Toxoplasmosis, worldwide protozoan disease, is usually benign, except when acute disease occurs in pregnant women, resulting in fetal infection with deaths or high morbidity after birth. Treatment blocks fetal infection or damage after infection, imposing a quick and effective diagnosis. Maternal infection is mostly asymptomatic thus regular serology are the main tool for detect seroconversion and acute infection in prenatal care. Screening test for specific anti T. gondii IgG, IgM and IgA must be quick, cheaper and available for prenatal care.
Fluorescent solid-phase assays appear as a good alternative as they allow one well detection of IgG and IgM aside to allow high throughput in 384 wells. Here, we standardize and analyze a single well anti-T. gondii IgG, IgM and IgA immunosorbent fluorescent assay in a large sample of a public hospital. We construct conjugates for each immunoglobulin with specific fluorophores, which allows concomitant detection in a microplate fluorimeter, with stability and reproducibility, allowing cheaper 384 wells use.
Tested in our 600 mother samples from a large public hospital, they presented the same reactivity as standard routine tests, but with adequate IgM and IgA screening, as adequately standardized in house ELISA, while the design of most commercial assays give false positive results. The few TFISA positive IgG, IgM and IgA samples also had low avidity IgG, confirming recent infection. TFISA will help a screening toxoplasmosis in pregnancy program in large cities, with , allowing testing large numbers of samples at low cost and must be considered for other serological purposes.

Elevated SARS-Cov-2-Specific IgM Levels Indicate Clinically Unfavorable Outcomes in Patients with COVID-19: A Retrospective Cohort Study

COVID-19 outbreak began in Wuhan and pandemics occur. Although SARS-CoV-2-specific immunoglobulins have been detected in serum of COVID-19 patients, their dynamics and association with outcomes have not been fully characterized.
Methods: This retrospective cohort study investigated the association between SARS-CoV-2-specific immunoglobulins and clinical outcomes of COVID-19 patients. We recruited 137 participants who were diagnosed with COVID-19 in four wards of the Tongji Hospital in Wuhan, China. Among the 137 participants, 81 patients were recovered, 23 patients died, and 33 patients remained hospitalized by the end of the study. SARS-CoV-2-specific immunoglobulins were analyzed by chemiluminescence assays. Laboratory and radiological characteristics, and clinical outcomes were compared between the recovered group and the deceased group. Furthermore, a matched cohort study was conducted in which each non-survivor was matched to two recovered patients of similar age.
Results: SARS-CoV-2-specific IgM levels peaked in the fourth week after the onset of COVID-19, while serum IgG levels rose earlier and remained high up to the eighth week. In the age-matched cohort study, the serum IgM, but not IgG levels, were higher among the non-survivors than in the recovered group (P = 0.006).
The area under the ROC curve for the IgM and IgG levels was 0.702 (95% CI: 0.560-0.845, P = 0.006) and 0.596 (95% confidence interval: 0.449-0.744, P = 0.194), respectively. We also showed that patients with COVID-19 who had high IgM or IgG levels (stratified according to best cut-off) exhibited significantly lower overall survival (Kaplan-Meier survival curves, P < 0.05).
Discussion: These results indicate the association between immunoglobulins and outcome in patients with COVID-19 and demonstrated that elevated serum IgM levels could indicate poor outcomes in patients with COVID-19. Further, the information about the profile of SARS-CoV-2-specific IgGs may be useful for the future epidemiological investigations of COVID-19 therapies.

P064 Association Between Crohn’s Disease (CD) and Transient IgM and IgG Immunodeficiency-Diagnostic and Management Challenges

Case: Background: CD is an inflammatory disease that predominantly affects the gastrointestinal tract and has a progressive course. Immunodeficiencies can occur by altering different components of innate or adaptive immune responses. Such changes put the patient at greater risk for infectious diseases or non-infectious complications. Among the non-infectious complications are cancer, autoimmune diseases and gastrointestinal diseases, which should be suspected in patients with recurrent infections, with clinical, radiological or histological features of the disease are atypical or in the occurrence of an unsatisfactory response to conventional therapy.
Objective: To describe a case report of a CD patient complicated with enteric alterations related to immunodeficiency.
Results: A 31-year-old male patient started presenting abdominal pain, diarrhea, and weight loss in October 2020. He underwent a colonoscopy that showed diffuse involvement of the colon suggestive of CD. He didn`t respond to therapy with prednisone and azathioprine and lost 40kg until January 2021, when he was referred to an IBD center. He had signs of malnutrition (eg, BMI = 15.4, Albumin = 1.8g/dL).
The MRI showed diffuse thickening of the walls of the entire length of the ileum, colon and rectum, associated with submucosal edema and more intense enhancement in the inner layer. Findings compatible with nonspecific diffuse ileocolitis. After negative infectious screening, hydrocortisone, azathioprine and infliximab were prescribed, but the patient remained clinical worsening.
During the 6 weeks of therapy use, the patient presented oral candidiasis, septic pyoarthritis, DNA testing for Clostridium difficile and PCR for Cytomegalovius were positive. Despite the treatment of infectious diseases, the patient continued to worsen and was considered a primary non-response to anti-TNF, opting for the initiation of Ustekinumab. There was improvement in endoscopic and fecal calprotectin, but continued progression of ileal disease and diarrhea, with no response to enteral or parenteral nutritional therapies.
An investigation for immunodeficiencies was carried out and IgM/IgG deficiency was noted, and parenteral immunoglobulin was started. He presented an intestinal subocclusion, requiring a loop ileostomy. In this surgery, it was decided to send a sample of ileal tissue for diagnostic differentiation, where ileitis was observed with intense plasma cell reaction and lymphoid nodular hyperplasia, uncharacteristic of DC, which may correspond to enteropathy secondary to immunodeficiency.
Despite the treatment, the patient presented fungal endocarditis, esophageal moniliasis and septic condition of undetermined origin. He remained in hospital for 120 days until complete improvement of the infectious complications. With the slow improvement in nutritional parameters, there was a reduction in the incidence of infections as well as an increase in immunoglobulins. After five months of combined nutrition and anti-interleukin therapy, the patient presented clinical improvement, weight gain and complete normalization of IgG and IgM. Ileal disease is in regression and in the colon, in endoscopic remission.

IgM to IgG Class Switching Is a Necessary Step for Pemphigus Phenotype Induction in Desmoglein 3-Specific B Cell Receptor Knock-in Mouse

Pemphigus vulgaris is an autoimmune blistering disease caused by IgG targeting desmoglein 3 (Dsg3), an adhesion molecule of keratinocytes. Anti-Dsg3 IgG production is prevented in healthy individuals, but it is unclear how Dsg3-specific B cells are regulated. To clarify the immunological condition regulating Dsg3-specific B cells, a pathogenic anti-Dsg3 Ig (AK23) knock-in mouse was generated. AK23 knock-in B cells developed normally without undergoing deletion or acquiring an anergic phenotype in vivo. The knock-in B cells showed Ca2+ influx upon IgM cross-linking and differentiated into AK23-IgG+ B cells after LPS and IL-4 stimulation in vitro that induced a pemphigus phenotype after adoptive transfer into Rag2 -/- mice.
However, the knock-in mouse itself produced AK23-IgM but little IgG without blisters in vivo. Dsg3 immunization and skin inflammation caused AK23-IgG production and a pemphigus phenotype in vivo. Furthermore, Fcgr2b deficiency or haploinsufficiency spontaneously induced AK23-IgG production and a pemphigus phenotype with poor survival rates in AK23 knock-in mice.

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NOVATest IgG/IgM Antibody Rapid Test Kit (NOVA Test)

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Japanese Encephalitis Differential IgG/IgM Test

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TOXO IgG/IgM Ab Rapid Test Kit

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Human Cytomegalovirus IgG / IgM, Toxoplasma IgG / IgM, Rubella IgG Rapid Test Kit

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SD Dengue Duo NS1 Ag and IgG/IgM test

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TOX (Toxoplasma Antibody IgM) ELISA test

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Human IgG Quick test strip

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Mouse IgG Quick test strip

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To assess Fcgr2b involvement in Ig class-switch efficiency, postswitch transcripts of B cells were quantified and significantly higher in Fcgr2b-/- and Fcgr2b+/- mice than wild-type mice in a gene dose-dependent manner. Finally, RNA sequencing revealed reduced expression of FCGR2B and FcγRIIB-related genes in patient B cells. These results indicated that Dsg3-specific B cells do not spontaneously perform pathogenic class switching in vivo, and pemphigus phenotype induction was prevented under normal conditions. Attenuated FcγRIIB signaling is also one of the drivers for pathogenic class switching and is consistent with immunological features identified from clinical samples. This study unveiled a characteristic immune state silencing autoreactive B cells in mice.