The highly dynamic CRISPR1 system of <i>Streptococcus agalactiae</i> controls the diversity of its mobilome

Lopez-Sanchez, Maria-José; Sauvage, Elisabeth; Cunha, Violette Da; Clermont, Dominique; Hariniaina, Elisoa Ratsima; González‐Zorn, Bruno; Poyart, Claire; Rosinski‐Chupin, Isabelle; Glaser, Philippe

doi:10.1111/j.1365-2958.2012.08172.x

Cited by 150 publications

(218 citation statements)

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“…The name of this technique is based on that of the original spoligotyping method, which was first used for Mycobacterium tuberculosis complex (MTC) (10). Indeed, spoligotyping can be used to design a generic method, since polymorphic CRISPR regions are described at an increased pace and for more and more pathogens (11)(12)(13). Spoligotyping gained great acceptance internationally as a first-line method to genotype MTC for molecular epidemiological studies and allowed an understanding of its global phylogeographical structure (14).…”

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confidence: 99%

Validation of a Microbead-Based Format for Spoligotyping of Legionella pneumophila

et al. 2014

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A 42-plex clustered regularly interspaced short palindromic repeat (CRISPR)-based typing technique (spoligotyping) was recently developed at the French National Reference Center for Legionella. It allows the subtyping of the Legionella pneumophila sequence type 1/Paris pulsotype. In this report, we present the transfer of the membrane-based spoligotyping technique to a microbead-based multiplexed format. This microbead-based high-throughput assay uses devices such as Luminex 200 or the recently launched Magpix system (Luminex Corp., Austin, TX). We designated this new technique LP-SPOL (for L. pneumophila spoligotyping). We used two sets of samples previously subtyped by the membrane-based spoligotyping method to set up and validate the transfer on the two microbead-based systems. The first set of isolates (n ‫؍‬ 56) represented the whole diversity of the CRISPR patterns known to date. These isolates were used for transfer setup (determination of spacer cutoffs for both devices). The second set of isolates (n ‫؍‬ 245) was used to validate the transfer to the two microbead-based systems. The results obtained by the Luminex 200 system were 100% concordant with those obtained by the Magpix system for the 2 sets of isolates. In total, 10 discrepant results were observed when comparing the membrane-based method to the microbead-based method. These discrepancies were further resolved by repeating either the membrane-based or the microbead-based assay. This new assay is expected to play an emerging role for surveillance of L. pneumophila, starting with one of the most frequent genotypes, the sequence type 1/Paris pulsotype. However, the generalization of this typing method to all L. pneumophila strains is not feasible, since not all L. pneumophila strains contain CRISPRs.

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confidence: 99%

Validation of a Microbead-Based Format for Spoligotyping of Legionella pneumophila

et al. 2014

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“…Under these circumstances, spacers against lysogenic phages serve to maintain diversity in the host strain population and limit the proportion of cells susceptible to prophage induction and lysis. This interpretation seems to be favored with respect to the role of CRISPR/ Cas in regulating the mobilome of the gut bacterium Streptococcus agalactiae, 39 and is akin to other instances of defense strategies serving to maintain population diversity by compartmentalizing host susceptibility such as have been documented in the ocean. 64 Alternatively, a spacer and an integrated phage may actually coexist in the same cell.…”

Section: Holding a Grudgementioning

confidence: 99%

“…While the rate of spacer insertion is not conclusively determined in these strains, it is unlikely that CRISPR arrays in so many different species are frozen, as evidenced by the diversity of leader-proximal ends of CRISPR arrays that harbor memory spacers. 28,39 Yet another possible explanation is that the retention of memory spacers across individuals is a consequence of phage selective pressure. Weinberger et al have recently studied conservation of spacers at the trailer-end of an archaeal CRISPR array using an elegant combination of computer simulation and metagenomic reconstruction from an acid-mine drainage biofilm.…”

Section: Holding a Grudgementioning

confidence: 99%

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Holding a grudge

2013

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T he CRISPR (clustered regularlyinterspaced short palindromic repeats)/Cas (CRISPR-associated) system of bacteria and archaea constitutes a mechanism of acquired adaptive immunity against phages, which is based on genome-encoded markers of previously infecting phage sequences ("spacers"). As a repository of phage sequences, these spacers make the system particularly suitable for elucidating phage-bacteria interactions in metagenomic studies. Recent metagenomic analyses of CRISPRs associated with the human microbiome intriguingly revealed conserved "memory spacers" shared by bacteria in multiple unrelated, geographically separated individuals. Here, we discuss possible avenues for explaining this phenomenon by integrating insights from CRISPR biology and phage-bacteria ecology, with a special focus on the human gut. We further explore the growing body of evidence for the role of CRISPR/Cas in regulating the interplay between bacteria and lysogenic phages, which may be intimately related to the presence of memory spacers and sheds new light on the multifaceted biological and ecological modes of action of CRISPR/Cas. IntroductionBacteriophages are known to play a crucial role in shaping the structure, diversity and evolution of microbial communities in various ecological niches. [1][2][3][4] The CRISPR (clustered regularly interspaced short palindromic repeats)/Cas Holding a grudgePersisting anti-phage CRISPR immunity in multiple human gut microbiomes (CRISPR-associated) system of bacteria and archaea constitutes a mechanism of acquired adaptive immunity against phages, which is based on genomeencoded markers of past infections. 5 The unique biology of this system has opened a window into the multifaceted interactions that take place in natural environments between microbial populations and their associated phages. At the same time, observation of these interactions raises intriguing questions about the underlying mechanistic activity of CRISPR/Cas. Here, we explore this reciprocity with a focus on the microbial community resident in the human gut.CRISPR loci are composed of a succession of short repeat sequences separated by unique "spacer" sequences, usually sized 24-50 bp. While the mechanism of action of the CRISPR/Cas system is not yet fully characterized and some specifics vary by subtype, it generally entails three processes: incorporation of fragments of phage or plasmid genomes as novel spacers in bacterial CRISPR arrays; transcription and processing of the array into small RNAs (crRNAs) and formation of a crRNA/Cas protein complex that recognizes foreign nucleic acids through sequence complementarity and interferes with phage replication. [6][7][8][9][10][11] Elucidation of the function of the CRISPR/Cas system has led to theoretical and experimental efforts at exploring the ecological impacts of CRISPR-mediated immunity, as well as the conditions under which its emergence would be favored. [12][13][14] However, this system provided an ©2012 Landes Bioscience. Do not distribute.

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“…This has raised some sanitary concerns, as it is possible that, in close contact with humans, this sequence type could adapt and become zoonotic. ST-261 has also been isolated from frogs from Louisiana in the United States (Elliott et al, 1990;Lopez-Sanchez et al, 2012) so is not constrained to teleost or chondrychthyan hosts. However, has not yet been associated with human infections.…”

Section: Australian Marine St-261 Gbs Isolates Are Not Closely Relatementioning

confidence: 99%

On the origin of Group B Streptococcus from disease outbreaks in wild marine fish in Australia

Deboutteville¹

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Since 2007, 96 wild Queensland groupers, Epinephelus lanceolatus, were found dead from Karumba (Gulf of Carpentaria) in Northern Territory to Brisbane in southern Queensland, Australia. In twelve cases, Streptoccocus agalactiae (Group B Streptococcus, GBS) was isolated in pure culture from eyes and internal organs. GBS has been also isolated from other marine species (Javelin grunter, Giant catfish and Squaretail mullet) and caused an outbreak in three species of wild stingrays (Eastern shovelnose ray, Mangrove whipray and Estuary ray) in a public aquarium also in Queensland. These findings have raised public and industry concern over threats to human health from potential transmission between fish and humans, over environmental impacts on other aquatic animals, and over the potential threat posed to the growing Australian aquaculture industry in which, to date, there have been no reported cases of this disease in spite of the devastation caused to the industry overseas. Therefore, my thesis set out to determine whether GBS isolated from dead wild fish in Australia is indeed virulent in Queensland grouper and causative of mortality, to try to identify from where these infections in wild fish originated, to determine potential risk to human health and to other aquatic animals, and to begin to identify mechanisms by which these Australian isolates may cause fatal disease in Queensland grouper. This information may be employed in future risk assessment of potential human transfer, in mitigating further outbreaks in wild fish through understanding origin of infection, and in future development of preventative measures such as vaccination for farmed fish.In order to confirm that S. agalactiae was the cause of the mortality in wild fish, experimental challenges were conducted by injection, immersion and through the oral route. To better understand why these strains of GBS are pathogenic in fish, we conducted a series of cellular immune assays using juvenile Queensland grouper head-kidney leucocytes in the presence of different piscine and terrestrial isolates. Whole-genome sequencing analysis of the piscine and terrestrial isolates provided information on potential origin and potential for transfer between species, and also informed us on critical virulence factors that could be used during pathogenesis of GBS. Moreover, genome analysis gave intriguing insight into the evolution and adaption of GBS to fish as a host and to life in the aquatic environment. To determine putative origin of GBS in wild fish in Australia, multilocus sequence typing (MLST) and capsular polysaccharide (CPS) were derived from whole-genome sequences and revealed that all piscine isolates from Australia were serotype Ib and fell into ST-261 lineage first identified in 1986 from Tilapia in Israel. Overseas, ST-261 has only been found in fish and poikilotherm animals, and has never been associated with humans. High similarity and identical point mutations in the capsular operon of the piscine isolates suggests a common source of infection...

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The highly dynamic CRISPR1 system of Streptococcus agalactiae controls the diversity of its mobilome

Cited by 150 publications

References 52 publications

Validation of a Microbead-Based Format for Spoligotyping of Legionella pneumophila

Validation of a Microbead-Based Format for Spoligotyping of Legionella pneumophila

Holding a grudge

On the origin of Group B Streptococcus from disease outbreaks in wild marine fish in Australia

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