Genomic island excisions in Bordetella petrii.

BACKGROUND: Among the members of the genus Bordetella B. petrii is unique, since it is the only species isolated from the environment, while the pathogenic Bordetellae are obligately associated with host organisms. Another feature distinguishing B. petrii from the other sequenced Bordetellae is the presence of a large number of mobile genetic elements including several large genomic regions with typical characteristics of genomic islands collectively known as integrative and conjugative elements (ICEs). These elements mainly encode accessory metabolic factors enabling this bacterium to grow on a large repertoire of aromatic compounds. RESULTS: During in vitro culture of Bordetella petrii colony variants appear frequently. We show that this variability can be attributed to the presence of a large number of metastable mobile genetic elements on its chromosome. In fact, the genome sequence of B. petrii revealed the presence of at least seven large genomic islands mostly encoding accessory metabolic functions involved in the degradation of aromatic compounds and detoxification of heavy metals. Four of these islands (termed GI1 to GI3 and GI6) are highly related to ICEclc of Pseudomonas knackmussii sp. strain B13. Here we present first data about the molecular characterization of these islands. We defined the exact borders of each island and we show that during standard culture of the bacteria these islands get excised from the chromosome. For all but one of these islands (GI5) we could detect circular intermediates. For the clc-like elements GI1 to GI3 of B. petrii we provide evidence that tandem insertion of these islands which all encode highly related integrases and attachment sites may also lead to incorporation of genomic DNA which originally was not part of the island and to the formation of huge composite islands. By integration of a tetracycline resistance cassette into GI3 we found this island to be rather unstable and to be lost from the bacterial population within about 100 consecutive generations. Furthermore, we show that GI3 is self transmissible and by conjugation can be transferred to B. bronchiseptica thus proving it to be an active integrative and conjugative element CONCLUSION: The results show that phenotypic variation of B. petrii is correlated with the presence of genomic islands. Tandem integration of related islands may contribute to island evolution by the acquisition of genes originally belonging to the bacterial core genome. In conclusion, B. petrii appears to be the first member of the genus in which horizontal gene transfer events have massively shaped its genome structure.

Genomic content of Bordetella pertussis clinical isolates circulating in areas of intensive children vaccination.

BACKGROUND: The objective of the study was to analyse the evolution of Bordetella pertussis population and the influence of herd immunity in different areas of the world where newborns and infants are highly vaccinated. METHODOLOGY: The analysis was performed using DNA microarray on 15 isolates, PCR on 111 isolates as well as GS-FLX sequencing technology on 3 isolates and the B. pertussis reference strain, Tohama I. PRINCIPAL FINDINGS: Our analyses demonstrate that the current circulating isolates are continuing to lose genetic material as compared to isolates circulating during the pre-vaccine era whatever the area of the world considered. The lost genetic material does not seem to be important for virulence. Our study confirms that the use of whole cell vaccines has led to the control of isolates that were similar to vaccine strains. GS-FLX sequencing technology shows that current isolates did not acquire any additional material when compared with vaccine strains or with isolates of the pre-vaccine era and that the sequenced strain Tohama I is not representative of the isolates. Furthermore, this technology allowed us to observe that the number of Insertion Sequence elements contained in the genome of the isolates is temporally increasing or varying between isolates. CONCLUSIONS: B. pertussis adaptation to humans is still in progress by losing genetic material via Insertion Sequence elements. Furthermore, recent isolates did not acquire any additional material when compared with vaccine strains or with isolates of the pre-vaccine era. Herd immunity, following intensive vaccination of infants and children with whole cell vaccines, has controlled isolates similar to the vaccine strains without modifying significantly the virulence of the isolates. With the replacement of whole cell vaccines by subunit vaccines, containing only few bacterial antigens targeting the virulence of the bacterium, one could hypothesize the circulation of isolates expressing less or modified vaccine antigens.