Kesesuaian Uji Antigen Capture Enzyme Linked Immunosorbant Assay dan Nested Multiplex Polymerase Chain Reaction untuk Diagnosis Rutin Bovine Viral Diarrhea

Bovine Viral Diarrhea (BVD) is one of the main causes of impaired productivity and reproduction of cows. Antigen capture Elisa (ACE) is one of the serological technique that is sensitive, reliable and used regularly for detecting persistent BVD infection individually which simpler than multiplex nested PCR. The aim of this study was to determine the agreement between ACE and multiplex nested PCR as a routine laboratory diagnostic technique to detect the presence of BVD infection. A total of 128 cow serum samples consisting of 63 positive and 65 negative samples based on ACE were used in this study. The samples were collected from active and passive surveillance in dairy and beef cattle conducted by Balai Besar Veteriner (BBVet) Wates. The serum samples were then tested molecularly using multiplex nested PCR against BVD. The result showed 48 out of 63 BVDV-1 positive samples were found positive BVD antigen whereas 57 of 65 BVDV-1 negative samples were negative using multiplex nested PCR, . The agreement value between the two different assays based on statistic analysis using Kappa method was 0.64 and classified a good one. The result concluded that the ACE BVD assay was equally suitable as routine diagnosis to determine BVD infected cattle in the farm. Keywords: Antigen capture ELISA; Bovine viral diarrhea; Kappa; Multiplex nested PCR. Abstrak Bovine Viral Diarrhea (BVD) merupakan salah satu penyebab gangguan produktivitas dan reproduksi sapi. Antigen capture ELISA (ACE) merupakan salah satu teknik serologis yang sensitif, dapat diandalkan dan digunakan secara teratur untuk mendeteksi infeksi BVD persisten secara individual yang lebih sederhana daripada multiplex nested PCR. Tujuan penelitian ini adalah untuk mengetahui kesesuaian antara uji ACE dan multiplex nested PCR sebagai teknik diagnostik laboratorium rutin untuk mendeteksi adanya infeksi BVD. Sebanyak 128 sampel serum sapi yang terdiri dari 63 sampel positif dan 65 negatif berdasarkan ACE BVDV Antigen Test Kit/Serum Plus (Idexx®) digunakan dalam kajian ini. Sampel serum sapi merupakan koleksi dari surveilans aktif dan pasif pada sapi perah dan potong yang dilakukan Balai Besar Veteriner (BBVet) Wates. Sampel serum kemudian diuji secara molekuler menggunakan multiplex nested PCR terhadap BVD. Hasil penelitian menunjukkan bahwa dengan teknik multiplex nested PCR, 48 dari 63 sampel positif BVDV-1 ditemukan positif untuk antigen BVD sedangkan 57 dari 65 sampel negatif BVDV-1 negatif untuk antigen BVD. Analisis statitik berdasarkan perhitungan metoda Kappa menunjukkan nilai kesesuaian antara dua uji sebesar 0,64 dan tergolong bagus. Hasil penelitian menunjukkan kesimpulan bahwa uji ACE BVD sesuai sebagai diagnosis rutin untuk menentukan ternak yang terinfeksi BVD di peternakan. Kata kunci: Antigen capture ELISA; Bovine viral diarrhea; Kappa; Multiplex nested PCR.

Risk Factor Assessment, Sero-Prevalence, and Genotyping of the Virus That Causes Foot-and-Mouth Disease on Commercial Farms in Ethiopia from October 2018 to February 2020

A cross-sectional serological, active outbreak search and a questionnaire-based survey were carried out to investigate foot-and-mouth disease (FMD) sero-epidemiology in Ethiopia. The circulating serotype of the FMD virus (FMDV) was identified, and the knowledge regarding FMD and husbandry practices was assessed. Using the questionnaire survey, a total of 237 individuals were interviewed, and the majority responded that there is no practice of reporting disease outbreak, free in and out movement of livestock, or share pastures, and they use traditional case management as a means of controlling the disease. A total of 1938 cattle, 490 domestic small ruminants, and 426 swine were sampled randomly for serological analysis using the 3ABC non-structural protein (NSP) enzyme-linked immunosorbant assay. An overall prevalence of 25% in cattle, 5% in domestic small ruminants, and 2% in swine was recorded. Multivariable logistic regression analysis revealed that cattle from the Oromia, Tigray, and Amhara regions had the highest probability of being sero-positive as compared with Addis Ababa odds ratio(OR)(OR: 4 (95% confidence interval (CI)(CI [3–6], 3 (95% CI [2,5]), and 2 (95% CI 2 [1,3]), respectively). Older cattle (older than three years) and domestic small ruminants (>18 months) had a higher chance of being seropositive (OR: 2, 95% CI [1.6,3]) and (OR: 6, 95% CI [2,18]), respectively). Female and older swine older than three years of age had a higher chance of being sero-positive (p < 0.05). Local breed cattle had the lowest chance (OR: 0.2. 95% CI [0.1–0.3]) of being sero-positive. A region, age, and breed proved to have a statistically significant association with sero-positivity (p < 0.05) in cattle. Swine from Bishoftu were less likely to test positive than swine from Addis Ababa (OR: 0.04, 95% CI [0.01–0.3]). From 96 herds, 72 pooled outbreak samples were analyzed by polymerase chain reaction (PCR), virus isolation, serotyping (antigen enzyme linked immunosorbant assay (ELISA)), sequencing, and phylogenetic tree analysis. Six serotype A (G-IV) FMD viruses and three serotype O east African (EA-3 and EA-4) FMDVs were identified. Thus, this study established the lack of disease outbreak reporting, poor husbandry problems, and the prevalence of FMD in three domestic species (cattle, small ruminant, and swine). In addition, continuous circulation of serotype A and O in the study area was confirmed.

Spectrum Bias and Individual Strengths of SARS-CoV-2 Serological Tests—A Population-Based Evaluation

Antibody testing for determining the SARS-CoV-2 serostatus was rapidly introduced in early 2020 and since then has been gaining special emphasis regarding correlates of protection. With limited access to representative samples with known SARS-CoV-2 infection status during the initial period of test development and validation, spectrum bias has to be considered when moving from a “test establishment setting” to population-based settings, in which antibody testing is currently implemented. To provide insights into the presence and magnitude of spectrum bias and to estimate performance measures of antibody testing in a population-based environment, we compared SARS-CoV-2 neutralization to a battery of serological tests and latent class analyses (LCA) in a subgroup (n = 856) of the larger population based TiKoCo-19 cohort (n = 4185). Regarding spectrum bias, we could proof notable differences in test sensitivities and specificities when moving to a population-based setting, with larger effects visible in earlier registered tests. While in the population-based setting the two Roche ELECSYS anti-SARS-CoV-2 tests outperformed every other test and even LCA regarding sensitivity and specificity in dichotomous testing, they didn’t provide satisfying quantitative correlation with neutralization capacity. In contrast, our in-house anti SARS-CoV-2-Spike receptor binding domain (RBD) IgG-ELISA (enzyme-linked-immunosorbant assay) though inferior in dichotomous testing, provided satisfactory quantitative correlation and may thus represent a better correlate of protection. In summary, all tests, led by the two Roche tests, provided sufficient accuracy for dichotomous identification of neutralizing sera, with increasing spectrum bias visible in earlier registered tests, while the majority of tests, except the RBD-ELISA, didn’t provide satisfactory quantitative correlations.

The correlation of syndecan-4 with disease activity and serological characteristics of rheumatoid arthritis

[Objectives]To investigate the expression of syndecan-4 in serum, synovial fluid (SF) and synovium in rheumatoid arthritis (RA) by comparing with osteoarthritis (OA) patients, and to analyze the correlation of syndecan-4 with disease activity of RA .[Methods]Syndecan-4 in sera of 60 RA patients, 20 OA patients, 20 healthy controls, and in paired SF of 23 RA patients were tested by enzyme linked immunosorbant assay (ELISA). The expressions of syndecan-4 in synovium of 5 RA patients and 5 OA patients were detected by immunohistochemistry. The expressions of syndecan-4 of cultured synovial fibroblasts from RA and OA patients were detected by immunofluorescence. The correlation between serum syndecan-4 concentration and disease activity were analyzed in 60 RA patients.[Results]The serum syndedcan-4 concentration was significantly higher in RA patients than in OA patients and healthy controls, and was higher in rheumatoid factor (RF)-positive RA patients than in RF-negative ones. Syndecan-4 concentration in SF of RA patients was comparable with OA patients. Syndecan-4 expression in synovial tissue was similar between RA and OA patients. The syndecan-4 concentration was significantly lower in synovial fluid than in serum, either in RA or in OA patients. The serum and SF syndecan-4 concentrations were both positively correlated with RA disease activity scores.[Conclusion]The serum syndecan-4 concentration is higher in RA patients than in OA patients, and significantly higher in RF-positive RA patients than in RF-negative ones. The serum and SF syndecan-4 concentrations were both positively correlated with disease activity of RA patients.

Australian Aedes aegypti mosquitoes are susceptible to infection with a highly divergent and sylvatic strain of dengue virus type 2 but are unlikely to transmit it

Background: Humans are the primary hosts of dengue viruses (DENV). However, sylvatic cycles of transmission can occur among non-human primates and human encroachment into forested regions can be a source of emergence of new strains such as the highly divergent and sylvatic strain of DENV2, QML22, recovered from a dengue fever patient returning to Australia from Borneo. The objective of the present study was to evaluate the vector competence of Australian Aedes aegypti (Ae. aegypti) mosquitoes for this virus. Methods: Four to five day old mosquitoes from two strains of Ae. aegypti from Queensland, Australia, were feed a meal of sheep blood containing 108 50% cell culture infectious dose per ml (CCID50/ml) of either QML22 or an epidemic strain of DENV serotype 2 (QML16) isolated from a dengue fever patient in Australia in 2015. Mosquitoes were maintained at 28°C, 75% relative humidity and sampled 7, 10 and 14 days post-infection (DPI). Live virions in mosquito bodies (abdomen/thorax), legs and wings and saliva expectorates from individual mosquitoes were quantified using a Cell Culture Enzyme-linked Immunosorbant Assay (CCELISA) to determine infection, dissemination and transmission rates. Findings: The infection and dissemination rates of the sylvatic DENV2 strain, QML22, were significantly lower than that for QML16. While the titres of virus in the bodies of mosquitoes infected with either of these viruses were similar, titres in legs and wings were significantly lower in mosquitoes infected with QML22 at most time points although they reached similar levels by 14 DPI. QML16 was detected in 16% (n = 25) and 28% (n = 25) of saliva expectorates at 10 and 14 DPI, respectively. In contrast, no virus was detected in the saliva expectorates of QML22 infected mosquitoes. Conclusions: Australia urban/peri-urban Ae. aegypti species are susceptible to infection by the sylvatic and highly divergent DENV 2 QML22 but replication of QML22 is attenuated relative to the contemporary strain, QML16. A salivary gland infection or escape barrier may be acting to prevent infection of saliva and would prevent onward transmission of this highly divergent virus in Australia.

Australian Aedes aegypti mosquitoes are susceptible to infection with a highly divergent and sylvatic strain of dengue virus type 2 but are unlikely to transmit it

Background: Humans are the primary hosts of dengue viruses (DENV). However, sylvatic cycles of transmission can occur among non-human primates and human encroachment into forested regions can be a source of emergence of new strains such as the highly divergent and sylvatic strain of DENV2, QML22, recovered from a dengue fever patient returning to Australia from Borneo. The objective of the present study was to evaluate the vector competence of Australian Aedes aegypti (A. aegypti) mosquitoes for this virus. Methods: Four day old mosquitoes from two strains of A. aegypti from Queensland, Australia, were feed a meal of sheep blood containing 10 8 50% cell culture infectious dose per ml (CCID 50 /ml) of either QML22 or an epidemic strain of DENV serotype 2 (QML16) isolated from a dengue fever patient in Australia in 2015. Mosquitoes were maintained at 28°C, 75% relative humidity and sampled at 7, 10 and 14 days post-infection (DPI). Live virions in mosquito bodies (abdomen/thorax), legs and wings and saliva expectorates from individual mosquitoes were quantified using a Cell Culture Enzyme-linked Immunosorbant Assay (CCELISA) to determine infection, dissemination and transmission rates. Findings: The infection and dissemination rates of the sylvatic DENV2 strain, QML22, were significantly lower than that for QML16. While the titres of virus in the bodies of mosquitoes infected with either of these viruses were similar, titres in legs and wings were significantly lower in mosquitoes infected with QML22 at most time points although they reached similar levels by 14 DPI. QML16 was detected in 16% (n = 25) and 28% (n = 25) of saliva expectorates at 10 and 14 DPI, respectively. In contrast, no virus was detected in the saliva expectorates of QML22 infected mosquitoes. Conclusions: Australia urban/peri-urban A. aegypti species are susceptible to infection by the sylvatic and highly divergent DENV2 virus QML22. Our results indicate that replication of QML22 is attenuated relative to the contemporary strain QML16. Alternatively a salivary gland infection or escape barrier acts to prevent infection of saliva, potentially preventing onward transmission of this highly divergent virus in Australia.

Australia Ae. aegypti mosquitoes are susceptible to a highly divergent and sylvatic dengue virus type 2 strain infection but are unlikely to transmit

Background: Humans are the primary hosts of the dengue virus; However, sylvatic cycles of transmission can occur among non-human primates and human encroachment to forested regions can be a source of emergence of new strains. We reported the isolation of a highly divergent and sylvatic DENV-2 strain (QML22) from a dengue fever patient returning Australia from Borneo. The objective of the present study was to evaluate the vector competence of Australian Ae. aegypti mosquitoes for this virus. Methods: Four-day old mosquitoes from two strains of Ae. aegypti from Queensland, Australia, were feed sheep blood meal containing 108 50% cell culture infectious dose per ml (CCID50/ml) of either QML22 or an Australian epidemic DENV serotype 2 strain (QML16) isolated from a dengue fever patient in 2015. Mosquitoes were maintained at 28°C, 75% relative humidity and sampled at 7, 10 and 14 days post-infection (DPI). Live virions in mosquito bodies (abdomen/thorax), legs and wings and saliva expectorates from individual mosquitoes were quantified using a Cell Culture Enzyme-linked Immunosorbant Assay (CCELISA) to determine infection, dissemination and transmission rates. Findings: The infection and dissemination rates of the sylvatic DENV2 strain, QML22, within mosquitoes were significantly lower than that for QML16. While the titres of virus in the bodies of mosquitoes infected with either of these viruses were similar, titres in legs and wings were significantly lower in mosquitoes infected with QML22 at most time points although they reached similar levels by 14 DPI. QML16 was detected in 16% (n = 25) and 28% (n = 25) of saliva expectorates at 10 and 14 DPI, respectively. In contrast, no virus was detected in the saliva expectorates of QML22 infected mosquitoes. Conclusions: Australia urban/peri-urban Ae.aegypti species are susceptible to infection by the sylvatic and highly divergent DENV-2 virus QML22. However, our results indicate that replication of QML22 is attenuated relative to the contemporary strain QML16 and/or a salivary gland infection or escape barrier acts to prevent infection of saliva, potentially preventing onward transmission of this highly divergent virus in Australia.

SCREENING OF PREGNANT WOMEN FOR PRESENCE OF IGG ANTIBODIES FOR RUBELLA, MUMPS, MEASLES AND VARICELLA VIRUSES IN TERTIARY CARE HOSPITAL , JAIPUR

Background : The aim of this study were assess the susceptible pregnant women for vaccine preventable infection like rubella, mumps, measles and varicella viruses. Infection of mothers with these viruses during pregnancy can be serious. They can cause congenital infections, miscarriage, stillbirth and death of fetuses. Material and Method : This study is cross sectional. To determine the presence of IgG antibodies for rubella, mumps, measles and varicella viruses, blood samples were collected, stored at -700 c. Serum was separated for detection of IgG antibodies for these viruses by using enzyme linked immunosorbant assay. Results : Of 277 samples evaluated for IgG antibodies. Susceptibility Of pregnant women for rubella, mumps measles and varicella viruses were 7.6%, 17.6%,7.2% and 19.5% respectively. Susceptibility rates for rubella and mumps were higher in rural population as compares to urban while for varicella urban population was more susceptible, but it was not statistically signicant. No correlation could be observed in susceptibility to different to different viruses and their education status and age of patients, but youngest age group was most susceptible to varicella and oldest group to rubella. Primigravida were more susceptible to rubella and varicella while multigravida were more susceptible to mumps and measles. Conclusion : Majority of the pregnant women had protective levels of IgG antibody although susceptibility to rubella, mumps measles and varicella were low. Intensication of MMRV immunization of all females of child-bearing age is advocated.