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Large scale immune profiling of infected humans and goats reveals differential recognition of Brucella melitensis antigens.

Liang L, Leng D, Burk C, Nakajima-Sasaki R, Kayala MA, Atluri VL, Pablo J, Unal B, Ficht TA, Gotuzzo E, Saito M, Morrow WJ, Liang X, Baldi P, Gilman RH, Vinetz JM, Tsolis RM, Felgner PL - PLoS Negl Trop Dis (2010)

Bottom Line: To better understand the universe of antibody responses that develop after B. melitensis infection, a protein microarray was fabricated containing 1,406 predicted B. melitensis proteins.Based on these results, a nitrocellulose line blot containing the human serodiagnostic antigens was fabricated and applied in a simple assay that validated the accuracy of the protein microarray results in the diagnosis of humans.These data demonstrate that an experimentally infected natural reservoir host produces a fundamentally different immune response than a naturally infected accidental human host.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, Department of Medicine, University of California Irvine, Irvine, California, USA.

ABSTRACT
Brucellosis is a widespread zoonotic disease that is also a potential agent of bioterrorism. Current serological assays to diagnose human brucellosis in clinical settings are based on detection of agglutinating anti-LPS antibodies. To better understand the universe of antibody responses that develop after B. melitensis infection, a protein microarray was fabricated containing 1,406 predicted B. melitensis proteins. The array was probed with sera from experimentally infected goats and naturally infected humans from an endemic region in Peru. The assay identified 18 antigens differentially recognized by infected and non-infected goats, and 13 serodiagnostic antigens that differentiate human patients proven to have acute brucellosis from syndromically similar patients. There were 31 cross-reactive antigens in healthy goats and 20 cross-reactive antigens in healthy humans. Only two of the serodiagnostic antigens and eight of the cross-reactive antigens overlap between humans and goats. Based on these results, a nitrocellulose line blot containing the human serodiagnostic antigens was fabricated and applied in a simple assay that validated the accuracy of the protein microarray results in the diagnosis of humans. These data demonstrate that an experimentally infected natural reservoir host produces a fundamentally different immune response than a naturally infected accidental human host.

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Related in: MedlinePlus

Probing a collection of B. melitensis human sera and discovery of human serodiagnostic antigens.Arrays were probed with human sera organized into 5 groups: Culture Positive, Culture Negative/Rose Bengal Positive, Rose Bengal Negative, USA Naïve, and Peruvian Naïve, as described in the text. (A). Heatmap showing normalized intensity with red strongest, bright green weakest and black in between. The antigens are in rows and are grouped to serodiagnostic and cross-reactive antigens. The human samples are in columns and sorted left to right by increasing average intensity to serodiagnostic antigens. (B) The mean sera reactivity of the 1406 antigens was compared between the Culture Positive and Peruvian Naive groups. Antigens with Benjamini Hochberg corrected p-value less than 0.05 are organized to the left and cross-reactive antigens to the right. The 13 most reactive serodiagnostic and 31 of the most reactive cross-reactive antigens are shown. C−/RB+, Culture Positive and Rose Bengal negative; RB−, Rose Bengal negative. Numbers in () are case numbers from each group.
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pntd-0000673-g003: Probing a collection of B. melitensis human sera and discovery of human serodiagnostic antigens.Arrays were probed with human sera organized into 5 groups: Culture Positive, Culture Negative/Rose Bengal Positive, Rose Bengal Negative, USA Naïve, and Peruvian Naïve, as described in the text. (A). Heatmap showing normalized intensity with red strongest, bright green weakest and black in between. The antigens are in rows and are grouped to serodiagnostic and cross-reactive antigens. The human samples are in columns and sorted left to right by increasing average intensity to serodiagnostic antigens. (B) The mean sera reactivity of the 1406 antigens was compared between the Culture Positive and Peruvian Naive groups. Antigens with Benjamini Hochberg corrected p-value less than 0.05 are organized to the left and cross-reactive antigens to the right. The 13 most reactive serodiagnostic and 31 of the most reactive cross-reactive antigens are shown. C−/RB+, Culture Positive and Rose Bengal negative; RB−, Rose Bengal negative. Numbers in () are case numbers from each group.

Mentions: Bm protein arrays were also probed with sera from acute brucellosis patients in Lima, Peru obtained within 1–3 weeks of the onset of symptoms. All patients in this study, as is true of virtually all patients from Lima [3], [59]–[62], were infected with B.. melitensis biovar 1. Sera from Bm culture-positive humans (Fig. 1b) showed pronounced reactivity against several antigens compared to unexposed individuals. A set of 33 antigens was identified to be serodominant among 1406 antigens tested (Fig. 3a, 3b). Of these, 13 antigens were serodiagnostic, and reacted differentially between naïve and culture positive patients from Peru (p-value<.05). The same antigens also reacted robustly with individuals diagnosed Rose Bengal positive but negative by blood culture for B. melitensis. For some of these subjects, treatment with antibiotics may have resulted in a negative blood culture for B. melitensis. The elevated antibody response from a few individuals in the Peruvian naïve group might be indicative of past exposure to similar proteins in environmental bacteria, or to a past subclinical Brucella infection. We also identified 20 cross-reactive antigens that reacted similarly among all human samples, whether from naïve individuals or individuals diagnosed to be infected and use of these antigens in serodiagnostic tests can therefore be selectively avoided.


Large scale immune profiling of infected humans and goats reveals differential recognition of Brucella melitensis antigens.

Liang L, Leng D, Burk C, Nakajima-Sasaki R, Kayala MA, Atluri VL, Pablo J, Unal B, Ficht TA, Gotuzzo E, Saito M, Morrow WJ, Liang X, Baldi P, Gilman RH, Vinetz JM, Tsolis RM, Felgner PL - PLoS Negl Trop Dis (2010)

Probing a collection of B. melitensis human sera and discovery of human serodiagnostic antigens.Arrays were probed with human sera organized into 5 groups: Culture Positive, Culture Negative/Rose Bengal Positive, Rose Bengal Negative, USA Naïve, and Peruvian Naïve, as described in the text. (A). Heatmap showing normalized intensity with red strongest, bright green weakest and black in between. The antigens are in rows and are grouped to serodiagnostic and cross-reactive antigens. The human samples are in columns and sorted left to right by increasing average intensity to serodiagnostic antigens. (B) The mean sera reactivity of the 1406 antigens was compared between the Culture Positive and Peruvian Naive groups. Antigens with Benjamini Hochberg corrected p-value less than 0.05 are organized to the left and cross-reactive antigens to the right. The 13 most reactive serodiagnostic and 31 of the most reactive cross-reactive antigens are shown. C−/RB+, Culture Positive and Rose Bengal negative; RB−, Rose Bengal negative. Numbers in () are case numbers from each group.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2864264&req=5

pntd-0000673-g003: Probing a collection of B. melitensis human sera and discovery of human serodiagnostic antigens.Arrays were probed with human sera organized into 5 groups: Culture Positive, Culture Negative/Rose Bengal Positive, Rose Bengal Negative, USA Naïve, and Peruvian Naïve, as described in the text. (A). Heatmap showing normalized intensity with red strongest, bright green weakest and black in between. The antigens are in rows and are grouped to serodiagnostic and cross-reactive antigens. The human samples are in columns and sorted left to right by increasing average intensity to serodiagnostic antigens. (B) The mean sera reactivity of the 1406 antigens was compared between the Culture Positive and Peruvian Naive groups. Antigens with Benjamini Hochberg corrected p-value less than 0.05 are organized to the left and cross-reactive antigens to the right. The 13 most reactive serodiagnostic and 31 of the most reactive cross-reactive antigens are shown. C−/RB+, Culture Positive and Rose Bengal negative; RB−, Rose Bengal negative. Numbers in () are case numbers from each group.
Mentions: Bm protein arrays were also probed with sera from acute brucellosis patients in Lima, Peru obtained within 1–3 weeks of the onset of symptoms. All patients in this study, as is true of virtually all patients from Lima [3], [59]–[62], were infected with B.. melitensis biovar 1. Sera from Bm culture-positive humans (Fig. 1b) showed pronounced reactivity against several antigens compared to unexposed individuals. A set of 33 antigens was identified to be serodominant among 1406 antigens tested (Fig. 3a, 3b). Of these, 13 antigens were serodiagnostic, and reacted differentially between naïve and culture positive patients from Peru (p-value<.05). The same antigens also reacted robustly with individuals diagnosed Rose Bengal positive but negative by blood culture for B. melitensis. For some of these subjects, treatment with antibiotics may have resulted in a negative blood culture for B. melitensis. The elevated antibody response from a few individuals in the Peruvian naïve group might be indicative of past exposure to similar proteins in environmental bacteria, or to a past subclinical Brucella infection. We also identified 20 cross-reactive antigens that reacted similarly among all human samples, whether from naïve individuals or individuals diagnosed to be infected and use of these antigens in serodiagnostic tests can therefore be selectively avoided.

Bottom Line: To better understand the universe of antibody responses that develop after B. melitensis infection, a protein microarray was fabricated containing 1,406 predicted B. melitensis proteins.Based on these results, a nitrocellulose line blot containing the human serodiagnostic antigens was fabricated and applied in a simple assay that validated the accuracy of the protein microarray results in the diagnosis of humans.These data demonstrate that an experimentally infected natural reservoir host produces a fundamentally different immune response than a naturally infected accidental human host.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, Department of Medicine, University of California Irvine, Irvine, California, USA.

ABSTRACT
Brucellosis is a widespread zoonotic disease that is also a potential agent of bioterrorism. Current serological assays to diagnose human brucellosis in clinical settings are based on detection of agglutinating anti-LPS antibodies. To better understand the universe of antibody responses that develop after B. melitensis infection, a protein microarray was fabricated containing 1,406 predicted B. melitensis proteins. The array was probed with sera from experimentally infected goats and naturally infected humans from an endemic region in Peru. The assay identified 18 antigens differentially recognized by infected and non-infected goats, and 13 serodiagnostic antigens that differentiate human patients proven to have acute brucellosis from syndromically similar patients. There were 31 cross-reactive antigens in healthy goats and 20 cross-reactive antigens in healthy humans. Only two of the serodiagnostic antigens and eight of the cross-reactive antigens overlap between humans and goats. Based on these results, a nitrocellulose line blot containing the human serodiagnostic antigens was fabricated and applied in a simple assay that validated the accuracy of the protein microarray results in the diagnosis of humans. These data demonstrate that an experimentally infected natural reservoir host produces a fundamentally different immune response than a naturally infected accidental human host.

Show MeSH
Related in: MedlinePlus