Role of the ESCRT‐III complex in controlling integrity of the Salmonella‐containing vacuole

Intracellular pathogens need to establish specialised niches for survival and proliferation in host cells. The enteropathogen Salmonella enterica accomplishes this by extensive reorganisation of the host endosomal system deploying the SPI2‐encoded type III secretion system (SPI2‐T3SS). Fusion events of endosomal compartments with the Salmonella‐containing vacuole (SCV) form elaborate membrane networks within host cells enabling intracellular nutrition. However, which host compartments exactly are involved in this process and how the integrity of Salmonella‐modified membranes is accomplished are not fully resolved. An RNA interference knockdown screen of host factors involved in cellular logistics identified the ESCRT (endosomal sorting complex required for transport) system as important for proper formation and integrity of the SCV in infected epithelial cells. We demonstrate that subunits of the ESCRT‐III complex are specifically recruited to the SCV and membrane network. To investigate the role of ESCRT‐III for the intracellular lifestyle of Salmonella, a CHMP3 knockout cell line was generated. Infected CHMP3 knockout cells formed amorphous, bulky SCV. Salmonella within these amorphous SCV were in contact with host cell cytosol, and the attenuation of an SPI2‐T3SS‐deficient mutant strain was partially abrogated. ESCRT‐dependent endolysosomal repair mechanisms have recently been described for other intracellular pathogens, and we hypothesise that minor damages of the SCV during bacterial proliferation are repaired by the action of ESCRT‐III recruitment in Salmonella‐infected host cells.     

Esterification of cholesterol by a type III secretion effector during intracellular Salmonella infection

Survival of Salmonella typhimurium within a vacuole in host cells depends on secreted virulence factors encoded by the Salmonella pathogenicity island 2 (SPI-2). High levels of cholesterol are detected at the Salmonella -containing vacuole (SCV). Here we show that the SPI-2 effector SseJ esterifies cholesterol in vitro , in cells and during infection. Intracellular infections with wild-type as compared with Δ sseJ bacteria led to higher levels of cholesterol ester production in HeLa cells and RAW macrophages and were shown to increase levels of lipid droplets (structures enriched in cholesterol esters). Ectopic expression of SseJ reduced cholesterol levels in cellular membranes and antagonized a major membrane activity of a second bacterial effector known to be important to the stability of the SCV. Previous studies in mouse models of infection have established a virulence defect in Δ sseJ bacteria and have suggested a role for SseJ in regulating SCV dynamics. Our data indicating the molecular activity of SseJ suggest that cholesterol and its esterification at the SCV are functionally important for intracellular bacterial survival.     

SopB‐ and SifA‐dependent shaping of the Salmonella‐containing vacuole proteome in the social amoeba Dictyostelium discoideum

The ability of Salmonella to survive and replicate within mammalian host cells involves the generation of a membranous compartment known as the Salmonella‐containing vacuole (SCV). Salmonella employs a number of effector proteins that are injected into host cells for SCV formation using its type‐3 secretion systems encoded in SPI‐1 and SPI‐2 (T3SS‐1 and T3SS‐2, respectively). Recently, we reported that S. Typhimurium requires T3SS‐1 and T3SS‐2 to survive in the model amoeba Dictyostelium discoideum. Despite these findings, the involved effector proteins have not been identified yet. Therefore, we evaluated the role of two major S. Typhimurium effectors SopB and SifA during D. discoideum intracellular niche formation. First, we established that S. Typhimurium resides in a vacuolar compartment within D. discoideum. Next, we isolated SCVs from amoebae infected with wild type or the ΔsopB and ΔsifA mutant strains of S. Typhimurium, and we characterised the composition of this compartment by quantitative proteomics. This comparative analysis suggests that S. Typhimurium requires SopB and SifA to modify the SCV proteome in order to generate a suitable intracellular niche in D. discoideum. Accordingly, we observed that SopB and SifA are needed for intracellular survival of S. Typhimurium in this organism. Thus, our results provide insight into the mechanisms employed by Salmonella to survive intracellularly in phagocytic amoebae.     

Presence of SopE and mode of infection result in increased Salmonella‐containing vacuole damage and cytosolic release during host cell infection by Salmonella enterica

Salmonella enterica serovar Typhimurium (STM) is an invasive, facultative intracellular pathogen that has evolved sophisticated molecular mechanisms to establish an intracellular niche within a specialised vesicular compartment, the Salmonella‐containing vacuole (SCV). The loss of the SCV and release of STM into the cytosol of infected host cells was observed, and a bimodal intracellular lifestyle of STM in the SCV versus life in the cytosol is currently discussed. We set out to investigate the parameters affecting SCV integrity and cytosolic release. A fluorescent protein‐based cytosolic reporter approach was established to quantify, time‐resolved, and on a single cell level, the release of STM into the cytosol of host cells. We observed that the extent of SCV damage and cytosolic release is highly dependent on experimental conditions such as multiplicity of infection, type of host cell line, and STM strain background. Trigger invasion mediated by the Salmonella Pathogenicity Island 1‐encoded type III secretion system (SPI1‐T3SS) and its effector proteins promoted cytosolic release, whereas cytosolic bacteria were rarely observed if entry was mediated by zipper invasion. Presence of SPI1‐T3SS effector SopE was identified as major factor for damage of the SCV in the early phase after STM invasion and sopE‐expressing strains showed higher levels of cytosolic release.     

Salmonella SPI1 effector SipA persists after entry and cooperates with a SPI2 effector to regulate phagosome maturation and intracellular replication

Salmonellae employ two type III secretion systems (T3SSs), SPI1 and SPI2, to deliver virulence effectors into mammalian cells. SPI1 effectors, including actin-binding SipA, trigger initial bacterial uptake, whereas SPI2 effectors promote subsequent replication within customized Salmonella-containing vacuoles (SCVs). SCVs sequester actin filaments and subvert microtubule-dependent motors to migrate to the perinuclear region. We demonstrate that SipA delivery continues after Salmonella internalization, with dosage being restricted by host-mediated degradation. SipA is exposed on the cytoplasmic face of the SCV, from where it stimulates bacterial replication in both nonphagocytic cells and macrophages. Although SipA is sufficient to target and redistribute late endosomes, during infection it cooperates with the SPI2 effector SifA to modulate SCV morphology and ensure perinuclear positioning. Our findings define an unexpected additional function for SipA postentry and reveal precise intracellular communication between effectors deployed by distinct T3SSs underlying SCV biogenesis.     

Inhibition of the PtdIns(5) kinase PIKfyve disrupts intracellular replication of Salmonella

3‐phosphorylated phosphoinositides (3‐PtdIns) orchestrate endocytic trafficking pathways exploited by intracellular pathogens such as Salmonella to gain entry into the cell. To infect the host, Salmonellae subvert its normal macropinocytic activity, manipulating the process to generate an intracellular replicative niche. Disruption of the PtdIns(5) kinase, PIKfyve, be it by interfering mutant, siRNA‐mediated knockdown or pharmacological means, inhibits the intracellular replication of Salmonella enterica serovar typhimurium in epithelial cells. Monitoring the dynamics of macropinocytosis by time‐lapse 3D (4D) videomicroscopy revealed a new and essential role for PI(3,5)P₂ in macropinosome‐late endosome/lysosome fusion, which is distinct from that of the small GTPase Rab7. This PI(3,5)P₂‐dependent step is required for the proper maturation of the Salmonella‐containing vacuole (SCV) through the formation of Salmonella‐induced filaments (SIFs) and for the engagement of the Salmonella pathogenicity island 2‐encoded type 3 secretion system (SPI2‐T3SS). Finally, although inhibition of PIKfyve in macrophages did inhibit Salmonella replication, it also appears to disrupt the macrophage's bactericidal response.     

Involvement of TIP60 acetyltransferase in intracellular <Emphasis Type="Italic">Salmonella</Emphasis>replication

Background  

Salmonella enterica is a facultative intracellular pathogen that replicates within a membrane-bound compartment termed Salmonella containing vacuole (SCV). The biogenesis of SCV requires Salmonella type III protein secretion/translocation system and their effector proteins which are translocated into host cells to exploit the vesicle trafficking pathways. SseF is one of these effectors required for SCV formation and Intracellular Salmonella replication through unknown mechanisms.     

The Salmonella Typhimurium effector protein SopE transiently localizes to the early SCV and contributes to intracellular replication

Salmonella enterica serovar Typhimurium (S. Tm) is a facultative intracellular pathogen that induces entry into non‐phagocytic cells by a Type III secretion system (TTSS) and cognate effector proteins. Upon host cell entry, S. Tm expresses a second TTSS and subverts intracellular trafficking to create a replicative niche – the Salmonella‐containing vacuole (SCV). SopE, a guanidyl exchange factor (GEF) for Rac1 and Cdc42, is translocated by the TTSS‐1 upon host cell contact and promotes entry through triggering of actin‐dependent ruffles. After host cell entry, the bulk of SopE undergoes proteasomal degradation. Here we show that a subfraction is however detectable on the nascent SCV membrane up to ～ 6 h post infection. Membrane localization of SopE and the closely related SopE2 differentially depend on the Rho‐GTPase‐binding GEF domain, and to some extent involves also the unstructured N‐terminus. SopE localizes transiently to the early SCV, dependent on continuous synthesis and secretion by the TTSS‐1 during the intracellular state. Mutant strains lacking SopE or SopE2 are attenuated in early intracellular replication, while complementation restores this defect. Hence, the present study reveals an unanticipated role for SopE and SopE2 in establishing the Salmonella replicative niche, and further emphasizes the importance of entry effectors in later stages of host‐cell manipulation.     

Single cell analyses reveal distinct adaptation of typhoidal and non-typhoidal Salmonella enterica serovars to intracellular lifestyle

Salmonella enterica is a common foodborne, facultative intracellular enteropathogen. Human-restricted typhoidal S. enterica serovars Typhi (STY) or Paratyphi A (SPA) cause severe typhoid or paratyphoid fever, while many S. enterica serovar Typhimurium (STM) strains have a broad host range and in human hosts usually lead to a self-limiting gastroenteritis. Due to restriction of STY and SPA to primate hosts, experimental systems for studying the pathogenesis of typhoid and paratyphoid fever are limited. Therefore, STM infection of susceptible mice is commonly considered as model system for studying these diseases. The type III secretion system encoded by Salmonella pathogenicity island 2 (SPI2-T3SS) is a key factor for intracellular survival of Salmonella. Inside host cells, the pathogen resides within the Salmonella-containing vacuole (SCV) and induces tubular structures extending from the SCV, termed Salmonella-induced filaments (SIF). This study applies single cell analyses approaches, which are flow cytometry of Salmonella harboring dual fluorescent protein reporters, effector translocation, and correlative light and electron microscopy to investigate the fate and activities of intracellular STY and SPA. The SPI2-T3SS of STY and SPA is functional in translocation of effector proteins, SCV and SIF formation. However, only a low proportion of intracellular STY and SPA are actively deploying SPI2-T3SS and STY and SPA exhibited a rapid decline of protein biosynthesis upon experimental induction. A role of SPI2-T3SS for proliferation of STY and SPA in epithelial cells was observed, but not for survival or proliferation in phagocytic host cells. Our results indicate that reduced intracellular activities are factors of the stealth strategy of STY and SPA and facilitate systemic spread and persistence of the typhoidal Salmonella.     

A role for the Salmonella Type III Secretion System 1 in bacterial adaptation to the cytosol of epithelial cells

Salmonella enterica serovar Typhimurium is a facultative intracellular pathogen that invades the intestinal epithelium. Following invasion of epithelial cells, Salmonella survives and replicates within two distinct intracellular niches. While all of the bacteria are initially taken up into a membrane bound vacuole, the Salmonella‐containing vacuole or SCV, a significant proportion of them promptly escape into the cytosol. Cytosolic Salmonella replicates more rapidly compared to the vacuolar population, although the reasons for this are not well understood. SipA, a multi‐function effector protein, has been shown to affect intracellular replication and is secreted by cytosolic Salmonella via the invasion‐associated Type III Secretion System 1 (T3SS1). Here, we have used a multipronged microscopy approach to show that SipA does not affect bacterial replication rates per se, but rather mediates intra‐cytosolic survival and/or initiation of replication following bacterial egress from the SCV. Altogether, our findings reveal an important role for SipA in the early survival of cytosolic Salmonella.     

Syntaxin 3 SPI-2 dependent crosstalk facilitates the division of Salmonella containing vacuole

Intracellular membrane fusion is mediated by membrane-bridging complexes of soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs). SNARE proteins are one of the key players in vesicular transport. Several reports shed light on intracellular bacteria modulating host SNARE machinery to establish infection successfully. The critical SNAREs in macrophages responsible for phagosome maturation are Syntaxin 3 (STX3) and Syntaxin 4 (STX4). Reports also suggest that Salmonella actively modulates its vacuole membrane composition to escape lysosomal fusion. Salmonella containing vacuole (SCV) harbours recycling endosomal SNARE Syntaxin 12 (STX12). However, the role of host SNAREs in SCV biogenesis and pathogenesis remains unclear. Upon knockdown of STX3, we observed a reduction in bacterial proliferation, which is concomitantly restored upon the overexpression of STX3. Live-cell imaging of Salmonella-infected cells showed that STX3 localises to the SCV membranes and thus might help in the fusion of SCV with intracellular vesicles to acquire membrane for its division. We also found the interaction STX3-SCV was abrogated when we infected with SPI-2 encoded Type 3 secretion system (T3SS) apparatus mutant (STM ∆ssaV) but not with SPI-1 encoded T3SS apparatus mutant (STM ∆invC). These observations were also consistent in the mice model of Salmonella infection. Together, these results shed light on the effector molecules secreted through T3SS encoded by SPI-2, possibly involved in interaction with host SNARE STX3, which is essential to maintain the division of Salmonella in SCV and help to maintain a single bacterium per vacuole.     

Rac and Rab GTPases dual effector Nischarin regulates vesicle maturation to facilitate survival of intracellular bacteria

The intracellular pathogenic bacterium Salmonella enterica serovar typhimurium (Salmonella) relies on acidification of the Salmonella‐containing vacuole (SCV) for survival inside host cells. The transport and fusion of membrane‐bound compartments in a cell is regulated by small GTPases, including Rac and members of the Rab GTPase family, and their effector proteins. However, the role of these components in survival of intracellular pathogens is not completely understood. Here, we identify Nischarin as a novel dual effector that can interact with members of Rac and Rab GTPase (Rab4, Rab14 and Rab9) families at different endosomal compartments. Nischarin interacts with GTP‐bound Rab14 and PI(3)P to direct the maturation of early endosomes to Rab9/CD63‐containing late endosomes. Nischarin is recruited to the SCV in a Rab14‐dependent manner and enhances acidification of the SCV. Depletion of Nischarin or the Nischarin binding partners—Rac1, Rab14 and Rab9 GTPases—reduced the intracellular growth of Salmonella. Thus, interaction of Nischarin with GTPases may regulate maturation and subsequent acidification of vacuoles produced after phagocytosis of pathogens.     

Take the tube: remodelling of the endosomal system by intracellular Salmonella enterica

Salmonella enterica is a facultative intracellular pathogen residing in a unique host cell‐derived membrane compartment, termed Salmonella‐containing vacuole or SCV. By the activity of effector proteins translocated by the SPI2‐endoced type III secretion system (T3SS), the biogenesis of the SCV is manipulated to generate a habitat permissive for intracellular proliferation. By taking control of the host cell vesicle fusion machinery, intracellular Salmonella creates an extensive interconnected system of tubular membranes arising from vesicles of various origins, collectively termed Salmonella‐induced tubules (SIT). Recent work investigated the dynamic properties of these manipulations. New host cell targets of SPI2‐T3SS effector proteins were identified. By applying combinations of live cell imaging and ultrastructural analyses, the detailed organization of membrane compartments inhabited and modified by intracellular Salmonella is now available. These studies provided unexpected new details on the intracellular environments of Salmonella. For example, one kind of SIT, the LAMP1‐positive Salmonella‐induced filaments (SIF), are composed of double‐membrane tubules, with an inner lumen containing host cell cytosol and cytoskeletal filaments, and an outer lumen containing endocytosed cargo. The novel findings call for new models for the biogenesis of SCV and SIT and give raise to many open questions we discuss in this review.     

Global Impact of Salmonella Pathogenicity Island 2-secreted Effectors on the Host Phosphoproteome

During the late stages of infection, Salmonella secretes numerous effectors through a type III secretion system that is encoded within Salmonella pathogenicity island 2 (SPI2). Despite the importance of SPI2 as a major virulence factor leading to the systemic spread of the bacteria and diseases, a global view of its effects on host responses is still lacking. Here, we measured global impacts of SPI2 effectors on the host phosphorylation and protein expression levels in RAW264.7 and in HeLa cells, as macrophage and nonphagocytic models of infection. We observe that SPI2 effectors differentially modulate the host phosphoproteome and cellular processes (e.g. protein trafficking, cytoskeletal regulation, and immune signaling) in a host cell-dependent manner. Our unbiased approach reveals the involvement of many previously unrecognized proteins, including E3 ligases (HERC4, RanBP2, and RAD18), kinases (CDK, SIK3, and WNK1), and histones (H2B1F, H4, and H15), in late stages of Salmonella infection. Furthermore, from this phosphoproteome analysis and other quantitative screens, we identified HSP27 as a direct in vitro and in vivo molecular target of the only type III secreted kinase, SteC. Using biochemical and cell biological assays, we demonstrate that SteC phosphorylates multiple sites in HSP27 and induces actin rearrangement through this protein. Together, these results provide a broader landscape of host players contributing to specific processes/pathways mediated by SPI2 effectors than was previously appreciated.Type III secretion systems (T3SSs)¹ are specialized virulence factors in Gram-negative pathogens that play an important role in delivering effector proteins to host cells. Salmonella enterica employs two distinct T3SSs encoded in Salmonella pathogenicity islands 1 and 2 (SPI1 and SPI2), with numerous effectors encoded around the genome, including a small number in SPI1 and SPI2 (1). SPI1 T3SS effectors are required for the bacterial internalization by intestinal epithelial cells at early stages of infection after oral ingestion. Although Salmonella is subsequently taken up by intestinal macrophages via phagocytosis, SPI2 T3SS effectors function to promote intracellular replication. Part of the role of SPI2 effectors is to control the maturation of the membrane-enclosed, Salmonella-containing vacuole (SCV) where Salmonella survives and replicates, eventually leading to a systemic infection known as typhoid fever (2, 3).Approximately 30 effectors are known to be translocated by the SPI2 T3SS but the actions and targets of most of these effectors are largely unknown (1, 3, 4). A recent systematic study using a single mutant collection of SPI2 genes showed particular virulence factors (e.g. SpvB, SifA, and SteC) play a dominant role in replication within macrophages (5). It is known that SpvB induces cytotoxicity through its ADP-ribosyltransferase activity (6), and SifA is required for maturation of the SCV and the formation of Salmonella-induced filaments (7). SteC has been identified as the sole serine/threonine protein kinase encoded in the Salmonella genome (8), but the target substrates of this kinase within the host are not fully understood, although it has been demonstrated that SteC partially targets the MAP kinase MEK (9). Interestingly, SteC is capable of promoting assembly of an F-actin meshwork around the SCV; this is dependent on its kinase activity but does not require activation of signaling pathways through Rho-associated protein kinase (8), Cdc42, Rac, N-WASP, Scar/WAVE, and Arp2/3 (10). These host signaling proteins are the main targets of T3SS-secreted effectors from many pathogens, including the SPI1 system in Salmonella (11) and Shigella (12). Therefore, SteC is thought to manipulate actin in a unique way through phosphorylation of host protein target(s).Recent advances in high throughput measurements allow us to characterize host gene expression profiles (13) and host phosphoproteme dynamics (14) dependent on the presence of SPI1 effectors in an unbiased, comprehensive manner. However, although it is clear that SPI2 T3SS is a major virulence factor contributing to systemic infection, our knowledge of its effects on host responses is limited. In this study, we used a mass spectrometry (MS)-based quantitative proteomics approach and measured global host phosphorylation changes as well as proteome abundance altered by SPI2 effectors. Furthermore, we explore a molecular target of SPI2 effector kinase SteC by integrating the phosphoproteomics data and other quantitative proteomics screens.     

沙门菌致病岛2 Ⅲ型分泌系统研究进展

沙门菌(Salmonella)是革兰氏阴性的兼性胞内菌,可引起其广泛宿主的一系列疾病,严重时可导致全身性感染,威胁生命安全。沙门菌致病岛2(SPI2)是与沙门菌全身性感染密切相关的重要毒力基因簇,其编码的Ⅲ型分泌系统2(T3SS2)在沙门菌侵入宿主细胞后开始组装合成,经该装置分泌的多种效应蛋白对沙门菌在宿主细胞内的生存和增殖起着重要作用。近些年来,与沙门菌T3SS2相关的研究一直都是病原微生物领域关注的焦点之一。本文简要综述了SPI2的基因特征、SPI2基因表达的调控、T3SS2的结构和组成、T3SS2的效应蛋白及与T3SS2相关的疫苗研究等方面的主要研究进展。     

The Salmonella effector SteA binds phosphatidylinositol 4‐phosphate for subcellular targeting within host cells

Many bacterial pathogens use specialized secretion systems to deliver virulence effector proteins into eukaryotic host cells. The function of these effectors depends on their localization within infected cells, but the mechanisms determining subcellular targeting of each effector are mostly elusive. Here, we show that the Salmonella type III secretion effector SteA binds specifically to phosphatidylinositol 4‐phosphate [PI(4)P]. Ectopically expressed SteA localized at the plasma membrane (PM) of eukaryotic cells. However, SteA was displaced from the PM of Saccharomyces cerevisiae in mutants unable to synthesize the local pool of PI(4)P and from the PM of HeLa cells after localized depletion of PI(4)P. Moreover, in infected cells, bacterially translocated or ectopically expressed SteA localized at the membrane of the Salmonella‐containing vacuole (SCV) and to Salmonella‐induced tubules; using the PI(4)P‐binding domain of the Legionella type IV secretion effector SidC as probe, we found PI(4)P at the SCV membrane and associated tubules throughout Salmonella infection of HeLa cells. Both binding of SteA to PI(4)P and the subcellular localization of ectopically expressed or bacterially translocated SteA were dependent on a lysine residue near the N‐terminus of the protein. Overall, this indicates that binding of SteA to PI(4)P is necessary for its localization within host cells.     

Ultrastructural insights into pathogen clearance by autophagy

Autophagy defends cells against proliferation of bacteria such as Salmonella in the cytosol. After escape from a damaged Salmonella‐containing vacuole (SCV) exposing luminal glycans that bind to Galectin‐8, the host cell ubiquitination machinery deposits a dense layer of ubiquitin around the cytosolic bacteria. The nature and spatial distribution of this ubiquitin coat in relation to other autophagy‐related membranes are unknown. Using transmission electron microscopy, we determined the exact localisation of ubiquitin, the ruptured SCV membrane and phagophores around cytosolic Salmonella. Ubiquitin was not predominantly present on the Salmonella surface, but enriched on the fragmented SCV. Cytosolic bacteria without SCVs were less efficiently targeted by phagophores. Single bacteria were contained in single phagophores but multiple bacteria could be within large autophagic vacuoles reaching 30 μm in circumference. These large phagophores followed the contour of the engulfed bacteria, they were frequently in close association with endoplasmic reticulum membranes and, within them, remnants of the SCV were seen associated with each engulfed particle. Our data suggest that the Salmonella SCV has a major role in the formation of autophagic phagophores and highlight evolutionary conserved parallel mechanisms between xenophagy and mitophagy with the fragmented SCV and the damaged outer mitochondrial membrane serving similar functions.     

The entry of Salmonella in a distinct tight compartment revealed at high temporal and ultrastructural resolution

Salmonella enterica induces membrane ruffling and genesis of macropinosomes during its interactions with epithelial cells. This is achieved through the type three secretion system‐1, which first mediates bacterial attachment to host cells and then injects bacterial effector proteins to alter host behaviour. Next, Salmonella enters into the targeted cell within an early membrane‐bound compartment that matures into a slow growing, replicative niche called the Salmonella Containing Vacuole (SCV). Alternatively, the pathogen disrupts the membrane of the early compartment and replicate at high rate in the cytosol. Here, we show that the in situ formed macropinosomes, which have been previously postulated to be relevant for the step of Salmonella entry, are key contributors for the formation of the mature intracellular niche of Salmonella. We first clarify the primary mode of type three secretion system‐1 induced Salmonella entry into epithelial cells by combining classical fluorescent microscopy with cutting edge large volume electron microscopy. We observed that Salmonella, similarly to Shigella, enters epithelial cells inside tight vacuoles rather than in large macropinosomes. We next apply this technology to visualise rupturing Salmonella containing compartments, and we use extended time‐lapse microscopy to establish early markers that define which Salmonella will eventually hyper replicate. We show that at later infection stages, SCVs harbouring replicating Salmonella have previously fused with the in situ formed macropinosomes. In contrast, such fusion events could not be observed for hyper‐replicating Salmonella, suggesting that fusion of the Salmonella entry compartment with macropinosomes is the first committed step of SCV formation.     

Typhoidal Salmonella: Distinctive virulence factors and pathogenesis

Although nontyphoidal Salmonella (NTS; including Salmonella Typhimurium) mainly cause gastroenteritis, typhoidal serovars (Salmonella Typhi and Salmonella Paratyphi A) cause typhoid fever, the treatment of which is threatened by increasing drug resistance. Our understanding of S. Typhi infection in human remains poorly understood, likely due to the host restriction of typhoidal strains and the subsequent popularity of the S. Typhimurium mouse typhoid model. However, translating findings with S. Typhimurium across to S. Typhi has some limitations. Notably, S. Typhi has specific virulence factors, including typhoid toxin and Vi antigen, involved in symptom development and immune evasion, respectively. In addition to unique virulence factors, both typhoidal and NTS rely on two pathogenicity‐island encoded type III secretion systems (T3SS), the SPI‐1 and SPI‐2 T3SS, for invasion and intracellular replication. Marked differences have been observed in terms of T3SS regulation in response to bile, oxygen, and fever‐like temperatures. Moreover, approximately half of effectors found in S. Typhimurium are either absent or pseudogenes in S. Typhi, with most of the remaining exhibiting sequence variation. Typhoidal‐specific T3SS effectors have also been described. This review discusses what is known about the pathogenesis of typhoidal Salmonella with emphasis on unique behaviours and key differences when compared with S. Typhimurium.