The WD Domain of Atg16l1 Crucial for LC3-Associated Phagocytosis Is Not Required for Preserving Skin Barrier Function in Mice.

The skin is a multifunctional organ, forming a barrier between the external and internal environment, thereby functioning as a safeguard against extrinsic factors. Autophagy has been implicated in epidermal differentiation and in preserving skin homeostasis. LC3-associated phagocytosis (LAP) uses some but not all components of autophagy. The Atg16l1 (Δ WD) mouse model lacks the WD40 domain required for LAP and has been widely used to study the effects of LAP deficiency and autophagy on tissue homeostasis and response to infection. In this study, the Δ WD model was used to study the relationship between LAP and skin homeostasis by determining whether LAP-deficient mice display a cutaneous phenotype. Skin histology of wild-type and Δ WD mice aged 1 year revealed minor morphological differences in the tail skin dermal layer. RT-qPCR and western blot analysis showed no differences in key keratin expression between genotypes. Skin barrier formation, assessed by dye permeation assays, demonstrated full and proper formation of the skin barrier at embryonic day 18.5 in both genotypes. Biomechanical analysis of the skin showed decreased skin elasticity in aged Δ WD but not wild-type mice. In summary, the LAP-deficient Δ WD mice displayed subtle alterations in dermal histology and age-related biomechanical changes.

The V-ATPase-ATG16L1 axis recruits LRRK2 to facilitate the lysosomal stress response.

Leucine-rich repeat kinase 2 (LRRK2), a Rab kinase associated with Parkinson's disease and several inflammatory diseases, has been shown to localize to stressed lysosomes and get activated to regulate lysosomal homeostasis. However, the mechanisms of LRRK2 recruitment and activation have not been well understood. Here, we found that the ATG8 conjugation system regulates the recruitment of LRRK2 as well as LC3 onto single membranes of stressed lysosomes/phagosomes. This recruitment did not require FIP200-containing autophagy initiation complex, nor did it occur on double-membrane autophagosomes, suggesting independence from canonical autophagy. Consistently, LRRK2 recruitment was regulated by the V-ATPase-ATG16L1 axis, which requires the WD40 domain of ATG16L1 and specifically mediates ATG8 lipidation on single membranes. This mechanism was also responsible for the lysosomal stress-induced activation of LRRK2 and the resultant regulation of lysosomal secretion and enlargement. These results indicate that the V-ATPase-ATG16L1 axis serves a novel non-autophagic role in the maintenance of lysosomal homeostasis by recruiting LRRK2.

Investigating the Human Intestinal DNA Virome and Predicting Disease-Associated Virus-Host Interactions in Severe Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS).

Understanding how the human virome, and which of its constituents, contributes to health or disease states is reliant on obtaining comprehensive virome profiles. By combining DNA viromes from isolated virus-like particles (VLPs) and whole metagenomes from the same faecal sample of a small cohort of healthy individuals and patients with severe myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS), we have obtained a more inclusive profile of the human intestinal DNA virome. Key features are the identification of a core virome comprising tailed phages of the class Caudoviricetes, and a greater diversity of DNA viruses including extracellular phages and integrated prophages. Using an in silico approach, we predicted interactions between members of the Anaerotruncus genus and unique viruses present in ME/CFS microbiomes. This study therefore provides a framework and rationale for studies of larger cohorts of patients to further investigate disease-associated interactions between the intestinal virome and the bacteriome.

Investigating Antibody Reactivity to the Intestinal Microbiome in Severe Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS): A Feasibility Study.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a multisystemic disease of unknown aetiology that is characterised by disabling chronic fatigue and involves both the immune and gastrointestinal (GI) systems. Patients display alterations in GI microbiome with a significant proportion experiencing GI discomfort and pain and elevated blood biomarkers for altered intestinal permeability compared with healthy individuals. To investigate a possible GI origin of ME/CFS we designed a feasibility study to test the hypothesis that ME/CFS pathogenesis is a consequence of increased intestinal permeability that results in microbial translocation and a breakdown in immune tolerance leading to generation of antibodies reactive to indigenous intestinal microbes. Secretory immunoglobulin (Ig) A and serum IgG levels and reactivity to intestinal microbes were assessed in five pairs of severe ME/CFS patients and matched same-household healthy controls. For profiling serum IgG, we developed IgG-Seq which combines flow-cytometry based bacterial cell sorting and metagenomics to detect mucosal IgG reactivity to the microbiome. We uncovered evidence for immune dysfunction in severe ME/CFS patients that was characterised by reduced capacity and reactivity of serum IgG to stool microbes, irrespective of their source. This study provides the rationale for additional studies in larger cohorts of ME/CFS patients to further explore immune-microbiome interactions.

Control of infection by LC3-associated phagocytosis, CASM, and detection of raised vacuolar pH by the V-ATPase-ATG16L1 axis.

The delivery of pathogens to lysosomes for degradation provides an important defense against infection. Degradation is enhanced when LC3 is conjugated to endosomes and phagosomes containing pathogens to facilitate fusion with lysosomes. In phagocytic cells, TLR signaling and Rubicon activate LC3-associated phagocytosis (LAP) where stabilization of the NADPH oxidase leads to sustained ROS production and raised vacuolar pH. Raised pH triggers the assembly of the vacuolar ATPase on the vacuole membrane where it binds ATG16L1 to recruit the core LC3 conjugation complex (ATG16L1:ATG5-12). This V-ATPase-ATG16L1 axis is also activated in nonphagocytic cells to conjugate LC3 to endosomes containing extracellular microbes. Pathogens provide additional signals for recruitment of LC3 when they raise vacuolar pH with pore-forming toxins and proteins, phospholipases, or specialized secretion systems. Many microbes secrete virulence factors to inhibit ROS production and/or the V-ATPase-ATG16L1 axis to slow LC3 recruitment and avoid degradation in lysosomes.

LC3-associated phagocytosis in bone marrow macrophages suppresses acute myeloid leukemia progression through STING activation.

The bone marrow (BM) microenvironment regulates acute myeloid leukemia (AML) initiation, proliferation, and chemotherapy resistance. Following cancer cell death, a growing body of evidence suggests an important role for remaining apoptotic debris in regulating the immunologic response to and growth of solid tumors. Here, we investigated the role of macrophage LC3-associated phagocytosis (LAP) within the BM microenvironment of AML. Depletion of BM macrophages (BMMs) increased AML growth in vivo. We show that LAP is the predominate method of BMM phagocytosis of dead and dying cells in the AML microenvironment. Targeted inhibition of LAP led to the accumulation of apoptotic cells (ACs) and apoptotic bodies (ABs), resulting in accelerated leukemia growth. Mechanistically, LAP of AML-derived ABs by BMMs resulted in stimulator of IFN genes (STING) pathway activation. We found that AML-derived mitochondrial damage-associated molecular patterns were processed by BMMs via LAP. Moreover, depletion of mitochondrial DNA (mtDNA) in AML-derived ABs showed that it was this mtDNA that was responsible for the induction of STING signaling in BMMs. Phenotypically, we found that STING activation suppressed AML growth through a mechanism related to increased phagocytosis. In summary, we report that macrophage LAP of apoptotic debris in the AML BM microenvironment suppressed tumor growth.

Large-scale genomic analysis of antimicrobial resistance in the zoonotic pathogen Streptococcus suis.

BACKGROUND: Antimicrobial resistance (AMR) is among the gravest threats to human health and food security worldwide. The use of antimicrobials in livestock production can lead to emergence of AMR, which can have direct effects on humans through spread of zoonotic disease. Pigs pose a particular risk as they are a source of zoonotic diseases and receive more antimicrobials than most other livestock. Here we use a large-scale genomic approach to characterise AMR in Streptococcus suis, a commensal found in most pigs, but which can also cause serious disease in both pigs and humans. RESULTS: We obtained replicated measures of Minimum Inhibitory Concentration (MIC) for 16 antibiotics, across a panel of 678 isolates, from the major pig-producing regions of the world. For several drugs, there was no natural separation into 'resistant' and 'susceptible', highlighting the need to treat MIC as a quantitative trait. We found differences in MICs between countries, consistent with their patterns of antimicrobial usage. AMR levels were high even for drugs not used to treat S. suis, with many multidrug-resistant isolates. Similar levels of resistance were found in pigs and humans from regions associated with zoonotic transmission. We next used whole genome sequences for each isolate to identify 43 candidate resistance determinants, 22 of which were novel in S. suis. The presence of these determinants explained most of the variation in MIC. But there were also interesting complications, including epistatic interactions, where known resistance alleles had no effect in some genetic backgrounds. Beta-lactam resistance involved many core genome variants of small effect, appearing in a characteristic order. CONCLUSIONS: We present a large dataset allowing the analysis of the multiple contributing factors to AMR in S. suis. The high levels of AMR in S. suis that we observe are reflected by antibiotic usage patterns but our results confirm the potential for genomic data to aid in the fight against AMR.

Autophagy impairment by African swine fever virus.

African swine fever is a devastating disease of domestic swine and wild boar caused by a large double-stranded DNA virus that encodes for more than 150 open reading frames. There is no licensed vaccine for the disease and the most promising current candidates are modified live viruses that have been attenuated by deletion of virulence factors. Like many viruses African swine fever virus significantly alters the host cell machinery to benefit its replication and viral genes that modify host pathways represent promising targets for development of gene deleted vaccines. Autophagy is an important cellular pathway that is involved in cellular homeostasis, innate and adaptive immunity and therefore is manipulated by a number of different viruses. Autophagy is regulated by a complex protein cascade and here we show that African swine fever virus can block formation of autophagosomes, a critical functional step of the autophagy pathway through at least two different mechanisms. Interestingly this does not require the A179L gene that has been shown to interact with Beclin-1, an important autophagy regulator.

Non-canonical autophagy functions of ATG16L1 in epithelial cells limit lethal infection by influenza A virus.

Influenza A virus (IAV) and SARS-CoV-2 (COVID-19) cause pandemic infections where cytokine storm syndrome and lung inflammation lead to high mortality. Given the high social and economic cost of respiratory viruses, there is an urgent need to understand how the airways defend against virus infection. Here we use mice lacking the WD and linker domains of ATG16L1 to demonstrate that ATG16L1-dependent targeting of LC3 to single-membrane, non-autophagosome compartments - referred to as non-canonical autophagy - protects mice from lethal IAV infection. Mice with systemic loss of non-canonical autophagy are exquisitely sensitive to low-pathogenicity IAV where extensive viral replication throughout the lungs, coupled with cytokine amplification mediated by plasmacytoid dendritic cells, leads to fulminant pneumonia, lung inflammation and high mortality. IAV was controlled within epithelial barriers where non-canonical autophagy reduced IAV fusion with endosomes and activation of interferon signalling. Conditional mouse models and ex vivo analysis showed that protection against IAV infection of lung was independent of phagocytes and other leucocytes. This establishes non-canonical autophagy in airway epithelial cells as a novel innate defence that restricts IAV infection and lethal inflammation at respiratory surfaces.

Regulation of cytokine signaling through direct interaction between cytokine receptors and the ATG16L1 WD40 domain.

ATG16L1, an autophagy mediator that specifies the site of LC3 lipidation, includes a C-terminal domain formed by 7 WD40-type repeats (WD40 domain, WDD), the function of which is unclear. Here we show that the WDD interacts with the intracellular domain of cytokine receptors to regulate their signaling output in response to ligand stimulation. Using a refined version of a previously described WDD-binding amino acid motif, here we show that this element is present in the intracellular domain of cytokine receptors. Two of these receptors, IL-10RB and IL-2Rγ, recognize the WDD through the motif and exhibit WDD-dependent LC3 lipidation activity. IL-10 promotes IL-10RB/ATG16L1 interaction through the WDD, and IL-10 signaling is suboptimal in cells lacking the WDD owing to delayed endocytosis and inefficient early trafficking of IL10/IL-10R complexes. Our data reveal WDD-dependent roles of ATG16L1 in the regulation of cytokine receptor trafficking and signaling, and provide a WDD-binding motif that might be used to identify additional WDD activators.

Noncanonical function of an autophagy protein prevents spontaneous Alzheimer's disease.

Noncanonical functions of autophagy proteins have been implicated in neurodegenerative conditions, including Alzheimer's disease (AD). The WD domain of the autophagy protein Atg16L is dispensable for canonical autophagy but required for its noncanonical functions. Two-year-old mice lacking this domain presented with robust ß-amyloid (Aß) pathology, tau hyperphosphorylation, reactive microgliosis, pervasive neurodegeneration, and severe behavioral and memory deficiencies, consistent with human disease. Mechanistically, we found this WD domain was required for the recycling of Aß receptors in primary microglia. Pharmacologic suppression of neuroinflammation reversed established memory impairment and markers of disease pathology in this novel AD model. Therefore, loss of the Atg16L WD domain drives spontaneous AD in mice, and inhibition of neuroinflammation is a potential therapeutic approach for treating neurodegeneration and memory loss. A decline in expression of ATG16L in the brains of human patients with AD suggests the possibility that a similar mechanism may contribute in human disease.

Pathotyping the Zoonotic Pathogen Streptococcus suis: Novel Genetic Markers To Differentiate Invasive Disease-Associated Isolates from Non-Disease-Associated Isolates from England and Wales.

Streptococcus suis is one of the most important zoonotic bacterial pathogens of pigs, causing significant economic losses to the global swine industry. S. suis is also a very successful colonizer of mucosal surfaces, and commensal strains can be found in almost all pig populations worldwide, making detection of the S. suis species in asymptomatic carrier herds of little practical value in predicting the likelihood of future clinical relevance. The value of future molecular tools for surveillance and preventative health management lies in the detection of strains that genetically have increased potential to cause disease in presently healthy animals. Here we describe the use of genome-wide association studies to identify genetic markers associated with the observed clinical phenotypes (i) invasive disease and (ii) asymptomatic carriage on the palatine tonsils of pigs on UK farms. Subsequently, we designed a multiplex PCR to target three genetic markers that differentiated 115 S. suis isolates into disease-associated and non-disease-associated groups, that performed with a sensitivity of 0.91, a specificity of 0.79, a negative predictive value of 0.91, and a positive predictive value of 0.79 in comparison to observed clinical phenotypes. We describe evaluation of our pathotyping tool, using an out-of-sample collection of 50 previously uncharacterized S. suis isolates, in comparison to existing methods used to characterize and subtype S. suis isolates. In doing so, we show our pathotyping approach to be a competitive method to characterize S. suis isolates recovered from pigs on UK farms and one that can easily be updated to incorporate global strain collections.

The ATG5-binding and coiled coil domains of ATG16L1 maintain autophagy and tissue homeostasis in mice independently of the WD domain required for LC3-associated phagocytosis.

Macroautophagy/autophagy delivers damaged proteins and organelles to lysosomes for degradation, and plays important roles in maintaining tissue homeostasis by reducing tissue damage. The translocation of LC3 to the limiting membrane of the phagophore, the precursor to the autophagosome, during autophagy provides a binding site for autophagy cargoes, and facilitates fusion with lysosomes. An autophagy-related pathway called LC3-associated phagocytosis (LAP) targets LC3 to phagosome and endosome membranes during uptake of bacterial and fungal pathogens, and targets LC3 to swollen endosomes containing particulate material or apoptotic cells. We have investigated the roles played by autophagy and LAP in vivo by exploiting the observation that the WD domain of ATG16L1 is required for LAP, but not autophagy. Mice lacking the linker and WD domains, activate autophagy, but are deficient in LAP. The LAP-/- mice survive postnatal starvation, grow at the same rate as littermate controls, and are fertile. The liver, kidney, brain and muscle of these mice maintain levels of autophagy cargoes such as LC3 and SQSTM1/p62 similar to littermate controls, and prevent accumulation of SQSTM1 inclusions and tissue damage associated with loss of autophagy. The results suggest that autophagy maintains tissue homeostasis in mice independently of LC3-associated phagocytosis. Further deletion of glutamate E230 in the coiled-coil domain required for WIPI2 binding produced mice with defective autophagy that survived neonatal starvation. Analysis of brain lysates suggested that interactions between WIPI2 and ATG16L1 were less critical for autophagy in the brain, which may allow a low level of autophagy to overcome neonatal lethality. Abbreviations: CCD: coiled-coil domain; CYBB/NOX2: cytochrome b-245: beta polypeptide; GPT/ALT: glutamic pyruvic transaminase: soluble; LAP: LC3-associated phagocytosis; LC3: microtubule-associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; NOD: nucleotide-binding oligomerization domain; NADPH: nicotinamide adenine dinucleotide phosphate; RUBCN/Rubicon: RUN domain and cysteine-rich domain containing Beclin 1-interacting protein; SLE: systemic lupus erythematosus; SQSTM1/p62: sequestosome 1; TLR: toll-like receptor; TMEM: transmembrane protein; TRIM: tripartite motif-containing protein; UVRAG: UV radiation resistance associated gene; WD: tryptophan-aspartic acid; WIPI: WD 40 repeat domain: phosphoinositide interacting.

Streptococcus suis contains multiple phase-variable methyltransferases that show a discrete lineage distribution.

Streptococcus suis is a major pathogen of swine, responsible for a number of chronic and acute infections, and is also emerging as a major zoonotic pathogen, particularly in South-East Asia. Our study of a diverse population of S. suis shows that this organism contains both Type I and Type III phase-variable methyltransferases. In all previous examples, phase-variation of methyltransferases results in genome wide methylation differences, and results in differential regulation of multiple genes, a system known as the phasevarion (phase-variable regulon). We hypothesized that each variant in the Type I and Type III systems encoded a methyltransferase with a unique specificity, and could therefore control a distinct phasevarion, either by recombination-driven shuffling between different specificities (Type I) or by biphasic on-off switching via simple sequence repeats (Type III). Here, we present the identification of the target specificities for each Type III allelic variant from S. suis using single-molecule, real-time methylome analysis. We demonstrate phase-variation is occurring in both Type I and Type III methyltransferases, and show a distinct association between methyltransferase type and presence, and population clades. In addition, we show that the phase-variable Type I methyltransferase was likely acquired at the origin of a highly virulent zoonotic sub-population.

Effects of Environmental and Management-Associated Factors on Prevalence and Diversity of Streptococcus suis in Clinically Healthy Pig Herds in China and the United Kingdom.

Streptococcus suis, a global zoonosis of pigs, shows regional differences in the prevalence of human-associated disease for Asian and non-Asian countries. The isolation rates and diversities of S. suis on tonsils of healthy slaughter pigs in China and the United Kingdom were studied for effects of geography, temperature, pig age, and farm type. Isolates underwent analysis of molecular serotype and multilocus sequence type and virulence-associated genotyping. Although we found no significant difference in positive isolation rates between Chinese and UK farms, the prevalences of serotypes previously associated with human disease were significantly greater in the Chinese collection (P = 0.003). A significant effect of temperature was found on the positive isolation rate of the Chinese samples and the prevalence of human disease-associated serotypes in the UK S. suis population (China, P = 0.004; United Kingdom, P = 0.024) and on the prevalence of isolates carrying key virulence genes in China (P = 0.044). Finally, we found marked diversity among S. suis isolates, with statistically significant temperature effects on detection of multiple strain types within individual pigs. This study highlighted the high carriage prevalence and diversity of S. suis among clinically healthy pig herds of China and the United Kingdom. The significant effect of temperature on prevalence of isolation, human disease-associated serotypes, and diversity carried by individual pigs may shed new light on geographic variations in human S. suis-associated disease.IMPORTANCEStreptococcus suis is a global zoonotic pathogen and also a normal colonizer mainly carried on the tonsil of pigs. Thus, it is important to study the effect of environmental and management-associated factors on the S. suis populations in clinically healthy pigs. In this research, we investigated the similarities and differences between the S. suis populations obtained from different pig ages, seasons, and farm management systems and discovered the relationship between high climatic temperature and the prevalence of S. suis.

The WD40 domain of ATG16L1 is required for its non-canonical role in lipidation of LC3 at single membranes.

A hallmark of macroautophagy is the covalent lipidation of LC3 and insertion into the double-membrane phagophore, which is driven by the ATG16L1/ATG5-ATG12 complex. In contrast, non-canonical autophagy is a pathway through which LC3 is lipidated and inserted into single membranes, particularly endolysosomal vacuoles during cell engulfment events such as LC3-associated phagocytosis. Factors controlling the targeting of ATG16L1 to phagophores are dispensable for non-canonical autophagy, for which the mechanism of ATG16L1 recruitment is unknown. Here we show that the WD repeat-containing C-terminal domain (WD40 CTD) of ATG16L1 is essential for LC3 recruitment to endolysosomal membranes during non-canonical autophagy, but dispensable for canonical autophagy. Using this strategy to inhibit non-canonical autophagy specifically, we show a reduction of MHC class II antigen presentation in dendritic cells from mice lacking the WD40 CTD Further, we demonstrate activation of non-canonical autophagy dependent on the WD40 CTD during influenza A virus infection. This suggests dependence on WD40 CTD distinguishes between macroautophagy and non-canonical use of autophagy machinery.

LC3-Associated Phagocytosis Is Required for Dendritic Cell Inflammatory Cytokine Response to Gut Commensal Yeast Saccharomyces cerevisiae.

The human fungal microbiota known as mycobiota is increasingly recognized as a critical factor in human gut health and disease. Non-pathogenic commensal yeasts such as Saccharomyces cerevisiae promote homeostasis in the gut, whereas dysbiosis of the gut mycobiota is associated with inflammation. Glycan-binding receptors (lectins) are key host factors in host-mycobiota interaction in the gut. They are expressed on immune cells such as dendritic cells (DCs) and recognize fungal polysaccharides. This interaction is imperative to mount appropriate immune responses for immune homeostasis in the gut as well as clearance of fungal pathogens. Recent studies demonstrate that microtubule-associated protein light-chain 3 (LC3)-associated phagocytosis (LAP) is involved in lectin-fungi interactions. Yet, the biological impact of LAP on the lectin function remains largely elusive. In this report, we demonstrate that in mouse LAP is linked to dendritic cell-associated lectin 2 (Dectin-2), a C-type lectin specific to fungal α-mannan polysaccharide. We found that mouse Dectin-2 recognizes commensal yeast S. cerevisiae and Kazachstania unispora. Mouse bone marrow-derived DCs (BMDCs) produced inflammatory cytokines TNFα and IL-1ß in response to the yeasts in a Dectin-2 and spleen tyrosine kinase (Syk)-dependent manner. We found that S. cerevisiae and K. unispora induced LAP in mouse BMDCs upon internalization. Furthermore, LC3 was activated by stimulation of BMDCs with the yeasts in a Dectin-2 and Syk-dependent manner. To address the biological impact of LAP on Dectin-2 yeast interaction, we established a knock-in mouse strain (Atg16L1E230, thereafter called E230), which BMDCs exhibit autophagy-active and LAP-negative phenotypes. When stimulated with yeasts, E230 BMDCs produced significantly less amounts of TNFα and IL-1ß. Taken together, we revealed a novel link between Dectin-2 and LAP that enables host immune cells to respond to mycobiota.

Patterns of antimicrobial resistance in Streptococcus suis isolates from pigs with or without streptococcal disease in England between 2009 and 2014.

Antimicrobial resistance in Streptococcus suis, a global zoonotic pathogen of pigs, has been mostly studied only in diseased animals using surveys that have not evaluated changes over time. We compared patterns of resistance between S. suis isolates from clinical cases of disease (CC) and non-clinical case (NCC) pigs in England, collected over two discrete periods, 2009-2011 and 2013-2014. Minimum inhibitory concentrations (MIC) of 17 antimicrobials (nine classes) were determined on 405 S. suis isolates categorised by sampling period and disease association to assess changes in resistance over time and association with disease. First, isolates were characterized as resistant or susceptible using published clinical breakpoints. Second, epidemiological cut-offs (ECOFF) were derived from MIC values, and isolates classified as wild type (WT) below the ECOFF and non-wild type (NWT) above the ECOFF. Finally, isolate subsets were analysed for shifts in MIC distribution. NCC isolates were more resistant than CC isolates to cephalosporins, penams, pleuromutilins, potentiated sulphonamides and tetracyclines in both study periods. Resistance levels among CC isolates increased in 2013-2014 relative to 2009-2011 for antimicrobials including aminoglycosides, cephalosporins, fluoroquinolones, pleuromutilins, potentiated sulphonamides and tetracyclines. The prevalence of isolates categorised as NWT for five or more classes of antimicrobials was greater among NCC than CC isolates for both time periods, and increased with time. This study used standardised methods to identify significant shifts in antimicrobial resistance phenotypes of S. suis isolated from pigs in England, not only over time but also between isolates from known clinical cases or disease-free pigs.

Emergence of host-adapted Salmonella Enteritidis through rapid evolution in an immunocompromised host.

Host adaptation is a key factor contributing to the emergence of new bacterial, viral and parasitic pathogens. Many pathogens are considered promiscuous because they cause disease across a range of host species, while others are host-adapted, infecting particular hosts(1). Host adaptation can potentially progress to host restriction, where the pathogen is strictly limited to a single host species and is frequently associated with more severe symptoms. Host-adapted and host-restricted bacterial clades evolve from within a broader host-promiscuous species and sometimes target different niches within their specialist hosts, such as adapting from a mucosal to a systemic lifestyle. Genome degradation, marked by gene inactivation and deletion, is a key feature of host adaptation, although the triggers initiating genome degradation are not well understood. Here, we show that a chronic systemic non-typhoidal Salmonella infection in an immunocompromised human patient resulted in genome degradation targeting genes that are expendable for a systemic lifestyle. We present a genome-based investigation of a recurrent blood-borne Salmonella enterica serotype Enteritidis (S. Enteritidis) infection covering 15 years in an interleukin-12 ß1 receptor-deficient individual that developed into an asymptomatic chronic infection. The infecting S. Enteritidis harboured a mutation in the mismatch repair gene mutS that accelerated the genomic mutation rate. Phylogenetic analysis and phenotyping of multiple patient isolates provides evidence for a remarkable level of within-host evolution that parallels genome changes present in successful host-restricted bacterial pathogens but never before observed on this timescale. Our analysis identifies common pathways of host adaptation and demonstrates the role that immunocompromised individuals can play in this process.

Emergence of host-adapted Salmonella Enteritidis through rapid evolution in an immunocompromised host.

Host adaptation is a key factor contributing to the emergence of new bacterial, viral and parasitic pathogens. Many pathogens are considered promiscuous because they cause disease across a range of host species, while others are host-adapted, infecting particular hosts1. Host adaptation can potentially progress to host restriction where the pathogen is strictly limited to a single host species and is frequently associated with more severe symptoms. Host-adapted and host-restricted bacterial clades evolve from within a broader host-promiscuous species and sometimes target different niches within their specialist hosts, such as adapting from a mucosal to a systemic lifestyle. Genome degradation, marked by gene inactivation and deletion, is a key feature of host adaptation, although the triggers initiating genome degradation are not well understood. Here, we show that a chronic systemic non-typhoidal Salmonella infection in an immunocompromised human patient resulted in genome degradation targeting genes that are expendable for a systemic lifestyle. We present a genome-based investigation of a recurrent blood-borne Salmonella enterica serotype Enteritidis (S. Enteritidis) infection covering 15 years in an interleukin (IL)-12 ß-1 receptor-deficient individual that developed into an asymptomatic chronic infection. The infecting S. Enteritidis harbored a mutation in the mismatch repair gene mutS that accelerated the genomic mutation rate. Phylogenetic analysis and phenotyping of multiple patient isolates provides evidence for a remarkable level of within-host evolution that parallels genome changes present in successful host-restricted bacterial pathogens but never before observed on this timescale. Our analysis identifies common pathways of host adaptation and demonstrates the role that immunocompromised individuals can play in this process.