Person: Croucher, Nicholas J
Loading...
Email Address
AA Acceptance Date
Birth Date
Research Projects
Organizational Units
Job Title
Last Name
Croucher
First Name
Nicholas J
Name
Croucher, Nicholas J
11 results
Search Results
Now showing 1 - 10 of 11
Publication Rapid phylogenetic analysis of large samples of recombinant bacterial whole genome sequences using Gubbins(Oxford University Press, 2015) Croucher, Nicholas J; Page, Andrew J.; Connor, Thomas R.; Delaney, Aidan J.; Keane, Jacqueline A.; Bentley, Stephen D.; Parkhill, Julian; Harris, Simon R.The emergence of new sequencing technologies has facilitated the use of bacterial whole genome alignments for evolutionary studies and outbreak analyses. These datasets, of increasing size, often include examples of multiple different mechanisms of horizontal sequence transfer resulting in substantial alterations to prokaryotic chromosomes. The impact of these processes demands rapid and flexible approaches able to account for recombination when reconstructing isolatesâ recent diversification. Gubbins is an iterative algorithm that uses spatial scanning statistics to identify loci containing elevated densities of base substitutions suggestive of horizontal sequence transfer while concurrently constructing a maximum likelihood phylogeny based on the putative point mutations outside these regions of high sequence diversity. Simulations demonstrate the algorithm generates highly accurate reconstructions under realistically parameterized models of bacterial evolution, and achieves convergence in only a few hours on alignments of hundreds of bacterial genome sequences. Gubbins is appropriate for reconstructing the recent evolutionary history of a variety of haploid genotype alignments, as it makes no assumptions about the underlying mechanism of recombination. The software is freely available for download at github.com/sanger-pathogens/Gubbins, implemented in Python and C and supported on Linux and Mac OS X.Publication Dominant Role of Nucleotide Substitution in the Diversification of Serotype 3 Pneumococci over Decades and during a Single Infection(Public Library of Science, 2013) Croucher, Nicholas J; Mitchell, Andrea M.; Gould, Katherine A.; Inverarity, Donald; Barquist, Lars; Feltwell, Theresa; Fookes, Maria C.; Harris, Simon R.; Dordel, Janina; Salter, Susannah J.; Browall, Sarah; Zemlickova, Helena; Parkhill, Julian; Normark, Staffan; Henriques-Normark, Birgitta; Hinds, Jason; Mitchell, Tim J.; Bentley, Stephen D.Streptococcus pneumoniae of serotype 3 possess a mucoid capsule and cause disease associated with high mortality rates relative to other pneumococci. Phylogenetic analysis of a complete reference genome and 81 draft sequences from clonal complex 180, the predominant serotype 3 clone in much of the world, found most sampled isolates belonged to a clade affected by few diversifying recombinations. However, other isolates indicate significant genetic variation has accumulated over the clonal complex's entire history. Two closely related genomes, one from the blood and another from the cerebrospinal fluid, were obtained from a patient with meningitis. The pair differed in their behaviour in a mouse model of disease and in their susceptibility to antimicrobials, with at least some of these changes attributable to a mutation that up-regulated the patAB efflux pump. This indicates clinically important phenotypic variation can accumulate rapidly through small alterations to the genotype.Publication Population genomics of post-vaccine changes in pneumococcal epidemiology(2013) Croucher, Nicholas J; Finkelstein, Jonathan; Pelton, Stephen I.; Mitchell, Patrick K.; Lee, Grace; Parkhill, Julian; Bentley, Stephen D.; Hanage, William; Lipsitch, MarcWhole genome sequencing of 616 asymptomatically carried pneumococci was used to study the impact of the 7-valent pneumococcal conjugate vaccine. Comparison of closely related isolates revealed the role of transformation in facilitating capsule switching to non-vaccine serotypes and the emergence of drug resistance. However, such recombination was found to occur at significantly different rates across the species, and the evolution of the population was primarily driven by changes in the frequency of distinct genotypes extant pre-vaccine. These alterations resulted in little overall effect on accessory genome composition at the population level, contrasting with the fall in pneumococcal disease rates after the vaccineâs introduction.Publication Variable recombination dynamics during the emergence, transmission and âdisarmingâ of a multidrug-resistant pneumococcal clone(BioMed Central, 2014) Croucher, Nicholas J; Hanage, William; Harris, Simon R; McGee, Lesley; van der Linden, Mark; de Lencastre, Herminia; SĂĄ-LeĂŁo, Raquel; Song, Jae-Hoon; Ko, Kwan Soo; Beall, Bernard; Klugman, Keith P; Parkhill, Julian; Tomasz, Alexander; Kristinsson, Karl G; Bentley, Stephen DBackground: Pneumococcal ÎČ-lactam resistance was first detected in Iceland in the late 1980s, and subsequently peaked at almost 25% of clinical isolates in the mid-1990s largely due to the spread of the internationally-disseminated multidrug-resistant PMEN2 (or Spain6B-2) clone of Streptococcus pneumoniae. Results: Whole genome sequencing of an international collection of 189 isolates estimated that PMEN2 emerged around the late 1960s, developing resistance through multiple homologous recombinations and the acquisition of a Tn5253-type integrative and conjugative element (ICE). Two distinct clades entered Iceland in the 1980s, one of which had acquired a macrolide resistance cassette and was estimated to have risen sharply in its prevalence by coalescent analysis. Transmission within the island appeared to mainly emanate from ReykjavĂk and the Southern Peninsular, with evolution of the bacteria effectively clonal, mainly due to a prophage disrupting a gene necessary for genetic transformation in many isolates. A subsequent decline in PMEN2âs prevalence in Iceland coincided with a nationwide campaign that reduced dispensing of antibiotics to children in an attempt to limit its spread. Specific mutations causing inactivation or loss of ICE-borne resistance genes were identified from the genome sequences of isolates that reverted to drug susceptible phenotypes around this time. Phylogenetic analysis revealed some of these occurred on multiple occasions in parallel, suggesting they may have been at least temporarily advantageous. However, alteration of âcoreâ sequences associated with resistance was precluded by the absence of any substantial homologous recombination events. Conclusions: PMEN2âs clonal evolution was successful over the short-term in a limited geographical region, but its inability to alter major antigens or âcoreâ gene sequences associated with resistance may have prevented persistence over longer timespans.Publication Diversification of bacterial genome content through distinct mechanisms over different timescales(Nature Pub. Group, 2014) Croucher, Nicholas J; Coupland, Paul G.; Stevenson, Abbie E.; Callendrello, Alanna; Bentley, Stephen D.; Hanage, WilliamBacterial populations often consist of multiple co-circulating lineages. Determining how such population structures arise requires understanding what drives bacterial diversification. Using 616 systematically sampled genomes, we show that Streptococcus pneumoniae lineages are typically characterized by combinations of infrequently transferred stable genomic islands: those moving primarily through transformation, along with integrative and conjugative elements and phage-related chromosomal islands. The only lineage containing extensive unique sequence corresponds to a set of atypical unencapsulated isolates that may represent a distinct species. However, prophage content is highly variable even within lineages, suggesting frequent horizontal transmission that would necessitate rapidly diversifying anti-phage mechanisms to prevent these viruses sweeping through populations. Correspondingly, two loci encoding Type I restriction-modification systems able to change their specificity over short timescales through intragenomic recombination are ubiquitous across the collection. Hence short-term pneumococcal variation is characterized by movement of phage and intragenomic rearrangements, with the slower transfer of stable loci distinguishing lineages.Publication Evidence for Soft Selective Sweeps in the Evolution of Pneumococcal Multidrug Resistance and Vaccine Escape(Oxford University Press, 2014) Croucher, Nicholas J; Chewapreecha, Claire; Hanage, William; Harris, Simon R.; McGee, Lesley; van der Linden, Mark; Song, Jae-Hoon; Ko, Kwan Soo; de Lencastre, Herminia; Turner, Claudia; Yang, Fan; SĂĄ-LeĂŁo, Raquel; Beall, Bernard; Klugman, Keith P.; Parkhill, Julian; Turner, Paul; Bentley, Stephen D.The multidrug-resistant Streptococcus pneumoniae Taiwan19F-14, or PMEN14, clone was first observed with a 19F serotype, which is targeted by the heptavalent polysaccharide conjugate vaccine (PCV7). However, âvaccine escapeâ PMEN14 isolates with a 19A serotype became an increasingly important cause of disease post-PCV7. Whole genome sequencing was used to characterize the recent evolution of 173 pneumococci of, or related to, PMEN14. This suggested that PMEN14 is a single lineage that originated in the late 1980s in parallel with the acquisition of multiple resistances by close relatives. One of the four detected serotype switches to 19A generated representatives of the sequence type (ST) 320 isolates that have been highly successful post-PCV7. A second produced an ST236 19A genotype with reduced resistance to ÎČ-lactams owing to alteration of pbp1a and pbp2x sequences through the same recombination that caused the change in serotype. A third, which generated a mosaic capsule biosynthesis locus, resulted in serotype 19A ST271 isolates. The rapid diversification through homologous recombination seen in the global collection was similarly observed in the absence of vaccination in a set of isolates from the Maela refugee camp in Thailand, a collection that also allowed variation to be observed within carriage through longitudinal sampling. This suggests that some pneumococcal genotypes generate a pool of standing variation that is sufficiently extensive to result in âsoftâ selective sweeps: The emergence of multiple mutants in parallel upon a change in selection pressure, such as vaccine introduction. The subsequent competition between these mutants makes this phenomenon difficult to detect without deep sampling of individual lineages.Publication Stability of the pneumococcal population structure in Massachusetts as PCV13 was introduced(BioMed Central, 2015) Chang, Qiuzhi; Stevenson, Abbie E; Croucher, Nicholas J; Lee, Grace; Pelton, Stephen I; Lipsitch, Marc; Finkelstein, Jonathan; Hanage, WilliamBackground: The success of 7-valent pneumococcal conjugate vaccination (PCV-7) introduced to the US childhood immunization schedule in 2000 was partially offset by increases in invasive pneumococcal disease (IPD) and pneumococcal carriage due to non-vaccine serotypes, in particular 19A, in the years that followed. A 13-valent conjugate vaccine (PCV-13) was introduced in 2010. As part of an ongoing study of the response of the Massachusetts pneumococcal population to conjugate vaccination, we report the findings from the samples collected in 2011, as PCV-13 was introduced. Methods: We used multilocus sequence typing (MLST) to analyze 367 pneumococcal isolates carried by Massachusetts children (aged 3 months-7 years) collected during the winter of 2010â11 and used eBURST software to compare the pneumococcal population structure with that found in previous years. Results: One hundred and four distinct sequence types (STs) were found, including 24 that had not been previously recorded. Comparison with a similar sample collected in 2009 revealed no significant overall difference in the ST composition (p = 0.39, classification index). However, we describe clonal dynamics within the important replacement serotypes 19A, 15B/C, and 6C, and clonal expansion of ST 433 and ST 432, which are respectively serotype 22F and 21 clones. Conclusions: While little overall change in serotypes or STs was evident, multiple changes in the frequency of individual STs and or serotypes may plausibly be ascribed to the introduction of PCV-13. This 2011 sample documents the initial impact of PCV-13 and will be important for comparison with future studies of the evolution of the pneumococcal population in Massachusetts. Electronic supplementary material The online version of this article (doi:10.1186/s12879-015-0797-z) contains supplementary material, which is available to authorized users.Publication Identification of pneumococcal colonization determinants in the stringent response pathway facilitated by genomic diversity(BioMed Central, 2015) Li, Yuan; Croucher, Nicholas J; Thompson, Claudette; TrzciĆski, Krzysztof; Hanage, William; Lipsitch, MarcBackground: Understanding genetic determinants of a microbial phenotype generally involves creating and comparing isogenic strains differing at the locus of interest, but the naturally existing genomic and phenotypic diversity of microbial populations has rarely been exploited. Here we report use of a diverse collection of 616 carriage isolates of Streptococcus pneumoniae and their genome sequences to help identify a novel determinant of pneumococcal colonization. Results: A spontaneously arising laboratory variant (SpnYL101) of a capsule-switched TIGR4 strain (TIGR4:19F) showed reduced ability to establish mouse nasal colonization and lower resistance to non-opsonic neutrophil-mediated killing in vitro, a phenotype correlated with in vivo success. Whole genome sequencing revealed 5 single nucleotide polymorphisms (SNPs) affecting 4 genes in SpnYL101 relative to its ancestor. To evaluate the effect of variation in each gene, we performed an in silico screen of 616 previously published genome sequences to identify pairs of closely-related, serotype-matched isolates that differ at the gene of interest, and compared their resistance to neutrophil-killing. This method allowed rapid examination of multiple candidate genes and found phenotypic differences apparently associated with variation in SP_1645, a RelA/ SpoT homolog (RSH) involved in the stringent response. To establish causality, the alleles corresponding to SP_1645 were switched between the TIGR4:19F and SpnYL101. The wild-type SP_1645 conferred higher resistance to neutrophil-killing and competitiveness in mouse colonization. Using a similar strategy, variation in another RSH gene (TIGR4 locus tag SP_1097) was found to alter resistance to neutrophil-killing. Conclusions: These results indicate that analysis of naturally existing genomic diversity complements traditional genetics approaches to accelerate genotype-phenotype analysis. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-1573-6) contains supplementary material, which is available to authorized users.Publication Selective and Genetic Constraints on Pneumococcal Serotype Switching(Public Library of Science, 2015) Croucher, Nicholas J; Kagedan, Lisa; Thompson, Claudette; Parkhill, Julian; Bentley, Stephen D.; Finkelstein, Jonathan; Lipsitch, Marc; Hanage, WilliamStreptococcus pneumoniae isolates typically express one of over 90 immunologically distinguishable polysaccharide capsules (serotypes), which can be classified into âserogroupsâ based on cross-reactivity with certain antibodies. Pneumococci can alter their serotype through recombinations affecting the capsule polysaccharide synthesis (cps) locus. Twenty such âserotype switchingâ events were fully characterised using a collection of 616 whole genome sequences from systematic surveys of pneumococcal carriage. Eleven of these were within-serogroup switches, representing a highly significant (p < 0.0001) enrichment based on the observed serotype distribution. Whereas the recombinations resulting in between-serogroup switches all spanned the entire cps locus, some of those that caused within-serogroup switches did not. However, higher rates of within-serogroup switching could not be fully explained by either more frequent, shorter recombinations, nor by genetic linkage to genes involved in ÎČâlactam resistance. This suggested the observed pattern was a consequence of selection for preserving serogroup. Phenotyping of strains constructed to express different serotypes in common genetic backgrounds was used to test whether genotypes were physiologically adapted to particular serogroups. These data were consistent with epistatic interactions between the cps locus and the rest of the genome that were specific to serotype, but not serogroup, meaning they were unlikely to account for the observed distribution of capsule types. Exclusion of these genetic and physiological hypotheses suggested future work should focus on alternative mechanisms, such as host immunity spanning multiple serotypes within the same serogroup, which might explain the observed pattern.Publication Distinct Effects on Diversifying Selection by Two Mechanisms of Immunity Against Streptococcus pneumoniae(Public Library of Science, 2012) Li, Yuan; Gierahn, Todd; Thompson, Claudette; TrzciĆski, Krzysztof; Ford, Christopher Burton; Croucher, Nicholas J; Gouveia, Paulo; Flechtner, Jessica B.; Malley, Richard; Lipsitch, MarcAntigenic variation to evade host immunity has long been assumed to be a driving force of diversifying selection in pathogens. Colonization by Streptococcus pneumoniae, which is central to the organism's transmission and therefore evolution, is limited by two arms of the immune system: antibody- and T cell- mediated immunity. In particular, the effector activity of CD4+ TH17 cell mediated immunity has been shown to act in trans, clearing co-colonizing pneumococci that do not bear the relevant antigen. It is thus unclear whether TH17 cell immunity allows benefit of antigenic variation and contributes to diversifying selection. Here we show that antigen-specific CD4+ TH17 cell immunity almost equally reduces colonization by both an antigen-positive strain and a co-colonized, antigen-negative strain in a mouse model of pneumococcal carriage, thus potentially minimizing the advantage of escape from this type of immunity. Using a proteomic screening approach, we identified a list of candidate human CD4+ TH17 cell antigens. Using this list and a previously published list of pneumococcal Antibody antigens, we bioinformatically assessed the signals of diversifying selection among the identified antigens compared to non-antigens. We found that Antibody antigen genes were significantly more likely to be under diversifying selection than the TH17 cell antigen genes, which were indistinguishable from non-antigens. Within the Antibody antigens, epitopes recognized by human antibodies showed stronger evidence of diversifying selection. Taken together, the data suggest that TH17 cell-mediated immunity, one form of T cell immunity that is important to limit carriage of antigen-positive pneumococcus, favors little diversifying selection in the targeted antigen. The results could provide new insight into pneumococcal vaccine design.