated urinary NGAL as compared to patients with lupus without kidney damage and controls (p <0.005), and the mean of uNGAL was (28.72 ± 4.53), (19.51 ± 4.72), (8.91 ± 3.37) respectively. Measurement of urinary NGAL levels showed a very good diagnostic performance for discriminating patients with Lupus nephritis from SLE without renal damage and of control individuals.

Keywords: lupus nephritis, biomarker, NGAL, urine samples

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Authors: Harika Atmaca, Emir Bozkurt, Latife Merve Oktay, Selim Uzunoglu, Ruchan Uslu, Burçak Karaca

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Microarrays have been developed for highly parallel enzyme-linked immunosorbent assay (ELISA) applications. The most common protein arrays are produced by using multiple monoclonal antibodies, since they are robust molecules which can be easily handled and immobilized by standard procedures without loss of activity. Protein expression profiling with protein array technology allows simultaneous analysis of the protein expression pattern of a large number of proteins. Trabectedin, a tetrahydroisoquinoline alkaloid derived from a Caribbean tunicate, Ecteinascidia turbinata, has been shown to have antitumor effects. Here, we used a novel proteomic approach to explore the mechanism of action of trabectedin in prostate cancer cell line PC-3 by apoptosis antibody microarray. XTT cell proliferation kit and Cell Death Detection Elisa Plus Kit (Roche) was used for measuring cytotoxicity and apoptosis. Human Apoptosis Protein Array (R&D Systems) which consists of 35 apoptosis related proteins was used to assess the omic protein expression pattern. Trabectedin induced cytotoxicity and apoptosis in prostate cancer cells in a time and concentration-dependent manner. The expression levels of the death receptor pathway molecules, TRAIL-R1/DR4, TRAIL R2/DR5, TNF R1/TNFRSF1A, FADD were significantly increased by 4.0-, 21.0-, 4.20- and 11.5-fold by trabectedin treatment in PC-3 cells. Moreover, mitochondrial pathway related pro-apoptotic proteins Bax, Bad, Cytochrome c, and Cleaved Caspase-3 expressions were induced by 2.68-, 2.07-, 2.8-, and 4.5-fold and the expression levels of anti-apoptotic proteins Bcl-2 and Bcl-XL were reduced by 3.5- and 5.2-fold in PC-3 cells. Proteomic (antibody microarray) analysis suggests that the mechanism of action of trabectedin may be exerted via the induction of both intrinsic and extrinsic apoptotic pathways. The antibody microarray platform can be utilised to explore the molecular mechanism of action of novel anticancer agents.

Keywords: trabectedin, prostate cancer, omic protein expression profile, apoptosis

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Authors: Ibrahim Djelloul Berkane

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A survey was conducted to assess the presence of the virus in Citrus tristeza one of the main citrus regions of Algeria, namely the Chlef region, using the technique of Direct Tissue Print Immunoprinting Assay (DTBIA) and the Double Sandwich ELISA antibodies. A nursery citrus, lumber yards, and commercial orchards, which are the main varieties cultivated citrus were subjected to samples collected samples for laboratory analysis. 0.91% of the plants tested orchards were infected with CTV, while no positive case was detected at the nursery the yard, however, it is reported that an alarming rate of 10,5% of orchards tested at the common Chettia were infected with tristeza virus. The investigation was launched to identify the vector species tristeza revealed the presence of a vector is important Aphis gossypii.

Keywords: aphis, chlef, citrus, DAS-ELISA, DTBIA, tristeza

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Authors: Ghebremedhin Tsegay, Weldu Tesfagaber, Yuanmao Zhu, Xijun He, Wan Wang, Zhenjiang Zhang, Encheng Sun, Jinya Zhang, Yuntao Guan, Fang Li, Renqiang Liu, Zhigao Bu, Dongming Zhao*

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African swine fever (ASF) is a highly infectious viral disease of pigs, resulting in significant economic loss worldwide. As there is no approved vaccines and treatments, the control of ASF entirely depends on early diagnosis and culling of infected pigs. Thus, highly specific and sensitive diagnostic assays are required for accurate and early diagnosis of ASF virus (ASFV). Currently, only a few recombinant proteins have been tested and validated for use as reagents in ASF diagnostic assays. The most promising ones for ASFV antibody detection were p72, p30, p54, and pp62. So far, three ELISA kits based on these recombinant proteins have been commercialized. Due to the complex nature of the virus and variety forms of the disease, robust serodiagnostic assays are still required. ASFV p22 protein, encoded by KP177R gene, is located in the inner membrane of viral particle and appeared transiently in the plasma membrane early after virus infection. The p22 protein interacts with numerous cellular proteins, involved in processes of phagocytosis and endocytosis through different cellular pathways. However, p22 does not seem to be involved in virus replication or swine pathogenicity. In this study, E.coli expressed recombinant p22 protein was used to generate a monoclonal antibody (mAb), and its potential use for the development of blocking ELISA (bELISA) was evaluated. A total of 806 pig serum samples were tested to evaluate the bELISA. Acording the ROC (Reciever operating chracteristic) analysis, 100% sensitivity and 98.10% of specificity was recorded when the PI cut-off value was set at 47%. The novel assay was able to detect the antibodies as early as 9 days post infection. Finaly, a highly sensitive, specific and rapid novel p22-mAb based bELISA assay was developed, and optimized for detection of antibodies against genotype I and II ASFVs. It is a promising candidate for an early and acurate detection of the antibodies and is highly expected to have a valuable role in the containment and prevention of ASF.

Keywords: ASFV, blocking ELISA, diagnosis, monoclonal antibodies, sensitivity, specificity

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Authors: P. O. Ughachukwu, P. O. Okonkwo, P. C. Unekwe, J. O. Ogamba

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Background: The prevalence of hepatitis B viral infection is high worldwide with liver cirrhosis and hepatocellular carcinoma as important complications. Cases of poor antibody response to hepatitis B vaccination abound. Immunosuppression, especially from glucocorticoids, is often cited as a cause of poor antibody response and there are documented evidences of irrational administration of glucocorticoids to children and adults. The study was, therefore, designed to find out if administration of glucocorticoids affects immune response to vaccination against hepatitis B in mice. Methods: Mice of both sexes were randomly divided into 2 groups. Daily intramuscular methylprednisolone injections, (15 mg kg-1), were given to the test group while sterile deionized water (0.1ml) was given to control mice for 30 days. On day 6 all mice were given 2 μg (0.1ml) hepatitis B vaccine and a booster dose on day 27. On day 34, blood samples were collected and analyzed for anti-HBs titres using enzyme-linked immunosorbent assay (ELISA). Statistical analysis was done using Graph Pad Prism 5.0 and the results taken as statistically significant at p value < 0.05. Results: There were positive serum anti-HBs responses in all mice groups but the differences in titres were not statistically significant. Conclusions: At the dosages and length of exposure used in this study, methylprednisolone injection did not significantly inhibit anti-HBs response in mice following immunization against hepatitis B virus. By extrapolation, methylprednisolone, when used in the usual clinical doses and duration of therapy, is not likely to inhibit immune response to hepatitis B vaccinations in man.

Keywords: anti-HBs, hepatitis B vaccine, immune response, methylprednisolone, mice

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Authors: Seyed Mehrzad Pahlavani, Mozaffar Haji Jafari Anaraki, Sayma Mohammadi

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A total of 34700 day-old broilers were exposed to green, blue and yellow light using a light-emitting diode system for 6 weeks to investigate the effects of light wave length on antibody responses to Newcastle disease by HI test and the correlation with ELISA. 3 poultry house broiler farms with the same conditions was selected and the lightening system of each was set according to the requirement. Blood samples were taken from 20 chicks on days 1, 24 and 46 and the Newcastle virus specific antibody was titered in serum using HI an ELISA test. On day 24, the probability value of more than 0/05 was observed in HI and ELISA tests of all groups while at the end of breeding period, the average HI serum antibody titer was more in the green light than the yellow one while the blue light was not significantly different from both. At the last titration, the green light has got the highest titer of Newcastle antibodies. There were no significant differences of Newcastle antibody titers between all groups and ages in broiler pullets in ELISA. According to the sampling and analysis of HI and ELISA serum tests, there were no significant relationships between all broiler pullets breeding in green, blue and yellow light on days 24 and 46 and the P-value was more than 0/05. It is suggested that the monochromatic light is effective on broilers immunity against Newcastle disease.

Keywords: monochromatic light, Newcastle disease, HI test, ELISA test

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Authors: Inikpi Ameh, Grace Kia, A. K. B. Sackey, Joy Atawodi, Richard Whittington

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Ranaviruses are belonging to the viral Family Iridoviridae, are a group of globally emerging pathogens recognized as major viral pathogens of cold-blooded vertebrates. They cause systemic infection in fishes, amphibians, and reptiles. Ranaviruses have been associated with numerous disease outbreaks in natural and cultured populations of fish, amphibians, and reptiles. To investigate the presence of the ranavirus in fish and edible frogs sourced from dams and ponds in Zaria, Kaduna State, Nigeria. A total of 425 frogs (Rana spp.) and fishes (n=215 and n=200, respectively) were randomly collected based on consent and availability. Liver, kidney, and spleen tissue samples from each animal were pooled and homogenized. The samples were screened for ranavirus using the Indirect Enzyme linked Immunosorbent assay (ELISA). An overall prevalence of 46.1% (196/425) was obtained from the study. Frogs had a prevalence of 51.2% (110/215) while fish had 43% (86/200). This is the first study on ranavirus in fish and edible frogs in Nigeria. This study has established that edible frogs (Rana spp) and fishes sold in Zaria, Nigeria were infected with ranavirus which may have great economic importance to the nation’s aquaculture. In view of occasional massive economic losses observed in fishery industry due to deaths of unknown origin, this preliminary investigation is useful in directing veterinarians, policy makers and researchers on need to survey for ranavirus and also enlighten the relevant stakeholders on its prevention and control in Nigeria.

Keywords: fish, frogs, Nigeria, Ranavirus

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Authors: Zakieh Rostamzadeh , Nariman Sepehrvand-Zahra Shirmohamadi

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Background: Cytomegalovirus (CMV) infection is one of the most common infectious problems following kidney transplantation. In this study, we are aimed to investigate the CMV infection in the setting of renal transplant recipients in Urmia-Iran, using both ELISA and polymerase chain reaction (PCR) methods. Methods: Ninety-six renal transplant recipients were selected randomly and enrolled in a cross-sectional study. Blood sampling was done via venipuncture, and all sera were investigated for anti-CMV IgM, and the seropositive cases in association with 14 randomly selected seronegative cases were investigated with PCR assay. Results: Thirty-three patients (34.3%) were seropositive for anti-CMV IgM, 3 patients (3.1%) were in borderline range, and 60 patients (62.5%) were seronegative. By considering the patients with borderline anti-CMV IgM levels as seropositive, 37.5% were seropositive for anti-CMV IgM. Among 36 seropositive cases, the CMV infection was confirmed in 19 (52.7%) of them using PCR. Age (P = 0.40), educational status (P = 0.77), history of pre-transplantation dialysis (0.52), history of blood transfusion (P = 0.52), and immunosuppressive regimen were not statistically different among recipients with positive versus negative CMV PCR study results. Conclusion: The seroprevalence of CMV infection was demonstrated to be high in renal transplant recipients of Urmia-Iran. The rate was higher compared to several previous reports in the literature. ELISA method has an appropriate sensitivity to screen the recipients for CMV infection but considering its relatively low specificity, the seropositive cases are better to be confirmed by further PCR study.

Keywords: cytomegalovirus, renal transplantation, ELISA, IgM, PCR

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Authors: Eva Tvrdá, Barbora Babečková, Michal Ďuračka, Róbert Kirchner, Július Árvay

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This study was designed to assess the in vitro effects of Origanum vulgare L. (oregano) extract on the testicular steroidogenesis. We focused on identifying major biomolecules present in the oregano extract, as well as to investigate its in vitro impact on the secretion of cholesterol, testosterone, dehydroepiandrosterone and androstenedione by murine testicular fragments. The extract was subjected to high performance liquid chromatography (HPLC) which identified cyranosid, daidzein, thymol, rosmarinic and trans-caffeic acid among the predominant biochemical components of oregano. For the in vitro experiments, testicular fragments from 20 sexually mature Institute of Cancer Research (ICR) mice were incubated in the absence (control group) or presence of the oregano extract at selected concentrations (10, 100 and 1000 μg/mL) for 24 h. Cholesterol levels were quantified using photometry and the hormones were assessed by ELISA (Enzyme-Linked Immunosorbent Assay). Our data revealed that the release of cholesterol and androstenedione (but not dehydroepiandrosterone and testosterone) by the testicular fragments was significantly impacted by the oregano extract in a dose-dependent fashion. Supplementation of the extract resulted in a significant decline of cholesterol (P < 0.05 in case of 100 μg/mL; P < 0.01 with respect 100 μg/mL extract), as well as androstenedione (P < 0.01 with respect to 100 and 1000 μg/mL extract). Our results suggest that the biomolecules present in Origanum vulgare L. could exhibit a dose-dependent impact on the secretion of male steroids, playing a role in the regulation of testicular steroidogenesis.

Keywords: mice, Origanum vulgare L., steroidogenesis, testes

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Authors: Elisangela Sobreira, Denise Teixeira

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Visceral leishmaniasis is a public health problem in Brazil, it is the main reservoir dog. In the period 2012-2016 78 diagnoses were performed in dogs suspected. Blood samples were collected from the cephalic vein obtaining serum used for the indirect immunofluorescence test and enzyme-linked immunosorbent assay, while it collected a drop of blood for the rapid chromatographic immunoassay. Obtained in 32 dogs positive. The test is important for the control of this disease and is used routinely in the Zoonoses Control Center.

Keywords: Brazil, dogs, Leismaniasis, Zoonoses center

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Authors: Mohammad Alkhatatbeh, Nehad Ayoub, Nizar Mhaidat, Nesreen Saadeh, Lisa Lincz

Abstract:

CD36 is involved in the development of atherosclerosis by enhancing macrophage endocytosis of oxidized-low density lipoproteins and foam cell formation. Soluble CD36 (sCD36) was found to be elevated in type 2 diabetic patients and was supposed to act as a marker of insulin resistance and atherosclerosis. In young subjects, sCD36 was associated with cardiovascular risk factors including obesity and hypertriglyceridemia. This study was conducted to further investigate the relationship between plasma sCD36 and cardiovascular risk factors among middle-aged patients with metabolic syndrome (MetS) and healthy controls. SCD36 concentrations were determined by enzyme-linked immunosorbent assays (ELISA) for 41 patients with MetS and 36 healthy controls. Data for other variables were obtained from patients' medical records. SCD36 concentrations were relatively low compared to most other studies and were not significantly different between the MetS group and controls (P-value=0.17). SCD36 was also not correlated with age, body mass index, glucose, lipid profile, serum electrolytes and blood counts. SCD36 was not significantly different between subjects with obesity, hyperglycemia, dyslipidemia, hypertension or cardiovascular disease and those without these abnormalities (P-value > 0.05). The inconsistency between results reported in this study and other studies may be unique to the study population or be a result of the lack of a reliable standardized method for determining absolute sCD36 concentrations. However, further investigations are required to assess CD36 tissue expression in the study population and to assess the accuracy of various commercially available sCD36 ELISA kits. Thus, the availability of a standardized simple sCD36 ELISA that could be performed in any basic laboratory would be more favorable to the specialized flow cytometry methods that detect CD36+ microparticles if it was to be used as a biomarker.

Keywords: metabolic syndrome, CD36, cardiovascular risk, obesity, type 2 diabetes mellitus

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Authors: Atefeh Esmailnejad, Mohammad Abbaszadeh Hasiri

Abstract:

Coxiella burnetii is a gram-negative obligatory intracellular bacterium that causes Q fever, a significant zoonotic disease. Sheep, cattle, and goats are the most commonly reported reservoirs for the bacteria, but infected cats and dogs have also been implicated in the transmission of the disease to human. The aim of present study was to investigate the presence of antibodies against Coxiella burnetii among companion dogs in Fars province, South of Iran. A total of 181 blood samples were collected from asymptomatic dogs, mostly referred to Veterinary Hospital of Shiraz University for regular vaccination. The IgG antibody detection against Coxiella burnetii was made by indirect Enzyme-linked Immunosorbent Assay (ELISA), employing phase I and II Coxiella burnetii antigens. A logistic regression model was developed to analyze multiple risk factors associated with seropositivity. An overall seropositivity of 7.7% (n=14) was observed. Prevalence was significantly higher in adult dogs above five years (18.18 %) compared with dogs between 1 and five years (7.86 %) and less than one year (6.17%) (P=0.043). Prevalence was also higher in male dogs (11.21 %) than in female (2.7 %) (P=0.035). There were no significant differences in the prevalence of positive cases and breed, type of housing, type of food and exposure to other farm animals (P>0.05). The results of this study showed the presence of Coxiella burnetii infection among the companion dogs population in Fars province. To our knowledge, this is the first study regarding Q fever in dogs carried out in Iran. In areas like Iran, where human cases of Q fever are not common or remain unreported, the public health implications of Q fever seroprevalence in dogs are quite significant.

Keywords: Coxiella burnetii, dog, Iran, Q fever

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Authors: Raviporn Madarasmi, Anjalee Vacharaksa, Pravej Serichetaphongse

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The host response in peri-implant tissue may differ from that in periodontal tissue in a healthy individual. The purpose of this study is to investigate the expression of inflammatory cytokines in peri-implant crevicular fluid (PICF) from single implant with different abutment types in comparison to healthy periodontal tissue. 19 participants with healthy implants and teeth were recruited according to inclusion and exclusion criteria. PICF and gingival crevicular fluid (GCF) was collected using sterile paper points. The expression level of inflammatory cytokines including IL-1α, IL-1β, TNF-α, IFN-γ, IL-6, and IL-8 was assessed using enzyme-linked immunosorbent assay (ELISA). Paired t test was used to compare the expression levels of inflammatory cytokines around natural teeth and peri-implant in PICF and GCF of the same individual. The Independent t-test was used to compare the expression levels of inflammatory cytokines in PICF from titanium and UCLA abutment. Expression of IL-6, TNF-α, and IFN-γ in PICF was not statistically different from GCF among titanium and UCLA abutment group. However, the level of IL-1α in the PICF from the implants with UCLA abutment was significantly higher than GCF (P=0.030). In addition, the level of IL-1β in PICF from the implants with titanium abutment was significantly higher than GCF (P=0.032). When different abutment types was compared, IL-8 expression in PICF from implants with UCLA abutment was significantly higher than titanium abutment (P=0.003).

Keywords: abutment, dental implant, gingival crevicular fluid and peri-implant crevicular fluid

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Authors: Mohamed A. Alhammad, Hadia A. Abou-Donia, Mona H. Hashish, Mohamed N. Massoud

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Background: Rotavirus is the leading etiologic agent of severe diarrheal disease in infants and young children worldwide. The present study was aimed 1) to detect rotavirus infection as a cause of diarrhoea among children under 5 years of age using the two serological methods (ELISA and LA) and the PCR technique (2) to evaluate the three methodologies used for human RV detection in stool samples. Materials and Methods: This study was carried out on 247 children less than 5 years old, diagnosed clinically as acute gastroenteritis and attending Alexandria University Children Hospital at EL-Shatby. Rotavirus antigen was screened by ELISA and LA tests in all stool samples, whereas only 100 samples were subjected to RT-PCR method for detection of rotavirus RNA. Results: Out of the 247 studied cases with diarrhoea, rotavirus antigen was detected in 83 (33.6%) by ELISA and 73 (29.6%) by LA, while the 100 cases tested by RT-PCR showed that 44% of them had rotavirus RNA. Rotavirus diarrhoea was significantly presented with a marked seasonal peak during autumn and winter (61.4%). Conclusion: The present study confirms the huge burden of rotavirus as a major cause of acute diarrhoea in Egyptian infants and young children. It was concluded that; LA is equal in sensitivity to ELISA, ELISA is more specific than LA, and RT-PCR is more specific than ELISA and LA in diagnosis of rotavirus infection.

Keywords: rotavirus, diarrhea, immunoenzyme techniques, latex fixation tests, RT-PCR

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Authors: Jacintha Menezes, Ramalingam Dharmalingam, Palaiahnakote Shivakumara

Abstract:

In the recent years, Omani acid lime cultivation and production has been affected by Citrus greening or Huanglongbing (HLB) disease. HLB disease is one of the most destructive diseases for citrus, with no remedies or countermeasures to stop the disease. Currently used Polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) HLB detection tests require lengthy and labor-intensive laboratory procedures. Furthermore, the equipment and staff needed to carry out the laboratory procedures are frequently specialized hence making them a less optimal solution for the detection of the disease. The current research uses hyperspectral imaging technology for automatic detection of citrus trees with HLB disease. Omani citrus tree leaf images were captured through portable Specim IQ hyperspectral camera. The research considered healthy, nutrition deficient, and HLB infected leaf samples based on the Polymerase chain reaction (PCR) test. The highresolution image samples were sliced to into sub cubes. The sub cubes were further processed to obtain RGB images with spatial features. Similarly, RGB spectral slices were obtained through a moving window on the wavelength. The resized spectral-Spatial RGB images were given to Convolution Neural Networks for deep features extraction. The current research was able to classify a given sample to the appropriate class with 92.86% accuracy indicating the effectiveness of the proposed techniques. The significant bands with a difference in three types of leaves are found to be 560nm, 678nm, 726 nm and 750nm.

Keywords: huanglongbing (HLB), hyperspectral imaging (HSI), · omani citrus, CNN

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Authors: Salman Akrokayan

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Synthetic cDNA (ScDNA) of granulocyte colony-stimulating factor (G-CSF) was constructed using a DNA synthesizer with the aim to increase its expression level. 5' end of the ScDNA of G-CSF coding region was modified by decreasing the GC content without altering the predicted amino acids sequence. The identity of the resulting protein from ScDNA was confirmed by the highly specific enzyme-linked immunosorbent assay. In conclusion, a synthetic G-CSF cDNA in combination with the recombinant DNA protocol offers a rapid and reliable strategy for synthesizing the target protein. However, the commercial utilization of this methodology requires rigorous validation and quality control.

Keywords: synthetic cDNA, recombinant G-CSF, cloning, gene expression

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Authors: Monika Szymańska-Czerwińska, Agnieszka Jodełko, Krzysztof Niemczuk

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Q fever (coxiellosis) is an infectious disease of animals and humans causes by C. burnetii and widely distributed throughout the world. Cattle and small ruminants are commonly known as shedders of C. burnetii. The aims of this study were the evaluation of seroprevalence and shedding of C. burnetii in cattle. Genotypes of the pathogen present in the tested specimens were also identified using MLVA (Multiple Locus Variable-Number Tandem Repeat Analysis) and MST (multispacer sequence typing) methods. Sampling was conducted in different regions of Poland in 2018-2021. In total, 2180 bovine serum samples from 801 cattle herds were tested by ELISA (enzyme-linked immunosorbent assay). 489 specimens from 157 cattle herds such as: individual milk samples (n=407), bulk tank milk (n=58), vaginal swabs (n=20), placenta (n=3) and feces (n=1) were subjected to C. burnetii specific qPCR. The qPCR (IS1111 transposon-like repetitive region) was performed using Adiavet COX RealTime PCR kit. Genotypic characterization of the strains was conducted utilizing MLVA and MST methods. MLVA was performed using 6 variable loci. The overall herd-level seroprevalence of C. burnetii infection was 36.74% (801/2180). Shedders were detected in 29.3% (46/157) cattle herds in all tested regions. ST 61 sequence type was identified in 10 out of 18 genotyped strains. Interestingly one strain represents sequence type which has never been recorded previously. MLVA method identified three previously known genotypes: most common was J but also I and BE were recognized. Moreover, a one genotype has never been described previously. Seroprevalence and shedding of C. burnetii in cattle is common and strains are genetically diverse.

Keywords: Coxiella burnetii, cattle, MST, MLVA, Q fever

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Authors: Dwitya Elvira, Eryati Darwin

Abstract:

Background: Graves’ disease (GD) is an autoimmune thyroid disease. Imbalance of Th1/Th2 cells and T-regulatory (Treg)/Th17 cells was thought to play pivotal role in the pathogenesis of GD. Treg FoxP3 produced TGF-β to maintain regulatory function, and Th17 cells produced IL-17 as cytokines that were thought in mediating several autoimmune diseases. The aim of this study is to assess the role of IL-17 and TGF-β in the pathogenesis of GD and to investigate its correlation with Thyroid Stimulating Hormone Receptor Antibody (TRAb) and Treg FoxP3 expression. Method: 30 GD patients and 27 age and sex-matched controls were enrolled in this study. Diagnosis of GD was based on clinical and biochemical of GD. Serum IL-17, TGF-β, TRAb, and FoxP3 were measured by enzyme-linked immunosorbent assay (ELISA). Data were analyzed by using SPSS 21.0 (SPSS Inc.). Spearman rank correlation test was used for assessment of correlation. The statistical significance was accepted as P<0.05. Result: There was no significant correlation between IL-17 and TGF-β serum with expression of FoxP3 level in GD, but there was significant correlation between TGF-β and TRAb serum level (P<0.05). Serum levels of IL-17 and TGF-β were found to be elevated in patient group compared to control, where mean values of IL-17 were 14.43±2.15 pg/mL and TGF-β were 10.44±3.19 pg/mL in patients group; and in control group, level of IL-17 were 7.1±1.45 pg/mL and TGF-β were 4.95±1.35 pg/mL. Conclusion: Serum Il-17 and TGF-β were elevated in GD patients that reflect the role of inflammatory and regulatory cytokines activation in pathogenesis of GD. There was significant correlation between TGF-β and TRAb, revealing that Treg cytokines may play a role in pathogenesis of GD.

Keywords: IL-17, TGF-B, FoxP3, TRAb, Graves’ disease

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Authors: Maia Kukhaleishvili, Ekaterine Bulauri, Iveta Megrelishvili, Tamar Shamatava, Tamar Chipashvili

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Plant viruses can cause loss of yield and quality in a lot of important crops. Symptoms of pathogens are variable depending on the cultivars and virus strain. Selection of resistant potato varieties would reduce the risk of virus transmission and significant economic impact. Other way to avoid reduced harvest yields is regular potato seed production sampling and testing for viral infection. The aim of this study was to determine the occurrence and distribution of viral diseases according potato cultivars for further selection of virus-free material in Georgia. During the summer 2015- 2016, 5 potato cultivars (Sante, Laura, Jelly, Red Sonia, Anushka) at 5 different farms located in Akhalkalaki were tested for 6 different potato viruses: Potato virus A (PVA), Potato virus M (PVM), Potato virus S (PVS), Potato virus X (PVX), Potato virus Y (PVY) and potato leaf roll virus (PLRV). A serological method, Double Antibody Sandwich-Enzyme linked Immunosorbent Assay (DASELISA) was used at the laboratory to analyze the results. The result showed that PVY (21.4%) and PLRV (19.7%) virus presence in collected samples was relatively high compared to others. Researched potato cultivars except Jelly and Laura were infected by PVY with different concentrations. PLRV was found only in three potato cultivars (Sante, Jelly, Red Sonia) and PVM virus (3.12%) was characterized with low prevalence. PVX, PVA and PVS virus infection was not reported. It would be noted that 7.9% of samples were containing PVY/PLRV mix infection. Based on the results it can be concluded that PVY and PLRV infections are dominant in all research cultivars. Therefore significant yield losses are expected. Systematic, long-term control of potato viral infection, especially seed-potatoes, must be regarded as the most important factor to increase seed productivity.

Keywords: virus, potato, infection, diseases

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Authors: Abdolreza Esmaeilzadeh, Mehri Mirzaei

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Objective: Rheumatic disease is a chronic disease that causes pain, stiffness, swelling and limited motion and function of many joints. RA is the most common form of autoimmune arthritis, affecting more than 1.3 million Americans. Of these, about 75% are women. Materials and Methods: This study was formed due to the misconception about ANA test, which is frequently performed with methods based upon solid phase as ELISA. This experiment was conducted on 430 patients, with clinical symptoms that are likely affected with rheumatic diseases, simultaneously by means of ANA and FANA. Results: 36 cases (8.37%) of patients, despite positive ANA, have demonstrated negative results via Indirect Immunofluorescence Assay (IIFA), (false positive). 116 cases (27%) have demonstrated negative ANA results, by means of the ELISA technique, although they had positive IIFA results. Conclusion: Other advantages of IIFA are antibody titration and specific pattern detection that have the capability of distinguishing positive dsDNA results. According to the restrictions and false negative cases, in patients, IIFA test is highly recommended for these disease's diagnosis.

Keywords: autoimmune disease, IIFA, EIA, rheumatic disease

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Authors: Belaynesh Tsegay Beyene, Teklay Gebrecherkos, Atsebaha Gebrekidan Kahsay, Mahmud Abdulkader

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Background: Hepatitis B and C viruses are of important health and socioeconomic problem of the globe with remarkable diseases and deaths in Sub-Saharan African countries. The burden of hepatitis is unknown in the prison settings of Tigrai. Therefore, we aimed to describe the seroprevalence and associated factors of hepatitis B and C viruses among prisoners of Tigrai, Ethiopia. Methods: A cross-sectional study was carried out from February 2020 to May 2020 at the prison facilities of Tigrai. Demographics and associated factors were collected from 315 prisoners prospectively. Five milliliter of blood was collected and tested using rapid tests kits of HBsAg (Zhejiang orient Gene Biotech Co., Ltd., China) and HCV antibodies (Volkan Kozmetik Sanayi Ve Ticaret Ltd. STI, Turkey). Positive samples were confirmed using enzyme-linked immunosorbent assay (ELISA) (Beijing Wantai Biological Pharmacy Enterprise Co. Ltd). Data were analyzed using Statistical Package for Social Sciences (SPSS) version 20 and p < 0.05 was considered statistically significant. Results: The overall seroprevalence of HBV and HCV were 25 (7.9%) and 1(0.3%), respectively. The majority of hepatitis B viral infections were identified from the age groups of 18-25 years (10.7%) and unmarried prisoners (11.8%). Prisoners greater than 100 per cell [AOR =3.95, 95% CI= (1.15, 13.6, p =0.029)] and having history of alcohol consumption [AOR =3.01, 95% CI= (1.17, 7.74, p =0.022)] were significantly associated with HBV infections. Conclusions: The seroprevalence of HBV among prisoners was nearly high or borderline (7.9%) with a very low HCV prevalence (0.3%). HBV was most prevalent among young adults, large number of prisoners per cell and those who had history of alcohol consumption. This study recommends that there should be prison-focused intervention including regular health education by emphasis on the mode of transmission and introducing HBV screening policy for prisoners especially when they enter to the prison.

Keywords: seroprevalence, HBV, HCV, prisoners, Tigrai

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