PUBLICATIONS
June 15, 2023
EVOLUTIONARY AND FUNCTIONAL HISTORY OF THE ESCHERICHIA COLI K1 CAPSULE
​
SERGIO ARREDONDO-ALONSO, GEORGE BLUNDELL-HUNTER, ZUYI FU, REBECCA A GLADSTONE, ALFRED FILLOL-SALOM, JESSICA LORAINE, ELAINE CLOUTMAN-GREEN, PÅL J JOHNSEN, ØRJAN SAMUELSEN, ANNA K PÖNTINEN, FRANÇOIS CLÉON, SUSANA CHAVEZ-BUENO, MIGUEL A DE LA CRUZ, MIGUEL A ARES, MANIVANH VONGSOUVATH, AGNIESZKA CHMIELARCZYK, CAROLYNE HORNER, NIGEL KLEIN, ALAN MCNALLY, JOICE N REIS, JOSÉ R PENADÉS, NICHOLAS R THOMSON, JUKKA CORANDER, PETER W TAYLOR, ALEX J MCCARTHY
Escherichia coli is a leading cause of invasive bacterial infections in humans. Capsule polysaccharide has an important role in bacterial pathogenesis, and the K1 capsule has been firmly established as one of the most potent capsule types in E. coli through its association with severe infections. However, little is known about its distribution, evolution and functions across the E. coli phylogeny, which is fundamental to elucidating its role in the expansion of successful lineages. Using systematic surveys of invasive E. coli isolates, we show that the K1-cps locus is present in a quarter of bloodstream infection isolates and has emerged in at least four different extraintestinal pathogenic E. coli (ExPEC) phylogroups independently in the last 500 years. Phenotypic assessment demonstrates that K1 capsule synthesis enhances E. coli survival in human serum independent of genetic background, and that therapeutic targeting of the K1 capsule re-sensitizes E. coli from distinct genetic backgrounds to human serum. Our study highlights that assessing the evolutionary and functional properties of bacterial virulence factors at population levels is important to better monitor and predict the emergence of virulent clones, and to also inform therapies and preventive medicine to effectively control bacterial infections whilst significantly lowering antibiotic usage.
​
https://pubmed.ncbi.nlm.nih.gov/37322051/
​
​
2023 - Nature Communications
HUMAN CEACAM1 IS TARGETED BY A STREPTOCOCCUS PYOGENES ADHESIN IMPLICATED IN PUERPERAL SEPSIS PATHOGENESIS
​
ERIN A. CATTON, DANIEL A. BONSOR, CAROLINA HERRERA, MARGARETHA STÃ…LHAMMAR-CARLEMALM, MYKOLA LYNDIN, CLAIRE E. TURNER, JO SODEN, JOS AG. VAN STRIJP, BERNHARD B. SINGER, NINA M. VAN SORGE, GUNNAR LINDAHL, ALEX J. MCCARTHY.
Life-threatening bacterial infections in women after childbirth, known as puerperal sepsis, resulted in classical epidemics and remain a global health problem. While outbreaks of puerperal sepsis have been ascribed to Streptococcus pyogenes, little is known about disease mechanisms. Here, we show that the bacterial R28 protein, which is epidemiologically associated with outbreaks of puerperal sepsis, specifically targets the human receptor CEACAM1. This interaction triggers events that would favor the development of puerperal sepsis, including adhesion to cervical cells, suppression of epithelial wound repair and subversion of innate immune responses. High-resolution structural analysis showed that an R28 domain with IgI3-like fold binds to the N-terminal domain of CEACAM1. Together, these findings demonstrate that a single adhesin-receptor interaction can drive the pathogenesis of bacterial sepsis and provide molecular insights into the pathogenesis of one of the most important infectious diseases in medical history.
​
https://pubmed.ncbi.nlm.nih.gov/37080973/
​
2022 - Journal of Immunology
GROUP B STREPTOCOCCUS SURFACE PROTEIN Î’: STRUCTURAL CHARACTERIZATION OF A COMPLEMENT FACTOR H-BINDING MOTIF AND ITS CONTRIBUTION TO IMMUNE EVASION
​
XIN XU, ALEXANDER L LEWIS MARFFY, ANDREW KEIGHTLEY, ALEX J MCCARTHY, BRIAN V GEISBRECHT
The β protein from group B Streptococcus (GBS) is a ∼132-kDa, cell-surface exposed molecule that binds to multiple host-derived ligands, including complement factor H (FH). Many details regarding this interaction and its significance to immune evasion by GBS remain unclear. In this study, we identified a three-helix bundle domain within the C-terminal half of the B75KN region of β as the major FH-binding determinant and determined its crystal structure at 2.5 Å resolution. Analysis of this structure suggested a role in FH binding for a loop region connecting helices α1 and α2, which we confirmed by mutagenesis and direct binding studies. Using a combination of protein cross-linking and mass spectrometry, we observed that B75KN bound to complement control protein (CCP)3 and CCP4 domains of FH. Although this binding site lies within a complement regulatory region of FH, we determined that FH bound by β retained its decay acceleration and cofactor activities. Heterologous expression of β by Lactococcus lactis resulted in recruitment of FH to the bacterial surface and a significant reduction of C3b deposition following exposure to human serum. Surprisingly, we found that FH binding by β was not required for bacterial resistance to phagocytosis by neutrophils or killing of bacteria by whole human blood. However, loss of the B75KN region significantly diminished bacterial survival in both assays. Although our results show that FH recruited to the bacterial surface through a high-affinity interaction maintains key complement-regulatory functions, they raise questions about the importance of FH binding to immune evasion by GBS as a whole.
2022 - Methods in Molecular Biology
ANALYSIS OF ESCHERICHIA COLI K1 VIRULENCE GENES BY TRANSPOSON-DIRECTED SEQUENCING
​
ALEX J MCCARTHY, PETER W TAYLOR
Transposon-directed insertion site sequencing (TraDIS) combines random transposon mutagenesis and massively parallel sequencing to shed light on bacterial gene function on a genome-wide scale and in a high-throughput manner. The technique has proven to be successful in the determination of the fitness contribution of every gene under specific conditions both in vitro and in vivo. In this contribution, we describe the procedure used for the identification of Escherichia coli K1 genes essential for in vitro growth, survival in pooled human serum and gastrointestinal colonisation in a rodent model of neonatal invasive infection. TraDIS has broad application for systems-level analysis of a wide range of pathogenic, commensular and saprophytic bacteria.
​
​https://pubmed.ncbi.nlm.nih.gov/34709618/
​
​
​
2021 - The EMBO Journal
BACTERIAL PROTEIN DOMAINS WITH A NOVEL IG-LIKE FOLD TARGET HUMAN CEACAM RECEPTORS
​
NINA M VAN SORGE, DANIEL A BONSOR, LIWEN DENG, ERIK LINDAHL, VERENA SCHMITT, MYKOLA LYNDIN, ALEXEJ SCHMIDT, OLOF R NILSSON, JAIME BRIZUELA, ELENA BOERO, ERIC J SUNDBERG, JOS A G VAN STRIJP, KELLY S DORAN, BERNHARD B SINGER, GUNNAR LINDAHL, ALEX J MCCARTHY
Streptococcus agalactiae, also known as group B Streptococcus (GBS), is the major cause of neonatal sepsis in humans. A critical step to infection is adhesion of bacteria to epithelial surfaces. GBS adhesins have been identified to bind extracellular matrix components and cellular receptors. However, several putative adhesins have no host binding partner characterised. We report here that surface-expressed β protein of GBS binds to human CEACAM1 and CEACAM5 receptors. A crystal structure of the complex showed that an IgSF domain in β represents a novel Ig-fold subtype called IgI3, in which unique features allow binding to CEACAM1. Bioinformatic assessment revealed that this newly identified IgI3 fold is not exclusively present in GBS but is predicted to be present in adhesins from other clinically important human pathogens. In agreement with this prediction, we found that CEACAM1 binds to an IgI3 domain found in an adhesin from a different streptococcal species. Overall, our results indicate that the IgI3 fold could provide a broadly applied mechanism for bacteria to target CEACAMs.
​
https://pubmed.ncbi.nlm.nih.gov/33522633/
​
2020 - Microorganisms
GENES ON THE MOVE: IN VITRO TRANSDUCTION OF ANTIMICROBIAL RESISTANCE GENES BETWEEN HUMAN AND CANINE STAPHYLOCOCCAL PATHOGENS
​
SIAN MARIE FROSINI, ROSS BOND, ALEX J MCCARTHY, CLAUDIA FEUDI, STEFAN SCHWARZ, JODI A LINDSAY, ANETTE LOEFFLER
Transmission of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus pseudintermedius (MRSP) between people and pets, and their co-carriage, are well-described. Potential exchange of antimicrobial resistance (AMR) genes amongst these staphylococci was investigated in vitro through endogenous bacteriophage-mediated transduction. Bacteriophages were UV-induced from seven donor isolates of canine (MRSP) and human (MRSA) origin, containing tet(M), tet(K), fusB or fusC, and lysates filtered. Twenty-seven tetracycline- and fusidic acid- (FA-) susceptible recipients were used in 122 donor-recipient combinations (22 tetracycline, 100 FA) across 415 assays (115 tetracycline, 300 FA). Bacteriophage lysates were incubated with recipients and presumed transductants quantified on antimicrobial-supplemented agar plates. Tetracycline resistance transduction from MRSP and MRSA to methicillin-susceptible S. pseudintermedius (MSSP) was confirmed by PCR in 15/115 assays. No FA-resistance transfer occurred, confirmed by negative fusB/fusC PCR, but colonies resulting from FA assays had high MICs (≥32 mg/L) and showed mutations in fusA, two at a novel position (F88L), nine at H457[Y/N/L]. Horizontal gene transfer of tetracycline-resistance confirms that resistance genes can be shared between coagulase-positive staphylococci from different hosts. Cross-species AMR transmission highlights the importance of good antimicrobial stewardship across humans and veterinary species to support One Health.
2020 - Frontiers in Immunology
LEUKOCYTE IMMUNOGLOBULIN-LIKE RECEPTORS (LILRS) ON HUMAN NEUTROPHILS: MODULATORS OF INFECTION AND IMMUNITY
​
ALEXANDER L LEWIS MARFFY, ALEX J MCCARTHY
Neutrophils have a crucial role in defense against microbes. Immune receptors allow neutrophils to sense their environment, with many receptors functioning to recognize signs of infection and to promote antimicrobial effector functions. However, the neutrophil response must be tightly regulated to prevent excessive inflammation and tissue damage, and regulation is achieved by expression of inhibitory receptors that can raise activation thresholds. The leukocyte immunoglobulin-like receptor (LILR) family contain activating and inhibitory members that can up- or down-regulate immune cell activity. New ligands and functions for LILR continue to emerge. Understanding the role of LILR in neutrophil biology is of general interest as they can activate and suppress antimicrobial responses of neutrophils and because several human pathogens exploit these receptors for immune evasion. This review focuses on the role of LILR in neutrophil biology. We focus on the current knowledge of LILR expression on neutrophils, the known functions of LILR on neutrophils, and how these receptors may contribute to shaping neutrophil responses during infection.
​
2020 - Journal of Immunology
THE ORPHAN IMMUNE RECEPTOR LILRB3 MODULATES FC RECEPTOR-MEDIATED FUNCTIONS OF NEUTROPHILS
​
YUXI ZHAO, ESTHER VAN WOUDENBERGH, JING ZHU, ALBERT J R HECK, KOK P M VAN KESSEL, CARLA J C DE HAAS, PIET C AERTS, JOS A G VAN STRIJP, ALEX J MCCARTHY
Neutrophils are critical to the generation of effective immune responses and for killing invading microbes. Paired immune receptors provide important mechanisms to modulate neutrophil activation thresholds and effector functions. Expression of the leukocyte Ig-like receptor (LILR)A6 (ILT8/CD85b) and LILRB3 (ILT5/CD85a) paired-receptor system on human neutrophils has remained unclear because of the lack of specific molecular tools. Additionally, there is little known of their possible functions in neutrophil biology. The objective of this study was to characterize expression of LILRA6/LILRB3 receptors during human neutrophil differentiation and activation, and to assess their roles in modulating Fc receptor-mediated effector functions. LILRB3, but not LILRA6, was detected in human neutrophil lysates following immunoprecipitation by mass spectrometry. We demonstrate high LILRB3 expression on the surface of resting neutrophils and release from the surface following neutrophil activation. Surface expression was recapitulated in a human PLB-985 cell model of neutrophil-like differentiation. Continuous ligation of LILRB3 inhibited key IgA-mediated effector functions, including production of reactive oxygen species, phagocytic uptake, and microbial killing. This suggests that LILRB3 provides an important checkpoint to control human neutrophil activation and their antimicrobial effector functions during resting and early-activation stages of the neutrophil life cycle.
​
2019 - Infection & Immunity
LOSS OF TREFOIL FACTOR 2 SENSITIZES RAT PUPS TO SYSTEMIC INFECTION WITH THE NEONATAL PATHOGEN ESCHERICHIA COLI K1
​
ALEX J MCCARTHY, GEORGE M H BIRCHENOUGH, PETER W TAYLOR
Gastrointestinal (GI) colonization of 2-day-old (P2) rat pups with Escherichia coli K1 results in translocation of the colonizing bacteria across the small intestine, bacteremia, and invasion of the meninges, with animals frequently succumbing to lethal infection. Infection, but not colonization, is strongly age dependent; pups become progressively less susceptible to infection over the P2-to-P9 period. Colonization leads to strong downregulation of the gene encoding trefoil factor 2 (Tff2), preventing maturation of the protective mucus barrier in the small intestine. Trefoil factors promote mucosal homeostasis. We investigated the contribution of Tff2 to protection of the neonatal rat from E. coli K1 bacteremia and tissue invasion. Deletion of tff2, using clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9, sensitized P9 pups to E. coli K1 bacteremia. There were no differences between tff2-/ - homozygotes and the wild type with regard to the dynamics of GI colonization. Loss of the capacity to elaborate Tff2 did not impact GI tract integrity or the thickness of the small-intestinal mucus layer but, in contrast to P9 wild-type pups, enabled E. coli K1 bacteria to gain access to epithelial surfaces in the distal region of the small intestine and exploit an intracellular route across the epithelial monolayer to enter the blood circulation via the mesenteric lymphatic system. Although primarily associated with the mammalian gastric mucosa, we conclude that loss of Tff2 in the developing neonatal small intestine enables the opportunistic neonatal pathogen E. coli K1 to enter the compromised mucus layer in the distal small intestine prior to systemic invasion and infection.
​
https://pubmed.ncbi.nlm.nih.gov/30833331/
​
2018 - Veterinary Research
MOLECULAR BASIS DETERMINING SPECIES SPECIFICITY FOR TLR2 INHIBITION BY STAPHYLOCOCCAL SUPERANTIGEN-LIKE PROTEIN 3 (SSL3)
​
KIRSTEN J KOYMANS, LOURIS J FEITSMA, ADINDA BISSCHOP, ERIC G HUIZINGA, JOS A G VAN STRIJP, CARLA J C DE HAAS, ALEX J MCCARTHY
Staphylococcus aureus is a versatile opportunistic pathogen, causing disease in human and animal species. Its pathogenicity is linked to the ability of S. aureus to secrete immunomodulatory molecules. These evasion proteins bind to host receptors or their ligands, resulting in inhibitory effects through high affinity protein-protein interactions. Staphylococcal evasion molecules are often species-specific due to differences in host target proteins between species. We recently solved the crystal structure of murine TLR2 in complex with immunomodulatory molecule staphylococcal superantigen-like protein 3 (SSL3), which revealed the essential residues within SSL3 for TLR2 inhibition. In this study we aimed to investigate the molecular basis of the interaction on the TLR2 side. The SSL3 binding region on murine TLR2 was compared to that of other species through sequence alignment and homology modeling, which identified interspecies differences. To examine whether this resulted in altered SSL3 activity on the corresponding TLR2s, bovine, equine, human, and murine TLR2 were stably expressed in HEK293T cells and the ability of SSL3 to inhibit TLR2 was assessed. We found that SSL3 was unable to inhibit bovine TLR2. Subsequent loss and gain of function mutagenesis showed that the lack of inhibition is explained by the absence of two tyrosine residues in bovine TLR2 that play a prominent role in the SSL3-TLR2 interface. We found no evidence for the existence of allelic SSL3 variants that have adapted to the bovine host. Thus, within this paper we reveal the molecular determinants of the TLR2-SSL3 interaction which adds to our understanding of staphylococcal host specificity.
​
2018 - Journal of Bacteriology
GENOME-WIDE IDENTIFICATION BY TRANSPOSON INSERTION SEQUENCING OF ESCHERICHIA COLI K1 GENES ESSENTIAL FOR IN VITRO GROWTH, GASTROINTESTINAL COLONIZING CAPACITY, AND SURVIVAL IN SERUM
​
ALEX J MCCARTHY, RICHARD A STABLER, PETER W TAYLOR
Escherichia coli K1 strains are major causative agents of invasive disease of newborn infants. The age dependency of infection can be reproduced in neonatal rats. Colonization of the small intestine following oral administration of K1 bacteria leads rapidly to invasion of the blood circulation; bacteria that avoid capture by the mesenteric lymphatic system and evade antibacterial mechanisms in the blood may disseminate to cause organ-specific infections such as meningitis. Some E. coli K1 surface constituents, in particular the polysialic acid capsule, are known to contribute to invasive potential, but a comprehensive picture of the factors that determine the fully virulent phenotype has not emerged so far. We constructed a library and constituent sublibraries of ∼775,000 Tn5 transposon mutants of E. coli K1 strain A192PP and employed transposon-directed insertion site sequencing (TraDIS) to identify genes required for fitness for infection of 2-day-old rats. Transposon insertions were lacking in 357 genes following recovery on selective agar; these genes were considered essential for growth in nutrient-replete medium. Colonization of the midsection of the small intestine was facilitated by 167 E. coli K1 gene products. Restricted bacterial translocation across epithelial barriers precluded TraDIS analysis of gut-to-blood and blood-to-brain transits; 97 genes were required for survival in human serum. This study revealed that a large number of bacterial genes, many of which were not previously associated with systemic E. coli K1 infection, are required to realize full invasive potential.
​
2017 - PNAS
IMMUNE EVASION BY A STAPHYLOCOCCAL INHIBITOR OF MYELOPEROXIDASE
​
NIENKE W M DE JONG, KASRA X RAMYAR, FERMIN E GUERRA, REINDERT NIJLAND, CINDY FEVRE, JOVANKA M VOYICH, ALEX J MCCARTHY, BRANDON L GARCIA, KOK P M VAN KESSEL, JOS A G VAN STRIJP, BRIAN V GEISBRECHT, PIETER-JAN A HAAS
Staphylococcus aureus is highly adapted to its host and has evolved many strategies to resist opsonization and phagocytosis. Even after uptake by neutrophils, S. aureus shows resistance to killing, which suggests the presence of phagosomal immune evasion molecules. With the aid of secretome phage display, we identified a highly conserved protein that specifically binds and inhibits human myeloperoxidase (MPO), a major player in the oxidative defense of neutrophils. We have named this protein "staphylococcal peroxidase inhibitor" (SPIN). To gain insight into inhibition of MPO by SPIN, we solved the cocrystal structure of SPIN bound to a recombinant form of human MPO at 2.4-Ã… resolution. This structure reveals that SPIN acts as a molecular plug that prevents H2O2 substrate access to the MPO active site. In subsequent experiments, we observed that SPIN expression increases inside the neutrophil phagosome, where MPO is located, compared with outside the neutrophil. Moreover, bacteria with a deleted gene encoding SPIN showed decreased survival compared with WT bacteria after phagocytosis by neutrophils. Taken together, our results demonstrate that S. aureus secretes a unique proteinaceous MPO inhibitor to enhance survival by interfering with MPO-mediated killing.
​
2017 - Scientific Reports
POSTNATAL DEVELOPMENT OF THE SMALL INTESTINAL MUCOSA DRIVES AGE-DEPENDENT, REGIO-SELECTIVE SUSCEPTIBILITY TO ESCHERICHIA COLI K1 INFECTION
​
GEORGE M H BIRCHENOUGH, FATMA DALGAKIRAN, LUCI A WITCOMB, MALIN E V JOHANSSON, ALEX J MCCARTHY, GUNNAR C HANSSON, PETER W TAYLOR
The strong age dependency of neonatal systemic infection with Escherichia coli K1 can be replicated in the neonatal rat. Gastrointestinal (GI) colonization of two-day-old (P2) rats leads to invasion of the blood within 48 h of initiation of colonization; pups become progressively less susceptible to infection over the P2-P9 period. We show that, in animals colonized at P2 but not at P9, E. coli K1 bacteria gain access to the enterocyte surface in the mid-region of the small intestine and translocate through the epithelial cell monolayer by an intracellular pathway to the submucosa. In this region of the GI tract, the protective mucus barrier is poorly developed but matures to full thickness over P2-P9, coincident with the development of resistance to invasion. At P9, E. coli K1 bacteria are physically separated from villi by the mucus layer and their numbers controlled by mucus-embedded antimicrobial peptides, preventing invasion of host tissues.
​
2016 - PLoS One
PATHOADAPTIVE MUTATIONS OF ESCHERICHIA COLI K1 IN EXPERIMENTAL NEONATAL SYSTEMIC INFECTION
​
ALEX J MCCARTHY, DAVID NEGUS, PATRICIA MARTIN, CATARINA PECHINCHA, ERIC OSWALD, RICHARD A STABLER, PETER W TAYLOR
​
Although Escherichia coli K1 strains are benign commensals in adults, their acquisition at birth by the newborn may result in life-threatening systemic infections, most commonly sepsis and meningitis. Key features of these infections, including stable gastrointestinal (GI) colonization and age-dependent invasion of the bloodstream, can be replicated in the neonatal rat. We previously increased the capacity of a septicemia isolate of E. coli K1 to elicit systemic infection following colonization of the small intestine by serial passage through two-day-old (P2) rat pups. The passaged strain, A192PP (belonging to sequence type 95), induces lethal infection in all pups fed 2-6 x 106 CFU. Here we use whole-genome sequencing to identify mutations responsible for the threefold increase in lethality between the initial clinical isolate and the passaged derivative. Only four single nucleotide polymorphisms (SNPs), in genes (gloB, yjgV, tdcE) or promoters (thrA) involved in metabolic functions, were found: no changes were detected in genes encoding virulence determinants associated with the invasive potential of E. coli K1. The passaged strain differed in carbon source utilization in comparison to the clinical isolate, most notably its inability to metabolize glucose for growth. Deletion of each of the four genes from the E. coli A192PP chromosome altered the proteome, reduced the number of colonizing bacteria in the small intestine and increased the number of P2 survivors. This work indicates that changes in metabolic potential lead to increased colonization of the neonatal GI tract, increasing the potential for translocation across the GI epithelium into the systemic circulation.
​
2016 - Journal of Infectious Diseases
THE BACTERIAL STRESS-RESPONSIVE HSP90 CHAPERONE (HTPG) IS REQUIRED FOR THE PRODUCTION OF THE GENOTOXIN COLIBACTIN AND THE SIDEROPHORE YERSINIABACTIN IN ESCHERICHIA COLI
​
CHRISTOPHE GARCIE, SOPHIE TRONNET, AMÉLIE GARÉNAUX, ALEX J MCCARTHY, ALEXANDER O BRACHMANN, MARIE PÉNARY, SÉBASTIEN HOULE, JEAN-PHILIPPE NOUGAYRÈDE, JÖRN PIEL, PETER W TAYLOR, CHARLES M DOZOIS, PIERRE GENEVAUX, ERIC OSWALD, PATRICIA MARTIN
​
The genotoxin colibactin, synthesized by Escherichia coli, is a secondary metabolite belonging to the chemical family of hybrid polyketide/nonribosomal peptide compounds. It is produced by a complex biosynthetic assembly line encoded by the pks pathogenicity island. The presence of this large cluster of genes in the E. coli genome is invariably associated with the high-pathogenicity island, encoding the siderophore yersiniabactin, which belongs to the same chemical family as colibactin. The E. coli heat shock protein HtpG (Hsp90Ec) is the bacterial homolog of the eukaryotic molecular chaperone Hsp90, which is involved in the protection of cellular proteins against a variety of environmental stresses. In contrast to eukaryotic Hsp90, the functions and client proteins of Hsp90Ec are poorly known. Here, we demonstrated that production of colibactin and yersiniabactin is abolished in the absence of Hsp90Ec We further characterized an interplay between the Hsp90Ec molecular chaperone and the ClpQ protease involved in colibactin and yersiniabactin synthesis. Finally, we demonstrated that Hsp90Ec is required for the full in vivo virulence of extraintestinal pathogenic E. coli This is the first report highlighting the role of heat shock protein Hps90Ec in the production of two secondary metabolites involved in E. coli virulence.
https://pubmed.ncbi.nlm.nih.gov/27412582/
2015 - Infection & Immunity
BIOLUMINESCENT IMAGING REVEALS NOVEL PATTERNS OF COLONIZATION AND INVASION IN SYSTEMIC ESCHERICHIA COLI K1 EXPERIMENTAL INFECTION IN THE NEONATAL RAT
​
LUCI A WITCOMB, JAMES W COLLINS, ALEX J MCCARTHY, GADI FRANKEL, PETER W TAYLOR
Key features of Escherichia coli K1-mediated neonatal sepsis and meningitis, such as a strong age dependency and development along the gut-mesentery-blood-brain course of infection, can be replicated in the newborn rat. We examined temporal and spatial aspects of E. coli K1 infection following initiation of gastrointestinal colonization in 2-day-old (P2) rats after oral administration of E. coli K1 strain A192PP and a virulent bioluminescent derivative, E. coli A192PP-lux2. A combination of bacterial enumeration in the major organs, two-dimensional bioluminescence imaging, and three-dimensional diffuse light imaging tomography with integrated micro-computed tomography indicated multiple sites of colonization within the alimentary canal; these included the tongue, esophagus, and stomach in addition to the small intestine and colon. After invasion of the blood compartment, the bacteria entered the central nervous system, with restricted colonization of the brain, and also invaded the major organs, in line with increases in the severity of symptoms of infection. Both keratinized and nonkeratinized surfaces of esophagi were colonized to a considerably greater extent in susceptible P2 neonates than in corresponding tissues from infection-resistant 9-day-old rat pups; the bacteria appeared to damage and penetrate the nonkeratinized esophageal epithelium of infection-susceptible P2 animals, suggesting the esophagus represents a portal of entry for E. coli K1 into the systemic circulation. Thus, multimodality imaging of experimental systemic infections in real time indicates complex dynamic patterns of colonization and dissemination that provide new insights into the E. coli K1 infection of the neonatal rat.
​
2015 - Infection & Immunity
THE GENOTOXIN COLIBACTIN IS A DETERMINANT OF VIRULENCE IN ESCHERICHIA COLI K1 EXPERIMENTAL NEONATAL SYSTEMIC INFECTION
​
ALEX J MCCARTHY, PATRICIA MARTIN, EMILIE CLOUP, RICHARD A STABLER, ERIC OSWALD, PETER W TAYLOR
Escherichia coli strains expressing the K1 capsule are a major cause of sepsis and meningitis in human neonates. The development of these diseases is dependent on the expression of a range of virulence factors, many of which remain uncharacterized. Here, we show that all but 1 of 34 E. coli K1 neonatal isolates carried clbA and clbP, genes contained within the pks pathogenicity island and required for the synthesis of colibactin, a polyketide-peptide genotoxin that causes genomic instability in eukaryotic cells by induction of double-strand breaks in DNA. Inactivation of clbA and clbP in E. coli A192PP, a virulent strain of serotype O18:K1 that colonizes the gastrointestinal tract and translocates to the blood compartment with very high frequency in experimental infection of the neonatal rat, significantly reduced the capacity of A192PP to colonize the gut, engender double-strand breaks in DNA, and cause invasive, lethal disease. Mutation of clbA, which encodes a pleiotropic enzyme also involved in siderophore synthesis, impacted virulence to a greater extent than mutation of clbP, encoding an enzyme specific to colibactin synthesis. Restoration of colibactin gene function by complementation reestablished the fully virulent phenotype. We conclude that colibactin contributes to the capacity of E. coli K1 to colonize the neonatal gastrointestinal tract and to cause invasive disease in the susceptible neonate.
​
2015 - Journal of Antimicrobial Chemotherapy
GENOMIC INSIGHTS INTO THE RAPID EMERGENCE AND EVOLUTION OF MDR IN STAPHYLOCOCCUS PSEUDINTERMEDIUS
​
ALEX J MCCARTHY, EWAN M HARRISON, KINGA STANCZAK-MROZEK, BERNADETTE LEGGETT, ANDREW WALLER, MARK A HOLMES, DAVID H LLOYD, JODI A LINDSAY, ANETTE LOEFFLER
Objectives: MDR methicillin-resistant Staphylococcus pseudintermedius (MRSP) strains have emerged rapidly as major canine pathogens and present serious treatment issues and concerns to public health due to their, albeit low, zoonotic potential. A further understanding of the genetics of resistance arising from a broadly susceptible background of S. pseudintermedius is needed.
Methods: We sequenced the genomes of 12 S. pseudintermedius isolates of varied STs and resistance phenotypes.
Results: Nine distinct clonal lineages had acquired either staphylococcal cassette chromosome (SCC) mec elements and/or Tn5405-like elements carrying up to five resistance genes [aphA3, sat, aadE, erm(B), dfrG] to generate MRSP, MDR methicillin-susceptible S. pseudintermedius and MDR MRSP populations. The most successful and clinically problematic MDR MRSP clones, ST68 SCCmecV(T) and ST71 SCCmecII-III, have further accumulated mutations in gyrA and grlA conferring resistance to fluoroquinolones. The carriage of additional mobile genetic elements (MGEs) was highly variable, suggesting that horizontal gene transfer is frequent in S. pseudintermedius populations.
Conclusions: Importantly, the data suggest that MDR MRSP evolved rapidly by the acquisition of a very limited number of MGEs and mutations, and that the use of many classes of antimicrobials may co-select for the spread and emergence of MDR and XDR strains. Antimicrobial stewardship will need to be comprehensive, encompassing human medicine and veterinary disciplines to successfully preserve antimicrobial efficacy.
​
https://pubmed.ncbi.nlm.nih.gov/25527273/
​
​
2014 - Journal of Visualized Experiments
NON-INVASIVE MODEL OF NEUROPATHOGENIC ESCHERICHIA COLI INFECTION IN THE NEONATAL RAT
​
FATMA DALGAKIRAN, LUCI A WITCOMB, ALEX J MCCARTHY, GEORGE M H BIRCHENOUGH, PETER W TAYLOR
Investigation of the interactions between animal host and bacterial pathogen is only meaningful if the infection model employed replicates the principal features of the natural infection. This protocol describes procedures for the establishment and evaluation of systemic infection due to neuropathogenic Escherichia coli K1 in the neonatal rat. Colonization of the gastrointestinal tract leads to dissemination of the pathogen along the gut-lymph-blood-brain course of infection and the model displays strong age dependency. A strain of E. coli O18:K1 with enhanced virulence for the neonatal rat produces exceptionally high rates of colonization, translocation to the blood compartment and invasion of the meninges following transit through the choroid plexus. As in the human host, penetration of the central nervous system is accompanied by local inflammation and an invariably lethal outcome. The model is of proven utility for studies of the mechanism of pathogenesis, for evaluation of therapeutic interventions and for assessment of bacterial virulence.
​
2013 - Genome Biology & Evolution
EXTENSIVE HORIZONTAL GENE TRANSFER DURING STAPHYLOCOCCUS AUREUS CO-COLONIZATION IN VIVO
​
ALEX J MCCARTHY, ANETTE LOEFFLER, ADAM A WITNEY, KATHERINE A GOULD, DAVID H LLOYD, JODI A LINDSAY
​
Staphylococcus aureus is a commensal and major pathogen of humans and animals. Comparative genomics of S. aureus populations suggests that colonization of different host species is associated with carriage of mobile genetic elements (MGE), particularly bacteriophages and plasmids capable of encoding virulence, resistance, and immune evasion pathways. Antimicrobial-resistant S. aureus of livestock are a potential zoonotic threat to human health if they adapt to colonize humans efficiently. We utilized the technique of experimental evolution and co-colonized gnotobiotic piglets with both human- and pig-associated variants of the lineage clonal complex 398, and investigated growth and genetic changes over 16 days using whole genome sequencing. The human isolate survived co-colonization on piglets more efficiently than in vitro. During co-colonization, transfer of MGE from the pig to the human isolate was detected within 4 h. Extensive and repeated transfer of two bacteriophages and three plasmids resulted in colonization with isolates carrying a wide variety of mobilomes. Whole genome sequencing of progeny bacteria revealed no acquisition of core genome polymorphisms, highlighting the importance of MGE. Staphylococcus aureus bacteriophage recombination and integration into novel sites was detected experimentally for the first time. During colonization, clones coexisted and diversified rather than a single variant dominating. Unexpectedly, each piglet carried unique populations of bacterial variants, suggesting limited transmission of bacteria between piglets once colonized. Our data show that horizontal gene transfer occurs at very high frequency in vivo and significantly higher than that detectable in vitro.
​
2014 - Cellular and Molecular Life Sciences
FRUCTOSE TRANSPORT-DEFICIENT STAPHYLOCOCCUS AUREUS REVEALS IMPORTANT ROLE OF EPITHELIAL GLUCOSE TRANSPORTERS IN LIMITING SUGAR-DRIVEN BACTERIAL GROWTH IN AIRWAY SURFACE LIQUID
​
JAMES P GARNETT, DANIELA BRAUN, ALEX J MCCARTHY, MATTHEW R FARRANT, EMMA H BAKER, JODI A LINDSAY, DEBORAH L BAINES
Hyperglycaemia as a result of diabetes mellitus or acute illness is associated with increased susceptibility to respiratory infection with Staphylococcus aureus. Hyperglycaemia increases the concentration of glucose in airway surface liquid (ASL) and promotes the growth of S. aureus in vitro and in vivo. Whether elevation of other sugars in the blood, such as fructose, also results in increased concentrations in ASL is unknown and whether sugars in ASL are directly utilised by S. aureus for growth has not been investigated. We obtained mutant S. aureus JE2 strains with transposon disrupted sugar transport genes. NE768(fruA) exhibited restricted growth in 10 mM fructose. In H441 airway epithelial-bacterial co-culture, elevation of basolateral sugar concentration (5-20 mM) increased the apical growth of JE2. However, sugar-induced growth of NE768(fruA) was significantly less when basolateral fructose rather than glucose was elevated. This is the first experimental evidence to show that S. aureus directly utilises sugars present in the ASL for growth. Interestingly, JE2 growth was promoted less by glucose than fructose. Net transepithelial flux of D-glucose was lower than D-fructose. However, uptake of D-glucose was higher than D-fructose across both apical and basolateral membranes consistent with the presence of GLUT1/10 in the airway epithelium. Therefore, we propose that the preferential uptake of glucose (compared to fructose) limits its accumulation in ASL. Pre-treatment with metformin increased transepithelial resistance and reduced the sugar-dependent growth of S. aureus. Thus, epithelial paracellular permeability and glucose transport mechanisms are vital to maintain low glucose concentration in ASL and limit bacterial nutrient sources as a defence against infection.
​
2013 - Infection, Genetics & Evolution
STAPHYLOCOCCUS AUREUS INNATE IMMUNE EVASION IS LINEAGE-SPECIFIC: A BIOINFOMATICS STUDY
​
ALEX J MCCARTHY, JODI A LINDSAY
​
Staphylococcus aureus is a major human pathogen, and is targeted by the host innate immune system. In response, S. aureus genomes encode dozens of secreted proteins that inhibit complement, chemotaxis and neutrophil activation resulting in successful evasion of innate immune responses. These proteins include immune evasion cluster proteins (IEC; Chp, Sak, Scn), staphylococcal superantigen-like proteins (SSLs), phenol soluble modulins (PSMs) and several leukocidins. Biochemical studies have indicated that genetic variants of these proteins can have unique functions. To ascertain the scale of genetic variation in secreted immune evasion proteins, whole genome sequences of 88 S. aureus isolates, representing 25 clonal complex (CC) lineages, in the public domain were analysed across 43 genes encoding 38 secreted innate immune evasion protein complexes. Twenty-three genes were variable, with between 2 and 15 variants, and the variants had lineage-specific distributions. They include genes encoding Eap, Ecb, Efb, Flipr/Flipr-like, Hla, Hld, Hlg, Sbi, Scin-B/C and 13 SSLs. Most of these protein complexes inhibit complement, chemotaxis and neutrophil activation suggesting that isolates from each S. aureus lineage respond to the innate immune system differently. In contrast, protein complexes that lyse neutrophils (LukSF-PVL, LukMF, LukED and PSMs) were highly conserved, but can be carried on mobile genetic elements (MGEs). MGEs also encode proteins with narrow host-specificities arguing that their acquisition has important roles in host/environmental adaptation. In conclusion, this data suggests that each lineage of S. aureus evades host immune responses differently, and that isolates can adapt to new host environments by acquiring MGEs and the immune evasion protein complexes that they encode. Cocktail therapeutics that targets multiple variant proteins may be the most appropriate strategy for controlling S. aureus infections.
​
2012 - Mobile Genetic Elements
SHUFFLING OF MOBILE GENETIC ELEMENTS (MGES) IN SUCCESSFUL HEALTHCARE-ASSOCIATED MRSA (HA-MRSA)
​
JODI A LINDSAY, GWENAN M KNIGHT, EMMA L BUDD, ALEX J MCCARTHY
Methicillin-resistant Staphylococcus aureus (MRSA) CC22 SCCmecIV is a successful hospital-associated (HA-) MRSA, widespread throughout the world, and now the dominant clone in UK hospitals. We have recently shown that MRSA CC22 is a particularly fit clone, and it rose to dominance in a UK hospital at the same time as it began acquiring an increased range of antibiotic resistances. These resistances were not accumulated by individual CC22 isolates, but appear to shuffle frequently between isolates of the MRSA CC22 population. Resistances are often encoded on mobile genetic elements (MGEs) that include plasmids, transposons, bacteriophage and S. aureus pathogenicity islands (SaPIs). Using multi-strain whole genome microarrays, we show that there is enormous diversity of MGE carried within a MRSA CC22 SCCmecIV population, even among isolates from the same hospital and time period. MGE profiles were so variable that they could be used to track the spread of variant isolates within the hospital. We exploited this to show that the majority of patients colonised with MRSA at hospital admission that subsequently became infected were infected with their own colonising isolate. Our studies reveal MGE spread, stability, selection and clonal adaptation to the healthcare setting may be key to the success of HA-MRSA clones, presumably by allowing rapid adaptation to antibiotic exposure and new hosts.
​
https://pubmed.ncbi.nlm.nih.gov/23446976/
​
​
2012 - PLoS One
COMPARATIVE HOST SPECIFICITY OF HUMAN- AND PIG- ASSOCIATED STAPHYLOCOCCUS AUREUS CLONAL LINEAGES
​
ARSHNEE MOODLEY, CARMEN ESPINOSA-GONGORA, SØREN S NIELSEN, ALEX J MCCARTHY, JODI A LINDSAY, LUCA GUARDABASSI
Bacterial adhesion is a crucial step in colonization of the skin. In this study, we investigated the differential adherence to human and pig corneocytes of six Staphylococcus aureus strains belonging to three human-associated [ST8 (CC8), ST22 (CC22) and ST36(CC30)] and two pig-associated [ST398 (CC398) and ST433(CC30)] clonal lineages, and their colonization potential in the pig host was assessed by in vivo competition experiments. Corneocytes were collected from 11 humans and 21 pigs using D-squame® adhesive discs, and bacterial adherence to corneocytes was quantified by a standardized light microscopy assay. A previously described porcine colonization model was used to assess the potential of the six strains to colonize the pig host. Three pregnant, S. aureus-free sows were inoculated intravaginally shortly before farrowing with different strain mixes [mix 1) human and porcine ST398; mix 2) human ST36 and porcine ST433; and mix 3) human ST8, ST22, ST36 and porcine ST398] and the ability of individual strains to colonize the nasal cavity of newborn piglets was evaluated for 28 days after birth by strain-specific antibiotic selective culture. In the corneocyte assay, the pig-associated ST433 strain and the human-associated ST22 and ST36 strains showed significantly greater adhesion to porcine and human corneocytes, respectively (p<0.0001). In contrast, ST8 and ST398 did not display preferential host binding patterns. In the in vivo competition experiment, ST8 was a better colonizer compared to ST22, ST36, and ST433 prevailed over ST36 in colonizing the newborn piglets. These results are partly in agreement with previous genetic and epidemiological studies indicating the host specificity of ST22, ST36 and ST433 and the broad-host range of ST398. However, our in vitro and in vivo experiments revealed an unexpected ability of ST8 to adhere to porcine corneocytes and persist in the nasal cavity of pigs.
​
2012 - PLoS One
SURVIVAL OF STAPHYLOCOCCUS AUREUS ST398 IN THE HUMAN NOSE AFTER ARTIFICIAL INOCULATION
​
BIBI C G C SLINGERLAND, MEHRI TAVAKOL, ALEX J MCCARTHY, JODI A LINDSAY, SUSAN V SNIJDERS, JAAP A WAGENAAR, ALEX VAN BELKUM, MARGREET C VOS, HENRI A VERBRUGH, WILLEM J B VAN WAMEL
There is evidence that MRSA ST398 of animal origin is only capable of temporarily occupying the human nose, and it is therefore, often considered a poor human colonizer.We inoculated 16 healthy human volunteers with a mixture of the human MSSA strain 1036 (ST931, CC8) and the bovine MSSA strain 5062 (ST398, CC398), 7 weeks after a treatment with mupirocin and chlorhexidine-containing soap. Bacterial survival was studied by follow-up cultures over 21 days. The human strain 1036 was eliminated faster (median 14 days; range 2-21 days) than the bovine strain 5062 (median 21 days; range 7-21 days) but this difference was not significant (p = 0.065). The bacterial loads were significantly higher for the bovine strain on day 7 and day 21. 4/14 volunteers (28.6%) showed elimination of both strains within 21 days. Of the 10 remaining volunteers, 5 showed no differences in bacterial counts between both strains, and in the other 5 the ST398 strain far outnumbered the human S. aureus strain. Within the 21 days of follow-up, neither human strain 1036 nor bovine strain 5062 appeared to acquire or lose any mobile genetic elements. In conclusion, S. aureus ST398 strain 5062 is capable of adequately competing for a niche with a human strain and survives in the human nose for at least 21 days.
​
https://pubmed.ncbi.nlm.nih.gov/23155425/
​
​
2012 - Applied & Environmental Microbiology
STAPHYLOCOCCUS AUREUS CC398 CLADE ASSOCIATED WITH HUMAN-TO-HUMAN TRANSMISSION
​
ALEX J MCCARTHY, WILLEM VAN WAMEL, STIEN VANDENDRIESSCHE, JESPER LARSEN, OLIVIER DENIS, CRISTINA GARCIA-GRAELLS, ANN-CATRIN UHLEMANN, FRANKLIN D LOWY, ROBERT SKOV, JODI A LINDSAY
Staphylococcus aureus clonal complex 398 (CC398) isolates colonize livestock and can spread to human contacts. Genetic analysis of isolates epidemiologically associated with human-to-human, but not livestock, transmission in multiple countries and continents identified a common clade that was negative for tet(M) and positive for bacteriophage 3. Another group of human-to-human-transmitted isolates belonged to the common livestock-associated clade but had acquired a unique 7 bacteriophage.
​
2012 - Frontiers in Cellular & Infection Microbiology
STAPHYLOCOCCUS AUREUS TEMPERATE BACTERIOPHAGE: CARRIAGE AND HORIZONTAL GENE TRANSFER IS LINEAGE ASSOCIATED
​
ALEX J MCCARTHY, ADAM A WITNEY, JODI A LINDSAY
Staphylococcus aureus is a major cause of human and animal infections. Bacteriophage are a class of mobile genetic element (MGE) that carry virulence genes and disseminate them horizontally, including Panton-Valentine leukocidin (PVL), the immune evasion cluster (IEC) associated with human specificity, and enterotoxin A the major toxin associated with food poisoning. S. aureus isolates group into major clonal complex (CC) lineages that largely evolve independently due to possession of different restriction-modification (RM) systems. We aimed to better understand the horizontal and vertical transmission dynamics of virulence and resistance genes by bacteriophage by using (i) bioinformatic approaches to analyze bacteriophage genomes from the first 79 sequenced S. aureus isolates and (ii) S. aureus microarrays to analyze the distribution of bacteriophage and virulence genes in S. aureus isolates from a broader range of lineages. The distribution of eight bacteriophage families was highly variable but lineage associated. Nevertheless, there was evidence of frequent acquisition and loss and not just vertical transmission. Most bacteriophage genes were dispensable, and extensive mosaicism was seen. Surprisingly, virulence genes were tightly associated with specific phage families. This data suggests S. aureus bacteriophage evolve rapidly, and the horizontal gene transfer (HGT) of virulence genes encoded by bacteriophage is restricted by bacteriophage family and the lineage of the host bacterium, delaying the evolution of fully resistant and virulent strains.
​
​https://pubmed.ncbi.nlm.nih.gov/22919598/
​
​
2012 - Veterinary Dermatology
ARE ALL METICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS (MRSA) EQUAL IN ALL HOSTS? EPIDEMIOLOGICAL AND GENETIC COMPARISON BETWEEN ANIMAL AND HUMAN MRSA
​
ALEX J MCCARTHY, JODI A LINDSAY, ANETTE LOEFFLER
Meticillin-resistant Staphylococcus aureus (MRSA) continues to pose a major threat to human health. In animals, MRSA has become established as a veterinary pathogen in pets and horses; in livestock, it presents a concern for public health as a reservoir that can infect humans and as a source of transferrable resistance genes. Genetic analyses have revealed that the epidemiology of MRSA is different in different animal hosts. While human hospital-associated MRSA lineages are most commonly involved in pet infection and carriage, horse-specific MRSA most often represent 'traditional' equine S. aureus lineages. A recent development in the epidemiology of animal MRSA is the emergence of pig-adapted strains, such as CC398 and CC9, which appear to have arisen independently in the pig population. Recent insight into the genome structure and the evolution of S. aureus has helped to explain key aspects of these three distinct epidemiological scenarios. This nonsystematic literature review summarizes the structure and variations of the S. aureus genome and gives an overview of the current distribution of MRSA lineages in various animal species. It also discusses present knowledge about the emergence and evolution of MRSA in animals, adaptation to different host species and response to selective pressure from animal-specific environments. An improved understanding of the genetics and selective pressure that underpin the adaptive behaviour of S. aureus may be used in the future to predict new developments in staphylococcal diseases and to investigate novel control strategies required at a time of increasing resistance to antimicrobial agents.
​
2012 - BMC Microbiology
THE DISTRIBUTION OF PLASMIDS THAT CARRY VIRULENCE AND RESISTANCE GENES IN STAPHYLOCOCCUS AUREUS IS LINEAGE ASSOCIATED
​
ALEX J MCCARTHY, JODI A LINDSAY
Background: Staphylococcus aureus is major human and animal pathogen. Plasmids often carry resistance genes and virulence genes that can disseminate through S. aureus populations by horizontal gene transfer (HGT) mechanisms. Sequences of S. aureus plasmids in the public domain and data from multi-strain microarrays were analysed to investigate (i) the distribution of resistance genes and virulence genes on S. aureus plasmids, and (ii) the distribution of plasmids between S. aureus lineages.
Results: A total of 21 plasmid rep gene families, of which 13 were novel to this study, were characterised using a previously proposed classification system. 243 sequenced plasmids were assigned to 39 plasmid groups that each possessed a unique combination of rep genes. We show some resistance genes (including ermC and cat) and virulence genes (including entA, entG, entJ, entP) were associated with specific plasmid groups suggesting there are genetic pressures preventing recombination of these genes into novel plasmid groups. Whole genome microarray analysis revealed that plasmid rep, resistance and virulence genes were associated with S. aureus lineages, suggesting restriction-modification (RM) barriers to HGT of plasmids between strains exist. Conjugation transfer (tra) complex genes were rare.
Conclusion: This study argues that genetic pressures are restraining the spread of resistance and virulence genes amongst S. aureus plasmids, and amongst S. aureus populations, delaying the emergence of fully virulent and resistant strains.
​
January 25, 2025
IDENTIFICATION OF A HIGHLY TRANSMISSIBLE ANIMAL-INDEPENDENT STAPHYLOCOCCUS AUREUS ST398 CLONE WITH DISTINCT GENOMIC AND CELL ADHESION PROPERTIES
​
ANNE-CATRIN UHLEMANN, STEPHEN F PORCELLA, SHEETAL TRIVEDI, SEAN B SULLIVAN, CORY HAFER, ADAM D KENNEDY, KENT D BARBIAN, ALEX J MCCARTHY, CRAIG STREET, DAVID L HIRSCHBERG, W IAN LIPKIN, JODI A LINDSAY, FRANK R DELEO, FRANKLIN D LOWY
A methicillin-resistant Staphylococcus aureus (MRSA) clone known as ST398 has emerged as a major cause of acute infections in individuals who have close contact with livestock. More recently, the emergence of an animal-independent ST398 methicillin-sensitive S. aureus (MSSA) clone has been documented in several countries. However, the limited surveillance of MSSA has precluded an accurate assessment of the global spread of ST398 and its clinical relevance. Here we provide evidence that ST398 is a frequent source of MSSA infections in northern Manhattan and is readily transmitted between individuals in households. This contrasts with the limited transmissibility of livestock-associated ST398 (LA-ST398) MRSA strains between humans. Our whole-genome sequence analysis revealed that the chromosome of the human-associated ST398 MSSA clone is smaller than that of the LA-ST398 MRSA reference strain S0385, due mainly to fewer mobile genetic elements (MGEs). In contrast, human ST398 MSSA isolates harbored the prophage Ï•3 and the human-specific immune evasion cluster (IEC) genes chp and scn. While most of the core genome was conserved between the human ST398 MSSA clone and S0385, these strains differed substantially in their repertoire and composition of intact adhesion genes. These genetic changes were associated with significantly enhanced adhesion of human ST398 MSSA isolates to human skin keratinocytes and keratin. We propose that the human ST398 MSSA clone can spread independent of animal contact using an optimized repertoire of MGEs and adhesion molecules adapted to transmission among humans.
​
2012 - Journal of Clinical Microbiology
DETECTION OF MOBILE-GENETIC-ELEMENT VARIATION BETWEEN COLONIZING AND INFECTING HOSPITAL-ASSOCIATED METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS ISOLATES
​
ALEX J MCCARTHY, AODHAN S BREATHNACH, JODI A LINDSAY
Whole-genome analysis by 62-strain microarray showed variation in resistance and virulence genes on mobile genetic elements (MGEs) between 40 isolates of methicillin-resistant Staphylococcus aureus (MRSA) strain CC22-SCCmecIV but also showed (i) detection of two previously unrecognized MRSA transmission events and (ii) that 7/8 patients were infected with a variant of their own colonizing isolate.
​
January 25, 2025
THE DISTRIBUTION OF MOBILE GENETIC ELEMENTS (MGES) IN MRSA CC398 IS ASSOCIATED WITH BOTH HOST AND COUNTRY
​
ALEX J MCCARTHY, ADAM A WITNEY, KATHERINE A GOULD, ARSHNEE MOODLEY, LUCA GUARDABASSI, ANDREAS VOSS, OLIVIER DENIS, ELS M BROENS, JASON HINDS, JODI A LINDSAY
Methicillin-resistant Staphylococcus aureus clonal complex (CC) 398 has emerged from pigs to cause human infections in Europe and North America. We used a new 62-strain S. aureus microarray (SAM-62) to compare genomes of isolates from three geographical areas (Belgium, Denmark, and Netherlands) to understand how CC398 colonizes different mammalian hosts. The core genomes of 44 pig isolates and 32 isolates from humans did not vary. However, mobile genetic element (MGE) distribution was variable including SCCmec. Ï•3 bacteriophage and human specificity genes (chp, sak, scn) were found in invasive human but not pig isolates. SaPI5 and putative ruminant specificity gene variants (vwb and scn) were common but not pig specific. Virulence and resistance gene carriage was host associated but country specific. We conclude MGE exchange is frequent in CC398 and greatest among populations in close contact. This feature may help determine epidemiological associations among isolates of the same lineage.
​
https://pubmed.ncbi.nlm.nih.gov/21920902/
​
2011 - Animal Genetics
NOVEL POLYMORPHISMS IN OVINE IMMUNE RESPONSE GENES AND THEIR ASSOCIATION WITH ABORTION
​
R J DARLAY, A J MCCARTHY, N E ILLOT, J E SMITH, M-A SHAW
​
The sheep has worldwide agricultural importance, yet the genetic control of the immune responses underlying susceptibility or resistance to ovine disease is little understood. Here, we identify six novel polymorphisms in the ovine immune response genes interferon-γ (IFNG), tumour necrosis factor-α (TNF), interleukin-1β (IL1B) and interleukin-4 (IL4) in pedigree Charollais flocks. We confirm the presence of previously reported polymorphisms in IFNG and IL1B in Charollais. Restriction fragment length polymorphism (RFLP) genotyping assays have been developed for four polymorphisms, IFNGg.168C>T, IFNGg.285A>G, IL1Bg.689C>T and TNFg.3UTRA>G, and a Taqman genotyping assay has been developed for IL4g.485C>T. The previously described IL2g.647C>T polymorphism is adapted for RFLP analysis. Allele frequencies are described in Charollais, Lleyn and Suffolk cross sheep. Polymorphisms are typed in both Charollais ewes and lambs and analysed against abortion phenotypes. A subset of animals have also been analysed for the presence of Toxoplasma gondii, an abortion-causing protozoan. The IFNGg.168T allele is shown to be associated with increased risk of a ewe having an abortion, while the IFNGg.285G allele is associated with increased risk of a lamb being aborted. These assays provide tools for the investigation of the genetic basis of other phenotypes in sheep, including infectious disease susceptibility.
​
2011 - Infection, Genetics & Evolution
VARIATION IN EUROPEAN HARBOUR SEAL IMMUNE RESPONSE GENES AND SUSCEPTIBILITY TO PHOCINE DISTEMPER VIRUS (PDV)
​
ALEX J MCCARTHY, MARIE-ANNE SHAW, PAUL D JEPSON, SOPHIE M J M BRASSEUR, PETER J H REIJNDERS, SIMON J GOODMAN
Phocine distemper virus (PDV) has caused two mass mortalities of European harbour seals (Phoca vitulina) in recent decades. Levels of mortality varied considerably among European populations in both the 1988 and 2002 epidemics, with higher mortality in continental European populations in comparison to UK populations. High levels of genetic differentiation at neutral makers among seal populations allow for the possibility that there could be potential genetic differences at functional loci that may account for some of the variation in mortality. Recent genome sequencing of carnivore species and development of genomic tools have now made it possible to explore the possible contribution of variation in candidate genes from harbour seals in relation to the differential mortality patterns. We assessed variation in eight genes (CD46, IFNG, IL4, IL8, IL10, RARa, SLAM and TLR2) encoding key proteins involved in host cellular interactions with Morbilliviruses and the relationship of variants to disease status. This work constitutes the first genetic association study for Morbillivirus disease susceptibility in a non-model organism, and for a natural mortality event. We found no variation in harbour seals from across Europe in the protein coding domains of the viral receptors SLAM and CD46, but SNPs were present in SLAM intron 2. SNPs were also present in IL8 p2 and RARa exon 1. There was no significant association of SLAM or RARa polymorphisms with disease status implying no role of these genes in determining resistance to PDV induced mortality, that could be detected with the available samples and the small number of polymorphisms indentified. However there was significant differentiation of allele frequencies among populations. PDV and other morbilliviruses are important models for wildlife epidemiology, host switches and viral evolution. Despite a negative result in this case, full sequencing of pinniped and other 'non-model' carnivore genomes will help in refining understanding the role of host genetics in disease susceptibility for these viruses.
​
2010 - BMC Microbiology
GENETIC VARIATION IN STAPHYLOCOCCUS AUREUS SURFACE AND IMMUNE EVASION GENES IS LINEAGE ASSOCIATED: IMPLICATIONS FOR VACCINE DESIGN AND HOST-PATHOGEN INTERACTIONS
​
ALEX J MCCARTHY, JODI A LINDSAY
Background: S. aureus is a coloniser and pathogen of humans and mammals. Whole genome sequences of 58 strains of S. aureus in the public domain and data from multi-strain microarrays were compared to assess variation in the sequence of proteins known or putatively interacting with host.
Results: These included 24 surface proteins implicated in adhesion (ClfA, ClfB, Cna, Eap, Ebh, EbpS, FnBPA, FnBPB, IsaB, IsdA, IsdB, IsdH, SasB, SasC, SasD, SasF, SasG, SasH, SasK, SdrC, SdrD, SdrE, Spa and SraP) and 13 secreted proteins implicated in immune response evasion (Coa, Ecb, Efb, Emp, EsaC, EsxA, EssC, FLIPr, FLIPr like, Sbi, SCIN-B, SCIN-C, VWbp) located on the stable core genome. Many surface protein genes were missing or truncated, unlike immune evasion genes, and several distinct variants were identified. Domain variants were lineage specific. Unrelated lineages often possess the same sequence variant domains proving that horizontal transfer and recombination has contributed to their evolution. Surprisingly, sequenced strains from four animal S. aureus strains had surface and immune evasion proteins remarkably similar to those found in human strains, yet putative targets of these proteins vary substantially between different hosts. This suggests these proteins are not essential for virulence. However, the most variant protein domains were the putative functional regions and there is biological evidence that variants can be functional, arguing they do play a role.
Conclusion: Surface and immune evasion genes are candidates for S. aureus vaccines, and their distribution and functionality is key. Vaccines should contain cocktails of antigens representing all variants or they will not protect against naturally occurring S. aureus populations.
​
January 25, 2025
REASSESSING CONFLICTING EVOLUTIONARY HISTORIES OF THE PARAMYXOVIRIDAE AND THE ORIGINS OF RESPIROVIRUSES WITH BAYESIAN MULTIGENE PHYLOGENIES
​
ALEX J MCCARTHY, SIMON J GOODMAN
The evolution of paramyxoviruses is still poorly understood since past phylogenetic studies have revealed conflicting evolutionary signals among genes, and used varying methods and datasets. Using Bayesian phylogenetic analysis of full length single and concatenated sequences for the 6 genes shared among paramyxovirus genera, we reassess the ambiguous evolutionary relationships within the family, and examine causes of varying phylogenetic signals among different genes. Relative to a pneumovirus outgroup, the concatenated gene phylogeny, splits the Paramyxovirinae into two lineages, one comprising the avulaviruses and rubulaviruses, and a second containing the respiroviruses basal to the henipaviruses, and morbilliviruses. Phylogenies for the matrix (M), RNA dependent RNA polymerase (L) and the fusion (F) glycoprotein genes, are concordant with the topology from the concatenated dataset. In phylogenies derived from the nucleocapsid (N) and phosphoprotein (P) genes, the respiroviruses form the most basal genus of the Paramyxovirinae subfamily, with the avulaviruses and rubulaviruses in one lineage, and the henipaviruses, and morbilliviruses in a second. The phylogeny of the hemagglutinin (H) gene places the respiroviruses basal to the avula-rubulavirus group, but the relationship of this lineage with henipa and morbillviruses is not resolved. Different genes may be under varying evolutionary pressures giving rise to these conflicting signals. Given the level of conservation in the M and L genes, we suggest that together with F gene, these or concatenated datasets for all six genes are likely to reveal the most reliable phylogenies at a family level, and should be used for future phylogenetic studies in this group. Split decomposition analysis suggests that recombination within genera, may have a contributed to the emergence of dolphin morbillivirus, and several species within respiroviruses. A partial L gene alignment, resolves the relationship of 25 unclassified paramxyoviruses into 4 clades (Chiopteran-, Salmon-, Rodentian- and Ophidian paramyxoviruses) which group with rubula-, respiro-, morbilliviruses, and within the paramxyovirinae respectively.
​
2007 - Proceedings of Biological Sciences
PATHOGEN EVOLUTION AND DISEASE EMERGENCE IN CARNIVORES
​
ALEX J MCCARTHY, MARIE-ANNE SHAW, SIMON J GOODMAN
Emerging infectious diseases constitute some of the most pressing problems for both human and domestic animal health, and biodiversity conservation. Currently it is not clear whether the removal of past constraints on geographical distribution and transmission possibilities for pathogens alone are sufficient to give rise to novel host-pathogen combinations, or whether pathogen evolution is also generally required for establishment in novel hosts. Canine distemper virus (CDV) is a morbillivirus that is prevalent in the world dog population and poses an important conservation threat to a diverse range of carnivores. We performed an extensive phylogenetic and molecular evolution analysis on complete sequences of all CDV genes to assess the role of selection and recombination in shaping viral genetic diversity and driving the emergence of CDV in non-dog hosts. We tested the specific hypothesis that molecular adaptation at known receptor-binding sites of the haemagglutinin gene is associated with independent instances of the spread of CDV to novel non-dog hosts in the wild. This hypothesis was upheld, providing compelling evidence that repeated evolution at known functional sites (in this case residues 530 and 549 of the haemagglutinin molecule) is associated with multiple independent occurrences of disease emergence in a range of novel host species.
​
https://pubmed.ncbi.nlm.nih.gov/17956850/
​