Acinetobacter baumannii and other Acinetobacter species are a growing public health concern because they commonly cause healthcare-associated infections. Acinetobacter spp. are often multi-drug resistant, leading to significant morbidity and mortality. Therefore, understanding the reservoirs for Acinetobacter spp. in healthcare settings is particularly important to infection control. Acinetobacter spp. are known to asymptomatically colonize the human gut, skin, and respiratory flora, and recent work has found the prevalence of Acinetobacter colonization is high in patients in healthcare environments, such as hospitals and long-term care facilities. Patients colonized by A. baumannii are at increased risk of developing clinical infections, including by the same strains. Additionally, multiple studies report transmission of A. baumannii strains to other patients in the same healthcare setting. Recent mechanistic studies on A. baumannii gut colonization have found that the microbiota, host immunity, and diet contribute to colonization resistance in animal models. Here, we review the epidemiologic evidence for asymptomatic colonization as a clinically important reservoir for A. baumannii and examine the nascent study of the molecular mechanisms of Acinetobacter colonization. We discuss future prospects for understanding Acinetobacter colonization that may help develop strategies to support colonization resistance and prevent infections.

Macrophages employ multiple strategies to dispatch phagocytosed microbes, including bombardment with toxic compounds and radicals, or the deprivation of key nutrients. The lethality of macrophages depends on their ability to regulate levels of microbicidal effectors and proper targeting of these effectors to the phagosome, where many intracellular pathogens are detained. Mitochondria play important roles in establishing an antimicrobial phagosomal environment in macrophages, playing roles in both the production and targeting of these effectors. In response to infection, the mitochondrial network undergoes dramatic structural and functional perturbations that modulate macrophage microbicidal potential. As central coordinators of cellular metabolism, mitochondria control the abundance of metabolites that can either be toxic to ingested microbes or necessary for their growth. Mitochondria are also prolific producers of reactive oxygen species (ROS), which play direct roles in killing microbes and serve as signaling molecules to coordinate antimicrobial responses. While mitochondria are also known to play extensive roles in regulating cytokine production and proinflammatory responses, this review will focus on highlighting mechanisms by which mitochondria manipulate the macrophage microbicidal milieu.

Aspergillus fumigatus is the primary etiologic agent of pulmonary mold infections, the most severe of which is invasive pulmonary aspergillosis (IPA). As the currently approved antifungal drugs are threatened by growing resistance rates and host toxicity, novel therapeutic options are urgently needed. Our recent work suggests that septation inhibition may be one such strategy, as aseptate strains of A. fumigatus are hypersusceptible to the echinocandin antifungal drugs and display significantly reduced virulence in animal models of IPA. In the current study, we expand upon our understanding of the anillin-like protein, Bud4, and Rho GTPase, Rho4, which regulate septation in Aspergillus nidulans and A. fumigatus, respectively. Whereas AnBud4 is not essential for septation in A. nidulans, deletion of the A. fumigatus ortholog (∆bud4) resulted in complete loss of septation. A similar phenotype was also observed in ∆rho4, consistent with previous studies. Supporting a critical role for hyphal septation in the establishment of invasive aspergillosis, we find that ∆bud4 and ∆rho4 display avirulence in a mouse model of IPA characterized by a loss of tissue-invasive growth. Furthermore, both deletion mutations were able to convert anti-Aspergillus echinocandin activity from static to cidal. Through titratable bud4 expression using a doxycycline-repressible promoter system, we also find that even partial repression of septation significantly increases echinocandin susceptibility. Taken together, these studies support a therapeutic strategy which could incorporate a septation inhibitor in combination with echinocandins to reduce tissue-invasive growth and improve antifungal susceptibility.

Chlamydia species are obligate intracellular bacteria with a biphasic developmental cycle. They cause trachoma, urogenital tract infection, community-acquired pneumonia, and zoonotic respiratory disease. A common feature of these conditions is not simply infection but the combination of persistence, recurrent inflammation, and tissue injury. Neutrophils arrive early at infected tissues and can initiate phagocytosis, degranulation, reactive oxygen species production, and neutrophil extracellular trap formation through pattern-recognition receptors, complement receptors, and Fc receptors. That response is not uniformly beneficial. When recruitment is excessive or poorly resolved, neutrophils can amplify inflammation, weaken epithelial barriers, and contribute to fibrosis and chronic immunopathology. Here, we review Chlamydia-neutrophil interactions as a staged biological process: recognition, recruitment, effector activation, apoptosis delay, inflammatory amplification, negative feedback, and tissue injury. Particular attention is given to TLRs, NF-κB, MAPK, PI3K/Akt, the NLRP3 inflammasome, mitochondrial reactive oxygen species, and IL-10/JAK-STAT-related feedback. We distinguish evidence obtained directly from neutrophils from evidence inferred from epithelial cells, macrophages, PBMCs, animal tissues, or other pathogen systems. This distinction is essential because mechanisms supported in one cell type or one chlamydial species cannot be transferred automatically to another. A more precise view of this network may refine current models of chlamydia-host interaction and pathogen persistence and help frame future studies on stage-specific immunomodulation.

The commensal bacteria of the intestinal microbiota inhabit one of the most densely populated and dynamic microbial ecosystems known, where fluctuations in nutrient availability, host-derived stresses, and microbial competition require highly coordinated regulatory responses to maintain colonization and fitness. As a stable, homeostatic microbiota performs essential functions for the host, understanding how commensals adapt to the fluctuating intestinal environment is important for maintaining intestinal homeostasis and informing the development of new microbiota-targeted therapeutics. Throughout diverse bacterial species, rapid adaptation to environmental perturbations is mediated in part by small regulatory RNAs (sRNAs). While most sRNA research has focused on a limited number of model organisms, particularly bacterial pathogens, recent studies have uncovered important roles for these noncoding RNAs in promoting the fitness and resilience of gut commensals. Advances in differential RNA sequencing, comparative genomics, and functional genetics have revealed diverse repertoires of sRNAs in prominent gut symbionts, with regulatory roles in nutrient acquisition, stress responses, and host colonization. This minireview highlights recent advances and outlines future directions to further elucidate sRNA-mediated regulation as a pillar of commensal resilience in the mammalian gut.

Clostridioides difficile infection (CDI) is a common cause of antibiotic-associated diarrhea and arises from disruption of the resident microbiota in the gastrointestinal tract. Despite being a major cause of morbidity and mortality worldwide, CDI presents as a wide spectrum of clinical manifestations, ranging from completely asymptomatic infection to severe and fulminant colitis. Recent research has revealed that CDI disease severity is influenced by the complex ecosystem of the gut, involving interactions between C. difficile, the host, and the resident gut microbiota. This review will summarize the current state of the field in understanding how the balance of interactions in the gut alters CDI disease severity using clinical data and mouse models of infection. In addition, external factors influencing these interactions, such as diet and pharmaceutical drugs, will be discussed. Ultimately, a comprehensive understanding of the ecological factors influencing patient outcome will be critical for the design of novel therapeutics for primary and recurrent CDI.

Previous research demonstrated that Campylobacter jejuni encodes a heme utilization system that facilitates heme-dependent growth under iron-limiting conditions, and that transcription of this system is induced during human infection. Despite these observations, it remained unknown whether the heme transport system is required for colonization and disease in a susceptible host. To address this, we created individual non-polar deletion mutants of each component of the heme transport system and examined their ability to promote heme-dependent growth and iron uptake. From this work, we found that only the heme receptor, ChuA, was required for heme-dependent growth and iron acquisition, which supports the earlier work of another group. Furthermore, we examined whether intestinal colonization, immune activation, and pathology were altered during infection with these mutants. After establishing that elevated heme and chuABCD expression occur during C. jejuni infection of IL-10[-/-] mice, we found that a mutant of the heme receptor, ChuA, exhibited significantly reduced colonization of the colon. In addition, we found that neutrophil and circulating monocyte recruitment were significantly reduced in the colon during infection with the ChuA mutant, but that populations of self-maintaining tissue macrophages remained high. Loss of ChuA reduced colonic colonization and was accompanied by diminished innate immune cell recruitment and intestinal pathology. Together, these findings identify ChuA-dependent heme acquisition as a key determinant of efficient colonic colonization and the associated inflammatory disease.

Urinary tract infections (UTIs) are a significant public health burden that impact millions of people every year and are highly prevalent among hospital-acquired infections. Klebsiella pneumoniae is globally the second most common cause of UTIs after uropathogenic Escherichia coli (UPEC). Thus far, the molecular mechanisms underlying pathogenesis are better understood in UPEC than in K. pneumoniae. UPEC is known to have virulence phenotypes such as fimbrial adhesion and evasion of complement-mediated killing. In other infection types, K. pneumoniae fitness has been associated with mucoidy and diverse capsular serotypes. To establish K. pneumoniae virulence phenotypes contributing to UTI, we examined how environmental cues regulate urovirulence-associated phenotypes in clinical K. pneumoniae UTI strains. These phenotypes included capsular polysaccharide properties, hemagglutination, serum resistance, adherence to bladder epithelial cells, and in vivo fitness. We found that clinical K. pneumoniae UTI isolates' phenotypes are highly heterogeneous and can change in response to human urine. Despite K. pneumoniae clinical isolates presenting heterogeneous fitness properties, all similarly colonize the urinary tract. These results suggest that additional fitness factors contribute to K. pneumoniae uropathogenesis. Identifying these shared fitness factors will provide mechanistic insights into Klebsiella uropathogenesis and reveal candidate therapeutic targets.

It is well established that within a bacterial species, genotypic and phenotypic variation leads to differences in virulence, antibiotic susceptibility, and metabolism. However, there is also increasing evidence that often multiple variants of the same species co-inhabit the same site in the human body, both during health and disease. This within-host microdiversity can develop over time for pathogens in chronic infections and has been observed for many members of the microbiota, for instance in the gut and on the skin. Research is just beginning to uncover how this within-host intra-species diversity develops and why it matters. Intra-species diversity is largely stable over time, with occasional selective sweeps of new genotypes, and the overall level of diversity is modulated by both microbe-microbe and microbe-host interactions. Furthermore, data indicate that within-host microdiversity can impact the stability of bacterial communities, their antimicrobial resistance, and the efficacy of probiotics and microbiota transplants. Here, we describe the evolutionary and ecological processes that promote high or low within-host microdiversity and the extent of this diversity in the human microbiota and in infections. In addition, we consider how microdiversity within individuals shapes the functional role of the human microbiota and the progression of disease. Together, within-host microdiversity is a newly developing way to think about the diversity of human-associated microbes, with implications in how we treat chronic infections and manipulate our microbiota to promote health.

In polymicrobial infections, how the host recognizes and responds to pathogens influences which species will persist to cause chronic infections. The human respiratory tract is a common anatomical site for viral-bacterial co-infections, where primary viral infections predispose to secondary bacterial infections, leading to increased morbidity and mortality. Additionally, co-infections are disproportionately prevalent in people with chronic lung diseases, such as chronic obstructive pulmonary disease and cystic fibrosis. We previously reported that primary viral infections and antiviral interferon (IFN) signaling stimulate Pseudomonas aeruginosa biofilm formation on airway epithelial cells (AECs). IFN signaling induces aerobic glycolysis in AECs and generates lactate as a cellular byproduct. Given that innate immune systems play an integral role in co-infection dynamics, we investigated the role of host-secreted metabolites (i.e., lactate) on innate immune cell activity during respiratory co-infections. We found that exposure to the apical secretions from IFNβ-treated AECs significantly compromised macrophage antibacterial activity, with the soluble metabolite lactate playing an important role. Macrophages used monocarboxylate transporters (MCTs) and G-protein receptors to transport and/or sense lactate, respectively, and this exposure to lactate diminished their bacterial-killing activity in a time exposure-dependent manner. Lactate exposure particularly reprogrammed macrophage cellular metabolism toward a metabolically uncoupled state by increasing oxidative phosphorylation, fatty acid oxidation, and downregulating the proinflammatory antimicrobial functions. Collectively, these findings provide insights into metabolites as complex regulators of trans-kingdom interactions and epithelial-macrophage crosstalk during respiratory co-infections.

Mycoplasma gallisepticum (MG) causes chronic respiratory disease, which is thought to be contributed to by its capacity to evade the immune response of infected birds. It possesses multiple copies of the genes encoding the mycoplasma immunoglobulin binding-protease (MIB-MIP) system, which binds and cleaves avian immunoglobulins (Igs), but also encodes homologs of the Ig-binding Protein-M of M. genitalium and M. pneumoniae and the Ig-cleaving CysP of M. synoviae. However, the interactions between these homologs, their host-specificity, and their interplay with the MIB-MIP system have not been defined. Here, the putative Protein-M and CysP in MG were expressed as recombinant glutathione-S-transferase (GST)-fusion proteins, functionally characterized, and their interactions with different MIB-MIP system components, as well as their impact on antibody-dependent complement-mediated killing of MG, examined. GST-Protein-M bound avian and mammalian immunoglobulins, but did not interact with the MIP proteases. GST-CysP selectively cleaved free chicken IgY and IgY complexed with GST-MIB or GST-Protein-M. GST-Protein-M, GST-CysP, and GST-MIB-MIP proteins all bound ATP, but only GST-CysP had ATPase activity. Sera from MG-infected chickens contained antibodies against a subset of MIP proteins, but not antibodies against Protein-M or CysP. The capacity of rabbit anti-MG antibodies to enhance complement-mediated killing of MG was inhibited by GST-MIB-MIP and GST-Protein-M. GST-CysP could not cleave rabbit IgG and had no detectable effect in the antibody-dependent complement-mediated killing assays. Collectively, these findings establish that Protein-M and CysP function as additional Ig-targeting systems in M. gallisepticum alongside the MIB-MIP system through distinct but complementary mechanisms, enhancing immune evasion and persistence of this important pathogen.

Amino acids are the fundamental building blocks of proteins. Dietary proteins and endogenous protein turnover are the main sources of amino acids in the body. Over the past decade, our understanding of the role of the gut microbiota in amino acid metabolism has expanded considerably. The gut microbiota metabolizes dietary amino acids, regulates intestinal amino acid bioavailability, and maintains host nutrient homeostasis. Besides nutrient regulation, emerging evidence has revealed that amino acid availability in the gut also plays a crucial role in modulating enteric infections. Amino acids can influence infection outcomes both directly, by affecting the virulence of enteric pathogens, and indirectly, by shaping host immune responses. While most studies have focused on bacterial metabolism of amino acids, recent findings highlight important contributions from non-bacterial members of the gut microbiota, including fungi and protists. Both fungi and protists can shape amino acid availability in the gut and, consequently, infection outcomes. In this review, we integrate current knowledge on how diet and amino acid metabolism, mediated by both bacterial and non-bacterial communities, collectively influence enteric infection dynamics.

Staphylococcus aureus deploys an arsenal of secreted proteases to remodel the host environment during infection. While traditional substrate-by-substrate characterization has revealed critical host proteins under protease modulation, these conventional strategies cannot reveal the full scope of proteolytic activity occurring at the host-pathogen interface. Degradomic and N-terminomic technologies have begun to bridge this gap by enabling an unbiased, global mapping of protease-generated neo-N-termini across complex host tissues and cell proteomes. Recent work applying these tools to define discrete S. aureus protease pathodegradomes has propelled the field toward a system-level understanding of proteolytic activity. In this minireview, we highlight how terminal amine isotopic labeling of substrates and a newer methodology, terminal amine guanidination of substrates-charge reversal, have captured a deeper understanding of the V8 protease and how this enzyme can dismantle host immune defenses and engender disease. These techniques offer a powerful framework for the future in characterizing the substrate repertoires of other S. aureus secreted proteases, such as aureolysin, staphopain A and B, and the Spls. Furthermore, these advances demonstrate the applicability of degradomic profiling to diverse infection niches, revealing previously unrecognized roles for proteolytic activity in varying host contexts. Finally, we consider how this comprehensive understanding of S. aureus protease activity can inform clinical applications such as protease inhibitor design, host-directed therapies, and pathodegradome-based diagnostics.