Progress 10/01/12 to 09/30/17
Outputs
Target Audience:The target audience for our work are physicians, veterinarians and public health officials interested in how food borne diseases cause long term chronic disease problems in humans. We mainly report to other scientists in these fields by publishing scientific manuscripts and by giving talks at scientific meetings. We also occasionally report directly to the public to explain risks to their health from food borne bacteria and antibiotic resistance. A main target audience are medical and veterinary medical students who Dr. Mansfield teaches. Here the main messages are targeting how bacteria cause food borne disease and how to recognize this disease in humans and animals. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?In a continuing process, several students were mentored in 2016-2017 for study of enteric diseases of food animals. Jean Brudvig, DVM (PhD candidate) will defend her PhD thesis in December 2017 and began a job at Michigan State University asa clincial pathologist/diagnostician. Phillip Brooks successfully dfended his doctoral defense in May 2016 with a focus on C. jejuni genomics and bioinformatics and the role of antibiotics in exacerbating the disease due to this bacterium. He has taken a postdoctoral position at the School of Veterinary Medicine, University of California at Davis and studying bioinformatics methods for study of bacterial organisms in ground water. Phillip isfrom minority groups (African American) underrepresented in the sciences. This shows the success of the Mansfield lab in mentoring students to stay in fields related to food safety. How have the results been disseminated to communities of interest?In the past year, we have disseminated our scientific results to communities of interest by publishing scientific papers, giving scientific talks at national and international meetings and presenting posters at scientific and international meetings. Three publications have been submitted or published in scientific journals, three talks have been given, and twelve abstracts were accepted and given as talks or posters at national or international meetings. Dr. Mansfield also gave instructional programs to medical students, veterinary students and practicing veterinarians designed to enhance their knowledge of microbiology and foodborne illness. Dr. Mansfield also gave instructional programs to veterinary residents or assistant professors designed to improve their ability to conduct scientific research and progress in their careers. What do you plan to do during the next reporting period to accomplish the goals?In the next project our long term goal is to develop effective inhibitors to decrease viability and fitness of C. jejuni in humans and its natural hosts to prevent disease and transmission. Our overarching hypothesis is that particular C. jejuni strains, microbial communities, and host innate and adaptive immune responses as well as suites of antibiotic resistant elements in the pathogen and microbial community interact to determine disease outcomes. Our short term goal is to examine these interactions by applying a systems-based approach to discover mechanisms of pathogenesis in AR C. jejuni in animal models of tolerance, enteric disease, and acute disease that elicits autoimmunity. Bioinformatics and core support will serve as a major discovery platform for the entire project. Mathematical models will be explored to identify key genes, transcriptional networks, and metabolic mediators of the pathogen, host and microbial communities that define these interactions, which may serve as novel drug targets. Our approaches will uncover new knowledge regarding AR C. jejuni and will serve as a guide for other investigators studying AR pathogens and their hosts. Based on extensive preliminary data for each of our long term goals, the team will carry out the following Specific Aims: • (Specific Aim 1). Determine the synergistic role of host response, microbiome and antibiotic treatment in enhancing colonization, enteritis and risk of Guillain Barré syndrome after infection with C. jejuni. We will study animal models to determine the interaction pathways between the host response, microbiome and antibiotic treatment in enhancing colonization, enteritis and risk of Guillain Barré syndrome after infection with C. jejuni. We will develop mathematical models to explain these interactions and make predictions, which form the foundation for later discovery efforts in Aims 2 and 3. • (Specific Aim 2). Identify molecular pathways important for the emergence of resistance and evolution of virulence in vivo to guide the discovery of novel therapeutics. We will define conditions and pathways important for the emergence of antibiotic resistance in vivo and develop a catalog of mutations associated with resistance and enhanced fitness. Networks identified in Aim 2 will be validated using deep-sequencing based genome interrogation approaches to identify novel targets for therapeutic discovery. This will set the stage for identifying compounds to test as novel therapeutics that have the ability to block AR infections. • (Specific Aim 3):Determine the impact of antibiotic resistant C. jejuni infections on the intestinal microbiome in patients during acute infections and post recovery. We will use metagenomics to compare microbial communities among patients with AR and susceptible C. jejuni infections. We will also examine differences in the abundance of resistance genes, microbes, and viruses, and identify targets that have a negative impact on patient outcomes or disease severity. This work forms the foundation for iterative experimental testing in the animal models and for the retrospective screening of humanintestinal metagenomes for therapeutic, resistance and fitness targets identified in Aims 1 and 2. Outcomes. We expect to identify molecular pathways in different pathogen-host-microbiome combinations that are important for disease caused by AR C. jejuni. We also expect to identify antimicrobial mechanisms that drive pathways involved in the emergence of AR C. jejuni. Information generated will inform antimicrobial practices to prevent severe disease and suggest microbiome amendments for testing as preventatives or treatments. These efforts also facilitate discovery of novel therapeutic compounds that can be applied to prevent and treat acute campylobacteriosis and the chronic autoimmune sequelae linked to it.
Impacts
What was accomplished under these goals?
Accomplishments 1. Focus on emerging diseases: we will identify, characterize and develop improved detection and prevention methods related to newly recognized, novel or emerging causes of zoonotic enteric disease and enteric pathogens of food animals. 2. Focus on preventions and interventions: we will develop and improve preventative measures and interventions to reduce the incidence and prevalence of infections of food animals with enteric and foodborne and waterborne pathogens. 3. Focus on disseminating knowledge: we will provide training or continuing education to disseminate new information to students, producers, veterinarians, diagnostic labs and others to implement interventions and preventative measures. Major goals of the project What was accomplished under these goals? Impact. The impact of the sciuentific results described below is that we have proven that the microbial communities that live in your gastrointestinal tract contribute to susceptibility to several foodborne diseases. We have also shown that antibiotic treatment can further enhance this sausceptibility due to the specific microbiome one harbors. Clearly, preventions and treatments for foodborne bacterial pathogens must be tested and shown to work in subjects (people or animals) with a wide variety of microbiome types. Antibiotic treatment chocies must also take into account what detrimental effects these treatments may have on the gut microbiome. 1) Focus on emerging diseases: we will identify, characterize and develop improved detection and prevention methods related to newly recognized, novel or emerging causes of zoonotic enteric disease and enteric pathogens of food animals. Transplanted Human Fecal Microbiota Enhanced Guillain Barré Syndrome Autoantibody Responses after Campylobacter jejuni Infection in C57BL/6 mice Background. Campylobacter jejuni is the leading antecedent infection to the autoimmune neuropathy Guillain-Barré Syndrome (GBS), which is accompanied by an autoimmune antiganglioside antibody attack on peripheral nerves. Previously we showed that contrasting immune responses mediate C. jejuni induced colitis and autoimmunity in interleukin-10 (IL-10) deficient mice, dependent upon the infecting strain. Strains from colitis patients elicited T helper 1 (TH1)-dependent inflammatory responses while strains from GBS patients elicited TH2-dependent autoantibody production. Both syndromes were exacerbated by antibiotic depletion of the microbiota, but other factors controlling susceptibility to GBS are unknown. Methods. Using 16S rRNA gene high-throughput sequencing, we examined whether structure of the gut microbial community alters host 1) gastrointestinal inflammation or 2) anti-ganglioside antibody responses after infection with C. jejuni strains from colitis or GBS patients. We compared these responses in C57BL/6 mice with either 1) stable human gut microbiota (Humicrobiota) transplants or 2) conventional mouse microbiota (Convmicrobiota). Results. Inoculating germ free C57BL/6 wild type (WT) mice with a mixed human fecal slurry provided a murine model that stably passed its microbiota over > 20 generations. Mice were housed in specific pathogen free (SPF) facilities, while extra precautions of having caretakers wear sterile garb along with limited access ensured that no mouse pathogens were acquired. Humicrobiota conferred many changes upon the WT model in contrast to previous results, which showed only colonization with no disease after C. jejuni challenge. When compared to Convmicrobiota mice for susceptibility to C. jejuni enteric or GBS patient strains, infected Humicrobiota mice had 1) 10-100 fold increases in C. jejuni colonization of both strains, 2) pathologic change in draining lymph nodes but not colon or cecal lamina propria, 3) significantly lower Th1/Th17-dependent anti-C. jejuni responses, 4) significantly higher IL-4 responses at 5 but not 7 weeks post infection (PI), 5) significantly higher Th2-dependent anti-C. jejuni responses, and 6) significantly elevated antiganglioside autoantibodies after C. jejuni infection. These responses in Humicrobiota mice were correlated with a dominant Bacteroidetes and Firmicutes microbiota. Conclusions. These data demonstrate that Humicrobiota altered host-pathogen interactions in infected mice, increasing colonization and Th-2 and autoimmune responses in a C. jejuni strain-dependent manner. Thus, microbiota composition is another factor controlling susceptibility to GBS. 2) Focus on preventions and interventions: we will develop and improve preventative measures and interventions to reduce the incidence and prevalence of infections of food animals with enteric and foodborne and waterborne pathogens. Antibiotic depletion drives severe Campylobacter jejuni-mediated Type 1/17 colitis with three Guillain-Barré Syndrome Associated strains Background. Campylobacter jejuni is a leading cause of bacterial diarrheal illness worldwide. Disease is most often selflimiting; however, in approximately 7 in 10,000 infected individuals the autoimmune neurological diseases Guillain-Barré Syndrome (GBS) or its variant Miller Fisher Syndrome arise. When GBS is preceded by C. jejuni infection, lipooligosaccharides resembling host gangliosides activate the immune system to produce neurotoxic anti-ganglioside antibodies. Our aim was to determine the role of the gut microbiota in inflammatory and autoimmune disease following infection with three C. jejuni strains in a murine model. We hypothesized that alteration of the microbiota with a broadspectrum antibiotic, Cefoperazone (CPZ), would increase C. jejuni colonization and the severity of gastroenteritis. Methods. C57BL/6 IL-10-/- mice were administered antibiotic-free water or 0.5 mg/mL CPZ in their drinking water for 7 days prior to inoculation with the vehicle, C. jejuni 260.94, C. jejuni D8942, or C. jejuni HB93-13. Results. Infection with antibiotic resistant Campylobacter jejuni strains from Guillain Barré Syndrome (GBS) patients produced both severe Type 1/17 colitis responses when C57BL/6 IL-10-/- mice were treated with a broad spectrum antibiotic that decreased complexity and abundance of the gut microbiota. Antibiotic depletion of microbiota was the main factor in inducing enhanced enteric disease and GBS associated phenotypes although the degree of severity was also dependent on the C. jejuni strain. Notably, antibiotic treatment drove severe colitis by C. jejuni strains isolated from GBS patients that had heretofore produced no or only mild colitis in this model. Antibiotic treated infected mice had high numbers of C. jejuni in the apical, basolateral and paracellular junctions of gut epithelium and within cells of the lamina propria, submucosa and lymph nodes indicating increased invasion and translocation from the gut. Conclusion. These results indicate that antibiotic depletion of gut microbiota alters immune responses to C. jejuni in a manner that exacerbates colitis with C. jejuni strains previously shown to promote predominantly Type 2 responses.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2017
Citation:
Brooks PT, Brakel KA, Bell JA, Bejcek CE, Gilpin T, Brudvig JM, Mansfield LS. 2017. Transplanted Human Fecal
Microbiota Enhanced Guillain Barr� Syndrome Autoantibody Responses after Campylobacter jejuni Infection in C57BL/6 mice. Microbiome (2017) 5:92, DOI 10.1186/s40168-017-0284-4.
- Type:
Journal Articles
Status:
Accepted
Year Published:
2017
Citation:
Brooks PT, Bell JA, Bejcek CE, Malik A, and Mansfield LS. 2017. ?Antibiotic Depletion Drives Severe Campylobacter
jejuni-Mediated Type 1/17 Colitis and Type 2 Autoimmunity. PLOS One, accepted.
|
Progress 10/01/15 to 09/30/16
Outputs
Target Audience:The target audience for our work are physicians, veterinarians and public health officials interested in how food borne diseases cause long term chronic disease problems in humans. We mainly report to other scientists in these fields by publishing scientific manuscripts and by giving talks at scientific meetings. We also occasionally report directly to the public to explain risks to their health from food borne bacteria and antibiotic resistance. A main target audience are medical and veterinary medical students who Dr. Mansfield teaches. Here the main messages are targeting how bacteria cause food borne disease and how to recognize this disease in humans and animals. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? In a continuing process, several students were mentored in 2015-2016 for study of enteric diseases of food animals. Barbie Gadsden, DVM (PhD candidate) successfully defended her PhD thesis in December 2015 and began a job at MPI pharmaceuticals in Matawan, Michigan as an Immunologist/ Pathologist. Phillip Brooks passed his doctoral defense in May 2016 with a focus on C. jejuni genomics and bioinformatics and the role of antibiotics in exacerbating the disease due to this bacterium. He has taken a postdoctoral position at the School of Veterinary Medicine, University of California at Davis and will study bioinformatics methods for study of bacterial organisms in ground water. Both Phillip and Barbie are from minority groups (African American) underrepresented in the sciences. This shows the success of the Mansfield lab in mentoring students to stay in the field of food safety. Three other students are pursuing PhD degrees in food safety related to these enteric pathogens including Jean Brudvig, DVM, Daniel Claiborne and Hinako Terauchi. Jean Brudvig passed her preliminary exam in 2015. Five undergraduate students worked in the lab on these projects including Christopher Bejcek, Alexander Ethridge, James Chen, Zachary Jansen and Keenan Odea. Dr. Linda Mansfield continued to act as the Principal Investigator and the Administrative Core Leader of the Michigan State University, Enterics Research Investigational Network, Cooperative Research Center (MSU ERIN CRC) in 2015-2016. This is a multidisciplinary, highly integrated research center to study the enteric microbiome in health and disease with the long-term goal of understanding and alleviating one of the most prevalent and important global health problems, diarrheal illness. The central overarching theme of the center is to explore and elucidate the relationship of the enteric microbiome to acute diarrheal illness. Dr. Mansfield gave talks at the Conference of Research Workers on Animal Diseases, the Twenty-fifth Anniversary of the David Hyde Allergy and Asthma Research Centre in the United Kingdom, the Bach Endowed lectureship at Western University, Kalamazoo, MI and at the American College of Veterinary Internal Medicine Forum in Denver Colorado among other talks. Dr. Mansfield along with Dr. Susan Ewart organized a grantsmanship workshop for assistant professors and residents in Large Animal Clinical Sciences at the College of Veterinary Medicine at MSU where preharvest food safety was prominently featured. Dr. Mansfield also attended the NC1202 annual meeting (December 3-4) and the Conference of Research Workers on Animal Diseases (December 4-6) both in Chicago, Illinois in 2016. Two of the undergraduates presented their work at the Undergraduate Research and Arts Forum (UURAF) at Michigan State University. Each of the three graduate students presented their work at the Phi Zeta Research Day, American Society for Microbiology and the Society for Neurology meetings in 2016. How have the results been disseminated to communities of interest? In the past year, we have disseminated our scientific results to communities of interest by publishing scientific papers, giving scientific talks at national and international meetings and presenting posters at scientific and international meetings. Three publications have been submitted or published in scientific journals, four talks have been given, and ten abstracts were accepted and given as talks or posters at national or international meetings. Dr. Mansfield also gave instructional programs to medical students, veterinary students and practicing veterinarians designed to enhance their knowledge of microbiology and foodborne illness. Dr. Mansfield also gave instructional programs to veterinary residents or assistant professors designed to improve their ability to conduct scientific research and progress in their careers. What do you plan to do during the next reporting period to accomplish the goals? Our future goals are to define C. jejuni gene expression controlling the different pathotypes that we have defined in the reported murine models. We will continue this study on the genetic diversity of human and animal isolates of C. jejuni that we have acquired through the US Centers for Diseases Control and Prevention and from a collaboration with Dr. Shannon Manning from an epidemiologic study in Michigan. Methods will include Illumina sequencing, screening in mice with human microbiota given by fecal transplant for Campylobacter gene expression, and selected gene knockouts followed by challenge in the murine models. We are especially interested in evolutionary mechanisms of C. jejuni and how it initiates autoimmune disease in its host.
Impacts
What was accomplished under these goals?
Guillain Barré Syndrome is induced in Non-Obese Diabetic (NOD) mice following Campylobacter jejuni infection and is exacerbated by antibiotics Background: Campylobacter jejuni is a leading cause of bacterial gastroenteritis linked to several serious autoimmune sequelae such as the peripheral neuropathies Guillain Barré syndrome (GBS) and Miller Fisher syndrome (MFS). We hypothesized that GBS and MFS can result in NOD wild type (WT) mice or their congenic interleukin (IL)-10 or B7-2 knockouts secondary to C. jejuni infection. Methods: Mice were gavaged orally with C. jejuni strains HB93-13 and 260.94 from patients with GBS or CF93-6 from a patient with MFS and assessed for clinical neurological signs and phenotypes, anti-ganglioside antibodies, and cellular infiltrates and lesions in gut and peripheral nerve tissues. Results: Significant increases in autoantibodies against single gangliosides (GM1, GQ1b, GD1a) occurred in infected NOD mice of all genotypes, although the isotypes varied (NOD WT had IgG1, IgG3; NOD B7-2-/- had IgG3; NOD IL-10-/- had IgG1, IgG3, IgG2a). Infected NOD WT and NOD IL-10-/- mice also produced anti-ganglioside antibodies of the IgG1 isotype directed against a mixture of GM1/GQ1b gangliosides. Phenotypic tests showed significant differences between treatment groups of all mouse genotypes. Peripheral nerve lesions with macrophage infiltrates were significantly increased in infected mice of NOD WT and IL-10-/- genotypes compared to sham-inoculated controls, while lesions with T cell infiltrates were significantly increased in infected mice of the NOD B7-2-/- genotype compared to sham-inoculated controls. In both infected and sham inoculated NOD IL-10-/- mice, antibiotic treatment exacerbated neurological signs, lesions and the amount and number of different isotypes of antiganglioside autoantibodies produced. Conclusions: Thus, inducible mouse models of post-C. jejuni GBS are feasible and can be characterized based on evaluation of three factors--onset of GBS clinical signs/phenotypes, anti-ganglioside autoantibodies and nerve lesions. Based on these factors we characterized 1) NOD B-7-/- mice as an acute inflammatory demyelinating polyneuropathy (AIDP)-like model, 2) NOD IL-10-/- mice as an acute motor axonal neuropathy (AMAN)-like model best employed over a limited time frame, and 3) NOD WT mice as an AMAN model with mild clinical signs and lesions. Taken together these data demonstrate that C. jejuni strain genotype, host genotype and antibiotic treatment affect GBS disease outcomes in mice and that many disease phenotypes are possible. Autoantibody-dependent mechanisms alone are not sufficient to mediate Guillain-Barré syndrome lesions in a mouse model Background: Guillain-Barré syndrome (GBS) is an auto-immune disorder of the peripheral nervous system that results in ascending flaccid paralysis. 7% of patients die and 20% have long term disability, yet no effective therapeutics exist. Antecedent infections with several pathogens, most prominently Campylobacter jejuni, have been linked to development of autoreactive antibodies thought to mediate GBS through cross-reaction with peripheral nerve gangliosides. Mouse models that develop GBS nerve lesions subsequent toC. jejuniinfection are needed to elucidate the pathogenesis and to establish targets for new treatments. We hypothesized that BALB/c interleukin (IL)-10deficient mice orally inoculated with a C. jejuni GBS patient strain will have higher numbers of macrophage infiltrates in peripheral nerves than infected wildtype BALB/c mice; a second hypothesis was that BALB/c IL-10-/-mice infected with a C. jejuni GBS patient strain will have higher nerve infiltration than C57BL/6 IL-10-/- mice infected with the same strain or the enteritis-associated strain C. jejuni 11168. Methods: For the first experiment, BALB/c wild-type (WT) and IL-10-/-mice were inoculated with a GBS-associatedC. jejunistrain 260.94 or sham inoculated (vehicle)(4 groups, 10 mice/group) and sacrificed after five weeks. For the second, BALB/c IL-10-/- and C57 IL-10-/- mice were inoculated with C. jejuni 11168, 260.94, or a vehicle (6 groups, 10 mice/group) and sacrificed after four weeks. Sciatic nerves and lumbar dorsal root ganglia were dissected from all 100 mice in these two experiments, embedded en bloc, sectioned and labeled immunohistochemically with an F4/80 macrophage marker. Results: Macrophage infiltration in DRG were scored quantitatively using morphometry. No significant differences were observed between any treatment groups in either experiment. Conclusions: Because infected mice had robust anti-C. jejuni specific antibodies and some groups in both experiments had elevated antiganglioside autoantibodies of several isotypes, the lack of nerve lesions suggests that autoantibody dependent mechanisms alone are not sufficient to mediate GBS lesions over this 4-5 week time course. Funded by NIAID, NIH, Department of Health and Human Services under ERIN CRC grant U19AI090872
Publications
- Type:
Journal Articles
Status:
Accepted
Year Published:
2016
Citation:
St. Charles JL, Bell JA, Gadsden BJ, Malik A, Cook H, Van de Grift LK, Kim HY, Smith EJ, Mansfield LS. 2015. Guillain Barr� Syndrome is induced in Non-Obese Diabetic (NOD) mice following Campylobacter jejuni infection and is exacerbated by antibiotics. Journal of Autoimmunity, accepted 09/04/2016, in press.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2016
Citation:
Brooks, P.T., Brakel K.A., Bell J.A., Malik A., Gilpin T., Mansfield L.S. 2016. Transplanted Human Fecal Microbiota Alters Adaptive Immune Responses to Campylobacter jejuni Infection in C57BL/6 mice, Microbiome, In Review.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2016
Citation:
Brooks, P.T., Bell J.A., Bejcek C.E., Malik A., Gadsden, B.J., Mansfield L.S. Antimicrobial resistant Campylobacter strains from Guillain-Barr� syndrome patients produced severe Type 1/17 colitis and Type 2 autoimmune responses in an antibiotic-depleted murine model. Journal of Experimental Medicine, In Review.
|
Progress 10/01/14 to 09/30/15
Outputs
Target Audience:The target audience for our work are physicians, veterinarians and public health officials interested in how food borne diseases cause long term chronic disease problems in humans. We mainly report to other scientists in these fields by publishing scientific manuscripts and by giving talks at scientific meetings. We also occasionally report directly to the public to explain risks to their health from food borne bacteria and antibiotic resistance. A main target audience are medical and veterinary medical students who Dr. Mansfield teaches. Here the main messages are targeting how bacteria cause food borne disease and how to recognize this disease in humans and animals. Changes/Problems:No scientific problems were encountered. What opportunities for training and professional development has the project provided?Three students are pursuing PhD degrees in food safety related to these enteric pathogens including Barbie Gadsden, DVM, Jean Brudvig, DVM, and Phillip Brooks. Phillip Brooks, an African American, passed his preliminary examination for the PhD with a focus on C. jejuni genomics and bioinformatics and the role of antibiotics in exacerbating the disease die to this bacterium. Jean Brudvig also passed her preliminary exam. Barbie Gadsden also an African American student will defend her thesis on December 8, 2015. Five undergraduate students worked in the lab on these projects including Christopher Bejcek Alexander Ethridge, James Chen, Zachary Jansen and Keenan Odea. How have the results been disseminated to communities of interest?The results were presented as talks and are being written up for publication as scientific journal articles. What do you plan to do during the next reporting period to accomplish the goals?Our future goals are to define C. jejuni gene expression controlling the different pathotypes that we have defined in the reported murine models. We will continue this study on the genetic diversity of human and animal isolates of C. jejuni that we have acquired through the US Centers for Diseases Control and Prevention and from a collaboration with Dr. Shannon Manning from an epidemiologic study in Michigan. Methods will include Illumina sequencing, screening in mice with human microbiota given by fecal transplant for Campylobacter gene expression, and selected gene knockouts followed by challenge in the murine models. We are especially interested in evolutionary mechanisms of C. jejuni and how it initiates autoimmune disease in its host.
Impacts
What was accomplished under these goals?
1. Focus on emerging diseases: we will identify, characterize and develop improved detection and prevention methods related to newly recognized, novel or emerging causes of zoonotic enteric disease and enteric pathogens of food animals. A. Campylobacter jejuni Specific Aim (1). Characterize definitively the neurological signs and disease lesions associated with Guillain Barré and Miller Fisher Syndromes in murine models using single blind methods Non-Obese Diabetic (NOD) mice serve as inducible models of Guillain Barré Syndrome following Campylobacter jejuni infection Background. Campylobacter jejuni is a leading cause of bacterial gastroenteritis linked to several serious autoimmune sequelae such as the peripheral neuropathies Guillain Barré syndrome (GBS) and Miller Fisher syndrome (MFS). These neuropathies are thought to be triggered by antibodies formed in response to sialylated lipooligosaccharides (LOS) produced by some C. jejuni strains that cross-react with gangliosides that are enriched on peripheral nerves. We hypothesized that GBS and MFS result in NOD mice (wild type or interleukin-10 (IL-10) or B7-2 deficient) secondary to infection with C. jejuni with sialylated LOS that elicits anti-ganglioside autoantibodies mediating peripheral neuropathy and nerve lesions. Methods. Mice were gavaged orally with C. jejuni strains HB93-13 and 260.94 from GBS patients or CF93-6 from an MFS patient and assessed for clinical neurological signs/ phenotypes, anti-ganglioside antibodies and cellular infiltrates and lesions in gut and peripheral nervous tissue. Trypticase soya broth sham inoculated mice of all genotypes served as controls. Results. Significant increases in autoantibodies against single gangliosides (GM1, GQ1b, GD1a) occurred in infected NOD mice of all genotypes, although the antibody isotypes varied. NOD WT mice inoculated with MFS-associated strain CF93-6 developed significantly increased antibodies reactive with a mixture of gangliosides GM1 and GQ1b. Phenotypic tests showed significant differences between treatment groups of all mouse genotypes. Antibiotic treatment of NOD IL-10-/- mice altered the array of antibody isotypes reactive with nerve gangliosides, suggesting a possible role for the gut microbiota in disease expression. Peripheral nerve lesions with macrophage infiltrates were significantly different between sham-inoculated and infected mice of NOD and NOD IL-10-/- genotypes and lesions with T cell infiltrates between sham-inoculated and infected mice of the NOD B7-2-/- genotype. Significance. Thus, mouse models of GBS are feasible and can be evaluated based on three factors--onset of GBS clinical signs/phenotypes, anti-ganglioside autoantibodies and nerve lesions. We ranked NOD mice for suitability as inducible GBS models as 1) NOD B-7-/- mice (most suitable), 2) NOD IL-10-/- (suitable over a limited time frame), and 3) NOD (suitable with mild clinical signs and lesions). Taken together, these data demonstrate that differences in C. jejuni strain genotypes and host genotypes affect the GBS/MFS disease outcome in mice and that many disease phenotypes are possible.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2015
Citation:
Kopper JA, Patterson JS and Mansfield LS. 2015. Metronidazolebut not IL-10 or prednisolonerescues Trichuris muris infected C57BL/6 IL-10 deficient mice from severe disease. Veterinary Parasitology, 2015 Aug 6. pii: S0304-4017(15)00380-5. doi: 10.1016/j.vetpar.2015.07.038.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Flies AS, Maksimoski MT, Mansfield LS, Weldele ML and Holekamp KE. 2014. Characterization of toll-like receptors 1-10 in spotted hyenas. Veterinary Research Communications, Veterinary Research Communications Jun;38(2):165-70. doi: 10.1007/s11259-014-9592-3. Epub 2014 Feb 2.
- Type:
Journal Articles
Status:
Accepted
Year Published:
2015
Citation:
Flies AS, Mansfield LS, Grant CK, Weldele ML, Holekamp KE. 2015. Markedly elevated antibody responses in wild versus captive spotted hyenas show that environmental and ecological factors are important modulators of immunity, PLOS ONE, in press.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2015
Citation:
Flies, A. S., Mansfield, L. S., Johnston, E.S., Grant, C. K., and Holekamp, K. E. 2015. Socioecological predictors of immune defenses in a wild spotted hyenas. Functional Ecology, Revision under review.
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2015
Citation:
Invited Talk. Mansfield, LS. Campylobacter jejuni induces acute and chronic disease leading to autoimmunity. Department of Microbiology and Molecular Genetics, Michigan State University, Dec 2015.
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2015
Citation:
Talk. Mansfield LS, Brakel KA, Malik A, Bell JA. 2015. A Human Microbiome Enhanced Campylobacter jejuni Induced Autoantibodies and Th-2 Skewing of Adaptive Immunity after Fecal Transplant American Academy of Allergy, Asthma and Immunology Meeting, Houston TX, February 20-24.
- Type:
Other
Status:
Other
Year Published:
2015
Citation:
Lay press article: Mansfield, L.S. 2014. Transplanting human gut microbes into mice causes the animals to have increased potentially dangerous autoimmune responses. Press release, 114th meeting of the American Society for Microbiology, Boston, MA, May 17-20.
|
Progress 10/01/13 to 09/30/14
Outputs
Target Audience: The target audience for our work are physicians, veterinarians and public health officials interested in how food borne diseases cause long term chronic disease problems in humans. We mainly report to other scientists in these fields by publishing scientific manuscripts and by giving talks at scientific meetings. We also occasionally report directly to the public to explain risks to their health from food borne bacteria and antibiotic resistance. A main target audience are medical and veterinary medical students who Dr. Mansfield teaches. Here the main messages are targeting how bacteria cause food borne disease and how to recognize this disease in humans and animals. Changes/Problems: Nothing to report What opportunities for training and professional development has the project provided? Once again, several students were mentored for study of enteric diseases of food animals. Jessica St. Charles (PhD candidate) successfully defended her PhD thesis in 2013 and began a job as a Immunologist/ Microbiologist for MPI. She is a supervisory microbiologist and leads their program in food safety. Ankit Malik graduated from the PhD program in October 2014 and has been hired as a postdoctoral candidate by St. Jude’s hospital in Memphis, TN. This shows the success of the Mansfield lab in mentoring students to stay in the field of food safety. Three other students are pursuing PhD degrees in food safety related to these enteric pathogens including Barbie Gadsden, DVM, Jean Brudvig, DVM, and Phillip Brooks. One of these students has passed their preliminary exam. Two of the graduate students are underrepresented minorities (African American). Four undergraduate students are working in the lab on these projects including Elizabeth Gensterbloom, Alexander Ethridge, James Chen, and Christopher Bejcek. How have the results been disseminated to communities of interest? Dr. Linda Mansfield continued to act as the Principal Investigator and the Administrative Core Leader of the Michigan State University, Enterics Research Investigational Network, Cooperative Research Center (MSU ERIN CRC) in 2013-2014. This is a multidisciplinary, highly integrated research center to study the enteric microbiome in health and disease with the long-term goal of understanding and alleviating one of the most prevalent and important global health problems, diarrheal illness. The central overarching theme of the center is to explore and elucidate the relationship of the enteric microbiome to acute diarrheal illness. During October 1, 2013 to September 30, 2014, two half day symposia were organized to discuss scientific progress. Dr. Mansfield also gave an invited talk to Merial-NIH Veterinary Scholars Symposium in 2013 where 400 veterinary students attended. Dr. Mansfield also participated in a round table at the Merial-NIH Scholars Symposium in 2013 entitled “Becoming Faculty: a short course on launching a scientific career”. Dr. Mansfield along with Dr. Paul Bartlett organized the Fall seminar series in Large Animal Clinical Sciences at the College of Veterinary Medicine at MSU where preharvest food safety was prominently featured. People attending came from the Agricultural, Veterinary Medicine, Microbiology and Food Science, and Human Nutrition departments. Also, Dr. Mansfield attended and presented at a scientific conference held by the National Institutes of Health in Ann Arbor, MI on enteric diseases. Dr. Mansfield also helped to organize and attended the NC1202 annual meeting in Chicago, Illinois on November 30tht and December 1st, 2013. What do you plan to do during the next reporting period to accomplish the goals? Work Planned Our future goals are to define C. jejuni gene expression controlling the different pathotypes that we have defined in the reported murine models. We will continue this study on the genetic diversity of human and animal isolates of C. jejuni that we have acquired through the US Centers for Diseases Control and Prevention and from a collaboration with Dr. Shannon Manning from an epidemiologic study in Michigan. Methods will include Illumina sequencing, screening in mice with human microbiota given by fecal transplant for Campylobacter gene expression, and selected gene knockouts followed by challenge in the murine models. We are especially interested in evolutionary mechanisms of C. jejuni and how it initiates autoimmune disease in its host.
Impacts
What was accomplished under these goals?
Aim (1). Characterize the neurological disease lesions in mice with Guillain Barré Syndrome Spontaneous Autoimmune Poly Peripheral neuropathywas first reported in NOD CD86-/- mice. Beginning at 20 weeks of age, NOD CD86-/- mice begin to exhibit hind limb weakness and have trouble walking. By 32 weeks of age, 100% of females and 30% of males are affected by SAPP, but are protected from diabetes by an unknown mechanism[1]. In humans with AIDP, the exact nerve “self” antigen has not been determined, but myelin P0 protein was implicated in NOD CD86-/-mice with SAPP[2]. Auto-antibodies to P0 are detectable in these mice beginning ~4 months of age. As in the human form of AIDP, cytokines play a role in the development of SAPP. Two key cytokines play vital roles in the onset and disease course of SAPP. Gene expression of IL-17, a cytokine produced by T helper 17 (TH17) cells, is known to be elevated in the spleens of CD86-/- mice at approximately 16 weeks of age, roughly 4 weeks before the earliest published reports of clinical disease onset[2]. IFNγ gene expression begins to rise between 16 and 24 weeks of age and reaches its peak at 32 weeks of age, which is the reported age at which all female mice and 30% of male mice are affected [1, 2]. The goal of Aim 1 was to clinically characterize the neurological disease in affected mice in detail to determine how well these models mimic human symptoms of GBS. Dr. Gadsden, a trained veterinarian, described, tested and scored the neurological signs in the mice. The peripheral nervous system was examined using histology, EM and IHC to classify the neurological disease to subtype (AIDP, AMAN, AMSAN, etc). Dr. Gadsden also developed a dissection protocol to recover the entire length of the sciatic nerve, nerves of the brachial plexus (en bloc), the dorsal nerve roots, the spinal cord and the brain. The entire length of the sciatic nerve, its dorsal roots, and the brachial plexus were histologically evaluated. A set of nerve scoring criteria was developed by Drs. Mansfield and Gadsden. Fecal samples were collected and evaluated for bacterial pathogens common in mouse colonies and a single blood sample was taken at necropsy, and, evaluated by ELISA for anti-ganglioside antibodies. Experiment (1). Inducible GBS in NOD CD86-/- mice. Recently we have shown that in young NOD CD86-/- mice, 60% of females and 25% of males 8-19 weeks of age develop two to three features of GBS after infection with C. jejuni strains from human GBS patients. These results show that this mouse model can be used to study both induced and spontaneous forms of GBS. Experiment (2). NOD CD86-/- mice develop SAPP in a time- and sex-dependent manner. Hypothesis 1: NOD CD86-/- mice will exhibit clinical signs of peripheral neuropathy similar to those seen in humans with GBS. Inflammatory infiltrates will be consistent with that seen in the Acute Inflammatory Demyelinating Polyneuropathy (AIDP) form of the syndrome. We expected SAPP to develop in our NOD CD86-/- mice as previously described [1]. The objective of this study was to determine whether NOD.129S4 CD86-/- mice in a specific pathogen free colony developed peripheral neuropathy and to characterize the clinical signs and lesions. We expected that NOD CD86-/- mice would begin to develop peripheral neuropathy at 20 weeks of age, with females being affected at a higher rate than males. The study consisted of a total of 82 mice: 60 females, and 22 males. The Open Field Test was used for phenotyping, along with gross clinical observation and photo documentation. We concluded that mice in our colony develop SAPP disease characteristics with some differences in disease phenotypes compared to the single published report. Specifically, 100% of females developed SAPP and <30% of male mice in our colony were affected. Severely affected mice exhibited flipped hind limbs and foot drag when walking. Newly identified features include occasional knuckling and crossing over of the forelimbs, overgrown toenails, flipped hind limbs, and an abnormal reaching reflex. SAPP was not fatal; however, mice were severely debilitated and no recovery phase was observed. Nerve histopathology showed varying degrees of inflammation, composed of lymphocytes, plasma cells, neutrophils, and some macrophage with intracytoplasmic myelin debris. Experiment (3) – NOD CD86-/- mice with clinical signs of SAPP have peripheral nerve lesions characteristic of an inflammatory disorder. We expected that inflammatory cells present within the nerve are responsible for the outward clinical manifestation of neurologic signs. We further proposed that inflammatory infiltrates would precede the clinical signs. The objective of this study was to determine if there are inflammatory lesions present within the nerve before 20 weeks of age and if lesion severity increases with time.The hypothesis was that CD86-/- mice would develop inflammation in their peripheral nerves (sciatic nerve and brachial plexus) shortly preceding onset of disease at 20 weeks of age. The study had 40 female and 11 male mice and used the Open Field and DigiGait tests for phenotyping. For postmortem neurologic evaluation, both forelimbs were removed from the carcass at the root of the brachial plexus and the forelimbs were placed in 10% neutral buffered formalin. Bilaterally, the muscle over the sciatic nerve was reflected back and the sciatic nerve exposed up to the level of the spinal cord. A dorsal laminectomy was performed to expose the spinal cord for fixation of the remainder of the carcass. After two days, the specimens were transferred to 60% ethanol until nerves and roots were harvested. The sciatic nerve was cut at its distal most aspect where it branches into the tibial and common peroneal nerves and the entire length of the nerve was removed (up to the dorsal roots) and placed in a histology cassette. The nerve roots were then removed. Nerves of the brachial plexus were removed en bloc from the axilla and also placed in a cassette. Peripheral nerve sections stained with H&E were scored in the following: dorsal root ganglion, brachial plexus, and sciatic nerve. Scoring was based on the amount of inflammation present in each section (mild, moderate, severe). Cells composing the infiltrate were recorded along with any other architectural changes, such as myelin loss. A Luxol fast blue stained section was used to quantify the percentage of myelin loss. Results: Histopathologic results of nerve dissection and lesion scoring revealed a significant difference in nerve lesions scores between mice that were less than 20 weeks of age, compared to those more than 20 weeks of age. Mice with phenotypic signs of neurological disease had lesions. All NOD CD86-/- female mice developed peripheral neuropathy. Histologic analysis showed that these mice developed inflammatory lesions consistent with GBS. Further, the sciatic nerves of three SAPP affected CD86-/- mice and 3 wild type mice were pooled and analyzed by flow cytometry for immune cell content. There were significant changes in the numbers of neutrophils, dendritic cells, and myeloid cells in the sciatic nerve of the CD86-/- mice, when compared to wild type mice using flow cytometry. Nerves were also scored based on histopathology. A standard operating procedure for nerve lesions scoring is presented at the end of this section of the report. Electron microscopy was also used to detect lesions in the sciatic nerves of NOD CD86-/- mice. We saw lesions in the mice that were consistent with GBS lesions. When compared to normal animals, mice with GBS-like disease had accessory/folded myelin in the center and macrophages with intracytoplasmic myelin accumulation.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Malik A., Sharma D., St. Charles J.L., Dybas L.A., Mansfield L.S. 2013. Contrasting immune responses mediate Campylobacter jejuni induced colitis and autoimmunity, Nature Mucosal immunology, 13 November 2013; doi: 10.1038/mi.2013.97.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Samuelson DR, Eucker TP, Bell JA, Dybas LA, Mansfield LS, Konkel ME. 2013. The Campylobacter jejuni CiaD effector protein activates MAP 1 kinase signaling pathways and is required for the development of acute disease. Cell Communication and Signaling 2013, 11:79, doi:10.1186/1478-811X-11-79.
- Type:
Book Chapters
Status:
Published
Year Published:
2014
Citation:
Jerome, JP and Mansfield LS. 2013 Within-host evolution of Campylobacter jejuni, In "Campylobacter Ecology and Evolution", Samuel K. Sheppard; Associate editor: Guillaume M�ric, Caister Academic Press, UK, pp. 1-25, Publication date: April 2014, ISBN: 978-1-908230-36-2.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Flies AS, Maksimoski MT, Mansfield LS, Weldele ML and Holekamp KE. 2014. Characterization of toll-like receptors 1-10 in spotted hyenas. Veterinary Research Communications, Veterinary Research Communications Jun;38(2):165-70. doi: 10.1007/s11259-014-9592-3. Epub 2014 Feb 2.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2014
Citation:
Flies AS, Mansfield LS, Grant CK, Weldele ML, Holekamp KE. 2014. Markedly elevated antibody responses in wild versus captive spotted hyenas show that environmental and ecological factors are important modulators of immunity, Developmental and
Comparative Immunology, in review.
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Progress 10/01/12 to 09/30/13
Outputs
Target Audience: The target audience are veterinary scientists and microbiologists that are trying to develop preventatives and treatments to combat food borne illness. The goal is to lessen carriage of foodborne bacteria in food animals to protect people. Changes/Problems: No changes were made to the project plan. Excellent progress was made on the goals for this year. What opportunities for training and professional development has the project provided? Professional medical doctors, veterinarians and basic scientists heard the presentations of the research presented by Dr. Mansfield. Further, 4 graduate students were trained in the Comparative Enteric Diseases Laboratory run by Dr. Mansfield. How have the results been disseminated to communities of interest? Dr. Mansfield presented the results of these studies at 3 scientific meetings. These included the Conference of Research Workers in Animal Diseases (Dec, 2012), the Enteric Research Integrated Network Conference in Traverse City Michigan (June3-4, 2013), and a workshop held at the National Institutes of Health in Bethesda, MD (September 3-4, 2013). What do you plan to do during the next reporting period to accomplish the goals? Continue to follow the experimental plan.
Impacts
What was accomplished under these goals?
Determine whether autoimmune sequelae vary with differences in C. jejuni LOS profiles and differences in the LOS region (identifying C. jejuni Guillain Barré Syndrome virulence elements through genotyping. Experiment 1. Multiple molecular typing schemes applied to define genetic relationships among C. jejuni isolates that share the same LOS classification Abstract. The Gram negative bacterium Campylobacter jejuni is the leading cause of bacterial gastroenteritis in the Western world. Symptoms of C. jejuni infection include fever, abdominal cramping, and watery or bloody diarrhea. Sequelae that can occur after infection include Guillain Barré Syndrome (GBS), Miller Fisher Syndrome (MFS), Reiter’s Arthritis, and both Irritable Bowel Syndrome and Inflammatory Bowel Disease. C. jejuni is the most common antecedent infection associated with development of GBS. The suggested mechanism of GBS is molecular mimicry of the C. jejuni lipo-oligosaccharide (LOS) and neurogangliosides resulting in development of anti-ganglioside antibodies. Gangliosides are sialylated glycolipid structures highly enriched in nervous tissue, and it has been demonstrated that C. jejuni sialylated LOS can mimic gangliosides. Serotyping and several molecular typing methods (LOS classification, multi-locus sequence typing (MLST), flaA SVR sequence typing, and porA allele typing) have been applied to define possible genetic relationships among C. jejuni isolates from GBS, MFS, and enteritis patients, but outcomes have been inconsistent. We hypothesized that there would be associations between MLST, flaA, or porA sequence types and C. jejuni strains belonging to LOS classes associated with GBS and MFS in a temporally and geographically limited set of strains. We applied these molecular typing schemes to 68 C. jejuni isolates from patients that presented with enteritis; a small set of unrelated strains, including strains from three patients known to have developed neuropathy were included in the analysis. We expected to see no genetic relationship between LOS classes and the porA allele types or flaA SVR sequence types. We did expect to find strains of some clonal complexes defined by MLST to contain one predominant LOS class. In our study we were able to classify 56/68 C. jejuni isolates into LOS classes A–F; however no isolate was determined to be in LOS class D. Our MLST studies indicated that the strain collection included 36 different sequence types that were clustered into 18 different clonal complexes. DNA sequencing of a portion of the porA gene and flaA SVR gene resulted in the assignment of 43 different porA alleles and 39 different sequence types respectively. Analysis of the LOS classification, MLST, porA gene and flaA SVR gene resulted in 49/68 isolates that were singletons; the remaining isolates were grouped into 8 different groups of 2 or more isolates based on allele assignments. These data demonstrate that C. jejuni populations associated with enteritis are highly diverse based on several genotyping systems. The only clonal complexes with three or more strains were CC 21 (16), CC 353 (7), CC 45 (5), and CC 206 (3). Furthermore, several clonal complexes had associations with particular LOS classes. Experiment 2 – Campylobacter jejuni isolates from calves have A, B, and C lipo-oligosaccharide (LOS) biosynthetic locus classes similar to human Guillain Barré Syndrome associated strains Abstract. Campylobacter jejuni, a Gram negative zoonotic bacterium, is a frequent cause of human gastroenteritis. The acute neuropathy Guillain Barré Syndrome (GBS) is a post-infectious polyradiculoneuropathy triggered by C. jejuni through molecular mimicry. The lipo-oligosaccharide (LOS) of C. jejuni is variable; some forms can mimic gangliosides enriched on peripheral nerves leading to autoimmunity. Classification of the LOS can be determined by PCR based on gene content of the complex LOS biosynthesis locus; 23 classes have been identified. Infection with C. jejuni strains of LOS classes A–C has been strongly associated with GBS. Members of a family managing a large southwest Michigan dairy operation reported a history of C. jejuni infections that lasted several days and recurred over the course of two years. Because up to 37.7% of dairy cattle have been found to shed C. jejuni, we sought to determine whether calves were the source of the family infections. Fecal samples were obtained from 25 randomly selected calves, 1 dog, and 1 family member and cultured for C. jejuni. Human and calf isolates were characterized by LOS biosynthesis locus class typing and by multi-locus sequence typing (MLST). C. jejuni was cultured from 15 (60%) of calves and one asymptomatic family member; the dog was negative. Campylobacter coli was cultured from 2 (1%) of calves. Some calves had diarrhea, but most were clinically normal. Typing of LOS biosynthetic loci showed that 10 of 15calf C. jejuni isolates fell into LOS classifications A, B, and C. The human isolate belonged to LOS class E, which is associated with enteric disease. Two calves had C. jejuni with LOS class E. Thus, the typing results suggest that some calf and human C. jejuni isolates have similar characteristics. Molecular typing with multi-locus sequence typing, porA allele typing, and flaA SVR typing indicated that one calf strain and the human isolate of the same LOS class share the same phylogenetic lineage, thereby providing direct support for zoonotic transmission. Furthermore, finding multiple MLST sequence types, flaA SVR sequence types, and porA alleles LOS classes in isolates from these calves demonstrates that there are diverse C. jejuni populations present on this farm. We also conducted a composite profile analysis. Here, the allele profiles derived from MLST, flaA SVR sequence typing, and porA allele typing were assessed together; results are shown in Figure 5.3. LIAN 3.5 was used to calculate linkage equilibrium parameters; the results indicated that the population is in linkage disequilibrium (Monte Carlo p<0.0001) with an index of association sIA of 0.43. The average genetic diversity, H, for the composite allele profiles from MLST (7 loci), flaA SVR, and porA was calculated using LIAN 3.5; H was 0.67 ± 0.05. It was also observed that within the 16 different isolates (15 calf and 1 family member) analyzed in this study, there were 13 distinct composite profiles based on MLST, porA alleles, and flaA SVR sequence type; a very diverse population of isolates.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Bell J. A., Jerome J.P., Plovanich-Jones A. E., Smith E. J., Gettings J. R., Kim H. Y., Landgraf , Lef�bure T., Kopper J. J., Rathinam V. A., St. Charles J. L., Eaton K. A., Stanhope M. J., Mansfield L. S. 2013. Outcome of infection of C57BL/6 IL-10-/- mice with Campylobacter jejuni strains is correlated with genome content of pathogen loci up- and down-regulated in vivo, Microbial Pathogenesis 54:1-19. PMID: 22960579.
- Type:
Journal Articles
Status:
Published
Year Published:
2012
Citation:
Jerome J.P., Klahn B.D., Bell J.A., Barrick J.E., Brown C.T., and Mansfield L.S. 2012 Draft Genome Sequences of Two Campylobacter jejuni Clinical Isolates, NW and D2600, Journal of Bacteriology 194(20):5707. DOI: 10.1128/JB.01338-12.
- Type:
Journal Articles
Status:
Published
Year Published:
2012
Citation:
Kim J.S., Artymovich K.A., Hall D.F., Smith E.J., Fulton R., Bell J.A., Dybas L., Mansfield L.S., Tempelman R., Wilson D.L., Linz J.E. 2012. Passage of Campylobacter jejuni through the chicken reservoir or mice promotes phase variation in contingency genes Cj0045 and Cj0170 that strongly associates with colonization and disease in a mouse model. Microbiology 158(Pt 5):1304-16. doi: 10.1099/mic.0.057158-0. Epub 2012 Feb 16.
- Type:
Book Chapters
Status:
Published
Year Published:
2013
Citation:
Jerome, J.P. and Mansfield, LS. 2013 Within-host evolution of Campylobacter jejuni, In "Campylobacter Ecology and Evolution", Sheppard, S.K. ed, Sanger Centre, England, pp. 1-25.
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