Quorum Sensing Controls Flagellar

Morphogenesis in Burkholderia
glumae
BY: M O O N S U N JA N G , E U N H Y E G O O , JA E H Y U N G A N , J I N W O O K I M , I N GY U
H WA N G
P R E S E N T E D BY: E R I C M AT H E W S A N D D E S E S PA N TA

Burkholderia
Burkholderia
glumae is the
glumae
causative agent of rice panicle

blight
Has multiple polar flagella
Has TofR/TofI QS system
Produces the phytotoxin toxoflavin

Rice panicle blight caused by B.

glumae
www.forestryimages.org

What is the goal?

The purpose of the study is to understand the effects of temperature and
quorum sensing on flagellar biogenesis and morphogenesis in B. glumae.

Figure 9 from Regulation of polar flagellum genes is

mediated by quorum sensing and FlhDC in Burkholderia
glumae

STRAINS

From Table 1 of Regulation of polar flagellum genes is mediated by quorum sensing and
FlhDC in Burkholderia glumae

Does temperature affect flagellar number

in wild-type B. glumae?
Cells were dipped into swim assay plates (of LB
and 0.3% agar) and incubated for 20 hours at
28oC and 37oC.
Cells were collected from the I, M and O regions
and observed by Transmission Electron
Microscopy.
Figure 1A
37o
C

28o
C

Wild-type B. glumae

Figure 1B

*n=100

At 37oC:
>77% had one or two polar flagella in all of the regions.
At 28oC,
>76% had two to four polar flagella in all of the regions.
Temperature does affect the number of flagella formed by wild-type B.
glumae cells. As temperature gets closer to the optimal temperature for B.
glumae growth (<30oC), flagellar number increases.

Does expression of flagellar biosynthesis

genes differ between 28oC and 37oC?
Fig. 2

By qRT-PCR, expression of flhC gene was

measured and found to be at statistically
equivalent levels for 28oC.

Fig. 2

Other flagellar genes, such as fliC and flgK,

were also measured (by use of glucuronidase assay) at 28oC and 37oC.
Results indicate higher expression levels for
both genes at 28oC.
In contrast, fliC and flgK expression levels
were down for the QS mutants at 37oC,
indicating that expression of these genes are
dependent on QS (at 37oC).

Does QS and temperature influence

flagellar biogenesis and morphogenesis?
Swim assays of BGR1, BGS2 and BGS9 were
performed at 28oC and 37oC.
At 37oC, QS mutants were largely aflagellate,
indicating that QS plays a large role in flagellar
biogenesis at this temperature.
At 28oC, QS mutants were predominantly
flagellated, with approximately half of these
exhibiting non-polar flagella.

Figure 3

QS and flagellar morphology

Previous studies indicate that QS affects flagellar biogenesis.
Whether QS is directly or indirectly involved in flagellar morphology is still
unclear.
QS both positively and negatively regulates flagellar biogenesis in other
organisms.
In B. glumae, although flagellar biogenesis is independent of QS at 28 oC, QS is
still required for normal flagellar morphology at this temperature, showing that:
QS and temperature work together to modulate biogenesis and
morphology of flagella in B. glumae.

Flagellar morphology observed under TEM

Figure 4

Fig. 6

Non-polar flagella (Peritrichous and

Lateral)

BGS2

BGS9
- Sites of flagellar formation

How does lack of QS affect directional

swimming movement?

Cell movement was recorded under a phasecontrast microscope for 30 seconds.

As expected, at 37oC, BGR1 exhibited normal

swimming movement. BGS2 and BGS9 were
largely aflagellate and were consequently nonmotile.
- Addition of C8-HSL to BGS2 recovered normal
swimming movement.

At 28oC, BGR1 showed normal motility at the I

region. BGS2 and BGS9 showed tumbling and
spinning motility.
- Addition of C8-HSL to BGS2 recovered normal
swimming movement.
At 28oC, all strains showed normal motility in the
O region.

Figure 5

What is FlhF and what is its role in B. glumae?

FlhF is an SRP (Signal Recognition Particle) type
GTPase that is essential in flagellar formation.

Figure 6

Fig. 6

Fig. 6
plasmid
carrying the
fliA gene
wt
phenotype
recovered

Using swim assays, swimming motility of

the BGR1 mutant fliA::Tn3-gusA45 was
examined at 28oC and 37oC.

At 28oC, expression of flhF gene was

analyzed by qrt-PCR. BGF41 (flhC
mutant) made little to no flhF.

FlhF in other organisms

The flhF deletion mutant:
is aflagellate in V. cholerae and C. jejuni.
has randomly arranged flagella and is non-motile in P. aeruginosa.
Conversely,
Overexpression of FlhF causes hyperflagellation in P. putida.
FlhF expression levels and localization are important for normal flagellar
formation (in polarly flagellated bacteria).

What is the relationship between QS and FlhF?

Hypothesis: QS deficiency causes mislocalization of proteins involved in flagellar
formation, like FlhF in B. glumae.
Hypothesis was supported by the mislocalization of FlhF in QS mutants.
Whether the dysfunctional FlhF directly causes abnormal flagellar formation or
mislocalizes other flagellar body proteins (thereby indirectly affecting flagellar
morphology) is yet to be determined.

Conclusion

As established in the 2007 paper, normal polar flagellar

formation is essential for the pathogenicity of B. glumae.

Abnormal flagellar morphology may impair swimming movement

of B. glumae and hinder invasion success in rice flowers.
Therefore, this characteristic may be used to develop methods for
reducing the spread of rice panicle blight and increase produce.