Brian P. Butler

Gut inflammation provides a respiratory electron acceptor for Salmonella

Salmonella enterica serotype Typhimurium (S. Typhimurium) causes acute gut inflammation by using its virulence factors to invade the intestinal epithelium and survive in mucosal macrophages. The inflammatory response enhances the transmission success of S. Typhimurium by promoting its outgrowth in the gut lumen through unknown mechanisms. Here we show that reactive oxygen species generated during inflammation react with endogenous, luminal sulphur compounds (thiosulphate) to form a new respiratory electron acceptor, tetrathionate. The genes conferring the ability to use tetrathionate as an electron acceptor produce a growth advantage for S. Typhimurium over the competing microbiota in the lumen of the inflamed gut. We conclude that S. Typhimurium virulence factors induce host-driven production of a new electron acceptor that allows the pathogen to use respiration to compete with fermenting gut microbes. Thus the ability to trigger intestinal inflammation is crucial for the biology of this diarrhoeal pathogen.

Lipocalin-2 Resistance Confers an Advantage to Salmonella enterica Serotype Typhimurium for Growth and Survival in the Inflamed Intestine

In response to enteric pathogens, the inflamed intestine produces antimicrobial proteins, a process mediated by the cytokines IL-17 and IL-22. Salmonella enterica serotype Typhimurium thrives in the inflamed intestinal environment, suggesting that the pathogen is resistant to antimicrobials it encounters in the intestinal lumen. However, the identity of these antimicrobials and corresponding bacterial resistance mechanisms remain unknown. Here, we report that enteric infection of rhesus macaques and mice with S. Typhimurium resulted in marked Il-17- and IL-22-dependent intestinal epithelial induction and luminal accumulation of lipocalin-2, an antimicrobial protein that prevents bacterial iron acquisition. Resistance to lipocalin-2, mediated by the iroBCDE iroN locus, conferred a competitive advantage to the bacterium in colonizing the inflamed intestine of wild-type but not of lipocalin-2-deficient mice. Thus, resistance to lipocalin-2 defines a specific adaptation of S. Typhimurium for growth in the inflamed intestine.

Both Hemolytic Anemia and Malaria Parasite-Specific Factors Increase Susceptibility to Nontyphoidal Salmonella enterica Serovar Typhimurium Infection in Mice

ABSTRACT Severe pediatric malaria is an important risk factor for developing disseminated infections with nontyphoidal Salmonella serotypes (NTS). While recent animal studies on this subject are lacking, early work suggests that an increased risk for developing systemic NTS infection during malaria is caused by hemolytic anemia, which leads to reduced macrophage microbicidal activity. Here we established a model for oral Salmonella enterica serotype Typhimurium challenge in mice infected with Plasmodium yoelii nigeriensis. Initial characterization of this model showed that 5 days after coinoculation, P. yoelii nigeriensis infection increased the recovery of S. Typhimurium from liver and spleen by approximately 1,000-fold. The increased bacterial burden could be only partially recapitulated by antibody-mediated hemolysis, which increased the recovery of S. Typhimurium from liver and spleen by 10-fold. These data suggested that both hemolysis and P. yoelii nigeriensis-specific factors contributed to the increased susceptibility to S. Typhimurium. The mechanism by which hemolysis impaired resistance to S. Typhimurium was further investigated. In vitro, S. Typhimurium was recovered 24 h after infection of hemophagocytic macrophages in 2-fold-higher numbers than after infection of mock-treated macrophages, making it unlikely that reduced macrophage microbicidal activity was solely responsible for hemolysis-induced immunosuppression during malaria. Infection with P. yoelii nigeriensis, but not antibody-mediated hemolysis, reduced serum levels of interleukin-12p70 (IL-12p70) in response to S. Typhimurium challenge. Collectively, studies establishing a mouse model for this coinfection suggest that multiple distinct malaria-induced immune defects contribute to increased susceptibility to S. Typhimurium.

Malaria parasite infection compromises control of concurrent systemic non-typhoidal Salmonella infection via IL-10-mediated alteration of myeloid cell function.

Non-typhoidal Salmonella serotypes (NTS) cause a self-limited gastroenteritis in immunocompetent individuals, while children with severe Plasmodium falciparum malaria can develop a life-threatening disseminated infection. This co-infection is a major source of child mortality in sub-Saharan Africa. However, the mechanisms by which malaria contributes to increased risk of NTS bacteremia are incompletely understood. Here, we report that in a mouse co-infection model, malaria parasite infection blunts inflammatory responses to NTS, leading to decreased inflammatory pathology and increased systemic bacterial colonization. Blunting of NTS-induced inflammatory responses required induction of IL-10 by the parasites. In the absence of malaria parasite infection, administration of recombinant IL-10 together with induction of anemia had an additive effect on systemic bacterial colonization. Mice that were conditionally deficient for either myeloid cell IL-10 production or myeloid cell expression of IL-10 receptor were better able to control systemic Salmonella infection, suggesting that phagocytic cells are both producers and targets of malaria parasite-induced IL-10. Thus, IL-10 produced during the immune response to malaria increases susceptibility to disseminated NTS infection by suppressing the ability of myeloid cells, most likely macrophages, to control bacterial infection.

Microbial amyloids induce interleukin 17A (IL-17A) and IL-22 responses via toll-like receptor 2 activation in the intestinal mucosa

ABSTRACT The Toll-like receptor 2 (TLR2)/TLR1 receptor complex responds to amyloid fibrils, a common component of biofilm material produced by members of the phyla Firmicutes, Bacteroidetes, and Proteobacteria. To determine whether this TLR2/TLR1 ligand stimulates inflammatory responses when bacteria enter intestinal tissue, we investigated whether expression of curli amyloid fibrils by the invasive enteric pathogen Salmonella enterica serotype Typhimurium contributes to T helper 1 and T helper 17 responses by measuring cytokine production in the mouse colitis model. A csgBA mutant, deficient in curli production, elicited decreased expression of interleukin 17A (IL-17A) and IL-22 in the cecal mucosa compared to the S. Typhimurium wild type. In TLR2-deficient mice, IL-17A and IL-22 expression was blunted during S. Typhimurium infection, suggesting that activation of the TLR2 signaling pathway contributes to the expression of these cytokines. T cells incubated with supernatants from bone marrow-derived dendritic cells (BMDCs) treated with curli fibrils released IL-17A in a TLR2-dependent manner in vitro. Lower levels of IL-6 and IL-23 production were detected in the supernatants of the TLR2-deficient BMDCs treated with curli fibrils. Consistent with this, three distinct T-cell populations—CD4+ T helper cells, cytotoxic CD8+ T cells, and γδ T cells—produced IL-17A in response to curli fibrils in the intestinal mucosa during S. Typhimurium infection. Notably, decreased IL-6 expression by the dendritic cells and decreased IL-23 expression by the dendritic cells and macrophages were observed in the cecal mucosa of mice infected with the curli mutant. We conclude that TLR2 recognition of bacterial amyloid fibrils in the intestinal mucosa represents a novel mechanism of immunoregulation, which contributes to the generation of inflammatory responses, including production of IL-17A and IL-22, in response to bacterial entry into the intestinal mucosa.

The Capsule-Encoding viaB Locus Reduces Intestinal Inflammation by a Salmonella Pathogenicity Island 1-Independent Mechanism

ABSTRACT Salmonella enterica serotype Typhimurium elicits acute neutrophil influx in the human intestinal mucosa within 1 or 2 days after infection, resulting in inflammatory diarrhea. In contrast, no overt symptoms are observed within the first 1 or 2 weeks after infection with S. enterica serotype Typhi. Here we show that introduction of the capsule-encoding viaB locus of serotype Typhi reduced the ability of serotype Typhimurium to elicit acute intestinal inflammation in a streptomycin-pretreated mouse model. Serotype Typhimurium requires a functional invasion-associated type III secretion system (type III secretion system 1 [T3SS-1]) to elicit cecal inflammation within 48 h after infection of streptomycin-pretreated mice, and the presence of the viaB locus reduced its invasiveness for human intestinal epithelial cells in vitro. However, a reduced activity of T3SS-1 could not account for the ability of the viaB locus to attenuate cecal inflammation, because introduction of the viaB locus into an invasion-deficient serotype Typhimurium strain (invA mutant) resulted in a significant reduction of pathology and inflammatory cytokine expression in the cecum 5 days after infection of mice. We conclude that a T3SS-1-independent mechanism contributes to the ability of the viaB locus to reduce intestinal inflammation.

The mucosal inflammatory response to non-typhoidal Salmonella in the intestine is blunted by IL-10 during concurrent malaria parasite infection

Coinfection can markedly alter the response to a pathogen, thereby changing its clinical presentation. For example, non-typhoidal Salmonella (NTS) serotypes are associated with gastroenteritis in immunocompetent individuals. In contrast, individuals with severe pediatric malaria can develop bacteremic infections with NTS, during which symptoms of gastroenteritis are commonly absent. Here we report that, in both a ligated ileal loop model and a mouse colitis model, malaria parasites caused a global suppression of gut inflammatory responses and blunted the neutrophil influx that is characteristic of NTS infection. Further, malaria parasite infection led to increased recovery of Salmonella enterica serotype Typhimurium from the draining mesenteric lymph node (MLN) of mice. In the mouse colitis model, blunted intestinal inflammation during NTS infection was independent of anemia but instead required parasite-induced synthesis of interleukin (IL)-10. Blocking of IL-10 in coinfected mice reduced dissemination of S. Typhimurium to the MLN, suggesting that induction of IL-10 contributes to development of disseminated infection. Thus IL-10 produced during the immune response to malaria in this model contributes to suppression of mucosal inflammatory responses to invasive NTS, which may contribute to differences in the clinical presentation of NTS infection in the setting of malaria.

Inflammation-associated alterations to the intestinal microbiota reduce colonization resistance against non-typhoidal Salmonella during concurrent malaria parasite infection

Childhood malaria is a risk factor for disseminated infections with non-typhoidal Salmonella (NTS) in sub-Saharan Africa. While hemolytic anemia and an altered cytokine environment have been implicated in increased susceptibility to NTS, it is not known whether malaria affects resistance to intestinal colonization with NTS. To address this question, we utilized a murine model of co-infection. Infection of mice with Plasmodium yoelii elicited infiltration of inflammatory macrophages and T cells into the intestinal mucosa and increased expression of inflammatory cytokines. These mucosal responses were also observed in germ-free mice, showing that they are independent of the resident microbiota. Remarkably, P. yoelii infection reduced colonization resistance of mice against S. enterica serotype Typhimurium. Further, 16S rRNA sequence analysis of the intestinal microbiota revealed marked changes in the community structure. Shifts in the microbiota increased susceptibility to intestinal colonization by S. Typhimurium, as demonstrated by microbiota reconstitution of germ-free mice. These results show that P. yoelii infection, via alterations to the microbial community in the intestine, decreases resistance to intestinal colonization with NTS. Further they raise the possibility that decreased colonization resistance may synergize with effects of malaria on systemic immunity to increase susceptibility to disseminated NTS infections.

Nasal Adenocarcinoma and Secondary Chronic Sinusitis in a Hyacinth Macaw (Anodorhynchus hyacinthinus)

Abstract An adult male hyacinth macaw (Anodorhynchus hyacinthinus) that presented for acute onset nasal discharge and dyspnea had purulent discharge from the right naris and serosanguineous discharge from the left naris on physical examination. Results of a complete blood count revealed severe leukocytosis with a mature heterophilia. Computed tomography scans showed a large amount of soft-tissue attenuating material within the infraorbital sinus and associated diverticula. Aerobic culture results of the nasal discharge showed a mixed population of Staphylococcus intermedius and Pasteurella species, including Pasteurella pneumotropica; all isolated bacteria were susceptible to enrofloxacin. Clinical signs did not resolve over the course of 9 weeks of antibiotic treatment. The macaw died after cardiopulmonary arrest while hospitalized. At necropsy, a 2 × 2 × 3–cm firm, tan, friable, space-occupying mass surrounded by a thick exudate was present in the left preorbital diverticulum of the infraorbital sinus. The cranioventral one-third of the trachea contained a 4 × 0.5–cm white-yellow plaque. On histologic examination, the sinus mass was diagnosed as a nasal adenocarcinoma, and the tracheal plaque was caused by fungal infection, most likely with an Aspergillus species.

Clinical Characteristics of Hepatocellular Carcinoma in 19 cats from a Single Institution (1980-2013)

Clinical features of feline hepatocellular carcinoma (HCA) have been poorly characterized. In this retrospective study, we describe the signalment, clinical features, clinicopathologic parameters, imaging characteristics, hepatic mass size and lobe distribution, concurrent disorders, and survival in 19 cats with HCA. HCA is a rare neoplasm in elderly cats often associated with weight loss, hyporexia, and increased hepatic transaminase activities. Concurrent disorders (e.g., hyperthyroidism, inflammatory bowel disease, cholangiohepatitis, copper-associated hepatopathy) often confounded interpretation of clinical and clinicopathologic findings; 42% of HCA were incidentally identified. Although an abdominal mass was palpated in only 21% of cats, many cats had masses identified on ultrasonographic imaging with 47% having lesions >4 cm. Tumors were nearly equally distributed between right and left liver lobes, and two cats had HCA in multiple liver lobes. Median survival of eight cats diagnosed antemortem was 1.7 (0.6 to 6.5) yr. Median survival of six cats undergoing HCA surgical resection was 2.4 (1.0 to 6.5) yr with two cats still alive at time of manuscript submission. Following surgical resection, one cat treated with carboplatin survived 4 yr. Two cats with HCA diagnosed antemortem without surgical resection survived for 0.6 and 1 yr.