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Proteus mirabilis

• gram negative, facultative anaerobe rod

• causes infections in human urinary tract (pyelonephritis, kidney stones) followed by septicaemia

• motile bacterium with peritrichous flagella

• Morphology change in swarming process

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Swarming proteus mirabilis• „bulls-eye“ swarming on solid surfaces & self

identity

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Boundary between P. mirabilis strain BB2000 and HI4320

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Aim of study

• Identification and characterization of the genetic mechanism that is responsible for bacterial self identity

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Construction of p. mirabilis mutant library

• randomized Tn5-transposon mutagenesis of rifampicin resistent p. miarabilis BB2000

Tn5-cat Shuttle vector

• conjugal transfer by mating of E.coli [pUTminiTn5-Cm] and BB2000

cat

tnp: Transposase; mobRP4: conjugal transfer domain

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Loss of self recognition I• Screening of 3600 transposon BB2000 mutants

revealed one mutant with phenotypic loss of self recognition

„identification of self“ (Ids)

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Loss of self recognition II• six gene cluster idsABCDEF carries information for

„identification of self“

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Phenotyping of individual idsABCDEF knock outs

idsB – idsF are required for self recognition !

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Phenotyping of individual idsABCDEF knock outs in

HI4320 background

idsD and idsE are essential for self recognition ! idsB, idsC and idsF can be complemented by allels of different strains

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idsD and idsE encode identity determinants

….but the ids gene cluster is not the sole determinant of boundary formation !

boundary between BB2000+pidsHI4320 and HI4320+pidsBB2000

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Conserved organization of ids gene cluster in other P.

mirabilis isolates

idsABCDEF with nearly identical length and ~96% identity to homolog idsD and idsE have variable domains in protein sequences

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Additional putative idsE genes in genome of P. mirabilis

HI4320

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Model of self recognitation

idsA is not required for self identity; idsF overtakes function distinct from idsB

and idsC in self-nonself recognition; idsD and idsE are molecular self identifiers

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Summary• gene locus idsABCDEF of P. mirabilis is important for

self-nonself discrimination and boundary formation• Conserved ids gene cluster has been found in other

proteus mirabilis strains• idsA is not required for parental recognition• idsB, idsC and idsF are interchangeable between

strains• idsD and idsE are specific molecular identifiers that

possess strain dependent variable domains • other ids homologs in bacterial genome may

contribute to the self recognition process

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Thank you for your attention

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P. mirabilis boundaries

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Transposition and pBBR-MCS2 shuttle vector for mutagenesis