Enzyme-linked immunosorbent assays for detection of immunoglobulin M to nontreponemal and treponemal antigens for the diagnosis of congenital syphilis.

Sera from newborn infants born of mothers with a high risk of syphilis were examined for immunoglobulin M (IgM) antibodies in two different enzyme-linked immunosorbent assays (ELISAs), using either purified flagella from Treponema phagedenis biotype Reiter or the Venereal Disease Research Laboratory (VDRL) antigen as the antigen. All sera were also examined by the fluorescent treponemal antibody absorption (FTA-ABS) test for IgM. Three different groups of patients were studied. Group 1 consisted of 84 women and their newborn infants from a high-risk population for syphilis. Congenital syphilis was diagnosed in one child who had an IgM-positive cord blood specimen in both the ELISA and the FTA-ABS test. Group 2 consisted of 10 mothers and their newborn children. All mothers had positive syphilis-screening tests, and all children had signs of congenital syphilis. All but one child had positive IgM tests. Group 3 consisted of 15 mothers and their newborn children. These mothers had been treated for syphilis late in pregnancy, and all had a positive screening test at delivery. Two of the children had positive IgM tests, probably caused by reactivity after late intrauterine treatment of congenital syphilis. The specificities of the IgM tests were high when evaluated with sera from newborn children without signs of congenital syphilis. Even though IgM rheumatoid factor was found in all of the children tested with definite congenital syphilis, the rheumatoid factor did not cause false-positive results in either the VDRL ELISA or the flagellum ELISA. No significant IgG-IgM competition was noticed in the ELISAs. This study also confirmed that IgA antibodies do not cross the placenta; most newborn children with congenital syphilis were positive in the VDRL ELISA for IgA. Both the VDRL ELISA and the flagellum ELISA are very useful in the diagnosis of congenital syphilis and may be substitute for the FTA-ABS test. The VDRL ELISA for IgM will be especially useful in developing countries with a high incidence of congenital syphilis.     

Next-Generation Sequencing Based Approach to Identify Underlying Genetic Defects of Glanzmann Thrombasthenia

Glanzmann thrombasthenia (GT) is an autosomal recessive platelet function disorder characterized by mucocutaneous bleeding as the most common clinical phenotype. Patients with GT have normal platelet counts, platelet morphology but reduced platelet aggregation in response to various agonists. Homozygosity or compound heterozygosity for variants in the ITGA2B/ITGB3 genes is the genetic basis for GT. Establishing a molecular diagnosis is definitive and is important for predictive testing. Using multi-gene panels is an accurate, faster, and cost-effective mode as compared to Sanger sequencing in large genes. We used a targeted resequencing based approach to identify pathogenic variants in eight cases in seven families. These variants were validated using Sanger sequencing in patients as well as family members and were predicted probably pathogenic using in-silico prediction tools. The variants include three missense (3/7 = 43%) (ITGA2B:c.1028 T > C, ITGA2B:c.1186G > A, ITGB3:c.1388G > C), two deletions (ITGA2B:c.559delG, ITGA2B:c.3092delT), one duplication (ITGA2B:c.1424_1427dupAGGT) and nonsense variant (ITGA2B:c.2578C > T, p.Gln860Ter). Except for one case which was compound heterozygous, the rest of the cases were homozygous. We found two novel variants that are reported for the first time in GT. The targeted resequencing based approach revealed varied genetic variants in North Indian patients, including two novels ones. The high yield of our panel indicates its suitability for usage in larger cohorts for the genetic diagnosis of GT patients. This approach is cost-effective and less cumbersome as compared to Sanger sequencing for these large size genes with multiple exons. The information so obtained is helpful in prenatal testing, carrier analysis, and genetic counseling.     

Long-term preservation of whole blood samples for flow cytometry analysis in normal and HIV-infected individuals from Africa.

A whole blood method requiring less than 4 ml of heparinized blood was developed to assess the practicality of preparing whole blood samples that could be easily stored, transported and readily used to determine the lymphocyte phenotypes and proliferation responses of individuals from remote areas who are infected with the human immunodeficiency virus. Minor modifications in standard whole blood procedure for lymphocyte phenotyping have significantly increased the stability of light scatter and fluorescence intensity of the cells for subsequent flow cytometry (FC) analysis. These changes include removal of lysis solution prior to fixation, fixation of monoclonal antibody-stained cells in 1% paraformaldehyde for 30 minutes and storage of fixed samples in medium containing 1% bovine serum albumin. Lymphocyte subsets and their functional subsets could reliably be determined on samples stored for up to 4 weeks. Further, blood samples could be kept at room temperature for up to 96 hours or at ambient temperature during transportation from Africa before staining for FC without affecting their quantitation. While samples could be processed for FC analysis under field-laboratory conditions, proliferation assays could only be performed on samples that were transported within 48 hours of their collection. The whole blood method saves time and expense and decreases the volumes of blood required to perform phenotypic analysis and functional assays on specimens collected in remote areas.     

Rat‐bite fever identified by polymerase chain reaction detection of Streptobacillus moniliformis DNA

A 74‐year‐old woman presented with erythema of the extremities, a high fever and arthralgia after being bitten by a rat. The patient was diagnosed as having rat‐bite fever based on the symptoms and clinical course, as well as the polymerase chain reaction detection of Streptobacillus moniliformis DNA in the crust of the bite site. This is the first case to be diagnosed using polymerase chain reaction on a crusted skin lesion specimen. Although clinical symptoms initially remitted with minocycline therapy, they relapsed. Subsequent administration of piperacillin sodium resulted in complete disappearance of the high fever and arthralgia.     

Cholecystokinin secretion induced by β-conglycinin peptone depends on Gαq-mediated pathways in enteroendocrine cells

Background  Intraduodenal administration of peptone prepared from soybean β-conglycinin (BconP) stimulates cholecystokinin (CCK) secretion from enteroendocrine cells, and suppresses food intake in rats. However, the sensing mechanism of BconP by CCK-producing cells is unknown. Aim of the study  We investigated signal transduction pathways mediating CCK secretion in response to BconP in the murine CCK-producing cell line, STC-1. Methods  STC-1 cells were seeded in 48-well culture plates until sub-confluent and CCK secretion was examined under various conditions. CCK concentration was determined by the enzyme immunoassay. Results  BconP dose-dependently induced CCK secretion in STC-1 cells. Treatment with BAPTA-AM, an intracellular Ca²⁺ chelator, reduced BconP-induced CCK secretion, however, removal of extracellular Ca²⁺ did not affect the secretory response. Treatment with 2-amino borate (2-APB) reduced CCK releasing responses, suggesting the involvement of IP₃. In addition, BconP failed to induce CCK secretion after treatment with the Gαq protein inhibitor (YM-254890). Conclusion  These results indicate that Gαq pathway is responsible for BconP-induced CCK secretion in STC-1 cells, and suggest the involvement of a Gαq-coupled GPCR(s) in dietary peptide sensing in enteroendocrine cells.     

Prevalence of concurrent Chlamydia trachomatis infections among men with gonococcal urethritis in Lusaka

The prevalence of Chlamydia trachomatis infections among male patients with gonococcal urethritis in the sexually transmitted diseases (STDs) clinic at UTH, Lusaka, was determined by two methods to be around 4.7%. Methods used were specific for C. trachomatis (Syva Microtrak), and second isolation of C. trachomatis. The results indicate that with a prevalence rate of 4-7% for concurrent gonococcal and chlamydial infection, it is not financially or medically practical to treat all cases of gonorrhoea in Zambia for possible chlamydial infection, as is advocated in some countries. The findings further suggest that the commercially available Syva Microtrak test should probably be considered positive when one or two monoclonal-stained elementary bodies are seen, rather than ten, to avoid false-negative results.     

Use of Nd-YAG laser in an emergency partial splenectomy

Traumatic rupture of the spleen is usually treated by splenectomy. This has immunological disadvantages for children and teenagers. We report a case where, after traumatic rupture, the splenic hilar remnant was preserved by using a neodymium-YAG laser.     

A single dose of self-transcribing and replicating RNA-based SARS-CoV-2 vaccine produces protective adaptive immunity in mice

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