Characterization of the Specific Interaction between Sialoadhesin and Sialylated Campylobacter jejuni Lipooligosaccharides

ABSTRACT In Campylobacter jejuni-induced Guillain-Barré syndrome (GBS), molecular mimicry between C. jejuni lipooligosaccharide (LOS) and host gangliosides leads to the production of cross-reactive antibodies directed against the peripheral nerves of the host. Currently, the presence of surface exposed sialylated LOS in C. jejuni is the single known bacterial pathogenesis factor associated with the development of GBS. Using a unique, well-characterized strain collection, we demonstrate that GBS-associated C. jejuni strains bind preferentially to sialoadhesin (Sn, Siglec-1, or CD169), a sialic acid receptor found on a subset of macrophages. In addition, using a whole-cell enzyme-linked immunosorbent assay (ELISA), C. jejuni strains with sialylated LOS bound exclusively to soluble Sn. Mass spectrometry revealed that binding was sialic acid-linkage specific with a preference for α(2,3)-linked sialic acid attached to the terminal galactose of the LOS chain as seen in the gangliosides GD1a, GM1b, and GM3. This molecular interaction was also related to functional consequences as a GBS-associated C. jejuni strain that bound Sn in a whole-cell ELISA adhered to surface-expressed Sn of Sn-transfected CHO cells but was unable to adhere to wild-type CHO cells. Moreover, a sialic acid-negative mutant of the same C. jejuni strain was unable to bind Sn-transfected CHO cells. This is the first report of the preferential binding of GBS-associated C. jejuni strains to the Sn immune receptor (P = 0.014). Moreover, because this binding is dependent on sialylated LOS, the main pathogenic factor in GBS progression, the present findings bring us closer to unraveling the mechanisms that lead to formation of cross-reactive antibodies in GBS disease.

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Putative Novel Surface-Exposed Streptococcus agalactiae Protein Frequently Expressed by the Group B Streptococcus from Zimbabwe

Clinical and Vaccine Immunology ◽

10.1128/cvi.00133-09 ◽

2009 ◽

Vol 16 (9) ◽

pp. 1302-1308 ◽

Cited By ~ 6

Author(s):

Rooyen T. Mavenyengwa ◽

Johan A. Maeland ◽

Sylvester R. Moyo

Keyword(s):

Rabbit Antiserum ◽

Periodate Oxidation ◽

Cross Reactivity ◽

Enzyme Linked Immunosorbent Assay ◽

Protein Antigens ◽

Group B Streptococci ◽

Phenotypic Marker ◽

Whole Cell ◽

Strain Collection ◽

Group B

ABSTRACTGroup B streptococci (GBS) express a variety of surface-exposed and strain-variable proteins which function as phenotypic markers and as antigens which are able to induce protective immunity in experimental settings. Among these proteins, the chimeric and immunologically cross-reacting alpha-like proteins are particularly important. Another protein, R3, which has been less well studied, occurred at a frequency of 21.5% in GBS from Zimbabwe and, notably, occurred in serotype V strains at a frequency of 75.9%. Working with rabbit antiserum raised against the R3 reference strain ATCC 49447 (strain 10/84; serotype V/R3) to detect the expression of the R3 protein, we recorded findings which suggested that strain 10/84 expressed a strain-variable protein antigen, in addition to R3. The antigen was detected by various enzyme-linked immunosorbent assay-based tests by using acid extract antigens or GBS whole-cell coats and by whole-cell-based Western blotting. We named the putative novel antigen the Z antigen. The Z antigen was a high-molecular-mass antigen that was susceptible to degradation by pepsin and trypsin but that was resistant tom-periodate oxidation and failed to show immunological cross-reactivity with any of a variety of other GBS protein antigens. The Z antigen was expressed by 33/121 (27.2%) of strains of a Zimbabwean GBS strain collection and by 64.2% and 72.4% of the type Ib and type V strains, respectively, and was occasionally expressed by GBS of other capsular serotypes. Thus, the putative novel GBS protein named Z showed distinct capsular antigen associations and presented as an important phenotypic marker in GBS from Zimbabwe. It may be an important antigen in GBS from larger areas of southern Africa. Its prevalence in GBS from Western countries is not known.

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Evaluation of Three Commercial Varicella-Zoster Virus IgG Enzyme-Linked Immunosorbent Assays in Comparison to the Fluorescent-Antibody-to-Membrane-Antigen Test

Clinical and Vaccine Immunology ◽

10.1128/cvi.00183-12 ◽

2012 ◽

Vol 19 (8) ◽

pp. 1261-1268 ◽

Cited By ~ 15

Author(s):

Andreas Sauerbrei ◽

Anna Schäfler ◽

Jörg Hofmann ◽

Michael Schacke ◽

Bernd Gruhn ◽

...

Keyword(s):

Varicella Zoster Virus ◽

Fluorescent Antibody ◽

Marrow Transplant ◽

Enzyme Linked Immunosorbent Assay ◽

Lower Sensitivity ◽

Serum Samples ◽

Reliable Determination ◽

Whole Cell ◽

Varicella Zoster ◽

Cell Enzyme

ABSTRACTCommercial serologic assays for varicella-zoster virus (VZV), which enable reliable determination of VZV immune status and are amenable to automation, are needed. The present study compares the automated performance of the VZV whole-cell enzyme-linked immunosorbent assay (ELISA) Enzygnost anti-VZV/IgG, the Euroimmun anti-VZV ELISA (IgG) based on highly purified viral proteins, and the VZV glycoprotein (gp)-based Serion ELISA Classic VZV IgG. The fluorescent-antibody-to-membrane-antibody (FAMA) test was used as a reference. A total of 638 serum samples from VZV-negative children, blood donors, varicella vaccinees, and bone marrow transplant recipients were included. The Enzygnost anti-VZV/IgG and the Serion ELISA Classic VZV IgG showed sensitivities of 99.6% and 99.2%, respectively, and the Euroimmun anti-VZV ELISA (IgG) had a significantly lower sensitivity of 90.5%. Specificity was calculated as 100% for both the Euroimmun anti-VZV ELISA (IgG) and for the Enzygnost anti-VZV/IgG, and the Serion ELISA Classic VZV IgG had a significantly lower specificity of 89.4%. Quantitative results of all ELISAs correlated well, but there was a poor quantitative correlation between the ELISAs and FAMA. In conclusion, this study does not show any superiority of a gp- and a protein-based ELISA compared to a whole-cell ELISA for the automated detection of VZV-specific IgG. The automated performance of the Enzygnost anti-VZV/IgG assay correlated best with the FAMA reference assay.

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Identification of a Haemophilus influenzae5′-Nucleotidase Protein: Cloning of the nucA Gene and Immunogenicity and Characterization of the NucA Protein

Infection and Immunity ◽

10.1128/iai.68.5.2525-2534.2000 ◽

2000 ◽

Vol 68 (5) ◽

pp. 2525-2534 ◽

Cited By ~ 19

Author(s):

Robert J. Zagursky ◽

Peggy Ooi ◽

Kevin F. Jones ◽

Michael J. Fiske ◽

Robert P. Smith ◽

...

Keyword(s):

Sodium Dodecyl ◽

Enzyme Linked Immunosorbent Assay ◽

Type B ◽

Whole Cell ◽

E Coli ◽

Log Reduction ◽

Native Proteins ◽

Infant Rat ◽

Cell Enzyme ◽

Nthi Strain

ABSTRACT We report on the identification of a surface-exposed, highly conserved, immunogenic nontypeable Haemophilus influenzae(NTHi) protein, which elicits cross-reactive bactericidal antibodies against NTHi. The protein was extracted from NTHi strain P860295 with KSCN and purified; it migrated as a single band on a sodium dodecyl sulfate-polyacrylamide gel with an apparent molecular mass of 63 kDa. Mouse antiserum generated against the purified protein was reactive on whole-cell enzyme-linked immunosorbent assay (ELISA) with seven NTHi strains and type b Eagan and Whittier strains and exhibited bactericidal activity to homologous and heterologous NTHi strains. However, the protein is made in small amounts in NTHi as corroborated by immunoelectron microscopy. To further study this protein, we cloned, sequenced, and expressed it recombinantly in Escherichia coli. The recombinant protein is localized in the periplasm ofE. coli and has been purified to homogeneity. Both the recombinant and native proteins possess 5′-nucleotidase activity; hence, the protein has been called NucA. Mouse antiserum directed against the recombinant NucA protein was reactive on Western immunoblots and whole-cell ELISA with all H. influenzaestrains tested including Eagan and was bactericidal for two heterologous strains tested. The antiserum also resulted in a log reduction in bacteremia, in an infant-rat protection study withH. influenzae type b as the challenge strain. These features suggest that NucA is a potential subunit vaccine candidate against NTHi disease.

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Direct Human Papillomavirus E6 Whole-Cell Enzyme-Linked Immunosorbent Assay for Objective Measurement of E6 Oncoproteins in Cytology Samples

Clinical and Vaccine Immunology ◽

10.1128/cvi.00388-12 ◽

2012 ◽

Vol 19 (9) ◽

pp. 1474-1479 ◽

Cited By ~ 6

Author(s):

Yi-Shan Yang ◽

Karen Smith-McCune ◽

Teresa M. Darragh ◽

Yvonne Lai ◽

Ju-Hwa Lin ◽

...

Keyword(s):

Human Papillomavirus ◽

High Risk ◽

Immunosorbent Assay ◽

Objective Measurement ◽

Enzyme Linked Immunosorbent Assay ◽

Whole Cell ◽

Hpv Dna ◽

Hpv E6 ◽

Cell Enzyme ◽

Grade 3

ABSTRACTA novel, whole-cell enzyme-linked immunosorbent assay (ELISA) based on a non-type-specific anti-human papillomavirus (HPV) E6 antibody was tested on 182 residual cytological specimens. For samples with a designation of more severe thancervicalintraepithelialneoplasia grade 3 (CIN3+), 83% tested positive for E6; in a subset with paired testing for E6 ELISA and HPV DNA, 72% tested E6 positive and 92% tested high-risk (HR)-HPV DNA positive (P= 0.2). Among the women with a less than CIN3 diagnosis, 31% and 47% tested positive for E6 and HR-HPV DNA, respectively (P= 0.0006).

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Use of Monoclonal Antibodies To Serotype Bordetella pertussis Isolates: Comparison of Results Obtained by Indirect Whole-Cell Enzyme-Linked Immunosorbent Assay and Bacterial Microagglutination Methods

Journal of Clinical Microbiology ◽

10.1128/jcm.43.5.2449-2451.2005 ◽

2005 ◽

Vol 43 (5) ◽

pp. 2449-2451 ◽

Cited By ~ 14

Author(s):

R. S. W. Tsang ◽

M. L. Sill ◽

A. Advani ◽

D. Xing ◽

P. Newland ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Immunosorbent Assay ◽

Bordetella Pertussis ◽

Enzyme Linked Immunosorbent Assay ◽

Whole Cell ◽

Comparison Of Results ◽

Cell Enzyme

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Is it time to administer acellular pertussis vaccine to childbearing age/pregnant women in all areas using whole-cell pertussis vaccination schedule?

Therapeutic Advances in Vaccines and Immunotherapy ◽

10.1177/25151355211015842 ◽

2021 ◽

Vol 9 ◽

pp. 251513552110158

Author(s):

Abdoulreza Esteghamati ◽

Shirin Sayyahfar ◽

Yousef Alimohamadi ◽

Sarvenaz Salahi ◽

Mahmood Faramarzi

Keyword(s):

Pregnant Women ◽

Age Groups ◽

Vaccination Schedule ◽

Immunoglobulin M ◽

Enzyme Linked Immunosorbent Assay ◽

Childbearing Age ◽

Cross Sectional ◽

Whole Cell ◽

Childbearing Age Women ◽

The Mean

Background: Whole-cell pertussis (wP) vaccine administration is still advocated for children under 7 years of age in Iran. However, there is no recommendation for the administration of a dose of tetanus, diphtheria, and acellular pertussis (Tdap) vaccine to childbearing age/pregnant women in the Iranian vaccination program and it has increased the risk of infection through waning immunity during women’s childbearing age life. The study aimed to assess the levels of anti- Bordetella pertussis antibodies in childbearing age women of different ages in Iran. Methods: A cross-sectional study was conducted on a total number of 360 childbearing age women divided into six age groups, with 5-year intervals from 15 to 45 years old, in 2018–2019. Then, the levels of immunoglobulin A (IgA), immunoglobulin M (IgM), and immunoglobulin G (IgG) antibodies against B. pertussis were evaluated using enzyme-linked immunosorbent assay (ELISA). The IBM SPSS Statistics software (version 16.0) (SPSS Inc., Chicago, IL, USA) was used for data analysis. Results: The mean age of the participants was 30.01 ± 8.35 years (range 14–45 years). All the cases were IgM negative, but two IgA-positive individuals (in the age groups of 14–19 and 30–34 years) were reported. Overall, 239 (66.4%) cases were IgG positive. The mean age of IgG-positive cases was 30.37 ± 8.37 years. The IgG-positive cases were mostly in the age groups of 30–34 and 35–39 years [43 (71.1%)]. The odds of IgG positivity were 1.97. The highest odds of IgG positivity were seen in 30–34 and 35–39 years groups (2.52) and the lowest odds were seen in the 20–24 and 25–29 years groups (1.60). Using the Jonckheere–Terpstra test, the increasing trend of IgG changes in different age groups was not statistically significant (Tπ=5.78, p = 0.09). Conclusion: The infants of women of childbearing age might be prone to pertussis in countries using the wP vaccination schedule. It is suggested to administer a dose of Tdap to women before or during pregnancy to increase the immunity of their infants against this disease during early infancy.

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Recombinant γY278H Fibrinogen Showed Normal Secretion from CHO Cells, but a Corresponding Heterozygous Patient Showed Hypofibrinogenemia

International Journal of Molecular Sciences ◽

10.3390/ijms22105218 ◽

2021 ◽

Vol 22 (10) ◽

pp. 5218

Author(s):

Tomu Kamijo ◽

Takahiro Kaido ◽

Masahiro Yoda ◽

Shinpei Arai ◽

Kazuyoshi Yamauchi ◽

...

Keyword(s):

Cell Lines ◽

Cho Cells ◽

Fibrinogen Level ◽

Culture Media ◽

Chinese Hamster ◽

Plasma Fibrinogen ◽

Enzyme Linked Immunosorbent Assay ◽

Cho Cell ◽

Accelerated Degradation ◽

Heterozygous Patient

We identified a novel heterozygous hypofibrinogenemia, γY278H (Hiroshima). To demonstrate the cause of reduced plasma fibrinogen levels (functional level: 1.12 g/L and antigenic level: 1.16 g/L), we established γY278H fibrinogen-producing Chinese hamster ovary (CHO) cells. An enzyme-linked immunosorbent assay demonstrated that synthesis of γY278H fibrinogen inside CHO cells and secretion into the culture media were not reduced. Then, we established an additional five variant fibrinogen-producing CHO cell lines (γL276P, γT277P, γT277R, γA279D, and γY280C) and conducted further investigations. We have already established 33 γ-module variant fibrinogen-producing CHO cell lines, including 6 cell lines in this study, but only the γY278H and γT277R cell lines showed disagreement, namely, recombinant fibrinogen production was not reduced but the patients’ plasma fibrinogen level was reduced. Finally, we performed fibrinogen degradation assays and demonstrated that the γY278H and γT277R fibrinogens were easily cleaved by plasmin whereas their polymerization in the presence of Ca2+ and “D:D” interaction was normal. In conclusion, our investigation suggested that patient γY278H showed hypofibrinogenemia because γY278H fibrinogen was secreted normally from the patient’s hepatocytes but then underwent accelerated degradation by plasmin in the circulation.

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Detection of antibodies to recombinant truncated flagellin and sonicated whole cell antigen of Burkholderia pseudomallei in acute melioidosis and in healthy Bangladeshi individuals

IMC Journal of Medical Science ◽

10.3329/imcjms.v14i1.47455 ◽

2020 ◽

Vol 14 (1) ◽

pp. 47-52

Author(s):

Md Shariful Alam Jilani ◽

Tang Thean Hock ◽

Sraboni Mazumder ◽

Fahmida Rahman ◽

Md Mohiuddin ◽

...

Keyword(s):

Rural Areas ◽

Burkholderia Pseudomallei ◽

Enzyme Linked Immunosorbent Assay ◽

Igg Antibodies ◽

Healthy Individuals ◽

Serum Samples ◽

Igg Antibody ◽

Whole Cell ◽

Cell Antigen ◽

The Mean

Background and objectives: Several types of Burkholderia pseudomallei antigens have been used to determine the antibody response in acute and asymptomatic cases. In the present study, we have detected immunoglobulin G (IgG) antibody to recombinant truncated flagellin antigen (RTFA) of B. pseudomallei in the sera of acute melioidosis cases and healthy individuals from melioidosis endemic areas of Bangladesh by indirect enzyme-linked immunosorbent assay (ELISA). In parallel, IgG antibody to sonicated whole cell antigen (SWCA) of B. pseudomallei was determined to compare with anti-RTFA antibody. Methodology: Serum samples from culture confirmed melioidosis cases and from healthy individuals aged 21 years and above residing in melioidosis endemic rural areas were included in the study. Serum IgG antibody to RTFA and SWCA of B. pseudomallei was determined by indirect ELISA. Results: Out of 8 culture confirmed acute melioidosis cases, 7 (87.5%) and 8 (100%) were positive for anti-B. pseudomallei IgG antibodies by RTFA and SWCA methods respectively. Among 361 healthy individuals, the rate of seropositivity by RTFA-ELISA was significantly less than that of SWCA-ELISA (16.1% versus 26.8%; p = 0.001). The mean optical density (OD) of RTFA-ELISA of positive cases was significantly less than that of SWCA-ELISA in both melioidosis and healthy individuals (0.79±0.11 versus 2.4±0.08, p = 0.0001; 0.67±0.01 versus 1.27±0.02, p = 0.0001). The sensitivity and specificity of RTFA-ELISA were 88.9% and 100% respectively. Conclusion: Findings of the study suggest that multiple or combination of antigens should be used to study the seroprevalence of B. pseudomallei infection in a community. Also, prospective study is necessary to find out the duration of persistence of antibodies to different antigenic components of B. pseudomallei after exposure. Ibrahim Med. Coll. J. 2020; 14(1): 47-52

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Serodiagnosis of Louse-Borne Relapsing Fever with Glycerophosphodiester Phosphodiesterase (GlpQ) fromBorrelia recurrentis

Journal of Clinical Microbiology ◽

10.1128/jcm.38.10.3561-3571.2000 ◽

2000 ◽

Vol 38 (10) ◽

pp. 3561-3571 ◽

Cited By ~ 54

Author(s):

Stephen F. Porcella ◽

Sandra J. Raffel ◽

Merry E. Schrumpf ◽

Martin E. Schriefer ◽

David T. Dennis ◽

...

Keyword(s):

Serological Test ◽

Geometric Mean ◽

Eastern Africa ◽

Enzyme Linked Immunosorbent Assay ◽

Relapsing Fever ◽

Serum Samples ◽

Serological Tests ◽

Whole Cell ◽

Convalescent Phase ◽

Glycerophosphodiester Phosphodiesterase

Human louse-borne relapsing fever occurs in sporadic outbreaks in central and eastern Africa that are characterized by significant morbidity and mortality. Isolates of the causative agent,Borrelia recurrentis, were obtained from the blood of four patients during a recent epidemic of the disease in southern Sudan. TheglpQ gene, encoding glycerophosphodiester phosphodiesterase, from these isolates was sequenced and compared with the glpQ sequences obtained from other relapsing-fever spirochetes. Previously we showed that GlpQ of Borrelia hermsii is an immunogenic protein with utility as a serological test antigen for discriminating tick-borne relapsing fever from Lyme disease. In the present work, we cloned and expressed theglpQ gene from B. recurrentis and used recombinant GlpQ in serological tests. Acute- and convalescent-phase serum samples obtained from 42 patients with louse-borne relapsing fever were tested with an indirect immunofluorescence assay (IFA) and an enzyme-linked immunosorbent assay (ELISA) that used whole cells ofB. recurrentis and with immunoblotting to whole-cell lysates of the spirochete and Escherichia coli producing recombinant GlpQ. The geometric mean titers of the acute- and convalescent-phase serum samples measured by IFA were 1:83 and 1:575, respectively. The immunoblot analysis identified a high level of reactivity and seroconversion to GlpQ, and the assay was more sensitive than the whole-cell IFA and ELISA using purified, recombinant histidine-tagged GlpQ. Serum antibodies to GlpQ and other antigens persisted for 27 years in one patient. We conclude that assessment of anti-GlpQ antibodies will allow serological confirmation of louse-borne relapsing fever and determination of disease prevalence.

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