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Reinhardt, Richard Lee

Overview:


My current research objectives are directed toward understanding immune responses in vivo. Much work has been invested in determining the molecular events that drive CD4+ T helper cell differentiation in vitro as well as the critical cellular players necessary for initiating an adaptive immune response within lymphoid tissues. However, it has been difficult to address true CD4+ T helper cell biology due to a lack of experimental systems designed to access CD4+ T cell function directly in vivo.

Thus, my specific interests are developing new methods to study CD4+ T cells during the effector and memory phase of the immune response in the context of different parasitic, bacterial, and viral models of infection. I am interested in comparing the phenotype and function of CD4+ T-helper cells residing in the secondary lymphoid tissues (such as spleen and lymph nodes) to those residing at sites of infection. In particular, my goal is to understand the relationship between cytokine-producing T follicular helper cells (Tfh) and conventional T-helper type 1 (Th1), T-helper type 2, and follicular regulatory T cells (Tfr). Particular attention will be focused on elucidating the role that these functionally distinct T-helper cells play in maintaining immunologic memory, and how dis-regulation of these subsets can impact B cell lymphoma development, asthma, and autoinflammatory disease. We are also committed to understanding how exposure to microbes in early-life can influence T-helper cell differentiation, regulatory function, and disease onset. Currently our models focus on understanding how helminth colonization impacts allergic disease onset and severity.

The broad, long term goals of this research program are to understand the cellular and molecular events driving the differentiation of naïve CD4+ T cells into different T-helper cell subsets, and to investigate the function, plasticity, and cell fate decisions of these cells during a primary and secondary immune response. Elucidating the signals and events that regulate T-helper cell differentiation (i.e. Th1, Th2,Tfh, Treg,Tfr) and function (i.e. cytokine production) will have broad implications in basic CD4+ T cell biology and provide important insights that will aid in the fight against infectious disease, cancer, allergic asthma, and autoinflammatory diseases. 

Positions:

Adjunct Assistant Professor in the Department of Immunology

Immunology
School of Medicine

Member of the Duke Cancer Institute

Duke Cancer Institute
School of Medicine

Education:

Ph.D. 2002

Ph.D. — University of Minnesota, Twin Cities

Grants:

Organization and Function of Cellular Structure

Administered By
Basic Science Departments
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
July 01, 1975
End Date
June 30, 2020

Modeling affinity maturation at molecular resolution

Administered By
Immunology
AwardedBy
Boston University
Role
Co Investigator
Start Date
April 15, 2015
End Date
March 31, 2017

The role of BATF in allergic inflammation and anti-helminth immunity

Administered By
Immunology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
July 01, 2015
End Date
June 30, 2016

Publications:

γδ T Cells and B Cells

© 2017 Elsevier Inc.γδ T cells constitute the third arm of a tripartite adaptive immune system in jawed vertebrates, besides αβ T cells and B cells. Like the other two lymphocyte-types, they express diverse antigen receptors, capable of specific ligand recognition. Functionally, γδ T cells represent a system of differentiated subsets, sometimes engaged in cross-regulation, which ultimately determines their effect on other components of the immune system, including B cells and antibodies. γδ T cells are capable of providing help to B cells in antibody production. More recently it became clear that γδ T cells influence B cell differentiation during the peripheral stages of B cell development, control levels of circulating immunoglobulin (all subclasses), and affect production of autoantibodies. Because of this relationship between γδ T cells and B cells, the extensive variation of γδ T cells among human individuals might be expected to modulate their humoral responsiveness.

Authors
Born, WK; Huang, Y; Reinhardt, RL; Huang, H; Sun, D; O'Brien, RL
MLA Citation
Born, WK, Huang, Y, Reinhardt, RL, Huang, H, Sun, D, and O'Brien, RL. "γδ T Cells and B Cells (Accepted)." Advances in Immunology (January 1, 2017).
Source
scopus
Published In
Advances in immunology
Publish Date
2017
DOI
10.1016/bs.ai.2017.01.002

BATF Modulates the Th2 Locus Control Region and Regulates CD4+ T Cell Fate during Antihelminth Immunity.

The AP-1 factor basic leucine zipper transcription factor, ATF-like (BATF) is important for CD4+ Th17, Th9, and follicular Th cell development. However, its precise role in Th2 differentiation and function remains unclear, and the requirement for BATF in nonallergic settings of type-2 immunity has not been explored. In this article, we show that, in response to parasitic helminths, Batf-/- mice are unable to generate follicular Th and Th2 cells. As a consequence, they fail to establish productive type-2 immunity during primary and secondary infection. Batf-/- CD4+ T cells do not achieve type-2 cytokine competency, which implies that BATF plays a key role in the regulation of IL-4 and IL-13. In contrast to Th17 and Th9 cell subsets in which BATF binds directly to promoter and enhancer regions to regulate cytokine expression, our results show that BATF is significantly enriched at Rad50 hypersensitivity site (RHS)6 and RHS7 of the locus control region relative to AP-1 sites surrounding type-2 cytokine loci in Th2 cells. Indeed, Batf-/- CD4+ T cells do not obtain permissive epigenetic modifications within the Th2 locus, which were linked to RHS6 and RHS7 function. In sum, these findings reveal BATF as a central modulator of peripheral and humoral hallmarks of type-2 immunity and begin to elucidate a novel mechanism by which it regulates type-2 cytokine production through its modification of the Th2 locus control region.

Authors
Bao, K; Carr, T; Wu, J; Barclay, W; Jin, J; Ciofani, M; Reinhardt, RL
MLA Citation
Bao, K, Carr, T, Wu, J, Barclay, W, Jin, J, Ciofani, M, and Reinhardt, RL. "BATF Modulates the Th2 Locus Control Region and Regulates CD4+ T Cell Fate during Antihelminth Immunity." Journal of immunology (Baltimore, Md. : 1950) 197.11 (December 2016): 4371-4381.
PMID
27798167
Source
epmc
Published In
Journal of immunology (Baltimore, Md. : 1950)
Volume
197
Issue
11
Publish Date
2016
Start Page
4371
End Page
4381

Cytokine expression by invariant natural killer T cells is tightly regulated throughout development and settings of type-2 inflammation.

Invariant natural killer T (iNKT) cells produce cytokines interleukin-4 (IL-4) and IL-13 during type-2 inflammatory responses. However, the nature in which iNKT cells acquire type-2 cytokine competency and the precise contribution of iNKT cell-derived IL-4 and IL-13 in vivo remains unclear. Using IL-13-reporter mice to fate-map cytokine-expressing cells in vivo, this study reveals that thymic iNKT cells express IL-13 early during development, and this IL-13-expressing intermediate gives rise to mature iNKT1, iNKT2, and iNKT17 subsets. IL-4 and IL-13 reporter mice also reveal that effector iNKT2 cells produce IL-4 but little IL-13 in settings of type-2 inflammation. The preferential production of IL-4 over IL-13 in iNKT2 cells results in part from their reduced GATA-3 expression. In summary, this work helps integrate current models of iNKT cell development, and further establishes non-coordinate production of IL-4 and IL-13 as the predominant pattern of type-2 cytokine expression among innate cells in vivo.

Authors
O'Brien, TF; Bao, K; Dell'Aringa, M; Ang, WXG; Abraham, S; Reinhardt, RL
MLA Citation
O'Brien, TF, Bao, K, Dell'Aringa, M, Ang, WXG, Abraham, S, and Reinhardt, RL. "Cytokine expression by invariant natural killer T cells is tightly regulated throughout development and settings of type-2 inflammation." Mucosal immunology 9.3 (May 2016): 597-609.
PMID
26349658
Source
epmc
Published In
Mucosal immunology
Volume
9
Issue
3
Publish Date
2016
Start Page
597
End Page
609
DOI
10.1038/mi.2015.78

The differential expression of IL-4 and IL-13 and its impact on type-2 immunity.

Allergic disease represents a significant global health burden, and disease incidence continues to rise in urban areas of the world. As such, a better understanding of the basic immune mechanisms underlying disease pathology are key to developing therapeutic interventions to both prevent disease onset as well as to ameliorate disease morbidity in those individuals already suffering from a disorder linked to type-2 inflammation. Two factors central to type-2 immunity are interleukin (IL)-4 and IL-13, which have been linked to virtually all major hallmarks associated with type-2 inflammation. Therefore, IL-4 and IL-13 and their regulatory pathways represent ideal targets to suppress disease. Despite sharing many common regulatory pathways and receptors, these cytokines perform very distinct functions during a type-2 immune response. This review summarizes the literature surrounding the function and expression of IL-4 and IL-13 in CD4+ T cells and innate immune cells. It highlights recent findings in vivo regarding the differential expression and non-canonical regulation of IL-4 and IL-13 in various immune cells, which likely play important and underappreciated roles in type-2 immunity.

Authors
Bao, K; Reinhardt, RL
MLA Citation
Bao, K, and Reinhardt, RL. "The differential expression of IL-4 and IL-13 and its impact on type-2 immunity." Cytokine 75.1 (September 2015): 25-37. (Review)
PMID
26073683
Source
epmc
Published In
Cytokine
Volume
75
Issue
1
Publish Date
2015
Start Page
25
End Page
37
DOI
10.1016/j.cyto.2015.05.008

A novel model for IFN-γ-mediated autoinflammatory syndromes.

Autoinflammatory disease and hyperinflammatory syndromes represent a growing number of diseases associated with inappropriately controlled inflammation in multiple organs. Systemic inflammation commonly results from dysregulated activation of innate immune cells, and therapeutic targeting of the IL-1β pathway has been used to ameliorate some of these diseases. Some hyperinflammatory syndromes, however, such as hemophagocytic lymphohistiocytosis and the newly classified proteasome disability syndromes, are refractory to such treatments, suggesting that other factors or environmental stressors may be contributing. In comparing two cytokine reporter mouse strains, we identify IFN-γ as a mediator of systemic autoinflammatory disease. Chronically elevated levels of IFN-γ resulted in progressive multiorgan inflammation and two copies of the mutant allele resulted in increased mortality accompanied by myeloproliferative disease. Disease was alleviated by genetic deletion of T-bet. These studies raise the possibility that therapeutics targeting the IFN-γ pathway might be effective in hyperinflammatory conditions refractory to IL-1β-targeted therapies.

Authors
Reinhardt, RL; Liang, H-E; Bao, K; Price, AE; Mohrs, M; Kelly, BL; Locksley, RM
MLA Citation
Reinhardt, RL, Liang, H-E, Bao, K, Price, AE, Mohrs, M, Kelly, BL, and Locksley, RM. "A novel model for IFN-γ-mediated autoinflammatory syndromes." Journal of immunology (Baltimore, Md. : 1950) 194.5 (March 2015): 2358-2368.
PMID
25637019
Source
epmc
Published In
Journal of immunology (Baltimore, Md. : 1950)
Volume
194
Issue
5
Publish Date
2015
Start Page
2358
End Page
2368
DOI
10.4049/jimmunol.1401992

Age-dependent hepatic lymphoid organization directs successful immunity to hepatitis B

Hepatitis B virus (HBV) is a major human pathogen that causes immune-mediated hepatitis. Successful immunity to HBV is age dependent: viral clearance occurs in most adults, whereas neonates and young children usually develop chronic infection. Using a mouse model of HBV infection, we sought mechanisms underpinning the age-dependent outcome of HBV and demonstrated that hepatic macrophages facilitate lymphoid organization and immune priming within the adult liver and promote successful immunity. In contrast, lymphoid organization and immune priming was greatly diminished in the livers of young mice, and of macrophage-depleted adult mice, leading to abrogated HBV immunity. Furthermore, we found that CXCL13, which is involved in B lymphocyte trafficking and lymphoid architecture and development, is expressed in an age-dependent manner in both adult mouse and human hepatic macrophages and plays an integral role in facilitating an effective immune response against HBV. Taken together, these results identify some of the immunological mechanisms necessary for effective control of HBV.

Authors
Publicover, J; Gaggar, A; Nishimura, S; Horn, CMV; Goodsell, A; Muench, MO; Reinhardt, RL; Rooijen, NV; Wakil, AE; Peters, M; Cyster, JG; Erle, DJ; Rosenthal, P; Cooper, S; Baron, JL
MLA Citation
Publicover, J, Gaggar, A, Nishimura, S, Horn, CMV, Goodsell, A, Muench, MO, Reinhardt, RL, Rooijen, NV, Wakil, AE, Peters, M, Cyster, JG, Erle, DJ, Rosenthal, P, Cooper, S, and Baron, JL. "Age-dependent hepatic lymphoid organization directs successful immunity to hepatitis B." Journal of Clinical Investigation 123.9 (2013): 3728-3739.
PMID
23925290
Source
scival
Published In
Journal of Clinical Investigation
Volume
123
Issue
9
Publish Date
2013
Start Page
3728
End Page
3739
DOI
10.1172/JCI68182

Marking and quantifying IL-17A-producing cells in vivo

Interleukin (IL)-17A plays an important role in host defense against a variety of pathogens and may also contribute to the pathogenesis of autoimmune diseases. However, precise identification and quantification of the cells that produce this cytokine in vivo have not been performed. We generated novel IL-17A reporter mice to investigate expression of IL-17A during Klebsiella pneumoniae infection and during experimental autoimmune encephalomyelitis, conditions previously demonstrated to potently induce IL-17A production. In both settings, the majority of IL-17A was produced by non-CD4+ T cells, particularly γδ T cells, but also invariant NKT cells and other CD4-CD3ε+ cells. As measured in dual-reporter mice, IFN-γ-producing Th1 cells greatly outnumbered IL-17A-producing Th17 cells throughout both challenges. Production of IL-17A by cells from unchallenged mice or by non-T cells under any condition was not evident. Administration of IL-1β and/or IL-23 elicited rapid production of IL-17A by γδ T cells, invariant NKT cells and other CD4-CD3ε+ cells in vivo, demonstrating that these cells are poised for rapid cytokine production and likely comprise the major sources of this cytokine during acute immunologic challenges. © 2012 Price et al.

Authors
Price, AE; Reinhardt, RL; Liang, H-E; Locksley, RM
MLA Citation
Price, AE, Reinhardt, RL, Liang, H-E, and Locksley, RM. "Marking and quantifying IL-17A-producing cells in vivo." PLoS ONE 7.6 (2012).
PMID
22768117
Source
scival
Published In
PloS one
Volume
7
Issue
6
Publish Date
2012
DOI
10.1371/journal.pone.0039750

Divergent expression patterns of IL-4 and IL-13 define unique functions in allergic immunity.

Interleukin 4 (IL-4) and IL-13 are critical for responses to parasitic helminthes. We used genetically engineered reporter mice to assess the temporal and spatial production of these cytokines in vivo. In lymph nodes, IL-4, but not IL-13, was made by follicular helper T cells (T(FH) cells). In contrast, tissue type 2 helper T cells (T(H)2 cells) produced both cytokines. There was also divergent production of IL-4 and IL-13 among cells of the innate immune system, whereby basophils produced IL-4, whereas innate helper type 2 cells (Ih2 cells) produced IL-13. IL-13 production by T(H)2 and Ih2 cells was dependent on the transcription factor GATA-3, which was present in large amounts in these cells, and in contrast to the small amount of GATA-3 in T(FH) cells and basophils. The distinct localization and cellular expression of IL-4 and IL-13 explains their unique roles during allergic immunity.

Authors
Liang, H-E; Reinhardt, RL; Bando, JK; Sullivan, BM; Ho, I-C; Locksley, RM
MLA Citation
Liang, H-E, Reinhardt, RL, Bando, JK, Sullivan, BM, Ho, I-C, and Locksley, RM. "Divergent expression patterns of IL-4 and IL-13 define unique functions in allergic immunity. (Published online)" Nat Immunol 13.1 (December 4, 2011): 58-66.
PMID
22138715
Source
pubmed
Published In
Nature Immunology
Volume
13
Issue
1
Publish Date
2011
Start Page
58
End Page
66
DOI
10.1038/ni.2182

Systemically dispersed innate IL-13-expressing cells in type 2 immunity.

Type 2 immunity is a stereotyped host response to allergens and parasitic helminths that is sustained in large part by the cytokines IL-4 and IL-13. Recent advances have called attention to the contributions by innate cells in initiating adaptive immunity, including a novel lineage-negative population of cells that secretes IL-13 and IL-5 in response to the epithelial cytokines IL-25 and IL-33. Here, we use IL-4 and IL-13 reporter mice to track lineage-negative innate cells that arise during type 2 immunity or in response to IL-25 and IL-33 in vivo. Unexpectedly, lineage-negative IL-25 (and IL-33) responsive cells are widely distributed in tissues of the mouse and are particularly prevalent in mesenteric lymph nodes, spleen, and liver. These cells expand robustly in response to exogenous IL-25 or IL-33 and after infection with the helminth Nippostrongylus brasiliensis, and they are the major innate IL-13-expressing cells under these conditions. Activation of these cells using IL-25 is sufficient for worm clearance, even in the absence of adaptive immunity. Widely dispersed innate type 2 helper cells, which we designate Ih2 cells, play an integral role in type 2 immune responses.

Authors
Price, AE; Liang, H-E; Sullivan, BM; Reinhardt, RL; Eisley, CJ; Erle, DJ; Locksley, RM
MLA Citation
Price, AE, Liang, H-E, Sullivan, BM, Reinhardt, RL, Eisley, CJ, Erle, DJ, and Locksley, RM. "Systemically dispersed innate IL-13-expressing cells in type 2 immunity." Proc Natl Acad Sci U S A 107.25 (June 22, 2010): 11489-11494.
PMID
20534524
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
107
Issue
25
Publish Date
2010
Start Page
11489
End Page
11494
DOI
10.1073/pnas.1003988107

Cytokine-secreting follicular T cells shape the antibody repertoire.

High-affinity antibodies are critical for host protection and underlie successful vaccines. The generation of such antibodies requires T cell-dependent help, which mediates germinal center reactions in which mutation and selection of B cells occurs. Using an interleukin 4-reporter system, we show here that CD4(+) follicular helper T cells constituted essentially all of the cytokine-secreting T cells in lymph nodes and were functionally distinct from T cells secreting the same cytokine in peripheral tissues. Follicular helper T cells with different cytokine profiles could be isolated as conjugates with B cells undergoing cytokine-specific immunoglobulin class switching with evidence of somatic hypermutation. Our findings support a model in which B cells compete for cytokines produced by follicular helper T cells that shape the affinity and isotype of the antibody response.

Authors
Reinhardt, RL; Liang, H-E; Locksley, RM
MLA Citation
Reinhardt, RL, Liang, H-E, and Locksley, RM. "Cytokine-secreting follicular T cells shape the antibody repertoire." Nat Immunol 10.4 (April 2009): 385-393.
PMID
19252490
Source
pubmed
Published In
Nature Immunology
Volume
10
Issue
4
Publish Date
2009
Start Page
385
End Page
393
DOI
10.1038/ni.1715

Erratum: Pluripotency of mesenchymal stem cells derived from adult marrow (Nature (2002) 418 (41-49) DOI: 10.1038/nature00870)

Authors
Jiang, Y; Jahagirdar, BN; Reinhardt, RL; Schwartz, RE; Keene, CD; Ortiz-Gonzalez, XR; Reyes, M; Lenvik, T; Lund, T; Blackstad, M; Du, J; Aldrich, S; Lisberg, A; Low, WC; Largaespada, DA; Verfaillie, CM
MLA Citation
Jiang, Y, Jahagirdar, BN, Reinhardt, RL, Schwartz, RE, Keene, CD, Ortiz-Gonzalez, XR, Reyes, M, Lenvik, T, Lund, T, Blackstad, M, Du, J, Aldrich, S, Lisberg, A, Low, WC, Largaespada, DA, and Verfaillie, CM. "Erratum: Pluripotency of mesenchymal stem cells derived from adult marrow (Nature (2002) 418 (41-49) DOI: 10.1038/nature00870)." Nature 447.7146 (2007): 879-880.
Source
scival
Published In
Nature
Volume
447
Issue
7146
Publish Date
2007
Start Page
879
End Page
880
DOI
10.1038/nature05812

Visualization of IL-12/23p40 in vivo reveals immunostimulatory dendritic cell migrants that promote Th1 differentiation.

IL-12p40 is induced in macrophages and dendritic cells (DC) after activation by microbial TLR ligands and cytokines and constitutes a component of IL-12 and IL-23. In an effort to understand the location and kinetics of these cytokines during the course of an immune response, we generated knockin (gene-targeted) mice that express the p40 gene linked via a viral internal ribosome entry site element with fluorescent reporters, eYFP or eGFP. Macrophages and DC from these mice faithfully reported biallelic p40 induction using the fluorescent marker. s.c. inoculation with Listeria monocytogenes or LPS led to a rapid, but transient, accumulation of p40-expressing DC in draining lymph nodes, which could be blocked by the addition of pertussis toxin. In situ analysis also revealed the accumulation of IL-12p40 protein around high endothelial venules located in close proximity to p40-expressing DC. Consistent with the in vivo findings, in vitro-activated DC that expressed p40 migrated to draining lymph nodes and promoted Th1 differentiation more efficiently than DC that did not express p40. Accordingly, these mice provide a valuable tool for tracking critical functions of DC in vivo and should bestow a useful reagent for exploring the effector biology of these cells in models of infectious disease, cancer immunity, and vaccine development.

Authors
Reinhardt, RL; Hong, S; Kang, S-J; Wang, Z-E; Locksley, RM
MLA Citation
Reinhardt, RL, Hong, S, Kang, S-J, Wang, Z-E, and Locksley, RM. "Visualization of IL-12/23p40 in vivo reveals immunostimulatory dendritic cell migrants that promote Th1 differentiation." J Immunol 177.3 (August 1, 2006): 1618-1627.
PMID
16849470
Source
pubmed
Published In
Journal of immunology (Baltimore, Md. : 1950)
Volume
177
Issue
3
Publish Date
2006
Start Page
1618
End Page
1627

Activation of the integrated stress response during T helper cell differentiation.

Adaptive immune responses require clonal expansion and differentiation of naive T cells into cytokine-secreting effector cells. After priming via signals through the T cell receptor, naive T helper cells express cytokine mRNA but do not secrete cytokine protein without additional T cell receptor stimulation. Here we show that primed T cells demonstrated phosphorylation of eukaryotic initiation factor 2-alpha (eIF2alpha), a 'collapsed' polysome profile, increased expression of stress-response genes and accumulation of cytoplasmic granules associated with RNA-binding proteins, all features of the integrated stress response. Restimulation of the cells resulted in rapid eIF2alpha dephosphorylation, ribosomal mRNA loading and cytokine secretion. Interference with the function of granule-associated proteins or accumulation of phosphorylated eIF2alpha enhanced release of interleukin 4 during T helper type 2 priming. Therefore, T lymphocytes require components of the integrated stress response to uncouple differentiation from the execution of effector functions.

Authors
Scheu, S; Stetson, DB; Reinhardt, RL; Leber, JH; Mohrs, M; Locksley, RM
MLA Citation
Scheu, S, Stetson, DB, Reinhardt, RL, Leber, JH, Mohrs, M, and Locksley, RM. "Activation of the integrated stress response during T helper cell differentiation." Nat Immunol 7.6 (June 2006): 644-651.
PMID
16680145
Source
pubmed
Published In
Nature Immunology
Volume
7
Issue
6
Publish Date
2006
Start Page
644
End Page
651
DOI
10.1038/ni1338

T helper cell effector fates--who, how and where?

CD4 helper T cells functionally organize the host immune response by elaborating cytokines, often in patterns that have overlapping effects on other cells. Much interest centers on understanding how these stereotyped cytokine patterns become elaborated and what mechanisms underlie the generation of distinct helper T cell subsets. The past two years have seen advances in understanding of additional subsets, including T helper follicular cells and IL-17-producing T helper cells. Progress has also been achieved in resolving some of the crosstalk that regulates effector fate at the level of distinct transcription factors and chromatin reorganization of the cytokine genes, and a crucial role for gene silencing has been exposed. Finally, the role of innate cells in influencing these processes has become increasingly realized.

Authors
Reinhardt, RL; Kang, S-J; Liang, H-E; Locksley, RM
MLA Citation
Reinhardt, RL, Kang, S-J, Liang, H-E, and Locksley, RM. "T helper cell effector fates--who, how and where?." Curr Opin Immunol 18.3 (June 2006): 271-277. (Review)
PMID
16617008
Source
pubmed
Published In
Current Opinion in Immunology
Volume
18
Issue
3
Publish Date
2006
Start Page
271
End Page
277
DOI
10.1016/j.coi.2006.03.003

Primary induction of CD4 T cell responses in nasal associated lymphoid tissue during group A streptococcal infection.

CD4 T cells are important for development of long-term immunity to bacterial infections. Here we describe construction of a group A streptococcus (GAS) strain that expresses the model ovalbumin epitope (OVA) on its surface, and the use of this strain in adoptive transfer experiments to study CD4 T cell response to bacterial infection in nasal-associated lymphoid tissue (NALT), which was previously shown to be a specific target for GAS colonization. The OVA(+) GAS, but not the wild-type strain was shown to activate CD4 T cells in an antigen-specific manner both in vitro and in vivo. After intranasal infection of mice with this strain, OVA-specific CD4 T cells were first activated in NALT, which is functionally equivalent to human tonsils, rather than in the cervical lymph nodes. During localized infection, OVA(+) GAS induced rapid and prolonged activation of CD4 T cells at higher magnitudes in the NALT than in draining lymph nodes and spleen, where CD4 T cells underwent little or no activation. In contrast, systemic infection induced significantly higher activation of CD4 T cells in both lymph nodes and spleens, compared to when the infection was localized in NALT. Further investigation of cellular immune responses in NALT during GAS infection using adoptive T cell transfer, combined with the model antigen on the pathogen may ultimately shed light on mechanisms for failure of children to develop protective immune responses following streptococcal tonsillitis.

Authors
Park, H-S; Costalonga, M; Reinhardt, RL; Dombek, PE; Jenkins, MK; Cleary, PP
MLA Citation
Park, H-S, Costalonga, M, Reinhardt, RL, Dombek, PE, Jenkins, MK, and Cleary, PP. "Primary induction of CD4 T cell responses in nasal associated lymphoid tissue during group A streptococcal infection." Eur J Immunol 34.10 (October 2004): 2843-2853.
PMID
15368301
Source
pubmed
Published In
European Journal of Immunology
Volume
34
Issue
10
Publish Date
2004
Start Page
2843
End Page
2853
DOI
10.1002/eji.200425242

Th2 cells: orchestrating barrier immunity.

Authors
Stetson, DB; Voehringer, D; Grogan, JL; Xu, M; Reinhardt, RL; Scheu, S; Kelly, BL; Locksley, RM
MLA Citation
Stetson, DB, Voehringer, D, Grogan, JL, Xu, M, Reinhardt, RL, Scheu, S, Kelly, BL, and Locksley, RM. "Th2 cells: orchestrating barrier immunity." Adv Immunol 83 (2004): 163-189. (Review)
PMID
15135631
Source
pubmed
Published In
Advances in immunology
Volume
83
Publish Date
2004
Start Page
163
End Page
189
DOI
10.1016/S0065-2776(04)83005-0

Constitutive cytokine mRNAs mark natural killer (NK) and NK T cells poised for rapid effector function.

Natural killer (NK) and NK T cells are tissue lymphocytes that secrete cytokines rapidly upon stimulation. Here, we show that these cells maintain distinct patterns of constitutive cytokine mRNAs. Unlike conventional T cells, NK T cells activate interleukin (IL)-4 and interferon (IFN)-gamma transcription during thymic development and populate the periphery with both cytokine loci previously modified by histone acetylation. Similarly, NK cells transcribe and modify the IFN-gamma gene, but not IL-4, during developmental maturation in the bone marrow. Lineage-specific patterns of cytokine transcripts predate infection and suggest evolutionary selection for invariant but distinct types of effector responses among the earliest responding lymphocytes.

Authors
Stetson, DB; Mohrs, M; Reinhardt, RL; Baron, JL; Wang, Z-E; Gapin, L; Kronenberg, M; Locksley, RM
MLA Citation
Stetson, DB, Mohrs, M, Reinhardt, RL, Baron, JL, Wang, Z-E, Gapin, L, Kronenberg, M, and Locksley, RM. "Constitutive cytokine mRNAs mark natural killer (NK) and NK T cells poised for rapid effector function." J Exp Med 198.7 (October 6, 2003): 1069-1076.
PMID
14530376
Source
pubmed
Published In
The Journal of Experimental Medicine
Volume
198
Issue
7
Publish Date
2003
Start Page
1069
End Page
1076
DOI
10.1084/jem.20030630

Whole-body analysis of T cell responses.

In vivo methods to detect antigen-specific T cell responses at the single-cell level have greatly increased our knowledge of how the immune system works. However, most of these approaches have been confined to analysis of lymphoid tissues. Recently, the development of imaging techniques capable of simultaneously monitoring all the tissues of the body has led to the realization that antigen-experienced T cells reside in non-lymphoid tissues and may play a vital role in protecting the host against pathogens. Therefore, single-cell imaging at the level of the whole organism is needed to fully understand the dynamics of protective immunity.

Authors
Reinhardt, RL; Jenkins, MK
MLA Citation
Reinhardt, RL, and Jenkins, MK. "Whole-body analysis of T cell responses." Curr Opin Immunol 15.4 (August 2003): 366-371. (Review)
PMID
12900265
Source
pubmed
Published In
Current Opinion in Immunology
Volume
15
Issue
4
Publish Date
2003
Start Page
366
End Page
371

Distinct dendritic cell populations sequentially present antigen to CD4 T cells and stimulate different aspects of cell-mediated immunity.

Peptide:MHC II complexes derived from a fluorescent antigen were detected in vivo to identify the cells that present subcutaneously injected antigen to CD4 T cells. Skin-derived dendritic cells (DCs) that acquired the antigen while in the draining lymph nodes were the first cells to display peptide:MHC II complexes. Presentation by these cells induced CD69, IL-2 production, and maximal proliferation by the T cells. Later, DCs displaying peptide:MHC II complexes migrated from the injection site via a G protein-dependent mechanism. Presentation by these migrants sustained expression of the IL-2 receptor and promoted delayed type hypersensitivity. Therefore, presentation of peptide:MHC II complexes derived from a subcutaneous antigen occurs in two temporally distinct waves with different functional consequences.

Authors
Itano, AA; McSorley, SJ; Reinhardt, RL; Ehst, BD; Ingulli, E; Rudensky, AY; Jenkins, MK
MLA Citation
Itano, AA, McSorley, SJ, Reinhardt, RL, Ehst, BD, Ingulli, E, Rudensky, AY, and Jenkins, MK. "Distinct dendritic cell populations sequentially present antigen to CD4 T cells and stimulate different aspects of cell-mediated immunity." Immunity 19.1 (July 2003): 47-57.
PMID
12871638
Source
pubmed
Published In
Immunity
Volume
19
Issue
1
Publish Date
2003
Start Page
47
End Page
57

Preferential accumulation of antigen-specific effector CD4 T cells at an antigen injection site involves CD62E-dependent migration but not local proliferation.

The migration of antigen-specific T cells to nonlymphoid tissues is thought to be important for the elimination of foreign antigens from the body. However, recent results showing the migration of activated T cells into many nonlymphoid tissues raised the possibility that antigen-specific T cells do not migrate preferentially to nonlymphoid tissues containing antigen. We addressed this question by tracking antigen-specific CD4 T cells in the whole body after a localized subcutaneous antigen injection. Antigen-specific CD4 T cells proliferated in the skin-draining lymph nodes and the cells that underwent the most cell divisions acquired the ability to bind to CD62P. As time passed, CD62P-binding antigen-specific CD4 T cells with interferon gamma production potential accumulated preferentially at the site of antigen injection but only in recipients that expressed CD62E. Surprisingly, these T cells did not proliferate in the injection site despite showing evidence of more cell divisions than the T cells in the draining lymph nodes. The results suggest that the most divided effector CD4 T cells from the lymph nodes enter the site of antigen deposition via recognition of CD62E on blood vessels and are retained there in a nonproliferative state via recognition of peptide-major histocompatibility complex II molecules.

Authors
Reinhardt, RL; Bullard, DC; Weaver, CT; Jenkins, MK
MLA Citation
Reinhardt, RL, Bullard, DC, Weaver, CT, and Jenkins, MK. "Preferential accumulation of antigen-specific effector CD4 T cells at an antigen injection site involves CD62E-dependent migration but not local proliferation." J Exp Med 197.6 (March 17, 2003): 751-762.
PMID
12629067
Source
pubmed
Published In
The Journal of Experimental Medicine
Volume
197
Issue
6
Publish Date
2003
Start Page
751
End Page
762
DOI
10.1084/jem.20021690

Pluripotency of mesenchymal stem cells derived from adult marrow.

We report here that cells co-purifying with mesenchymal stem cells--termed here multipotent adult progenitor cells or MAPCs--differentiate, at the single cell level, not only into mesenchymal cells, but also cells with visceral mesoderm, neuroectoderm and endoderm characteristics in vitro. When injected into an early blastocyst, single MAPCs contribute to most, if not all, somatic cell types. On transplantation into a non-irradiated host, MAPCs engraft and differentiate to the haematopoietic lineage, in addition to the epithelium of liver, lung and gut. Engraftment in the haematopoietic system as well as the gastrointestinal tract is increased when MAPCs are transplanted in a minimally irradiated host. As MAPCs proliferate extensively without obvious senescence or loss of differentiation potential, they may be an ideal cell source for therapy of inherited or degenerative diseases.

Authors
Jiang, Y; Jahagirdar, BN; Reinhardt, RL; Schwartz, RE; Keene, CD; Ortiz-Gonzalez, XR; Reyes, M; Lenvik, T; Lund, T; Blackstad, M; Du, J; Aldrich, S; Lisberg, A; Low, WC; Largaespada, DA; Verfaillie, CM
MLA Citation
Jiang, Y, Jahagirdar, BN, Reinhardt, RL, Schwartz, RE, Keene, CD, Ortiz-Gonzalez, XR, Reyes, M, Lenvik, T, Lund, T, Blackstad, M, Du, J, Aldrich, S, Lisberg, A, Low, WC, Largaespada, DA, and Verfaillie, CM. "Pluripotency of mesenchymal stem cells derived from adult marrow." Nature 418.6893 (July 4, 2002): 41-49.
PMID
12077603
Source
pubmed
Published In
Nature
Volume
418
Issue
6893
Publish Date
2002
Start Page
41
End Page
49
DOI
10.1038/nature00870

Tracking salmonella-specific CD4 T cells in vivo reveals a local mucosal response to a disseminated infection.

A novel adoptive transfer system was used to track the fate of naive Salmonella-specific CD4 T cells in vivo. These cells showed signs of activation in the Peyer's patches as early as 3 hr after oral infection. The activated CD4 T cells then produced IL-2 and proliferated in the T cell areas of these tissues before migrating into the B cell-rich follicles. In contrast, Salmonella-specific CD4 T cells were not activated in the spleen and very few of these cells migrated to the liver, despite the presence of bacteria in both organs. These results show that the T cell response to pathogenic Salmonella infection is localized to the gut-associated lymphoid tissue and does not extend efficiently to the major sites of late infection.

Authors
McSorley, SJ; Asch, S; Costalonga, M; Reinhardt, RL; Jenkins, MK
MLA Citation
McSorley, SJ, Asch, S, Costalonga, M, Reinhardt, RL, and Jenkins, MK. "Tracking salmonella-specific CD4 T cells in vivo reveals a local mucosal response to a disseminated infection." Immunity 16.3 (March 2002): 365-377.
PMID
11911822
Source
pubmed
Published In
Immunity
Volume
16
Issue
3
Publish Date
2002
Start Page
365
End Page
377

Thermo-oscillatory convection

Average thermal convection of an incompressible fluid performing high-frequency oscillations in a straight channel at rest is considered. It is shown that the Stokes layers play an important role in the excitation of average flows, in addition to the classical thermovibrational mechanism. Skin-layer flow generation is identified as a mechanism additional to the thermovibrational mechanism whose contribution is determined by the relative oscillation amplitude: it can have both a destabilizing and a stabilizing effect, depending on the amplitude.

Authors
Jiang, Y; Jahagirdar, BN; Reinhardt, RL; Schwartz, RE; Keene, CD; Ortiz-Gonzalez, XR; Reyes, M; Lenvik, T; Lund, T; Blackstad, M; Du, J; Aldrich, S; Lisberg, A; Low, WC; Lergaespada, DA; Verfaillie, CM
MLA Citation
Jiang, Y, Jahagirdar, BN, Reinhardt, RL, Schwartz, RE, Keene, CD, Ortiz-Gonzalez, XR, Reyes, M, Lenvik, T, Lund, T, Blackstad, M, Du, J, Aldrich, S, Lisberg, A, Low, WC, Lergaespada, DA, and Verfaillie, CM. "Thermo-oscillatory convection." Fluid Dynamics 37.4 (January 1, 2002): 536-544.
Source
scopus
Published In
Fluid Dynamics / Izvestiya Akademii Nauk - Mekhanika Zhidkosti i Gaza
Volume
37
Issue
4
Publish Date
2002
Start Page
536
End Page
544
DOI
10.1023/A:1020633101346

Cutting edge: in vivo identification of TCR redistribution and polarized IL-2 production by naive CD4 T cells.

TCR aggregation at the point of contact with an APC is thought to play an important role in T cell signal transduction. However, this potentially important phenomenon has never been documented during an immune response in vivo. Here we used immunohistology to show that the TCR on naive Ag-specific CD4 T cells in the lymph nodes of mice injected with Ag redistributed to one side of the cell. In cases where the APC could be identified, the TCR was concentrated on the side of the T cell facing the APC. In those T cells that produced IL-2, the TCR and IL-2 localized to the same side of the cell. In vivo IL-2 production depended on costimulatory signaling through CD28, whereas TCR redistribution did not. These results show that Ag-stimulated CD4 T cells produce IL-2 in a polarized fashion and undergo CD28-independent TCR redistribution in vivo.

Authors
Reichert, P; Reinhardt, RL; Ingulli, E; Jenkins, MK
MLA Citation
Reichert, P, Reinhardt, RL, Ingulli, E, and Jenkins, MK. "Cutting edge: in vivo identification of TCR redistribution and polarized IL-2 production by naive CD4 T cells." J Immunol 166.7 (April 1, 2001): 4278-4281.
PMID
11254679
Source
pubmed
Published In
Journal of immunology (Baltimore, Md. : 1950)
Volume
166
Issue
7
Publish Date
2001
Start Page
4278
End Page
4281

Visualizing the generation of memory CD4 T cells in the whole body.

It is thought that immunity depends on naive CD4 T cells that proliferate in response to microbial antigens, differentiate into memory cells that produce anti-microbial lymphokines, and migrate to sites of infection. Here we use immunohistology to enumerate individual naive CD4 T cells, specific for a model antigen, in the whole bodies of adult mice. The cells resided exclusively in secondary lymphoid tissues, such as the spleen and lymph nodes, in mice that were not exposed to antigen. After injection of antigen alone into the blood, the T cells proliferated, migrated to the lungs, liver, gut and salivary glands, and then disappeared from these organs. If antigen was injected with the microbial product lipopolysaccharide, proliferation and migration were enhanced, and two populations of memory cells survived for months: one in the lymph nodes that produced the growth factor interleukin-2, and a larger one in the non-lymphoid tissues that produced the anti-microbial lymphokine interferon-gamma. These results show that antigen recognition in the context of infection generates memory cells that are specialized to proliferate in the secondary lymphoid tissues or to fight infection at the site of microbial entry.

Authors
Reinhardt, RL; Khoruts, A; Merica, R; Zell, T; Jenkins, MK
MLA Citation
Reinhardt, RL, Khoruts, A, Merica, R, Zell, T, and Jenkins, MK. "Visualizing the generation of memory CD4 T cells in the whole body." Nature 410.6824 (March 1, 2001): 101-105.
PMID
11242050
Source
pubmed
Published In
Nature
Volume
410
Issue
6824
Publish Date
2001
Start Page
101
End Page
105
DOI
10.1038/35065111

Visualization of immunotoxin-mediated tumor cell death in vivo.

We present a novel methodology to visualize tumor cells directly in a whole mouse. This technique combines immunohistochemistry with whole mouse sectioning. It lets one see the exact distribution of tumor cells throughout an animal and how effectively these cells are eliminated by cancer therapeutics. We used this technique to assess the efficacy of a T cell-specific immunotoxin in a severe combined immunodeficient mouse model of human T-cell leukemia. Severe combined immunodeficient mice were injected with one of two human T-cell acute lymphoblastic leukemia cell lines (Molt 3 and Molt 13) and were either left untreated or were treated with DA7, an immunotoxin specific for the T cell-associated antigen CD7. Mice were sacrificed after tumor cell injection and immunotoxin therapy, whole mouse cross-sections were prepared, and tumor cells in the sections were visualized by immunohistochemistry. No tumor cells were detected in DA7-treated mice injected with Molt 3, consistent with the long-term survival of this group and the sensitivity of Molt 3 to DA7 in vitro. In contrast, DA7 treatment did not visibly eliminate tumor cells in mice challenged with Molt 13, nor did it result in their long-term survival. Furthermore, tumor cells were detected in areas that may have otherwise been overlooked, and their distribution differed from that of mice injected with Molt 13 alone. These analyses indicate that whole mouse sectioning will be a valuable tool for assessing residual disease in the preclinical evaluation of cancer therapeutics.

Authors
Erickson, HA; Reinhardt, RL; Hermanson, JB; Panoskaltsis-Mortari, A; Pennell, CA
MLA Citation
Erickson, HA, Reinhardt, RL, Hermanson, JB, Panoskaltsis-Mortari, A, and Pennell, CA. "Visualization of immunotoxin-mediated tumor cell death in vivo." Clin Cancer Res 7.3 Suppl (March 2001): 890s-894s.
PMID
11300488
Source
pubmed
Published In
Clinical cancer research : an official journal of the American Association for Cancer Research
Volume
7
Issue
3 Suppl
Publish Date
2001
Start Page
890s
End Page
894s

In vivo activation of antigen-specific CD4 T cells.

Physical detection of antigen-specific CD4 T cells has revealed features of the in vivo immune response that were not appreciated from in vitro studies. In vivo, antigen is initially presented to naïve CD4 T cells exclusively by dendritic cells within the T cell areas of secondary lymphoid tissues. Anatomic constraints make it likely that these dendritic cells acquire the antigen at the site where it enters the body. Inflammation enhances in vivo T cell activation by stimulating dendritic cells to migrate to the T cell areas and display stable peptide-MHC complexes and costimulatory ligands. Once stimulated by a dendritic cell, antigen-specific CD4 T cells produce IL-2 but proliferate in an IL-2--independent fashion. Inflammatory signals induce chemokine receptors on activated T cells that direct their migration into the B cell areas to interact with antigen-specific B cells. Most of the activated T cells then die within the lymphoid tissues. However, in the presence of inflammation, a population of memory T cells survives. This population is composed of two functional classes. One recirculates through nonlymphoid tissues and is capable of immediate effector lymphokine production. The other recirculates through lymph nodes and quickly acquires the capacity to produce effector lymphokines if stimulated. Therefore, antigenic stimulation in the presence of inflammation produces an increased number of specific T cells capable of producing effector lymphokines throughout the body.

Authors
Jenkins, MK; Khoruts, A; Ingulli, E; Mueller, DL; McSorley, SJ; Reinhardt, RL; Itano, A; Pape, KA
MLA Citation
Jenkins, MK, Khoruts, A, Ingulli, E, Mueller, DL, McSorley, SJ, Reinhardt, RL, Itano, A, and Pape, KA. "In vivo activation of antigen-specific CD4 T cells." Annu Rev Immunol 19 (2001): 23-45. (Review)
PMID
11244029
Source
pubmed
Published In
Annual Review of Immunology
Volume
19
Publish Date
2001
Start Page
23
End Page
45
DOI
10.1146/annurev.immunol.19.1.23

Visualization of immunotoxin-mediated tumor cell death in vivo

We present a novel methodology to visualize tumor cells directly in a whole mouse. This technique combines immunohistochemistry with whole mouse sectioning. It lets one see the exact distribution of tumor cells throughout an animal and how effectively these cells are eliminated by cancer therapeutics. We used this technique to assess the efficacy of a T cell-specific immunotoxin in a severe combined immunodeficient mouse model of human T-cell leukemia. Severe combined immunodeficient mice were injected with one of two human T-cell acute lymphoblastic leukemia cell lines (Molt 3 and Molt 13) and were either left untreated or were treated with DA7, an immunotoxin specific for the T cell-associated antigen CD7. Mice were sacrificed after tumor cell injection and immunotoxin therapy, whole mouse cross-sections were prepared, and tumor cells in the sections were visualized by immunohistochemistry. No tumor cells were detected in DA7-treated mice injected with Molt 3, consistent with the long-term survival of this group and the sensitivity of Molt 3 to DA7 in vitro. In contrast, DA7 treatment did not visibly eliminate tumor cells in mice challenged with Molt 13, nor did it result in their long-term survival. Furthermore, tumor cells were detected in areas that may have otherwise been overlooked, and their distribution differed from that of mice injected with Molt 13 alone. These analyses indicate that whole mouse sectioning will be a valuable tool for assessing residual disease in the preclinical evaluation of cancer therapeutics.

Authors
Erickson, HA; Reinhardt, RL; Hermanson, JB; Panoskaltsis-Mortari, A; Pennell, CA
MLA Citation
Erickson, HA, Reinhardt, RL, Hermanson, JB, Panoskaltsis-Mortari, A, and Pennell, CA. "Visualization of immunotoxin-mediated tumor cell death in vivo." Clinical Cancer Research 7.11 SUPPL. (2001): 890s-894s.
Source
scival
Published In
Clinical Cancer Research
Volume
7
Issue
11 SUPPL.
Publish Date
2001
Start Page
890s
End Page
894s

Antigen-experienced CD4 T cells display a reduced capacity for clonal expansion in vivo that is imposed by factors present in the immune host.

It is thought that protective immunity is mediated in part by Ag-experienced T cells that respond more quickly and vigorously than naive T cells. Using adoptive transfer of OVA-specific CD4 T cells from TCR transgenic mice as a model system, we show that Ag-experienced CD4 T cells accumulate in lymph nodes more rapidly than naive T cells after in vivo challenge with Ag. However, the magnitude of clonal expansion by Ag-experienced T cells was much less than that of naive T cells, particularly at early times after primary immunization. Ag-experienced CD4 T cells quickly reverted to the slower but more robust clonal expansion behavior of naive T cells after transfer into a naive environment. Conversely, the capacity for rapid clonal expansion was acquired by naive CD4 T cells after transfer into passively immunized recipients. These results indicate that rapid in vivo response by Ag-experienced T cells is facilitated by Ag-specific Abs, whereas the limited capacity for clonal expansion is imposed by some other factor in the immune environment, perhaps residual Ag.

Authors
Merica, R; Khoruts, A; Pape, KA; Reinhardt, RL; Jenkins, MK
MLA Citation
Merica, R, Khoruts, A, Pape, KA, Reinhardt, RL, and Jenkins, MK. "Antigen-experienced CD4 T cells display a reduced capacity for clonal expansion in vivo that is imposed by factors present in the immune host." J Immunol 164.9 (May 1, 2000): 4551-4557.
PMID
10779756
Source
pubmed
Published In
Journal of immunology (Baltimore, Md. : 1950)
Volume
164
Issue
9
Publish Date
2000
Start Page
4551
End Page
4557

The effects of ultraviolet radiation on the moderate halophile Halomonas elongata and the extreme halophile Halobacterium salinarum.

Both the moderately halophilic bacterium, Halomonas elongata, and the extremely halophilic archaea, Halobacterium salinarum, can be found in hypersaline environments (e.g., salterns). On complex media, H. elongata grows over a salt range of 0.05-5.2 M, whereas, H. salinarum multiplies over a salt range of 2.5-5.2 M. The purpose of this study was to illustrate the effect that solar (UV-A and UV-B) and germicidal radiation (UV-C) had on the growth patterns of these bacteria at varied salt concentrations. Halomonas elongata grown on a complex medium at 0.05, 1.37, and 4.3 M NaCl was found to be more sensitive to UV-A and UV-B radiation, as the salt concentration of the medium increased. Halobacterium salinarum grown on a complex medium at 3.0 and 4.3 M NaCl did not show a significant drop in viability after 39.3 kJ.m-2 of UV-A and UV-B exposure. When exposed to UV-C, H. elongata exhibited substantially more sensitivity than H. salinarum. In H. elongata, differential sensitivity to UV-C was observed. At 0.05 M NaCl, H. elongata was less sensitive to UV-C than at 1.37 and 4.3 M NaCl. Both bacteria showed some photoreactivation when incubated under visible light following both UV-A, UV-B, and UV-C exposure. Mutagenesis following UV-C exposure was demonstrated by both organisms.

Authors
Martin, EL; Reinhardt, RL; Baum, LL; Becker, MR; Shaffer, JJ; Kokjohn, TA
MLA Citation
Martin, EL, Reinhardt, RL, Baum, LL, Becker, MR, Shaffer, JJ, and Kokjohn, TA. "The effects of ultraviolet radiation on the moderate halophile Halomonas elongata and the extreme halophile Halobacterium salinarum." Can J Microbiol 46.2 (February 2000): 180-187.
PMID
10721487
Source
pubmed
Published In
Canadian Journal of Microbiology
Volume
46
Issue
2
Publish Date
2000
Start Page
180
End Page
187
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