IgG Avidity in Samples Collected on Filter Paper: Importance of The Early Diagnosis of Congenital Toxoplasmosis

Objective The purpose of the present study is to standardize and evaluate the use of the immunoglobulin G (IgG) antibody avidity test on blood samples from newborns collected on filter paper to perform the heel test aiming at its implementation in ongoing programs. Methods Blood samples from newborns were collected on filter paper simultaneously with the heel prick test. All samples were subjected to immunoglobulin M IgM and IgG enzyme-linked immunosorbent assays (ELISA). Peripheral blood was collected again in the traditional way and on filter paper from newborns with high IgG levels (33). Three types of techniques were performed, the standard for measuring IgG in serum, adapted for filter paper and the technique of IgG avidity in serum and on filter paper. The results of the avidity test were classified according to the Rahbari protocol. Results Among the 177 samples, 17 were collected in duplicate from the same child, 1 of peripheral blood and 1 on filter paper. In this analysis, 1 (5.88%) of the 17 samples collected in duplicate also exhibited low IgG avidity, suggesting congenital infection. In addition, the results obtained from serum and filter paper were in agreement, that is, 16 (94.12%) samples presented high avidity, with 100% agreement between the results obtained from serum and from filter paper. Conclusion The results of the present study indicate that the avidity test may be another valuable method for the diagnosis of congenital toxoplasmosis in newborns.

Detection of Acute and Chronic Toxoplasma gondii Infection among Women with History of Abortion in the Southwest of Iran

Background. Toxoplasma gondii (T. gondii) is one of the most common intracellular protozoan parasites, which can infect humans and a wide range of mammals and birds. The current study is aimed at investigating the occurrence of T. gondii infection in women with a history of abortion in Khuzestan, Iran. Materials and Methods. A total of 480 women with an abortion history, as well as 200 pregnant women with a normal delivery, were examined in this study. The blood, placenta, and umbilical cord blood samples were assessed by the enzyme-linked immunosorbent assay (ELISA) and nested-polymerase chain reaction (PCR) assay. Results. Based on the results of ELISA assay, the prevalence of toxoplasmosis was 30.83% in women with a history of abortion (25.62% with T. gondii IgG and 5.20% with T. gondii IgM). According to the IgG avidity test, 60.16% of IgG-positive samples showed high avidity, while 27.64% showed low avidity. On the other hand, the prevalence of toxoplasmosis in women with a normal delivery was 23% (21.5% with T. gondii IgG and 1.5% with T. gondii IgM). According to the IgG avidity test, 81.39% of these women showed high avidity, while only 4.65% showed low avidity. Based on the nested-PCR method, T. gondii DNA was detected in 14.18% of blood samples, 4.69% of placental samples, and 1.34% of umbilical cord samples, collected from 148 seropositive women with a history of abortion. Besides, using this method, the parasite DNA was identified in 4.34% of blood samples, collected from 46 seropositive women with a normal delivery, but not in any of the umbilical cord or placenta samples. Conclusion. The present results showed that T. gondii infection contributes to abortion in Khuzestan Province, Iran. Therefore, it is essential to investigate toxoplasmosis in pregnant women, especially in those who are seronegative, using molecular and serological methods and inform them about their disease and the associated risks.

Prevalence and Recurrence Rates of Cytomegalovirus Infection Among Patients With Hematological Diseases in the Western Brazilian Amazon: A Cross-Sectional Study

Cytomegalovirus (CMV) is a worldwide distributed pathogen that may cause serious complications in patients with hematological diseases. This study aimed to serologically characterize CMV infection in patients suffering from hematological diseases in Amazonas state, Brazil. Serum samples from 323 patients were tested for the presence of anti-CMV IgM or IgG antibodies using an enzyme-linked immunosorbent assay. Positive samples for IgM were submitted to the IgG avidity test to differentiate primary infection from recurrent infection. An epidemiological questionnaire was administered to collect the sociodemographic information of the study population. The overall prevalence of CMV infection verified in this study was 91.3%. The highest rates were found in patients suffering from platelet disorders (94.5%), anemia (93.3%), or leukemia (91%). The study population was predominantly composed of individuals with low socioeconomic status. Blood transfusions were more common in patients with anemia or leukemia, but this variable was not correlated with the seropositivity for CMV infection. Measurement of IgG avidity in patients positive for anti-CMV IgM demonstrated a recurrent infection rate of 5.2% (17/323). Over 80% of recurrent infections occurred in patients with acute lymphocytic leukemia (ALL) or anemia. Our findings indicated that CMV infection is highly prevalent in patients from the western Brazilian Amazon who have hematological diseases. The prevalence observed progressively rose with increasing age, whereas anemia or ALL figured as risk factors for the recurrence of CMV infection.

Longitudinal Study of SARS-CoV-2 Antibody Characteristics Using Label-Free Immunoassays

Introduction/Objective Since the start of the COVID-19 pandemic, much research has focused on the kinetics and magnitude of humoral immune response. With the advantages of monitoring real-time immunoreactions, label-free immunoassay (LFIA) is becoming a powerful tool in serology studies. We have developed LFIAs to measure SARS- CoV-2 antibody avidity and neutralization activity in a cohort of COVID-19 patients and determine if they correlate with antibody concentration. Serial serum samples collected from mild to severe COVID-19 patients were measured out to 8 months post-symptom onset to determine the durability of the neutralizing antibody response. Methods/Case Report Based on thin-film interferometry technology, we established a label-free IgG avidity assay and a label-free surrogate virus neutralization test (LF-sVNT). For measurement, sensing probes pre-coated with receptor-binding domain (RBD) of SARS-CoV-2 spike protein are applied to serum samples containing SARS-CoV-2 antibodies. The label-free IgG avidity assay measures the binding strength between RBD and IgG under urea dissociation. The LF-sVNT analyzes the binding ability of RBD to ACE2 after neutralizing RBD with antibodies. Results (if a Case Study enter NA) IgG avidity indices and neutralizing antibody titers (IC50) were determined from serum samples (n=246) from COVID-19 patients (n=113). IgG concentrations were measured using a fluorescent immunoassay. The neutralizing antibody titers showed a weak correlation with IgG concentrations and no correlation with IgG avidity indices. Over the time course up to 8 months post-symptom onset, IgG concentrations and neutralizing antibody titers presented similar trends: an initial rise, plateau and then in some cases a gradual decline after 40 days. The IgG avidity indices, in the same cases, plateaued after the initial rise. Conclusion The results demonstrated that LFIA could be used an excellent solution in the determination of SARS- CoV-2 antibody characteristics. The study found that IgG concentration and neutralizing antibody titer declined over time, while IgG avidity index remained constant after reaching a plateau. The decline of antibody neutralization activity can be attributed to the reduction in antibody quantity rather than the deterioration of antibody quality, as measured by antibody avidity.

Clinical, Virologic and Immunologic Correlates of Breast Milk Acquired Infections in Very Low Birth Weight (VLBW) Infants in a Newborn Intensive Care Unit (NICU) Setting

Cytomegalovirus (CMV) infections acquired by very-low-birthweight (VLBW) infants are incompletely characterized. To examine CMV transmission in VLBW infants, we evaluated maternal DNAlactia, infant DNAemia, and presence of clinical disease in a blinded study in VLBW infants in our newborn intensive care unit (NICU). To examine these issues, 200 VLBW infants were enrolled in a surveillance study, with weekly breast milk and infant whole blood samples collected, as available. Virologic (breast milk and infant whole blood real time PCR) and immunologic (IgG, IgM, and IgG avidity) correlates were evaluated. A chart review examined whether infants had symptoms compatible with CMV disease. DNAlactia was identified in 65/150 (43%) of lactating mothers. Nine CMV infections were identified in 9/75 CMV-exposed infants (12% of exposed infants). A higher median breast milk viral load (DNAlactia) correlated with an increased likelihood of DNAemia (p = 0.05). Despite potential symptoms compatible with CMV infection, clinicians had not considered the diagnosis of CMV in 6/9 cases (66%). All of these infants had chronic lung disease at discharge. There was no correlation between IgG antibody titer or IgG avidity index and the likelihood of transmission or CMV disease. In conclusion, in VLBW infants receiving milk from seroposi-tive mothers, CMV infections are commonly acquired, and are frequently unrecognized. Future studies are needed to determine whether routine surveillance for CMV of either breast milk or infant plasma is beneficial in preventing or recognizing infection.

Serodiagnosis of Toxoplasmosis by Using ELISA and IgG Avidity Test in Relation to Some Risk Factors among Women at Childbearing Age in Zakho City, Duhok Province/ Iraq.

This study intended to evaluate the seroprevalence of anti-Toxoplasma IgG and IgM antibodies in the sera of 630 women at childbearing age, and to link the outcomes with some risk factors. The enrolled women visited Zakho Maternity Hospital from July 2018 to July 2019. Their ages ranged from 15 to 45 years. All samples were examined using ELISA to detect immunoglobulin G and M, in addition to performing IgG Avidity test for seropositive pregnant women. The differences between seropositivity and age was significant (p<0.05), the highest rate (20.43%) for anti-Toxoplasma IgG antibodies in the age group 33-38 years. Women who had more contact with cats showed higher IgG and IgM seropositivity rates (16.45% and 1.26%, respectively). Married women had higher IgG Abs seropositivity than single ones (12.52% vs 6.31%, respectively), moreover, only married women were seropositive for IgM Abs. Pregnant women presented higher IgG Abs seropositivity than non-pregnant (15.21% versus 10.49%), with almost equal seropositivity for IgM Abs (0.65% and 0.86%, respectively). Anti-Toxoplasma IgG Abs seropositivity was higher in women underwent miscarriages than those with normal pregnancies (18.44 vs. 8.81%), however IgM Abs was only found among women who had miscarriages (0.97%). Women with triple miscarriages presented the highest IgG Abs seropositivity (37.03%). Chronic infection was found in 68.75% of pregnant women, whereas acute infection was found in 31.25 %. Following up the pregnancy resulted in 15 healthy births, 9 miscarriages, and 10 women did not show up. The findings of this study demonstrate the relationship between toxoplasmosis and risk factors in women at childbearing age, with the aim of decreasing infection rates through the health education and application of hygienic measures.

Evolution of Anti-SARS-CoV-2 IgG Antibody and IgG Avidity Post Pfizer and Moderna mRNA Vaccinations

Messenger RNA (mRNA) based vaccines (Pfizer/BioNTech and Moderna) are highly effective at providing immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, there is uncertainty about the duration of immunity, evolution of IgG antibody levels and IgG avidity (an index of antibody-antigen binding strength), and differences in the immune responses between vaccines. Here we performed a prospective pilot study of 71 previously COVID-19 free subjects upon receiving both doses of either the Pfizer (n = 54) or Moderna (n = 17) mRNA vaccine. Anti-spike protein receptor binding domain (RBD) IgG antibodies were measured longitudinally using a qualitative finger stick MidaSpot rapid test at the point-of-care for initial screening and a quantitative dry blood spot-based pGOLD laboratory test over ~ four months post-vaccination. The average anti-RBD IgG antibody levels peaked at ~ two weeks after the second dose vaccine and declined thereafter, while antibody avidity increased, suggesting antibody maturation. Moderna vaccine recipients compared to Pfizer vaccine recipients exhibited higher side effect severity, higher peak anti-RBD IgG antibody levels, and higher avidity up to the 90 days period. Differences in antibody levels diminished at ~ 120 days post-vaccination, in line with the similar efficacy observed in the two vaccines. The MidaSpot rapid test detected 100% anti-SARS-CoV-2 RBD positivity for fully vaccinated subjects in both Pfizer and Moderna cohorts post full vaccination but turned negative greater than 90 days post-vaccination for 5.4% of subjects in the Pfizer cohort, whose quantitative anti-IgG were near the minimum levels of the group. Immune responses were found to vary greatly among vaccinees. Personalized longitudinal monitoring of antibodies could be necessary to assessing the immunity duration of vaccinated individuals.

Assessment of an in-house ELISA and IgG avidity test based on SAG1 and GRA7 proteins for discriminating between acute and chronic toxoplasmosis in humans

To improve serodiagnostic methods for diagnosis of acute from chronic toxoplasmosis, an economical in-house ELISA for measuring Toxoplasma -specific IgG, IgM and IgG avidity has been developed and assessed based on use of various T. gondii antigens, including SAG1, GRA7 and a combination of SAG1 and GRA7 (SAG1+GRA7) as well as Toxoplasma lysate antigens (TLAs). Performances of in-house IgM, IgG and IgG avidity assays were compared to those of ELISA commercial kits and VIDAS Toxo IgG avidity. A set of 138 sera from patients with acquired T. gondii infection and seronegative people were assessed. Receiver operating characteristic (ROC) analysis revealed an area under curve (AUC) of 0.98, 0.97, 0.99 and 0.99 for IgM-TLAs, IgM-SAG1, IgM-GRA7, and IgM-SAG1+GRA7, respectively. Furthermore, AUC was calculated as 0.99, 0.99, 0.98 and 0.99 for IgG-TLAs, IgG-SAG1, IgG-GRA7 and IgG-SAG1+GRA7, respectively. The current study showed that GRA7 included 100% sensitivity for the detection of Toxo IgM, while SAG1 included 89.7% sensitivity. Furthermore, the highest specificity (97.2%) to detect Toxo IgM was achieved using SAG1+GRA7 antigen. For the detection of Toxo IgG, the highest sensitivity (100%) was recorded for SAG1+GRA7 followed by TLAs (97.9%). The SAG1+GRA7 showed the greatest potential for assessing avidity of IgG antibodies with 97.1% of sensitivity and 96.6% of specificity, compared to VIDAS Toxo IgG avidity. The preliminary results have promised better discriminations between acute and chronic infections using a combination of SAG1 and GRA7 recombinant antigens, compared to TLAs.