'One Health' investigation: outbreak of human Salmonella Braenderup infections traced to a mail-order hatchery - United States, 2012-2013.

Human salmonellosis linked to contact with live poultry is an increasing public health concern. In 2012, eight unrelated outbreaks of human salmonellosis linked to live poultry contact resulted in 517 illnesses. In July 2012, PulseNet, a national molecular surveillance network, reported a multistate cluster of a rare strain of Salmonella Braenderup infections which we investigated. We defined a case as infection with the outbreak strain, determined by pulsed-field gel electrophoresis, with illness onset from 25 July 2012-27 February 2013. Ill persons and mail-order hatchery (MOH) owners were interviewed using standardized questionnaires. Traceback and environmental investigations were conducted. We identified 48 cases in 24 states. Twenty-six (81%) of 32 ill persons reported live poultry contact in the week before illness; case-patients named 12 different MOHs from eight states. The investigation identified hatchery D as the ultimate poultry source. Sampling at hatchery D yielded the outbreak strain. Hatchery D improved sanitation procedures and pest control; subsequent sampling failed to yield Salmonella. This outbreak highlights the interconnectedness of humans, animals, and the environment and the importance of industry knowledge and involvement in solving complex outbreaks. Preventing these infections requires a 'One Health' approach that leverages expertise in human, animal, and environmental health.

Severe histiolymphocytic and heterophilic bronchopneumonia as a reaction to in ovo fowlpox vaccination in broiler chicks.

Broiler chickens on several farms from a single poultry company experienced neurological signs and mortality in chicks between 3 days and 10 days of age over a 3-week period after use of a fowlpox-vectored infectious laryngotracheitis virus vaccine in ovo. At necropsy the lungs contained numerous tan or gray, opaque to translucent, 0.5- to 2.0-mm nodules in the parenchyma. Microscopic lesions were a multifocal severe lymphohistiocytic and heterophilic bronchopneumonia. Immunohistochemistry was positive for fowlpox virus in macrophages and lymphocytes, and polymerase chain reaction on paraffin-embedded lung tissues was positive for a fowlpox vector virus commonly used as a vaccine. The cause of the neurological signs was not determined.

Differential metabolic effects of glucosamine and N-acetylglucosamine in human articular chondrocytes.

OBJECTIVE: Aminosugars are commonly used to treat osteoarthritis; however, molecular mechanisms mediating their anti-arthritic activities are still poorly understood. This study analyzes facilitated transport and metabolic effects of glucosamine (GlcN) and N-acetylglucosamine (GlcNAc) in human articular chondrocytes. METHODS: Human articular chondrocytes were isolated from knee cartilage. Facilitated transport of glucose, GlcN and GlcNAc was measured by uptake of [3H]2-deoxyglucose, [3H]GlcN and [3H]GlcNAc. Glucose transporter (GLUT) expression was analyzed by Western blotting. Production of sulfated glycosaminoglycans (SGAG) was measured using [(35)S]SO4. Hyaluronan was quantified using hyaluronan binding protein. RESULTS: Chondrocytes actively import and metabolize GlcN but not GlcNAc and this represents a cell-type specific phenomenon. Similar to facilitated glucose transport, GlcN transport in chondrocytes is accelerated by cytokines and growth factors. GlcN non-competitively inhibits basal glucose transport, which in part depends on GlcN-mediated depletion of ATP stores. In IL-1beta-stimulated chondrocytes, GlcN inhibits membrane translocation of GLUT1 and 6, but does not affect the expression of GLUT3. In contrast to GlcN, GlcNAc accelerates facilitated glucose transport. In parallel with the opposing actions of these aminosugars on glucose transport, GlcN inhibits hyaluronan and SGAG synthesis while GlcNAc stimulates hyaluronan synthesis. GlcNAc-accelerated hyaluronan synthesis is associated with upregulation of hyaluronan synthase-2. CONCLUSION: Differences in GlcN and GlcNAc uptake, and their subsequent effects on glucose transport, GLUT expression and SGAG and hyaluronan synthesis, indicate that these two aminosugars have distinct molecular mechanisms mediating their differential biological activities in chondrocytes.

Cytokine regulation of facilitated glucose transport in human articular chondrocytes.

Glucose serves as the major energy substrate and the main precursor for the synthesis of glycosaminoglycans in chondrocytes. Facilitated glucose transport represents the first rate-limiting step in glucose metabolism. This study examines molecular regulation of facilitated glucose transport in normal human articular chondrocytes by proinflammatory cytokines. IL-1beta and TNF-alpha, and to a lesser degree IL-6, accelerate facilitated glucose transport as measured by [(3)H]2-deoxyglucose uptake. IL-1beta induces an increased expression of glucose transporter (GLUT) 1 mRNA and protein, and GLUT9 mRNA. GLUT3 and GLUT8 mRNA are constitutively expressed in chondrocytes and are not regulated by IL-1beta. GLUT2 and GLUT4 mRNA are not detected in chondrocytes. IL-1beta stimulates GLUT1 protein glycosylation and plasma membrane incorporation. IL-1beta regulation of glucose transport in chondrocytes depends on protein kinase C and p38 signal transduction pathways, and does not require phosphoinositide 3-kinase, extracellular signal-related kinase, or c-Jun N-terminal kinase activation. IL-1beta-accelerated glucose transport in chondrocytes is not mediated by endogenous NO or eicosanoids. These results demonstrate that stimulation of glucose transport represents a component of the chondrocyte response to IL-1beta. Two classes of GLUTs are identified in chondrocytes, constitutively expressed GLUT3 and GLUT8, and the inducible GLUT1 and GLUT9.

The osteoprotegerin/receptor activator of nuclear factor kappaB/receptor activator of nuclear factor kappaB ligand system in cartilage.

OBJECTIVE: The receptor activator of nuclear factor kappaB (RANK) is a member of the tumor necrosis factor receptor family. It is activated by the secreted or cell surface-bound RANK ligand (RANKL). Osteoprotegerin (OPG) is a soluble nonsignaling receptor for RANKL and interferes with RANK activation. This receptor-ligand system regulates the differentiation of osteoclasts and dendritic cells. The present study examined human articular cartilage for the expression of these molecules and the role of RANKL in the regulation of chondrocyte function. METHODS: Normal and osteoarthritic (OA) human articular cartilage was used for explant tissue culture or for isolation of chondrocytes and cell culture. Expression of RANK, RANKL, and OPG was analyzed by immunohistochemistry, Western blotting, or reverse transcription-polymerase chain reaction. Recombinant RANKL was added to cartilage or chondrocyte cultures, and gene expression, collagenase and nitric oxide production, and NF-kappaB activation were determined. RESULTS: RANK, RANKL, and OPG messenger RNA (mRNA) were expressed in normal cartilage. By immunohistochemistry, RANK, RANKL, and OPG were detected in the superficial zone of normal cartilage. OA cartilage contained increased levels of OPG mRNA, and expression of the 3 proteins extended into the midzone of OA cartilage. OPG was detected by Western blotting, and was increased in response to interleukin-1beta stimulation. OPG, RANK, and RANKL protein were also detected in cultured chondrocytes. Addition of exogenous RANKL did not activate NF-kappaB, induce expression of genes encoding proinflammatory mediators in chondrocytes, or stimulate the production of collagenase and nitric oxide. CONCLUSION: These results demonstrate the expression of OPG, RANK, and RANKL in cartilage. However, RANKL does not activate human articular chondrocytes.

Profile of glycosaminoglycan-degrading glycosidases and glycoside sulfatases secreted by human articular chondrocytes in homeostasis and inflammation.

OBJECTIVE: To determine enzymatic activities of the 8 key glycosaminoglycan-degrading glycosidases and glycoside sulfatases in cultured human articular chondrocytes and in synovial fluid from patients with osteoarthritis. METHODS: The following enzymes were analyzed: hexosaminidase and its isoenzyme A, N-acetyl-alpha-D-glucosaminidase, beta-galactosidase, beta-glucuronidase, alpha-L-iduronidase, aryl sulfatase, and galactose-6-sulfate sulfatase. Activity of the selected enzymes was analyzed by fluorometry with the aid of 4-methylumbelliferryl derivatives of the appropriate monosaccharides. RESULTS: Hexosaminidase was found to be the dominant enzyme released by chondrocytes into the extracellular compartment. Stimulation of chondrocytes with interleukin-1beta resulted in a selective increase of the extracellular hexosaminidase activity and, to a lesser degree, of the extracellular beta-galactosidase activity, without significant changes in the activity of the other studied enzymes. Analysis of the pH dependency of the enzymatic activities revealed that even at neutral pH, hexosaminidase expressed a measurable activity, much higher than the activity of the other studied enzymes. Chondrocyte apoptosis did not result in increased extracellular glycosidase activities, including hexosaminidase activity. The spectrum of glycosidase and glycoside sulfatase activities in the synovial fluid from patients with osteoarthritis was similar to that in cultured human articular chondrocytes. CONCLUSION: These data support the concept that lysosomal glycosidases, in particular hexosaminidase, represent a distinct subset of cartilage matrix-degrading enzymes that are activated by proinflammatory stimuli.

Human rheumatoid factor production is dependent on CD40 signaling and autoantigen.

High-affinity pathologic rheumatoid factor (RF) B cells occur in autoimmune diseases such as rheumatoid arthritis, but are deleted in healthy individuals. The reasons for the survival and differentiation of these autoreactive B cells in rheumatoid arthritis are not known. Previous studies in mice transgenic for a human IgM RF have shown that peripheral encounter with soluble human IgG leads to deletion of high-affinity RF B cells; however, deletion can be prevented when concomitant T cell help is provided. This study aimed to further discern the minimal factors necessary not only for the in vivo survival of RF B cells, but also for their differentiation into Ab-secreting cells. The combination of MHC class II-reactive T cells and Ag induced the production of RF in human IgM RF transgenic mice, while either stimulus alone was ineffective. Neutralizing Abs against CD40 ligand (CD40L), but not against IL-4 or IL-15, abrogated IgM-RF production. Moreover, blockade of CD40L-CD40 allowed IgG to delete the RF precursor cells. Most importantly, activating Abs to CD40 could substitute entirely for T cell help in promoting the survival of RF precursors and in stimulating RF synthesis in T cell deficient animals. The data indicate that CD40 signaling alone can prevent deletion of RF B cells by Ag and in the presence of IgG is sufficient to trigger RF synthesis. The results suggest that selective induction of apoptosis in high-affinity RF B cells may be achieved by blockade of CD40L-CD40 interaction.

Rheumatoid factor B cell tolerance via autonomous Fas/FasL-independent apoptosis.

Normal individuals do not express the high-affinity autoantibodies specific for self-IgG (rheumatoid factors, RF) that are commonly seen in rheumatoid arthritis patients. Studies of transgenic mice expressing a human IgM rheumatoid factor have shown that one mechanism by which higher affinity RF B cells are tolerized to IgG is through abortive RF B cell activation followed by deletion in the absence of T cell help. We show that RF B cell deletion occurs through an intrinsic apoptotic mechanism that is independent of the Fas/FasL pathway and does not involve active killing by T cells, as it occurs in RAG-1-deficient RF transgenic mice to the same extent as in the parental RF transgenic line.

Costimulation provided by DNA immunization enhances antitumor immunity.

The interaction of the TCR with MHC class I-bound Ag is insufficient for the priming of CTL unless secondary costimulatory signals are provided. To ascertain the minimum elements required to activate an Ag-specific CTL response in vivo, we injected mice intradermally or i.m. with plasmid DNA encoding a MHC class I-restricted peptide Ag (minigene) and different membrane-bound costimulatory ligands. The minigene-encoded epitope only primed a specific CTL response if injected in the vicinity of an ectopically expressed costimulatory ligand. Vector encoding B7-1 was repeatedly more potent at stimulating a cytolytic response than vector encoding B7-2. In contrast the B7-2-encoding plasmid preferentially enhanced Ag-specific Ab responses when injected with either protein or a cDNA expression vector. Gene vaccination with plasmids encoding OVA and B7-1, but not B7-2, prolonged survival in mice challenged with an OVA-transfected tumor. These results show that functional B7-1 transfection can be achieved in vivo and induces the selective induction of CTL. The data suggest that B7-1 plasmids should be coadministered with naked DNA vaccines that aim to induce tumor-specific cellular immunity.

Peripheral deletion of rheumatoid factor B cells after abortive activation by IgG.

Rheumatoid factor (RF) B cells proliferate during secondary immune responses to immune complexed antigen and antigen specific T cells, but higher affinity RFs are not detected except in patients with rheumatoid arthritis and other autoimmune diseases. Consequently, there must exist highly efficient mechanisms for inactivation of these higher-affinity RF B cell clones under normal circumstances. Exposure of transgenic mice expressing a human IgM RF to soluble human IgG in the absence of T cell help causes antigen specific B cell deletion in 2-3 days. The deletion is independent of the Fas/Fas ligand (FasL) pathway of apoptosis and is preceded by a phase of partial activation involving increase in cell size and expression of B7 and ICAM-1, and transient release of low levels of immunoglobulin. Complete B cell activation involving the formation of germinal centers and sustained high level RF secretion only occurs if T cell help is provided simultaneously. RF B cells exposed to tolerogen remain competent to secrete RF in vitro if provided with an appropriate antigenic stimulus and T cell help. Consequently, death of these cells is not preceded by anergy. Abortive activation/deletion of B cells by antigen in the absence of T cell-derived survival signals may represent the major mechanism for maintaining peripheral tolerance in B cells expressing higher affinity RF. The lack of anergy, and the potential for reactivation before death, provide a means for maintaining RF production under pathologic circumstances, such as may occur in the inflamed rheumatoid synovium.

Treatment of experimental gram-negative bacterial sepsis with murine monoclonal antibodies directed against lipopolysaccharide.

Monoclonal antibodies (MAbs) directed against gram-negative bacterial lipopolysaccharide (endotoxin, LPS) are currently being evaluated as an adjunctive form of therapy for lethal gram-negative bacterial sepsis and shock. The exact binding site within the LPS molecule against which antibody should be directed in order to maximize both cross-reactivity among bacterial strains and protective capacity has not been established. By developing a panel of MAbs that bound to various regions of the LPS molecule (O saccharide; outer, intermediate, and inner core; lipid A), we were able to determine that some epitopes in the inner core/lipid A region of LPS were broadly shared among different genera of gram-negative microorganisms, on the basis of immunoblot analysis of MAb binding to LPS. Pretreatment with lower doses of O saccharide-specific MAbs (2 micrograms per animal) provided protection against a lethal intraperitoneal challenge of viable Salmonella minnesota bacteria, compared with core LPS-specific MAbs, which required at least 1.0 mg of MAb per mouse to provide a similar degree of immunoprotection. Although inner core LPS-specific MAbs are less protective than O saccharide-specific MAbs, these MAbs will probably be more useful in the treatment of gram-negative sepsis because of their ability to bind to many types of LPS and enhance survival during infection, which is caused by a wide variety of gram-negative bacteria.

Comparison of O antigen-specific urinary immunoglobulins to Escherichia coli in normal women and women prone to Escherichia coli cystitis.

The enzyme-linked immunosorbent assay (ELISA) measured urine and serum levels of O antigen-specific IgA against Escherichia coli in five women prone to recurrent Esch. coli cystitis and in 20 normal women. Serial urine and serum samples were taken from the five women during Esch. coli infections and during uninfected intervals, while a single urine and serum sample was obtained from the 20 normal women. Elevation of any class of serum immunoglobulins for patients with uncomplicated Esch. coli cystitis was absent. A local antigen-specific immunological response, predominantly of the IgA class, was present during Esch. coli cystitis. There was no significant difference (P greater than 0.1) in the urinary or serum levels of any class of immunoglobulins between the recurrent cystitis group when uninfected and the normal women. There was a significant increase (P less than 0.001) in the levels of urinary IgA and secretory IgA during Esch. coli cystitis as compared with the same women when uninfected or with normal women. A deficiency in the local immunological response of women prone to recurrent Esch. coli cystitis is not supported by our data.

Measurement of human antibody activity against Escherichia coli and Pseudomonas aeruginosa using formalin treated whole organisms in an ELISA technique.

Using an ELISA technique, specific IgG and specific IgM antibodies to several strains of Escherichia coli and Pseudomonas aeruginosa were measured in 100 normal adults. The distribution of antibody activity to E. coli was narrow, with mean values less than 0.50 OD units. The one exception was in IgG activity to E. coli O.. Mean values for activity against P. aeruginosa ranged from 0.35 to 0.79 OD units. Significant rank order correlations were found for IgM activity among all E. coli and P. aeruginosa strains. The correlations were less consistent for the IgG activity. This baseline data will be used to monitor antibody activity to these common microbes along with several other parameters in a group of ill surgical patients.