A Laboratory Exercise Simulating Antibody and Antigen Reactions of the Ouchterlony Double Immunodiffusion Assay Using Inorganic Salts

The Ouchterlony double immunodiffusion assay is a serological technique used in the detection of antibodies and antigens for diagnostic purposes and also used in immunology laboratory courses as a common teaching assay where students observe the geometrical precipitation line patterns that form in the agarose, elucidating degrees of homology between antigens. In this classical technique, students must wait several hours to days to obtain results when protein antigens and antibodies are used.

Formation of Wind Gusts in an Extratropical Cyclone in Light of Doppler Lidar Observations and Large-Eddy Simulations

Damaging gusts in windstorms are represented by crude subgrid-scale parameterizations in today’s weather and climate models. This limitation motivated the Wind and Storms Experiment (WASTEX) in winter 2016–17 in the Upper Rhine Valley over southwestern Germany. Gusts recorded at an instrumented tower during the passage of extratropical cyclone “Thomas” on 23 February 2017 are investigated based on measurements of radial wind with ≈70-m along-beam spacing from a fast-scanning Doppler lidar and realistic large-eddy simulations with grid spacings down to 78 m using the Icosahedral Nonhydrostatic model. Four wind peaks occur due to the storm onset, the cold front, a precipitation line, and isolated showers. The first peak is related to a sudden drop in dewpoint and results from the downward mixing of a low-level jet and a dry layer within the warm sector characterized by extremely high temperatures for the season. While operational convection-permitting forecasts poorly predict the storm onset overall, a successful ensemble member highlights the role of upstream orography. Lidar observations reveal the presence of long-lasting wind structures that result from a combination of convection- and shear-driven instability. Large-eddy simulations contain structures elongated in the wind direction that are qualitatively similar but too coarse compared to the observed ones. Their size is found to exceed the effective model resolution by one order of magnitude due to their elongation. These results emphasize the need for subkilometer-scale measuring and modeling systems to improve the representation of gusts in windstorms.

The activity of polyclonal IgY derived from Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis in inhibiting colonization of Fusobacterium nucleatum and Streptococcus sanguinis

Background: Fusobacterium nucleatum (F. nucleatum) and Streptococcus sanguinis (S. sanguinis) play a role in dental plaque formation which leads to periodontitis. Immunoglobulin Y (IgY) is present in both serum and egg yolk and can bind to the surface components of bacteria. F. nucleatum and S. sanguinis feature the same type of IV pili as Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans). Saliva binding protein (SsaB) in S. sanguinis is a FimA homolog. FimA constitutes a surface element of Porphyromonas gingivalis (P. gingivalis). F. nucleatum and P. gingivalis possess the same outer membrane protein (OMP) molecular mass. Purpose: The study aimed to determine the activity of A. actinomycetemcomitans and P. gingivalis polyclonal IgY present in serum and egg yolk that can inhibit colonization of F. nucleatum and S. sanguinis. Methods: IgY samples were diluted with phosphate buffer saline (PBS). Several holes were made in the nutrient medium with 10 μl antigen (F. nucleatum/S. sanguinis) being inserted into the center hole. 10 μl PBS, 1:1, 1:2, 1:4, 1:8, 1:16 A. actinomycetemcomitans or P. gingivalis polyclonal IgY were subsequently introduced into the surrounding holes. The results of incubation at 37°C were observed after 24-48 hours. Kruskal Wallis and MannWhitney tests were administered to analyse the data. Results: A. actinomycetemcomitans and P. gingivalis polyclonal IgY groups in serum showed a precipitation line at dilution ratios of 1:1 and 1:2, whereas in egg yolk this occurred only at a 1:1 dilution ratio with F. nucleatum and S. sanguinis bacteria in this study. No significant differences were evident between each dilution (p>0.05) and none existed between serum and egg yolk (p>0.05). Conclusion: IgY polyclonal of A. actinomycetemcomitans and P. gingivalis in both serum and egg yolk initiate activities that can inhibit colonization of F. nucleatum and S. sanguinis.

Extraction and characterisation of Brucella abortus strain RB51 rough lipopolysaccharide

Brucellosis is an important zoonotic disease with considerable impacts on human and animal health. Brucella abortus strain RB51 vaccine is used for prevention of bovine brucellosis in Iran. Due to strain roughness, available serological tests cannot detect vaccinated animals. Detection of serological responses to the vaccine is important to monitor accurate vaccination implementation. Rough lipopolysaccharide (RLPS) of RB51 strain was extracted and characterised to develop serological tests for diagnosis of vaccinated animals. RLPS was extracted using phenol-chloroform-petroleum ether and evaluated by limulus amebocyte lysate (LAL) assay, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and agar gel immunodiffusion (AGID). According to our results, the extracted RLPS caused positive reaction in LAL assay. In SDS-PAGE, a band with a molecular weight around 14 kDa was identified after specific staining using silver nitrate. Double AGID of the RLPS with a hyperimmune serum resulted in a precipitation line formation. Our study showed that the method can be successfully used to extract RLPS from Brucella abortus strain RB51 as confirmed by LAL assay, PAGE and AGID.

Analysis of anti-Streptococcus sanguinis IgY ability to inhibit Streptococcus sanguinis adherence

Background: Streptococcus sanguinis (S. sanguinis), an oral commensal bacterium, is often implicated in infective endocarditis. Its adherence to the tooth surface is the initial step in dental plaque formation. In addition to the important role of S. sanguinis in systemic disease and antimicrobial resistance, it is necessary to develop methods to control dental plaque formation. Immunoglobulin Y (IgY) has been used to prevent bacterial infection. Purpose: The purpose of this study is to analyze the ability of anti-S. sanguinis IgY antibodies to inhibit S. sanguinis adherence to hydroxyapatite (HA) discs as a model of the tooth surface. Methods: Antibodies were produced by immunizing hens with S. sanguinis suspension. Boosters were given three times following the first injection. An agar gel precipitation test (AGPT) was used to detect the presence of anti-S. sanguinis IgY. A bacterial adherence assay was performed twice to analyze the ability of IgY and the optimal concentration required to inhibit bacterial adherence. Results: The formation of a precipitation line using AGPT confirmed the presence of the antibody. In addition, it was shown that the anti-S. sanguinis IgY antibody could inhibit bacterial adherence to HA. Statistical analysis using One-way ANOVA revealed a significant difference in the optical density (OD) value between the groups (p<0.05). The results of electron microscopy scanning confirmed the quantitative analysis by means of a bacterial adherence test. Conclusion: Anti-S. sanguinis IgY has the ability to inhibit adherence of S. sanguinis to HA discs at an optimal concentration of 30%. The inhibitive effect was stronger in the presence of saliva.

A Spatial Pattern Analysis of Frontier Passes in China’s Northern Silk Road Region Using a Scale Optimization BLR Archaeological Predictive Model

In China’s Northern Silk Road (CNSR) region, dozens of frontier passes built and fortified at critical intersections were exploited starting at approximately 114 B.C. to guarantee caravan safety. Understanding the pattern of these pass sites is helpful in understanding the defense and trading system along the Silk Road. In this study, a scale optimization Binary Logistic Regression (BLR) archaeological predictive model was proposed to study the spatial pattern of CNSR frontier passes for understanding the critical placement of ancient defense and trading pass sites. Three hundred and fifty sample locations and 17 natural proxies were input into the model. Four strongly correlated factors were reserved as independent variables to construct the model, which was validated by 150 surveyed data and Kvamme’s Gain statistics. According to the variable selection and model optimization, the best spatial scale varies with the stability of the variables, such as 50 m and 1000 m, respectively, for the terrain and non-terrain variables. Clustering characteristics were identified with division overlapped with a 400 mm precipitation line using the site sensibility map. The high and medium probability areas were assembled along the Great Wall and the CNSR routes, especially in the western part, revealing that the model is also helpful to reconstruct the Silk Road routes.

Stable Isotope Study of Precipitation in Jinan

The 8 water samples of rain in Jinan were collected in the summer. A local meteoric line (LMWL) was got. And the derived equation is that δD = 8.19δ18O+ 8.89, which represents the local precipitation line. The precipitation water in Jinan is characterized by the high mean deuterium excess (d=10.9‰). The isotopic compositions in Xi’an were also be collected. And the local meteoric line in Xi’an is that δD=6.97δ18O+3.01, with a low slope The slopes of the meteoric line were different for the two sites, which can reveals some evaporation information about the precipitation.

Stable Isotopes Analysis of Precipitation in Subtropical Plateau of Yunnan

To elucidate the relationship between stable isotopes of precipitation (SIP) and the extreme drought in Kunming area, based on the stable isotopes data of the GNIP in Kunming site from 1986 to 2003, the precipitation line equation is brought forward and the seasonal change rule of stable isotopes are discussed. The stable isotopic compositions of precipitation exhibit great diversities in different seasons during to influences of multiple factors, such as monsoon, rainfall amount moisture source and others. The δ18O values in rainwater exhibit significant seasonal variations, the average of-10.12‰ in rainy season, the dry season is-4.5‰, having lower values in the rainy season and higher one in the dry season. The amount effect of precipitation is very distinct, that concealed the temperature effect. Got the special geographical position,dvalues present unique characteristics, the average ofdvalues is 10.78‰ in rainy season, and is 4.86‰ in dry season, the mean value is generally lower than most parts of the world.

Genetics and Mapping of the Cl Gene for Circumlineated Pattern in Common Bean Using AFLP-based Bulk Segregant Analysis and SNP-based Bidirectional Selective Genotyping

Circumlineatus (cl) in common bean (Phaseolus vulgaris L.) is identified by a precipitation line in the seedcoat at the boundary of the white and colored zones. Cl is recessive and is expressed in partly colored seedcoats (t) with restricted patterns such as virgarcus. In this study, amplified fragment length polymorphism (AFLP) and single nucleotide polymorphism (SNP) markers, and the common bean genome sequence were used in combination with bulk segregant analysis and bidirectional selective genotyping to identify the genetic location of Cl. Markers were identified that cosegregated with Cl using Cl/Cl and cl/cl F3 and F5 progeny bulks from the cross t z cl G b v virgarcus BC3 5-593 × t zsel Cl G b v sellatus BC3 5-593. Two bands from an AFLP primer combination, which yielded unambiguous polymorphisms between the bulks, were cloned and sequenced. The two sequences were used to interrogate the common bean whole genome sequence identifying a region also found through cosegregation analysis using bidirectional selective genotyping with SNPs. Thus, the Cl gene was localized on Pv09 using cosegregating AFLP and SNP markers, and the physical location was confirmed with the whole genome sequence.