An Outer Membrane Vesicle-Adjuvanted Oral Vaccine Protects Against Lethal, Oral Salmonella Infection.

Non-typhoidal salmonellosis, caused by Salmonella enterica serovar Typhimurium is a common fecal-oral disease characterized by mild gastrointestinal distress resulting in diarrhea, chills, fever, abdominal cramps, head and body aches, nausea, and vomiting. Increasing incidences of antibiotic resistant invasive non-typhoidal Salmonella infections makes this a global threat requiring novel treatment strategies including next-generation vaccines. The goal of the current study was to formulate a novel vaccine platform against Salmonella infection that could be delivered orally. To accomplish this, we created a Salmonella-specific vaccine adjuvanted with Burkholderia pseudomallei outer membrane vesicles (OMVs). We show that adding OMVs to a heat-killed oral Salmonella vaccine (HKST + OMVs) protects against a lethal, oral challenge with Salmonella. Further, we show that opsonizing anti-Salmonella antibodies are induced in response to immunization and that CD4 T cells and B cells can be induced when OMVs are used as the oral adjuvant. This study represents a novel oral vaccine approach to combatting the increasing problem of invasive Salmonella infections.

Recent Advances in the Pursuit of an Effective Acinetobacter baumannii Vaccine.

Acinetobacter baumannii has been a major cause of nosocomial infections for decades. The absence of an available vaccine coupled with emerging multidrug resistance has prevented the medical community from effectively controlling this human pathogen. Furthermore, the ongoing pandemic caused by SARS-CoV-2 has increased the risk of hospitalized patients developing ventilator-associated pneumonia caused by bacterial opportunists including A. baumannii. The shortage of antibiotics in the development pipeline prompted the World Health Organization to designate A. baumannii a top priority for the development of new medical countermeasures, such as a vaccine. There are a number of important considerations associated with the development of an A. baumannii vaccine, including strain characteristics, diverse disease manifestations, and target population. In the past decade, research efforts have revealed a number of promising new immunization strategies that could culminate in a safe and protective vaccine against A. baumannii. In this review, we highlight the recent progress in the development of A. baumannii vaccines, discuss potential challenges, and propose future directions to achieve an effective intervention against this human pathogen.

Arcanobacterium haemolyticum Utilizes Both Phospholipase D and Arcanolysin To Mediate Its Uptake into Nonphagocytic Cells.

Arcanobacterium haemolyticum is an emerging human pathogen that causes pharyngitis and wound infections. A few studies have suggested that A. haemolyticum is able to induce its uptake into nonphagocytic epithelial cells, but the bacterial factors associated with host cell invasion and the host cell processes involved have yet to be studied. We investigated how two A. haemolyticum virulence factors, arcanolysin (ALN) and phospholipase D (PLD), affect the ability of the bacteria to adhere to and subsequently invade Detroit 562 pharyngeal epithelial cells. The sphingomyelinase activity of phospholipase D was necessary to increase bacterial adherence, while the absence of a functional arcanolysin had no effect on A. haemolyticum adherence but did lead to a decrease in A. haemolyticum invasion into Detroit 562 cells. Because of the known roles of cholesterol-dependent cytolysins in disrupting calcium gradients and inducing F-actin-mediated bacterial internalization, we sought to determine whether ALN and PLD played a similar role in the ability of A. haemolyticum to invade nonphagocytic cells. Elimination of extracellular calcium and inhibition of the Arp2/3 complex or F-actin polymerization also caused a decrease in the ability of A. haemolyticum to invade Detroit 562 cells. Overall, our findings suggest that A. haemolyticum utilizes phospholipase D primarily for adherence and utilizes arcanolysin primarily for invasion into Detroit 562 cells in a process dependent on extracellular calcium and F-actin polymerization. Our work marks the first insight into how the individual activities of arcanolysin and phospholipase D affect A. haemolyticum host-pathogen interactions using the biologically relevant Detroit 562 cell line.

Arcanobacterium haemolyticum Phospholipase D Enzymatic Activity Promotes the Hemolytic Activity of the Cholesterol-Dependent Cytolysin Arcanolysin.

Arcanolysin, produced by the human pathogen Arcanobacterium haemolyticum, is a cholesterol-dependent cytolysin. To mediate the pore-formation process, arcanolysin is secreted by A. haemolyticum and then must interact with cholesterol embedded within a host membrane. However, arcanolysin must compete with membrane components, such as the phospholipid sphingomyelin, to interact with cholesterol and form pores. Cholesterol forms transient hydrogen bonds with the extracellular portion of sphingomyelin, shielding cholesterol from extracellular factors, including arcanolysin. A. haemolyticum also produces a sphingomyelin-specific phospholipase D, which removes the choline head from sphingomyelin, leaving cyclic-ceramide phosphate and eliminating the potential for cholesterol sequestration. We hypothesized that the enzymatic activity of phospholipase D decreases sphingomyelin-mediated cholesterol sequestration and increases cholesterol accessibility for arcanolysin. Using purified arcanolysin and phospholipase D, we demonstrate that the enzymatic activity of phospholipase D is necessary to promote arcanolysin-mediated hemolysis in both time- and concentration-dependent manners. Phospholipase D promotion of arcanolysin-mediated cytotoxicity was confirmed in Detroit 562 epithelial cells. Furthermore, we determined that incubating phospholipase D with erythrocytes corresponds with an increase in the amount of arcanolysin bound to host membranes. This observation suggests that phospholipase D promotes arcanolysin-mediated cytotoxicity by increasing the ability of arcanolysin to bind to a host membrane.

Multiple intraintestinal signals coordinate the regulation of Vibrio cholerae virulence determinants.

Vibrio cholerae is a Gram-negative motile bacterium capable of causing fatal pandemic disease in humans via oral ingestion of contaminated water or food. Within the human intestine, the motile vibrios must evade the innate host defense mechanisms, penetrate the mucus layer covering the small intestine, adhere to and multiply on the surface of the microvilli and cause disease via the action of cholera toxin. The explosive diarrhea associated with V. cholerae intestinal colonization leads to dissemination of the vibrios back into the environment to complete this phase of the life cycle. The host phase of the vibrio life cycle is made possible via the concerted action of a signaling cascade that controls the synthesis of V. cholerae colonization determinants. These virulence proteins are coordinately synthesized in response to specific host signals that are still largely undefined. A more complete understanding of the molecular events involved in the V. cholerae recognition of intraintestinal signals and the subsequent transcriptional response will provide important information regarding how pathogenic bacteria establish infection and provide novel methods for treating and/or preventing bacterial infections such as Asiatic cholera. This review will summarize what is currently known in regard to host intraintestinal signals that inform the complex ToxR regulatory cascade in order to coordinate in a spatial and temporal fashion virulence protein synthesis within the human small intestine.