Learning by doing and creation of the shared discovery curriculum.

BACKGROUND: The Michigan State College of Human Medicine began as an experiment to teach medical students in community-based settings and to create a primary care workforce for the state. Decades later, CHM faced internal and external challenges that spurred creation of a new curriculum - the Share Discovery Curriculum - founded on learning by doing and other learning theories. METHODS: A curricular design group (CDG) developed guiding principles for reform. Based on this, pedagogies and structures were selected to achieve this vision and developed into a curricular structure. Components of the first-year curriculum were piloted with a group of students and faculty members. RESULTS: Six guiding principles were endorsed, grounded in learning theories such as Dewey's Learning by Doing. Based upon these, several key features of the new curriculum emerged: learning communities; one-on-one coaches for students; symptom-based presentations for content; simulation, authentic clinical tasks, flipped classrooms, and modified practice-based learning as primary teaching modalities; early, integrated clinical and scientific learning; milestones as course learning objectives; and a multidimensional, competency-based assessment system. DISCUSSION: The process and outcomes described here are intended as an exemplar for schools undertaking curricular change. Early stakeholder engagement, faculty development, sustainable administrative systems, and managing complexity are core to the success of such endeavors.

Emotional intelligence in undergraduate medical students: a scoping review.

PURPOSE: To perform a scoping review to determine what is known about emotional intelligence (EI) in undergraduate medical education (UME). Two main questions were asked: A. What medical student characteristics are associated with EI? Are there correlations with demographic or other factors? B. What research studies have been done on EI in UME? For example, is there evidence EI changes over time as a result of personal experiences? Should EI be used as an admission criterion? Can EI improve as a result of experiences or deliberate interventions? METHOD: The authors searched four databases (PubMed, PsycInfo, Education Resources Information Center, and Web of Science) for all papers published up to and including December 2020. Two reviewers independently screened articles to determine if they met inclusion criteria. All authors extracted and analyzed data. RESULTS: A set of 1520 papers on the topic of emotional intelligence was identified, with 119 papers meeting inclusion criteria. Most studies were done at international locations with only 17 done at US medical schools. Seventy-five were cohort or cross-sectional studies. Study populations were mixed among the studies, with year of medical study, inclusion of other healthcare students, and participation rates among the inter-study differences noted. CONCLUSIONS: Numerous gaps in the literature on EI exist with several points being clear: (1) there is disagreement on the definition of EI, (2) it is undetermined whether EI is a trait or an ability, and (3) there is marked variability among the instruments used to measure EI. It is also becoming apparent that using EI determination may be helpful as a component of the admission process, higher EI is likely related to improved clinical reasoning, and higher EI contributes to more effective stress management.

Investing in success: student experiences in a structured, decelerated preclinical medical school curriculum.

PURPOSE: Many students in the Michigan State University College of Human Medicine (CHM) are non-traditional with unique needs and experiences. To meet these needs, in 1988 CHM developed a structured Extended Curriculum Program (ECP), which allows students to take longer than 2 years to complete the preclinical curriculum. This work examined the reasons why students extended their programs, their perceptions of that experience, and the outcome with respect to satisfaction and success in their careers after graduation. METHODS: The analysis used data from the college database, follow-up surveys of residency directors and graduates, surveys of graduates who extended, and the AMA Physician Masterfile. RESULTS: Graduates who responded to the survey were evenly split between those who extended for academic reasons and those who extended for other reasons. Although feelings about extending were mixed at the time of extension, nearly all respondents agreed that extending was the right decision in the long run. Extended students continued to face academic challenges having lower basic science averages, lower USMLE Step 1 and 2 first attempt pass rates, and more instances of repeated clerkships compared to those who did not extend, however, most were able to secure a residency in the specialty they desired and had comparable career satisfaction ratings. CONCLUSIONS: The ECP allows some students to complete medical school who otherwise may not have been able to do so. This analysis has provided valuable information that was used to improve the program, allowing CHM to continue its mission of training a diverse set of students to be exemplary physicians.

A Moonlighting Enolase from Lactobacillus gasseri does not Require Enzymatic Activity to Inhibit Neisseria gonorrhoeae Adherence to Epithelial Cells.

Enolases are generally thought of as cytoplasmic enzymes involved in glycolysis and gluconeogenesis. However, several bacteria have active forms of enolase associated with the cell surface and these proteins are utilized for functions other than central metabolism. Recently, a surface-associated protein produced by Lactobacillus gasseri ATCC 33323 with homology to enolase was found to inhibit the adherence of the sexually transmitted pathogen, Neisseria gonorrhoeae, to epithelial cells in culture. Here, we show that the protein is an active enolase in vitro. A recombinantly expressed, C-terminal His-tagged version of the protein, His6-Eno3, inhibited gonococcal adherence. Assays utilizing inhibitors of enolase enzymatic activity showed that this inhibitory activity required the substrate-binding site to be in an open conformation; however, the enolase enzymatic activity of the protein was not necessary for inhibition of gonococcal adherence. An L. gasseri strain carrying an insertional mutation in eno3 was viable, indicating that eno3 is not an essential gene in L. gasseri 33323. This observation, along with the results of the enzyme assays, is consistent with reports that this strain encodes more than one enolase. Here we show that the three L. gasseri genes annotated as encoding an enolase are expressed. The L. gasseri eno3 mutant exhibited reduced, but not abolished, inhibition of gonococcal adherence, which supports the hypothesis that L. gasseri inhibition of gonococcal adherence is a multifactorial process.

Crystal structure of an efficacious gonococcal adherence inhibitor: an enolase from Lactobacillus gasseri.

Enolases are highly conserved metalloenzymes ubiquitous to cellular metabolism. While these enzymes share a large degree of sequence and structural similarity, they have been shown to possess a wide range of moonlighting functions. Recent studies showed that an enolase from Lactobacillus gasseri impedes the ability of Neisseria gonorrhoeae to adhere to epithelial cells. We present the crystal structure of this enolase, the first from Lactobacillus, with one of its Mg(2+) cofactors. Determined using molecular replacement to 2.08Å, the structure has a flexible and surface exposed catalytic loop containing lysines, and may play a role in the inhibitory function.

The College of Human Medicine at Michigan State University: expansion and reinvention.

The College of Human Medicine (CHM) at Michigan State University, which graduated its first class in 1972, was one of the first community-based medical schools in the country. It was established as a state-funded medical school with specific legislative directives to educate primary care physicians who would serve the needs of the state, particularly those of underserved areas. However, the model has proved challenging to sustain with the many changes to the health care system and the economic climate of Michigan. In 2006, a two-phase expansion plan was implemented, and in 2010, CHM permanently expanded the matriculating class from 106 to 200 students with the establishment of a second four-year site for medical education in Grand Rapids. This article describes what school leaders and faculty have learned as they look back at the opportunity provided by expansion as well as the growing pains and lessons learned. The community-based model met many of the mission-related goals for CHM's graduates, who represent a diverse group of practitioners whose values resonate with the school's mission. Expansion has offered an opportunity to explore new research and clinical opportunities as well as to more fully realize the potential of community partners to meet local health care needs and reinvent a robust future for community-integrated medical education.

Expression of a Clostridium perfringens type IV pilin by Neisseria gonorrhoeae mediates adherence to muscle cells.

Clostridium perfringens is an anaerobic, Gram-positive bacterium that causes a range of diseases in humans, including lethal gas gangrene. We have recently shown that strains of C. perfringens move across the surface of agar plates by a unique type IV pilus (TFP)-mediated social motility that had not been previously described. Based on sequence homology to pilins in Gram-negative bacteria, C. perfringens appears to have two pilin subunits, PilA1 and PilA2. Structural prediction analysis indicated PilA1 is similar to the pseudopilin found in Klebsiella oxytoca, while PilA2 is more similar to true pilins found in the Gram-negative pathogens Pseudomonas aeruginosa and Neisseria gonorrhoeae. Strains of N. gonorrhoeae that were genetically deficient in the native pilin, PilE, but supplemented with inducible expression of PilA1 and PilA2 of C. perfringens were constructed. Genetic competence, wild-type twitching motility, and attachment to human urogenital epithelial cells were not restored by expression of either pilin. However, attachment to mouse and rat myoblast (muscle) cell lines was observed with the N. gonorrhoeae strain expressing PilA2. Significantly, wild-type C. perfringens cells adhered to mouse myoblasts under anaerobic conditions, and adherence was 10-fold lower in a pilT mutant that lacked functional TFP. These findings implicate C. perfringens TFP in the ability of C. perfringens to adhere to and move along muscle fibers in vivo, which may provide a therapeutic approach to limiting this rapidly spreading and highly lethal infection.

Lactobacilli at the front line of defense against vaginally acquired infections.

Probiotics are microorganisms that provide a health benefit to the host and are promoted as alternatives for the treatment and prevention of infectious diseases and other conditions. One of the most rapidly developing areas of probiotic research is in the management of vaginally acquired infections. Several Lactobacillus species produce compounds that kill or inhibit the growth of vaginally acquired pathogens. Other lactobacilli reduce the adherence of pathogens to urogenital epithelial cells in culture. This article discusses the mechanisms by which vaginal lactobacilli prevent pathogen colonization of the urogenital tract, and potential mechanisms that warrant investigation. Animal models and clinical studies, while limited, are discussed with the idea that these are the next critical steps to advance the study of probiotics for the treatment and prevention of vaginally acquired infections.

Expression, purification, crystallization and preliminary X-ray studies of Lactobacillus jensenii enolase.

Recombinant Lactobacillus jensenii enolase fused to a C-terminal noncleavable His tag was expressed in Escherichia coli, purified and crystallized by sitting-drop vapor diffusion. A complete data set was collected to 3.25 A resolution. The crystals belonged to space group I4, with unit-cell parameters a=b=145.31, c=99.79 A. There were two protein subunits in the asymmetric unit, which gave a Matthews coefficient VM of 2.8 A3 Da(-1), corresponding to 55.2% solvent content.

Lactobacillus jensenii surface-associated proteins inhibit Neisseria gonorrhoeae adherence to epithelial cells.

High numbers of lactobacilli in the vaginal tract have been correlated with a decreased risk of infection by the sexually transmitted pathogen Neisseria gonorrhoeae. We have previously shown that Lactobacillus jensenii, one of the most prevalent microorganisms in the healthy human vaginal tract, can inhibit gonococcal adherence to epithelial cells in culture. Here we examined the role of the epithelial cells and the components of L. jensenii involved in the inhibition of gonococcal adherence. L. jensenii inhibited the adherence of gonococci to glutaraldehyde-fixed epithelial cells like it inhibited the adherence of gonococci to live epithelial cells, suggesting that the epithelial cells do not need to be metabolically active for the inhibition to occur. In addition, methanol-fixed L. jensenii inhibited gonococcal adherence to live epithelial cells, indicating that L. jensenii uses a constitutive component to inhibit gonococcal interactions with epithelial cells. Proteinase K treatment of methanol-fixed lactobacilli eliminated the inhibitory effect, suggesting that the inhibitory component contains protein. Released surface components (RSC) isolated from L. jensenii were found to contain at least two inhibitory components, both of which are protease sensitive. Using anion-exchange and size exclusion chromatography, an inhibitory protein which exhibits significant similarity to the enzyme enolase was isolated. A recombinant His6-tagged version of this protein was subsequently produced and shown to inhibit gonococcal adherence to epithelial cells in a dose-dependent manner.

Increased expression of the type IV secretion system in piliated Neisseria gonorrhoeae variants.

Neisseria gonorrhoeae produces a type IV secretion system that secretes chromosomal DNA. The secreted DNA is active in the transformation of other gonococci in the population and may act to transfer antibiotic resistance genes and variant alleles for surface antigens, as well as other genes. We observed that gonococcal variants that produced type IV pili secreted more DNA than variants that were nonpiliated, suggesting that the process may be regulated. Using microarray analysis, we found that a piliated strain showed increased expression of the gene for the putative type IV secretion coupling protein TraD, whereas a nonpiliated variant showed increased expression of genes for transcriptional and translational machinery, consistent with its higher growth rate compared to that of the piliated strain. These results suggested that type IV secretion might be controlled by either traD expression or growth rate. A mutant with a deletion in traD was found to be deficient in DNA secretion. Further mutation and complementation analysis indicated that traD is transcriptionally and translationally coupled to traI, which encodes the type IV secretion relaxase. We were able to increase DNA secretion in a nonpiliated strain by inserting a gene cassette with a strong promoter to drive the expression of the putative operon containing traI and traD. Together, these data suggest a model in which the type IV secretion system apparatus is made constitutively, while its activity is controlled through regulation of traD and traI.

Attenuated enzootic (pestoides) isolates of Yersinia pestis express active aspartase.

It is established that Yersinia pestis, the causative agent of bubonic plague, recently evolved from enteropathogenic Yersinia pseudotuberculosis by undergoing chromosomal degeneration while acquiring two unique plasmids that facilitate tissue invasion (pPCP) and dissemination by fleabite (pMT). Thereafter, plague bacilli spread from central Asia to sylvatic foci throughout the world. These epidemic isolates exhibit a broad host range including man as opposed to enzootic (pestoides) variants that remain in ancient reservoirs where infection is limited to muroid rodents. Cells of Y. pseudotuberculosis are known to express glucose-6-phosphate dehydrogenase (Zwf) and aspartase (AspA); these activities are not detectable in epidemic Y. pestis due to missense mutations (substitution of proline for serine at amino position 155 of Zwf and leucine for valine at position 363 of AspA). In this study, functional Zwf was found in pestoides strains E, F and G but not seven other enzootic isolates; enzymic activity was associated with retention of serine at amino acid position 155. Essentially, full AspA activity occurred in pestoides isolates where valine (pestoides A, B, C and D) or serine (pestoides E, F, G and I) occupied position 363. Reduced activity occurred in strains Angola and A16, which contained phenylalanine at this position. The kcat but not Km of purified AspA from strain Angola was significantly reduced. In this context, aspA of the recently described attenuated enzootic microtus biovar encodes active valine at position 363, further indicating that functional AspA is a biomarker for avirulence of Y. pestis in man.

Midpregnancy vaginal fluid defensins, bacterial vaginosis, and risk of preterm delivery.

OBJECTIVE: To assess relations among midpregnancy vaginal defensin levels, a component of the host innate immune response, bacterial vaginosis, and risk of preterm delivery. These relations are compared across race groups because previous studies have repeatedly shown that the prevalence of bacterial vaginosis and the risk of preterm delivery are greater in African-American women compared with that in white women. METHODS: Data are from a prospective study that enrolled pregnant women from 52 clinics in five Michigan communities. In the study subcohort, defensins (human neutrophil peptides 1, 2 and 3) and bacterial vaginosis (Nugent criteria) were measured in vaginal fluid collected at enrollment (15th through 27th week of pregnancy) from 1,031 non-Hispanic white and African-American women (787 term, 244 preterm). Preterm deliveries were categorized by clinical circumstances, ie, spontaneous and medically indicated. RESULTS: Among African Americans, vaginal human neutrophil peptides 1-3 levels greater than or equal to the median were associated with bacterial vaginosis and specifically with spontaneous preterm delivery only (adjusted odds ratio 2.3, 95% confidence interval 1.2-4.3). Once African-American women were stratified by human neutrophil peptide 1-3 levels, bacterial vaginosis added nothing to the prediction of spontaneous preterm delivery risk. None of the above associations were observed in non-Hispanic whites. CONCLUSION: The relations among human neutrophil peptide 1-3 levels, bacterial vaginosis, and preterm delivery vary by race group. In African Americans, midpregnancy human neutrophil peptide 1-3 levels were more informative to preterm delivery risk than was bacterial vaginosis, suggesting an important role for host response. In addition, elevated human neutrophil peptide 1-3 levels may be a marker for particular high-risk vaginal milieus that are not distinguished by the current bacterial vaginosis Nugent scoring system.

A missense mutation causes aspartase deficiency in Yersinia pestis.

It is established that cells of Yersinia pestis, the causative agent of bubonic plague, excrete l-aspartic acid at the expense of exogenous l-glutamic acid during expression of the low-calcium response. Results of enzymic analysis provided here suggest that a previously defined deficiency of aspartase (AspA) accounts for this phenomenon rather than an elevated oxaloacetate pool. The only known distinction between most sequenced isolates of aspA from Y. pestis and the active gene in Yersinia pseudotuberculosis (the immediate progenitor of Y. pestis) is a single base transversion (G.C-->T.A) causing replacement of leucine (encoded by UUG) for valine (encoded by GUG) at amino acid position 363. The gene from Y. pestis KIM possesses a unique second transversion (G.C-->T.A) at amino acid 146 causing substitution of aspartic acid (encoded by GAU) with tyrosine (encoded by UAU). We show in this study that Y. pestis expresses aspA as cross-reacting immunological material (CRIM). Functional and inactive aspA of Y. pseudotuberculosis PB1 and Y. pestis KIM, respectively, were then cloned and expressed in AspA-deficient Escherichia coli. After purification to near homogeneity, the products were subjected to biochemical analysis and found to exhibit similar secondary, tertiary and quaternary (tetrameric) structures as well as comparable Michaelis constants for l-aspartic acid. However, the k(cat) of the Y. pestis CRIM of strain KIM is only about 0.1 % of that determined for the active AspA of Y. pseudotuberculosis. Return of valine for leucine at position 363 of the Y. pestis enzyme restored normal turnover (k(cat) 86+/-2 s(-1)) provided that the amino acid substitution at position 146 was also reversed. These observations have important implications for understanding the nature of the stringent low-calcium response of Y. pestis and its role in promoting acute disease.

Inhibition of Neisseria gonorrhoeae epithelial cell interactions by vaginal Lactobacillus species.

High levels of Lactobacillus, the dominant genus of the healthy human vaginal microbiota, have been epidemiologically linked to a reduced risk of infection following exposure to the sexually transmitted pathogen Neisseria gonorrhoeae. In this work, a cell culture model of gonococcal infection was adapted to examine the effects of lactobacilli on gonococcal interactions with endometrial epithelial cells in vitro. Precolonization of epithelial cells with Lactobacillus jensenii, Lactobacillus gasseri ATCC 33323, or L. gasseri ATCC 9857 reduced gonococcal adherence by nearly 50%. Lactobacilli also inhibited gonococcal invasion of epithelial cells by more than 60%, which was independent of the effect on adherence. Furthermore, lactobacilli were able to displace adherent gonococci from epithelial cells, suggesting that these organisms have potential as a postexposure prophylactic. Thus, vaginal lactobacilli have the ability to inhibit gonococci at two key steps of an infection, which might have a significant effect in determining whether the gonococcus will be able to successfully establish an infection following exposure in vivo.

Two ABC transporter operons and the antimicrobial resistance gene mtrF are pilT responsive in Neisseria gonorrhoeae.

Retraction of type IV pili is mediated by PilT. We show that loss of pilT function leads to upregulation of mtrF (multiple transferable resistance) and two operons encoding putative ABC transporters in Neisseria gonorrhoeae MS11. This effect occurs indirectly through the transcriptional regulator FarR, which until now has been shown to regulate only farAB. L-Glutamine can reverse pilT downregulation of the ABC transporter operons and mtrF.

RpoH mediates the expression of some, but not all, genes induced in Neisseria gonorrhoeae adherent to epithelial cells.

Neisseria gonorrhoeae (gonococcus [GC]), is highly adapted to the human host, the only known reservoir for gonococcal infection. However, since it is sexually transmitted, infection of a new host likely requires a regulatory response on the part of the gonococcus to respond to this significant change in environment. We previously showed that adherence of gonococci to epithelial cells results in changes of gene expression in the bacteria that presumably prepare them for subsequent steps in the infection process. Expression of the heat shock sigma factor gene, rpoH, was shown to be important for the invasion step, as gonococci depleted for rpoH were reduced in their ability to invade epithelial cells. Here, we show that of the genes induced in adherent gonococci, two are part of the gonococcal RpoH regulon. When RpoH is depleted, expression of these genes is no longer induced by host cell contact, indicating that RpoH is mediating the host cell induction response of these genes. One RpoH-dependent gene, NGO0376, is shown to be important for invasion of epithelial cells, consistent with earlier observations that RpoH is necessary for this step of infection. Two genes, NGO1684 and NGO0340, while greatly induced by host cell contact, were found to be RpoH independent, indicating that more than one regulator is involved in the response to host cell contact. Furthermore, NGO0340, but not NGO1684, was shown to be important for both adherence and invasion of epithelial cells, suggesting a complex regulatory network in the response of gonococci to contact with host cells.

Expression capable library for studies of Neisseria gonorrhoeae, version 1.0.

BACKGROUND: The sexually transmitted disease, gonorrhea, is a serious health problem in developed as well as in developing countries, for which treatment continues to be a challenge. The recent completion of the genome sequence of the causative agent, Neisseria gonorrhoeae, opens up an entirely new set of approaches for studying this organism and the diseases it causes. Here, we describe the initial phases of the construction of an expression-capable clone set representing the protein-coding ORFs of the gonococcal genome using a recombination-based cloning system. RESULTS: The clone set thus far includes 1672 of the 2250 predicted ORFs of the N. gonorrhoeae genome, of which 1393 (83%) are sequence-validated. Included in this set are 48 of the 61 ORFs of the gonococcal genetic island of strain MS11, not present in the sequenced genome of strain FA1090. L-arabinose-inducible glutathione-S-transferase (GST)-fusions were constructed from random clones and each was shown to express a fusion protein of the predicted size following induction, demonstrating the use of the recombination cloning system. PCR amplicons of each ORF used in the cloning reactions were spotted onto glass slides to produce DNA microarrays representing 2035 genes of the gonococcal genome. Pilot experiments indicate that these arrays are suitable for the analysis of global gene expression in gonococci. CONCLUSION: This archived set of Gateway entry clones will facilitate high-throughput genomic and proteomic studies of gonococcal genes using a variety of expression and analysis systems. In addition, the DNA arrays produced will allow us to generate gene expression profiles of gonococci grown in a wide variety of conditions. Together, the resources produced in this work will facilitate experiments to dissect the molecular mechanisms of gonococcal pathogenesis on a global scale, and ultimately lead to the determination of the functions of unknown genes in the genome.

Global gene expression and the role of sigma factors in Neisseria gonorrhoeae in interactions with epithelial cells.

Like many bacterial pathogens, Neisseria gonorrhoeae must adapt to environmental changes in order to successfully colonize and proliferate in a new host. Modulation of gene expression in response to environmental signals is an efficient mechanism used by bacteria to achieve this goal. Using DNA microarrays and a tissue culture model for gonococcal infection, we examined global changes in gene expression in N. gonorrhoeae in response to adherence to host cells. Among those genes induced upon adherence to human epithelial cells in culture was rpoH, which encodes a homolog of the heat shock sigma factor, sigma(32) (RpoH), as well as genes of the RpoH regulon, groEL and groES. Attempts to construct an rpoH null mutant in N. gonorrhoeae were unsuccessful, suggesting that RpoH is essential for viability of N. gonorrhoeae. The extracytoplasmic sigma factor, RpoE (sigma(E)), while known to regulate rpoH in other bacteria, was found not to be necessary for the up-regulation of rpoH in gonococci upon adherence to host cells. To examine the role of RpoH in host cell interactions, an N. gonorrhoeae strain conditionally expressing rpoH was constructed. The results of our experiments showed that while induction of rpoH expression is not necessary for adherence of gonococci to epithelial cells, it is important for the subsequent invasion step, as gonococci depleted for rpoH invade cells two- to threefold less efficiently than a wild-type strain. Taken together, these results indicate that sigma(32), but not sigma(E), is important for the response of gonococci in the initial steps of an infection.

Purification, characterization and preliminary X-ray crystallographic studies on Neisseria gonorrhoeae Gly1ORF1.

Gly1ORF1 is a protein produced by the two pathogenic Neisseria species, N. gonorrhoeae and N. meningitidis, but not by commensal Neisseria, suggesting that it may be involved in pathogenesis. The protein has a signal sequence that is cleaved, is associated with outer membrane fractions of N. gonorrhoeae (GC) and is found in spent media and in outer-membrane fractions when expressed in Escherichia coli. GC strains with null mutations of the gly1 locus have increased toxicity to human fallopian tubes in organ culture, suggesting that Gly1ORF1 may alter the amount or properties of toxic moieties produced by GC [Arvidson et al. (1999), infect. Immun. 67, 643-652]. In an effort to understand the function of Gly1ORF1 and its role in pathogenesis, structural biology studies have been initiated. Here, the purification, characterization by dynamic light scattering, crystallization and preliminary X-ray crystallographic studies of recombinant Gly1ORF1 are reported. Dynamic light scattering indicated the protein to be a dimer in solution. The crystals belonged to space group P6(3), with unit-cell parameters a = 95.2, b = 95.2, c = 83.7 A and two molecules per asymmetric unit. The crystals diffracted to 2.4 A using a conventional X-ray source.