IgG and IgA Antibodies Post SARS-CoV-2 Vaccine in the Breast Milk and Sera of Breastfeeding Women

The COVID-19 pandemic has carried massive global health and economic burden that is currently counteracted by a challenging anti-COVID-19 vaccination campaign. Indeed, mass vaccination against COVID-19 is expected to be the most efficacious intervention to mitigate the pandemic successfully. The primary objective of the present study is to test the presence of neutralizing anti-SARS-CoV-2 antibodies (IgA and IgG) in the breast milk and sera samples from vaccinated women at least 20 days after the complete vaccine cycle. A secondary aim is to compare the IgG antibodies level in maternal serum and breast milk. The third target is to evaluate the presence of the IgG antibodies in breast milk after several weeks from the vaccination. Finally, we collected information on the health status of infants in the days following maternal vaccination. Forty-two mothers were enrolled in the study. Thirty-six received the Pfizer/BioNTech vaccine, four the Astra Zeneca vaccine, one the Moderna vaccine and another woman Astra Zeneca in the first dose and Pfizer/BioNTech in the second dose. All 42 milk samples confirmed the presence of anti-SARS-CoV-2 IgG, and none showed IgA presence. Regarding the matched 42 sera samples, 41 samples detected IgG presence, with one sample testing negative and only one positive for seric IgA. None of the 42 infants had fever or changes in sleep or appetite in the seven days following the maternal vaccination. The level of IgG antibodies in milk was, on average, lower than that in maternal serum. According to our analysis, the absence of IgA could suggest a rapid decrease after vaccination even if frequent breastfeeding could favour its persistence. IgG were present in breast milk even 4 months after the second vaccine dose. Information on the immunological characteristics of breast milk could change mothers’ choices regarding breastfeeding.

RADIOIMMUNOASSAY FOR DETECTION OF ANTI-RINDERPEST ANTIBODIES IN CATTLES

Liquid Phase Radioimmunoassay (RIA) was developed to detect and measure anti-rinderpest immunoglobulins in field animals sera, within two hours. Rinderpest virus adapted on Vero cell line culture and antigen purified by treatment with Triton-Gentron 13 Butanol, and, labeled with I isotope, using chloramin T iodination method. Comparative studies for detecting anti-rinderpest immunoglobulin in 80 calves sera samples, using the developed assay in parallel with virus neutralization test (VNT). The study showed 58.75 % agreement between the two methods. However, 71 % of seven months old, non-vaccinated calves showed anti rinderpest antibodies in their sera, also 81 % of 10 months old vaccinated calves were developed antibodies in their blood. These results demonstrate the development of sensitive, specific and rapid quantitative / qualitative radioimmunoassay, necessary for screening the development of immunity against rinderpest in cattle.

A serological survey of canine leptospirosis in the city of Belgrade, Serbia

Canine leptospirosis is a zoonosis caused by bacteria belonging to the genus Leptospira. Dogs are one of the animal species involved in the cycle of preservation and transmission of leptospirosis in urban areas. Serological testing for the presence of specific antibodies against Leptospira spp. in dogs was continuously performed between 2010 and 2020 in the city of Belgrade. At the request of the owners themselves, other veterinary laboratories or laboratory clinics, 179 blood sera from 179 dogs were examined in the Laboratory for Immunology, Scientific Institute of Veterinary Medicine of Serbia. Blood sera samples from dogs were examined using the standard microscopic agglutination test (MAT) for the presence of specific antibodies against seven different serovars of Leptospira: Pomona, Icterohaemorrhagiae, Grippotyphosa, Sejroe, Canicola, Bataviae, and Australis. The number of seropositive dogs was 17/179 (9.5%). Among all examined sera, the highest titre of seropositive samples was to serovar Icterohaemorrhagiae (10/17, 58.8%), followed by Pomona (4/17, 23.5%), and serovar Canicola (3/17, 17.6%). Specific antibodies for serovars Grippotyphosa, Sejroe, Bataviae and Australis were not detected in any of the dog sera. Cross-reaction (the presence of two or three titres with different values where one of them was higher than others) between different serovars was diagnosed in a low number of sera (n=4), with the following serovars: Icterohaemorrhagiae and Pomona (n=3) and Pomona and Canicola (n=1). The confirmed specific antibody titres for Leptospira spp. were between 1:100 to 1:3000 (5 sera had titres of 1:100, 7 had titres of 1:300, 4 had titres of 1:1000, and 1 serum had a titre 1:3000). Monitoring canine leptospirosis is a useful tool in preventing leptospirosis in Belgrade.

Epidemiological investigation for brucellosis in dogs of Thrissur

India is endemic to bovine brucellosis, and there is a high potential for transmission of disease from ruminants to dogs. A total of 18 bitches belonging to five different breeds at different stage of abortion (30 days to 65 days of gestation) were selected for this study. Majority of them were showing abortion (88.89 per cent) at 45 to 65 days of the gestation. Microscopic examination of Stamp stained smear obtained from the aborted foetal stomach contents revealed red coccobacillary organisms suggestive of Brucella spp.in three cases. RBPT on paired sera samples on day of presentation and three weeks after abortion showed agglutination within four minutes in five out of 18 female dogs. DNA extracted from the aborted tissues of a RBPT positive Labrador dog yielded amplicons of 193 base pair specific for Brucella spp. on PCR. The results obtained from this study stress the need for screening dogs for canine brucellosis in the current brucellosis surveillance and control programmes.

Pre-Existing Cross-Reactive Antibody Responses Do Not Significantly Impact Inactivated COVID-19 Vaccine-Induced Neutralization

Recent exposure to seasonal coronaviruses (sCoVs) may stimulate cross-reactive antibody responses against severe acute respiratory syndrome CoV 2 (SARS-CoV-2). However, previous studies have produced divergent results regarding protective or damaging immunity induced by prior sCoV exposure. It remains unknown whether pre-existing humoral immunity plays a role in vaccine-induced neutralization and antibody responses. In this study, we collected 36 paired sera samples from 36 healthy volunteers before and after immunization with inactivated whole-virion SARS-CoV-2 vaccines for COVID-19, and analyzed the distribution and intensity of pre-existing antibody responses at the epitope level pre-vaccination as well as the relationship between pre-existing sCoV immunity and vaccine-induced neutralization. We observed large amounts of pre-existing cross-reactive antibodies in the conserved regions among sCoVs, especially the S2 subunit. Excep t for a few peptides, the IgG and IgM fluorescence intensities against S, M and N peptides did not differ significantly between pre-vaccination and post-vaccination sera of vaccinees who developed a neutralization inhibition rate (%inhibition) <40 and %inhibition ≥40 after two doses of the COVID-19 vaccine. Participants with strong and weak pre-existing cross-reactive antibodies (strong pre-CRA; weak pre-CRA) had similar %inhibition pre-vaccination (10.9% ± 2.9% vs. 12.0% ± 2.2%, P=0.990) and post-vaccination (43.8% ± 25.1% vs. 44.6% ± 21.5%, P=0.997). Overall, the strong pre-CRA group did not show a significantly greater increase in antibody responses to the S protein linear peptides post-vaccination compared with the weak pre-CRA group. Therefore, we found no evidence for a significant impact of pre-existing antibody responses on inactivated vaccine-induced neutralization and antibody responses. Our research provides an important basis for inactivated SARS-CoV-2 vaccine use in the context of high sCoV seroprevalence.

Development and validation of a novel counter-immunoelectrophoresis assay for the detection of antibodies against extractable nuclear antigens

<p>The identification of autoantibodies is a primary diagnostic marker for the diagnosis of some autoimmune diseases. The sera of patients with connective tissue diseases commonly contain autoantibodies that target nuclear antigens. As such these antibodies are called antinuclear antibodies (ANAs). Furthermore, the clinical identification of ANAs in patient sera to specific nuclear antigens is a primary tool for the diagnosis of connective tissue diseases. For this purpose specific extractable nuclear antigens (ENAs) are used. The most commonly employed laboratory technique for the detection of ENA antibodies is the enzyme linked immunoabsorbant assay (ELISA). ELISA is a simple technique that can be automated and provides high diagnostic sensitivity for the detection of ENA antibodies comparatively to a counter immunoelectrophoresis (CIE) assay. However, compared to CIE ELISA lacks diagnostic specificity. Therefore there is a demand for an assay with high diagnostic specificity as a secondary diagnostic test for the detection of ENA antibodies. The central aim of this project was to develop and validate a novel CIE assay that used fluorescently labelled ENAs to detect ENA antibodies in patient serum. Development of the assay began with comparative testing of fluorescently labelled and unlabelled antigens to confirm that the immunochemical properties of the antigen remained intact. The CIE assay conditions were optimised for the detection of four ANAs SSA, SSB, RNP/Sm, and Sm using their corresponding ENAs. Assay conditions optimised were flurophore type, gel composition, buffer composition, antigen concentration, the running time, and analytical specificity. Evaluation of 281 clinical sera samples known to have previously tested positive test by ANA indirect immunofluorescence were performed using the optimised CIE assay and ELISA. The inter-rater agreement between the CIE assay and ELISA results was determined. Clinical details were used to retrospectively classify the clinical sera samples into clinical categories and case groups. This data was used for the diagnostic comparison of the CIE assay and ELISA results. There was strong inter-rater agreement between the SSA CIE assay and ELISA results. The diagnostic specificity and positive likelihood ratio values of the SSA CIE assay were superior to those of the ELISA, while diagnostic sensitivity and the negative likelihood ratio values were approximately the same. There was strong inter-rater agreement between the RNP/Sm CIE assay and ELISA. However, it was observed that the statistical measures of assay accuracy (diagnostic specificity and sensitivity, positive likelihood ratio, and negative likelihood ratio) for the RNP/Sm ELISA were superior to those of CIE assay. The diagnostic specificity of the SSB and Sm CIE assays were higher than that of their respective ELISAs. This was also true for the positive likelihood ratio values of the SSB and Sm CIE assays when compared to their respect ELISAs. The SSA CIE assay results suggest that this assay maybe a suitable replacement for ELISA as a diagnostic tool for the identification of anti-SSA antibodies and Sjogren’s syndrome. The SSB and Sm CIE assay results suggest that these assays would be suitable as secondary confirmatory diagnostic assays for the identification of their respective ENA antibodies. However, further performance evaluation of the assays within a routine diagnostic laboratory is required.</p>

Development and validation of a novel counter-immunoelectrophoresis assay for the detection of antibodies against extractable nuclear antigens

<p>The identification of autoantibodies is a primary diagnostic marker for the diagnosis of some autoimmune diseases. The sera of patients with connective tissue diseases commonly contain autoantibodies that target nuclear antigens. As such these antibodies are called antinuclear antibodies (ANAs). Furthermore, the clinical identification of ANAs in patient sera to specific nuclear antigens is a primary tool for the diagnosis of connective tissue diseases. For this purpose specific extractable nuclear antigens (ENAs) are used. The most commonly employed laboratory technique for the detection of ENA antibodies is the enzyme linked immunoabsorbant assay (ELISA). ELISA is a simple technique that can be automated and provides high diagnostic sensitivity for the detection of ENA antibodies comparatively to a counter immunoelectrophoresis (CIE) assay. However, compared to CIE ELISA lacks diagnostic specificity. Therefore there is a demand for an assay with high diagnostic specificity as a secondary diagnostic test for the detection of ENA antibodies. The central aim of this project was to develop and validate a novel CIE assay that used fluorescently labelled ENAs to detect ENA antibodies in patient serum. Development of the assay began with comparative testing of fluorescently labelled and unlabelled antigens to confirm that the immunochemical properties of the antigen remained intact. The CIE assay conditions were optimised for the detection of four ANAs SSA, SSB, RNP/Sm, and Sm using their corresponding ENAs. Assay conditions optimised were flurophore type, gel composition, buffer composition, antigen concentration, the running time, and analytical specificity. Evaluation of 281 clinical sera samples known to have previously tested positive test by ANA indirect immunofluorescence were performed using the optimised CIE assay and ELISA. The inter-rater agreement between the CIE assay and ELISA results was determined. Clinical details were used to retrospectively classify the clinical sera samples into clinical categories and case groups. This data was used for the diagnostic comparison of the CIE assay and ELISA results. There was strong inter-rater agreement between the SSA CIE assay and ELISA results. The diagnostic specificity and positive likelihood ratio values of the SSA CIE assay were superior to those of the ELISA, while diagnostic sensitivity and the negative likelihood ratio values were approximately the same. There was strong inter-rater agreement between the RNP/Sm CIE assay and ELISA. However, it was observed that the statistical measures of assay accuracy (diagnostic specificity and sensitivity, positive likelihood ratio, and negative likelihood ratio) for the RNP/Sm ELISA were superior to those of CIE assay. The diagnostic specificity of the SSB and Sm CIE assays were higher than that of their respective ELISAs. This was also true for the positive likelihood ratio values of the SSB and Sm CIE assays when compared to their respect ELISAs. The SSA CIE assay results suggest that this assay maybe a suitable replacement for ELISA as a diagnostic tool for the identification of anti-SSA antibodies and Sjogren’s syndrome. The SSB and Sm CIE assay results suggest that these assays would be suitable as secondary confirmatory diagnostic assays for the identification of their respective ENA antibodies. However, further performance evaluation of the assays within a routine diagnostic laboratory is required.</p>

A study on serodiagnosis of scrub typhus in a Teaching Hospital of South India

Background: crub typhus is caused by Orientia tsutsugamushi (rickettsial disease) commonly transmitted by the bite of larval chiggers of trombiculid mites. It has been one of the important causes of febrile illness, especially in south India. The clinical diagnosis is difficult owing to the non-specific presentation. We in the current study tried to evaluate the serodiagnosis of scrub typhus with the Weil Felix test and IgM ELISA. Methods: This study was conducted in the Department of Microbiology, Prathima Institute of Medical Sciences, Naganoor, Karimnagar. All the sera samples were subjected to the Weil Felix test using Proteus OX2, OX19, OX-K strain agglutination test, and subsequently, Scrub typhus IgM ELISA test. Results: All the samples were subjected to the Weil Felix test n=4(6.06%) were positive for scrub typhus (OXK antigen) n=11(16.67%) were positive for the spotted group of fever (OX2 antigen) and n=10 (15.15%) were positive of typhus group (OX19 antigen). N=5 sera samples were positive for more than one type of antigens. All the n=66 serum samples were subjected to IgM ELISA for scrub typhus. Out of n=66, only two serum samples (3.03%) were positive by IgM ELISA. Conclusion: Scrub typhus is emerging as an important public health issue. It is one of the important causes of acute febrile illness. Although it is difficult to distinguish scrub typhus based on the clinical symptoms alone a simple test such as Weil Felix was found to be promising in the diagnosis of scrub typhus. ELISA IgM test may be performed additionally in laboratories with adequate facilities. Hence for clinicians, any case with a fever of unknown origin should arouse suspicion of scrub typhus

Pilot Study of Risk Group Human Seroprevalence to Coxiella burnetii (Q Fever) in Latvia

Q fever is an important zoonotic disease worldwide. The main sources of human infection are inhalation of aerosols containing Coxiella burnetii bacteria and exposition to infected materials during parturition or slaughtering. The high-risk group includes people who work directly with infected livestock, such as farmers, veterinarians, veterinary medicine students, slaughterhouse and laboratory staff. Here we present a pilot study of risk-group human seroprevalence to C. burnetii in Latvia. The study included 240 sera samples — 190 from the risk groups and 50 from the control group. Samples were tested with Coxiella burnetii (Q-fever) Phase 1 and 2 IgG ELISA kits. All sera from the control group were negative. The seroprevalence among risk group persons was 8.04–11.54%. No statistically significant differences were observed between genders. The highest percentage of seropositive and equivocal sera samples (25%) were detected in age categories 39–48 years and 49–58 years. Working as a practicing veterinarian or former veterinarian was the only risk factor identified as statistically significant, and belonging to the risk group in general. The geographical distribution of seropositive risk group participants indicated that they tend to located more in the northern, central, and eastern part of the country.

Seroprevalence and associated risk factors of mycoplasmosis in layer chickens at southern region of Bangladesh

Mycoplasma gallisepticum (MG) is the most important pathogenic Mycoplasma spp. causing avian mycoplasmosis and brought about huge economic losses to poultry industry in Bangladesh. The present study was undertaken to know the seroprevalence of MG in layer birds in three different geographical areas of southern Barishal division, Bangladesh. Total 310 sera samples were collected from wing vein of 30 farms for this study. Sera samples were tested with Rapid Serum Agglutination (RSA) for MG using commercial Antigen Kit (manufactured by Lillidale Diagnostic) to detect the presence of antibodies against MG. The overall seroprevalence of MG by RSA was 36.13%. Seroprevalence of MG infection was dominant in winter season (45.54%) and significantly highest occurrence was recorded in age groups from 20-40 weeks of layer chickens (51.79%). Serological investigation in three different upazila of Barishal division showed the highest infection rate (45.26%) in medium scale flocks (1000-3000) in comparison to (21.43%) small (<1000) flocks. The seroprevalence of MG was highest in Swarupkathi (44.38%) than in Barishal Sadar (26%) and Banaripara upazila (28%). Biosecurity and managemental failure is the overall risk factor in all types of farm due to lack of proper knowledge among farmer. This study reveals the current scenario of mycoplasmosis in layer birds of three different areas of Barishal division. Asian J. Med. Biol. Res. 2021, 7 (3), 292-297