P108 and T109 on E2 Glycoprotein Domain I Are Critical for the Adaptation of Classical Swine Fever Virus to Rabbits but Not for Virulence in Pigs.

The classical swine fever virus (CSFV) live attenuated vaccine C-strain is adaptive to rabbits and attenuated in pigs, in contrast with the highly virulent CSFV Shimen strain. Previously, we demonstrated that P108 and T109 on the E2 glycoprotein (E2P108-T109) in domain I (E2DomainI) rather than R132, S133, and D191 in domain II (E2DomainII) determine C-strain's adaptation to rabbits (ATR) (Y. Li, L. Xie, L. Zhang, X. Wang, C. Li, et al., Virology 519:197-206, 2018). However, it remains elusive whether these critical amino acids affect the ATR of the Shimen strain and virulence in pigs. In this study, three chimeric viruses harboring E2P108-T109, E2DomainI, or E2DomainII of C-strain based on the non-rabbit-adaptive Shimen mutant vSM-HCLVErns carrying the Erns glycoprotein of C-strain were generated and evaluated. We found that E2P108-T109 or E2DomainI but not E2DomainII of C-strain renders vSM-HCLVErns adaptive to rabbits, suggesting that E2P108-T109 in combination with the Erns glycoprotein (E2P108-T109-Erns) confers ATR on the Shimen strain, creating new rabbit-adaptive CSFVs. Mechanistically, E2P108-T109-Erns of C-strain mediates viral entry during infection in rabbit spleen lymphocytes, which are target cells of C-strain. Notably, pig experiments showed that E2P108-T109-Erns of C-strain does not affect virulence compared with the Shimen strain. Conversely, the substitution of E2DomainII and Erns of C-strain attenuates the Shimen strain in pigs, indicating that the molecular basis of the CSFV ATR and that of virulence in pigs do not overlap. Our findings provide new insights into the mechanism of adaptation of CSFV to rabbits and the molecular basis of CSFV adaptation and attenuation.IMPORTANCE Historically, live attenuated vaccines produced by blind passage usually undergo adaptation in cell cultures or nonsusceptible hosts and attenuation in natural hosts, with a classical example being the classical swine fever virus (CSFV) lapinized vaccine C-strain, which was developed by hundreds of passages in rabbits. However, the mechanism of viral adaptation to nonsusceptible hosts and the molecular basis for viral adaptation and attenuation remain largely unknown. In this study, we demonstrated that P108 and T109 on the E2 glycoprotein together with the Erns glycoprotein of the rabbit-adaptive C-strain confer adaptation to rabbits on the highly virulent CSFV Shimen strain by affecting viral entry during infection but do not attenuate the Shimen strain in pigs. Our results provide vital information on the different molecular bases of CSFV adaptation to rabbits and attenuation in pigs.

Expression of immunoproteasome subunits in the brains of Toxoplasma gondii-infected mice.

The immunoproteasomes are specific proteasomes that clear oxidant-damaged proteins under inflammatory conditions in various diseases. Toxoplasma gondii (T. gondii) infects the central nervous system and causeencephalitis. However, the relationship between the immunoproteasomes and brain inflammation during T. gondii infection is not well characterized. In this study, we established an in vivo mouse model of T. gondii PLK strain infection via intraperitoneal injection and evaluated the expression of immunoproteasome subunits in the brains of infected mice. The results demonstrated that first, pathological changes in the brains of infected mice increase in severity over time. Second, following T. gondii infection, activated microglia and astrocytes undergo a series of functional alterations and morphological transformations, including proliferation and migration. Third, T. gondii infection induces expression of inflammatory cytokines, including IFN-γ, IL-1ß, TNF-α, and IL-6. Fourth, the immunoproteasome subunits low-molecular-weight polypeptide 2 (LMP2), LMP7, and LMP10 mRNA and protein levels are significantly upregulated in T. gondii-infected mouse brains, as shown by RT-qPCR and western blot analysis, compared with that in vehicle-treated brains, and their expression is localized in the microglia, astrocytes, and neurons of T. gondii-infected brains, as determined via immunofluorescence staining. Furthermore, the western blot mean gray value for the immunoproteasome subunits and the positive microglia and astrocyte immunohistochemical signals in the brains of T. gondii-infected mice were positively correlated, indicating that the observed relationships were highly significant. Therefore, it was concluded that the induction of the immunoproteasomes is a pathogenic mechanism underlying T. gondii infection-induced inflammation.

A Pedagogy of Preparation: Helping Underprepared Students Succeed in College-Level Coursework in Community Colleges.

This paper presents an overall educational philosophy of working with students underprepared for college-level work, which we term "a pedagogy of preparation." We consider how instructors scaffolded instruction to foster college readiness in students who were now able to enroll in college-level work regardless of academic preparation after state-level legislation (SB 1720) that dramatically altered the delivery of developmental education in the Florida College System (FCS). We also consider how collaboration increased among campus personnel after the legislation to foster college readiness in students underprepared for college-level work.

Institutional Transformation Reflected: Engagement in Sensemaking and Organizational Learning in Florida's Developmental Education Reform.

Following a major statewide developmental education reform in Florida, we explored institutional transformation among Florida College System institutions. We used statewide survey data to examine lead administrators' perceptions of challenges encountered during the planning process, ways in which colleges engaged in sensemaking (i.e., social processes for developing shared understanding) and organizational learning, and perceptions of the institutional transformation processes and outcomes following the reform. We found that institutions engaged in numerous types of sensemaking and organizational learning practices to promote change. Yet, despite different approaches taken to institutional transformation, almost all respondents reported that the change process was highly collaborative and involved a broad range of stakeholders.

Transcriptome of porcine alveolar macrophages activated by interferon-gamma and lipopolysaccharide.

The molecular repertoire of porcine alveolar macrophages (PAMs) is greatly affected by the microenvironment they are exposed to, and specifically by inflammatory cytokines, such as interferon gamma (IFN-γ) released by activated lymphocytes, and microbial products, such as lipopolysaccharide (LPS). In our previous study, we found that IFN-γ- and LPS-activated PAMs (M1) could inhibit porcine reproductive and respiratory syndrome virus (PRRSV) replication. In this study, comprehensive analysis of the expression profiles of the genes associated with the polarization of M0-type PAMs (resting) toward M1 phenotypes (activated by IFN-γ and LPS) led to the following main results: 1) 1551 and 1823 genes were upregulated or downregulated in M1-type PAMs, respectively, compared with M0-type PAMs; 2) Among these, genes encoding ASS1 and CRTAM were the most upregulated and downregulated, respectively; 3) Genes involved in cytokine-cytokine receptor interaction and the JAK/STAT signaling pathway were significantly upregulated, suggesting their critical role in cellular activation; and 4) Genes involved in antigen proteolysis and presentation (immunoproteasome subunits), and inhibition of virus replication (host restriction factors) were significantly upregulated, emphasizing the critical role of these cytokines in immunity. Thus, our results provide important information for future studies on the role of PAM polarization in modulation of infection.

Anti-viral immune response in the lung and thymus: Molecular characterization and expression analysis of immunoproteasome subunits LMP2, LMP7 and MECL-1 in pigs.

Both the lung and the thymus are vital target organ for pathogens including viruses. The immunoproteasome (i-proteasome) enhances antigen presentation for MHC class I molecules to activate CD8+T lymphocyte. These facilitate antiviral adaptive immune response. Our previous study found that, expression of i-proteasome subunits in porcine lung was altered during normal and inflammatory conditions. To date, the expression of i-proteasome subunits in porcine thymus to viruses has not been investigated. In the present study, LMP2, LMP7, and MECL-1 were cloned, identified and their sequences encoded predicted proteins of 216, 275, and 278 amino acids, respectively. Expression of LMP2, LMP7, and MECL-1, in the cytoplasm and nucleus, was markedly altered in the porcine reproductive and respiratory syndrome virus (PRRSV)-infected lung and thymus. And dendritic cells and epithelial cells readily expressed the i-proteasome subunit LMP2 in the thymus of PRRSV-infected pigs compared to that in mock-infected pigs. Additionally, the in vitro stimulation of a PAM cell line with PolyI:C for 12 and 24 h resulted in increased LMP2, LMP7, and MECL-1 expression. These results suggest a central role for these complexes in the activation of an antiviral immune response in pigs. A better understanding of the role of the i-proteasome in different cell types, tissues, and hosts could improve vaccine design and facilitate the development of effective treatment strategies for viral infections.

Solitary eosinophilic granulocytic sarcoma in a dog.

A 6-year-old male golden retriever presented with swelling of the left upper eyelid of 2 months duration, which did not improve following a course of antibiotics. Routine serum biochemistry, complete blood count and diagnostic imaging identified no clinically significant abnormalities. The mass was surgically excised, and histopathologic examination was performed. Eosinophilic granulocytic sarcoma (GS) was diagnosed based on the results of histopathology and immunohistochemistry. This is the first report of GS affecting the eyelid of a dog.

Bile duct hamartoma in a dog.

An 11-year-old female Collie presented with a significantly increased abdominal circumference. Computed tomography of the abdomen revealed that the left lateral lobe of the liver contained a large mass, which was excised via laparotomy. Histologically, many small, dilated, cystic luminal structures were anastomosed and connected to a net-like structure. Immunohistochemistry revealed cytokeratin 19-immunopositive areas, representing bile duct structures in the cystic lumen. Based on these results, the tumour was diagnosed as a bile duct hamartoma. To our knowledge, this is the first report of a bile duct hamartoma in a dog.

Do Colleges Perform the Same Following Developmental Education Reform? The Case of Florida's Senate Bill 1720.

Developmental education (DE) reform took place among the 28 Florida College System (FCS) institutions in 2014. In this study, we examine how cohort-based passing rates in college-level English and math courses changed at different colleges for pre- and post-policy period and explore what institutional characteristics were related with various institutional trajectories of cohort-based course passing rates in the post-policy period. Employing longitudinal data analysis, we found that colleges performed similarly regarding cohort-based passing rates in both college-level English and combined math courses before DE reform and had a similar elevation in the cohort-based English course passing rates when DE reform took place in 2014. However, colleges experienced different change patterns in the years following DE reform. Specifically, colleges located in rural areas and with more White students experienced relatively lower college-level English passing rates in the post-policy period than their counterparts. Different colleges had slight differences in the trajectory of college-level math passing rates by cohort after SB 1720 in 2014, but institutional characteristics in this study did not adequately capture inter-institutional differences.

Paraganglioma of the Urinary Bladder in a Dog.

A 3-year-old male Bichon Frise developed lethargy, anorexia and haematuria. B-scan ultrasonography examination revealed a small, irregular, soft-textured mass in the bladder. Histopathologically, there was an incomplete fibrous pseudocapsule around the tumour tissue and although there was clear demarcation from the surrounding tissue, there was invasion of the capsule. Tumour cells proliferated in nests or cords of variable size, separated by fibrovascular tissue. The neoplastic cells were immunopositive for chromogranin A, synaptophysin and neuron-specific enolase, and electron microscopy revealed that they contained cytoplasmic secretory granules. On the basis of these findings, the tumour was diagnosed as a primary paraganglioma of the urinary bladder.

Experimental infection of Bama miniature pigs with a highly virulent classical swine fever virus.

BACKGROUND: Currently, larger domestic pigs are only animals widely used in vaccine evaluation and pathogenicity study of classical swine fever virus (CSFV). This study was aimed to create an alternative animal experimental infection model of CSFV. RESULTS: Twenty specific-pathogen-free Bama miniature pigs were randomly divided into two groups and rooms, infected and non-infected, and the pigs in the infected group were inoculated intramuscularly with 104, 105 or 106 TCID50 (median tissue culture infective dose) CSFV Shimen strain (n = 5 × 3) or left uninoculated to serve as in-contact pigs (n = 3). The uninfected control pigs (n = 2) were housed in a separate room. Clinical signs, body temperature, viraemia, tissue antigen distribution, pathological changes and seroconversion were monitored. Clinical signs were observed as early as 2 days post-inoculation (dpi) in all infected pigs (though mild in contact pigs), but not non-infected control pigs. All inoculated pigs showed viraemia by 6 dpi. The in-contact pigs showed lower levels of viraemia. At 10 dpi, seroconversion was noted in five of the 15 inoculated pigs. All inoculated or one in-contact pigs died by 15 dpi. CONCLUSIONS: These results show that Bama miniature pigs support productive CSFV infection and display clinical signs and pathological changes consistent with CSFV infections observed in larger domestic pigs.

Strain Characterization of Streptococcus suis Serotypes 28 and 31, Which Harbor the Resistance Genes optrA and ant(6)-Ia.

Streptococcus suis causes disease in pigs and is implicated increasingly in human disease worldwide. Although most clinical cases are associated with serotype 2, infections by other serotypes have sometimes been reported. Here, we sequenced the genome of a multidrug-resistant S. suis serotype 28 (strain 11313) and a multidrug-resistant S. suis serotype 31 (strain 11LB5). Strain 11313 was apathogenic in mouse infection models, whereas strain 11LB5 displayed ganglion demyelination, meningeal thickening, congestion, mononuclear cell infiltration, massive proliferation of cortical glial cells, and bacteria (>104 CFU/g) in the spinal cord and ganglia in mice. Furthermore, immunohistochemistry found that the heavily infiltrated glial cells were astrocytes. Strain 11313 harbored the resistance genes ant(6)-Ia, erm(B), optrA, tet(l), tet(o), and strain 11LB5 harbored the resistance genes ant(6)-Ia, erm(B), tet(40), tet(o/w/32/o), aac(6')-aph(2″). Mouse studies showed that strain 11LB5 exhibited a similar virulence to serotype 2 strain 700794, highlighting the need for surveillance of the other serotype S. suis isolates, in addition to serotype 2, in farms. This is the first report of the aminoglycoside resistance gene ant(6)-Ia in S. suis from animals. This suggests that S. suis might serve as an antibiotic resistance reservoir, which spreads the resistance gene ant(6)-Ia or optrA to other streptococcal pathogens on farms.

Exploring Institutional Change in the Context of a Statewide Developmental Education Reform in Florida.

We explore institutional change and organizational learning in higher education in the context of a large-scale reform of developmental education in Florida. We use statewide survey data to examine administrators' perceptions of the use and effectiveness of accountability metrics, methods to identify solutions to improve student outcomes, and challenges responding to data. We find that institutions most commonly use accountability data to track their own performance, but find it less effective for making comparisons across colleges. Institutions use a variety of methods to identify solutions for improvement; with the most common method being presentations at professional association conferences. The greatest challenges in reviewing and responding to data are finding resources to implement solutions and understanding underlying problems affecting student outcomes. We conclude with implications for policymakers and state agency staff designing large-scale reforms in order to encourage organizational learning and promote meaningful change.

Fusion of C3d with hemagglutinin enhances protective immunity against swine influenza virus.

H1N1 and H3N2 are the dominant subtypes causing swine influenza in China and other countries. It is important to develop effective vaccines against both H1N1 and H3N2 subtypes of swine influenza virus (SIV). We examined the effects of a DNA vaccine expressing an influenza HA fused to three copies of murine complement C3d in mice. Plasmids encoding soluble HA (sHA), complete HA (tmHA), or a soluble fused form of HA (sHA-mC3d3) were constructed from the H3N2 subtype of SIV. The immune response was monitored by an enzyme-linked immunosorbent assay (ELISA), hemagglutination inhibition (HI) assays, and virus neutralization tests. Analysis of antibody titers indicated that immunization with HA-mC3d3 resulted in higher titers of anti-HA antibodies and higher antibody affinities, compared with serum from mice immunized with sHA or tmHA. Furthermore, the C3d fusion increased the Th2-biased immune response, by inducing IL-4 production. Splenocytes from mice immunized with sHA-mC3d3 produced about three-fold more IL-4 than did splenocytes from mice immunized with sHA or tmHA. Seven days post-challenge with homologous virus (H3N2), no virus was isolated from the mice immunized with HA-expressing plasmids. However, 10 days post-challenge with heterologous virus (H1N1), only mice immunized with sHA-mC3d3 had no virus or microscopic lesions in the kidneys and cerebrum. In conclusion, C3d enhanced antibody responses to hemagglutinin and protective immunity against SIV of different subtypes.

"The Panhandle is Different than the Peninsula": How Rural Colleges in Florida Implemented Education Reform.

When it comes to the creation of higher education policy, state legislators are challenged with addressing the diverse academic needs of college students enrolled across a spectrum of institutions. In this paper, we explore how 203 individuals at five rural-serving colleges in Florida engaged in state-wide developmental education (DE) reform using the theoretical framework of "situated cognition." Specifically, we ask whether the colleges implemented DE reform in unique ways that may have differed from their non-rural counterparts and what their rationale was for doing so. Our work indicates that institutional culture and capacity generated a combination of strengths and constraints in how reform played out in a rural context. In some ways, a rural-serving identity made colleges more flexible and better able to adjust their advising and curricular structures to comply with the mandates of SB 1720. But in other ways, rural-serving institutions lacked the resources necessary to adequately support students and campus personnel through this dramatic transition toward new ways of providing DE. In looking toward the passage and implementation of future education reform, we call upon policy-makers to consider more deeply the design of policies to ensure they support rural and non-rural colleges alike.

Reading, Writing, and English Course Pathways when Developmental Education is Optional: Course Enrollment and Success for Underprepared First-time-in-College Students.

Academic underpreparedness is an issue for many first-time-in-college students, particularly those entering community colleges. Whereas many underprepared students enroll in developmental education, research has indicated that traditional remediation may not increase students' chances for success. Therefore, states and colleges have begun to implement new course placement strategies to increase the accuracy of initial course placement and new instructional approaches to better serve their developmental students. Specifically, in 2013, the state of Florida passed Senate Bill 1720 which redesigned developmental coursework and placement policies across the Florida College System. The reform lifted developmental education placement exam testing and course enrollment requirements for certain exempt students, irrespective of prior academic preparation or achievement. The current study focuses on these exempt students-those who had the option to bypass developmental education-who were also underprepared, and their initial course selection and subsequent success in their gateway (introductory college-level) English course. Using statewide student-level data and logistic regression techniques, the results indicated that level of preparation was related to students' course enrollment and gateway English course success. Students slightly underprepared in reading or writing were more likely than severely underprepared students to enroll in the gateway English class, relative to a developmental reading or writing course. In reading and writing, slightly underprepared students were more likely to pass English, relative to severely underprepared students. The authors consider the findings in light of recent national changes to developmental education and offer recommendations for policy and practice.

Equality, Efficiency, and Developmental Education Reform: The Impact of SB 1720 on the Mission of the Florida College System.

OBJECTIVE: Community colleges have long made higher education more accessible to students from diverse academic backgrounds, particularly those who are academically underprepared and require remediation. In light of developmental education (DE) reform, our article answers the following questions: How do campus personnel articulate the unique mission of Florida's state colleges, formerly known as community colleges? Furthermore, how do they perceive the mandates of reform to have shaped their ability to carry out this mission? METHOD: This work is based on an embedded case study of 10 Florida College System institutions. Qualitative data were gathered between 2014 and 2018 from 544 college presidents, administrators, faculty, staff through 92 focus groups and 8 interviews. RESULTS: Campus personnel strongly affirmed the mission of the Florida Colleges System as one of democratic equality. However, many were concerned that DE reform, namely Senate Bill 1720, prioritized efficiency over equality in the pursuit of cost savings. Specifically, participants expressed frustration that reforms accelerated DE coursework to an unmanageable pace and ignored the presence of a digital divide. Opinions of DE reform improved in the 4 years following implementation, but some concern persisted. CONTRIBUTIONS: Our findings highlight the centrality of democratic equality to the community college mission for campus personnel. They also suggest that equality and efficiency need not always be opposing goals in education reform. Finally, they call into question social policy that universally promotes accelerated and computer-based courses without considering that some students may require accommodations.

Establishment of BV2 microglia polarization model and its effect on Toxoplasma gondii proliferation.

Toxoplasma gondii is an intracellular opportunistic, parasitic protozoan. Microglia have been classified into two main types: M1 (classically activated macrophages) and M2 (alternatively activated macrophages). BV2 cells were used in this study, together with lipopolysaccharide (LPS) and interferon (IFN)-γ or interleukin (IL)-4, which were used to induce resting microglia. Expression levels of M1/M2 markers were determined at both mRNA and protein levels, using PCR, western blot, and flow cytometry. Furthermore, cells were infected with T. gondii PLK strain, and the dynamic changes in M1/M2 marker expression levels were determined. An in vitro polarization model was successfully established. Expression of Nos2 and M1-associated markers was significantly upregulated at 12 h post-infection in BV2 cells. Further, the JAK/STAT1 and NF-κB signaling pathways were also activated following T. gondii infection. This demonstrated that T. gondii infection induces M1-type microglial polarization in vitro. The present study demonstrated that T. gondii infection affects microglial activation in vitro and elucidated the effects of activated microglia on T. gondii proliferation. This data may serve as a useful reference for more detailed elucidation of interactions between T. gondii and the innate immune system.

What Happens to Underprepared First-Time-in-College Students When Developmental Education is Optional? The Case of Developmental Math and Intermediate Algebra in the First Semester.

In 2014, developmental education became optional for many college students in Florida, regardless of prior academic preparation. This study investigated first-semester math course enrollment patterns for underprepared first-time-in-college (FTIC) students who would have previously been required to take developmental math and the passing rates for the students electing to take Intermediate Algebra (the most common gateway math course in Florida). We found that roughly a 3rd of underprepared students enrolled in developmental math, a 3rd enrolled in Intermediate Algebra, and roughly a 3rd enrolled in no math course whatsoever, with preparation level being related to enrollment pathways. Among those who enrolled in Intermediate Algebra, a small percentage also enrolled in developmental math in the same semester, either through a compressed or corequisite course, and FTIC students who received same-semester developmental support were more likely to pass Intermediate Algebra compared with similar underprepared students who took Intermediate Algebra without developmental support.

Microglia Are Derived from Peripheral Blood Mononuclear Cells After Pseudorabies Infection in Mice.

Pseudorabies virus (PRV) can spread along the peripheral nerves near the site of infection in the animals, and gradually migrates into the central nervous system, where it leads to the development of brain lesions. The aim of this study was to investigate the dynamics of microglia after PRV inoculation. A mouse model inoculated with PRV was established to study the interactions between PRV and microglia, microglial recruitment, and polarization effects. The mice were subcutaneously inoculated with different doses of PRV-Bartha K61 vaccine strain. The obtained results showed that mouse mortality rates increased with the applied doses of virus, and brain lesions, located in the brain tail and brain stem, were observed in each investigated group. Inflammatory cells were shown to infiltrate through the vasculature into perivascular cuff, and the number of microglia was increased as well. Mouse group treated with a medium infection dose demonstrated a high survival rate while developing serious brain lesions, and therefore, this dose was selected for further experiments. Immunohistochemistry, flow cytometry, and confocal laser scanning microscopy were used to analyze PRV-microglia interactions. After PRV inoculation, proliferating cell nuclear antigen (Pcna) and Iba1 double-positive cells were observed in the brain lesions, together with the activated microglia, suggesting that PRV can induce microglial proliferation and activation. Furthermore, 5-bromo-deoxy-uridine (BrdU) labeling demonstrated that microglial cells did not proliferate in situ and the proliferating cells originated from peripheral blood monocytes, mainly from the inflammatory monocytes (Ly6Chigh). In addition, microglia polarized into both M1 and M2 phenotypes by PRV infection. The results obtained in this study may help understand the development of pseudorabies infection and help improve the treatment, by recruiting and enhancing immune response.