Identification and characterization of episomal forms of integrative genomic islands in the genus Francisella

Rydzewski, Kerstin; Tlapák, Hana; Niehaus, Indra; Dabrowski, Piotr Wojtek; Grunow, Roland; Heuner, Klaus

doi:10.1016/j.ijmm.2015.08.037

Cited by 4 publications

(8 citation statements)

References 27 publications

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“…Na análise bacteriológica foram coletadas, assepticamente, amostras do cérebro e rim para inoculação em meio Àgar Sangue, assim como amostras do rim e baço foram inoculadas em Ágar Cistina Coração (TYC, Difco, USA) suplementado com 1% de hemoglobina bovina. Todas as amostras foram incubadas por 168h a 28°C (Rydzewski et al, 2015).…”

Section: Relato De Casounclassified

PRIMEIRO RELATO DE Francisella noatunensis SUBESPÉCIE ORIENTALIS NO SUL DO BRASIL – RELATO DE CASO

Jatobá¹,

Klipp²,

Hoppe³

2016

Acta Vet Bras

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RESUMOABSTRACT -The systemic visceral granuloma or franciselose is caused by Francisella noatunensis subspecies orientalis and F. noatunensis subspecies noatunensis. In Brazil, the first Francisella outbreak of Nile tilapia (Oreochromis niloticus) was recorded in the winters of 2012 and 2013 in the state of Minas Gerais. However, nothing was observed in the other producing regions. The aim of this study was to diagnose the causative agent of mortality Nile tilapia (Oreochromis niloticus) in a production unit in the state of Santa Catarina. For a mortality outbreak in September 2015, we were evaluated 35 Tilapia for the presence of parasites, microbiological evaluation and analysis of the polymerase chain reaction. After analysis, it was registered the presence of F. noatunensis subspecies orientalis, the first reported in southern Brazil

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Section: Relato De Casounclassified

PRIMEIRO RELATO DE Francisella noatunensis SUBESPÉCIE ORIENTALIS NO SUL DO BRASIL – RELATO DE CASO

Jatobá¹,

Klipp²,

Hoppe³

2016

Acta Vet Bras

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“…Recently, we demonstrated that the att -sites of FhaGI-1 of Fhis AS02-814 in combination with the site-specific integrase are sufficient to generate the episomal form FIV-Val of the vector in E. coli and that after transformation into Fth LVS the element integrates site specifically into the tRNA-Val gene of transformants (Rydzewski et al, 2015 ). We now utilized this information to develop phage integration vectors to be used in Francisella research, with the idea that the constructs would be integrated site specifically and stable into the genome.…”

Section: Resultsmentioning

confidence: 99%

“…The different constructs were then used to generate pFIV-Val vectors 1, 2, and pFIV1- iglC , using pUC57-Kana as the back-bone (for details, see Table 1 and Figure 1 ). Construct 1: FhaGI-gfp-CmR is composed of 4,644 bp, exhibiting the tRNA Val gene of FhaGI-1 (Rydzewski et al, 2015 ), followed by restriction sites for NotI, BclI, and SnaBI, a gfp gene with promotor from vector pKK289KmGFP (Bönquist et al, 2008 ), restriction sites for NotI and SacII, the GroES promotor of Fth LVS (pGroES) (Ericsson et al, 1997 ), the iglA promotor with the PigR response element (PRE, underlined bps in PRE * ), (PRE * : AGCTGTA TAA ACATTGTGTT ATTGGCGTTA TTAAGGTAAC TT) (Ramsey et al, 2015 ), the GroES promotor from strain Francisella sp. strain W12-1067 (Rydzewski et al, 2014 ), followed by a Cm resistance cassette (952 bp) with promotor GroES from vector pKK289KmGFP (Bönquist et al, 2008 ), the integrase of FhaGI-1 (FN3523_1033), and the phage integration site attR (47 bp) (Rydzewski et al, 2015 ).…”

Section: Methodsmentioning

confidence: 99%

“…We could show that the GI integrates site specifically into the tRNA Val gene of the genome and that it generates an episomal form in an integrase-dependent manner. Furthermore, we could demonstrate that small variants of FhaGI-1 are able to integrate site specifically into the genome of other Francisella species (Rydzewski et al, 2015 ). Therefore, we decided to create the first Francisella phage integration vector on the basis of this GI.…”

Section: Introductionmentioning

confidence: 96%

“…Both systems produce transformants that are stable without selective pressure, but they also require helper plasmids or multiple rounds of transformation and selection (Ludu et al, 2008 ; LoVullo et al, 2009a , b ). Phage integration vectors have not been generated for Francisella since phages for this organism have not been described before (LoVullo et al, 2009a ; Rydzewski et al, 2015 ). Further cryptic plasmids and a putative conjugative element have been described recently and may be used to generate further plasmids for Francisella in the future (Siddaramappa et al, 2014 ; Challacombe et al, 2017b ).…”

Section: Introductionmentioning

confidence: 99%

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Construction of a New Phage Integration Vector pFIV-Val for Use in Different Francisella Species

Tlapák

Köppen

Rydzewski

et al. 2018

Front. Cell. Infect. Microbiol.

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We recently identified and described a putative prophage on the genomic island FhaGI-1 located within the genome of Francisella hispaniensis AS02-814 (F. tularensis subsp. novicida-like 3523). In this study, we constructed two variants of a Francisella phage integration vector, called pFIV1-Val and pFIV2-Val (Francisella Integration Vector-tRNAVal-specific), using the attL/R-sites and the site-specific integrase (FN3523_1033) of FhaGI-1, a chloramphenicol resistance cassette and a sacB gene for counter selection of transformants against the vector backbone. We inserted the respective sites and genes into vector pUC57-Kana to allow for propagation in Escherichia coli. The constructs generated a circular episomal form in E. coli which could be used to transform Francisella spp. where FIV-Val stably integrated site specifically into the tRNAVal gene of the genome, whereas pUC57-Kana is lost due to counter selection. Functionality of the new vector was demonstrated by the successfully complementation of a Francisella mutant strain. The vectors were stable in vitro and during host-cell infection without selective pressure. Thus, the vectors can be applied as a further genetic tool in Francisella research, expanding the present genetic tools by an integrative element. This new element is suitable to perform long-term experiments with different Francisella species.

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The hidden life of integrative and conjugative elements

Delavat

Miyazaki

Carraro

et al. 2017

FEMS Microbiology Reviews

177

201

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Integrative and conjugative elements (ICEs) are widespread mobile DNA that transmit both vertically, in a host-integrated state, and horizontally, through excision and transfer to new recipients. Different families of ICEs have been discovered with more or less restricted host ranges, which operate by similar mechanisms but differ in regulatory networks, evolutionary origin and the types of variable genes they contribute to the host. Based on reviewing recent experimental data, we propose a general model of ICE life style that explains the transition between vertical and horizontal transmission as a result of a bistable decision in the ICE–host partnership. In the large majority of cells, the ICE remains silent and integrated, but hidden at low to very low frequencies in the population specialized host cells appear in which the ICE starts its process of horizontal transmission. This bistable process leads to host cell differentiation, ICE excision and transfer, when suitable recipients are present. The ratio of ICE bistability (i.e. ratio of horizontal to vertical transmission) is the outcome of a balance between fitness costs imposed by the ICE horizontal transmission process on the host cell, and selection for ICE distribution (i.e. ICE ‘fitness’). From this emerges a picture of ICEs as elements that have adapted to a mostly confined life style within their host, but with a very effective and dynamic transfer from a subpopulation of dedicated cells.

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Identification and characterization of episomal forms of integrative genomic islands in the genus Francisella

Cited by 4 publications

References 27 publications

PRIMEIRO RELATO DE Francisella noatunensis SUBESPÉCIE ORIENTALIS NO SUL DO BRASIL – RELATO DE CASO

PRIMEIRO RELATO DE Francisella noatunensis SUBESPÉCIE ORIENTALIS NO SUL DO BRASIL – RELATO DE CASO

Construction of a New Phage Integration Vector pFIV-Val for Use in Different Francisella Species

The hidden life of integrative and conjugative elements

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