2019sacls585 Remerciemen

J ' Aimerais Également Remercier

Satoko Yoshizawa

Philippe Bouloc

Yoshiharu Yamaichi

Sofian Ouchane

Anne-Soisig Steunou

Virginia Lioy

Stéphane Duigou

Bénédicte Michel

Christophe Possoz

Elena Espionosa

Claire Morvan

Brigitte Meunier

Nara Figueroa-Bossi

Lionello Bossi

Mickael Poidevin

Laurent Kuras

Une

Stéphanie Bury-Moné

Structure of aminogl cosides

Sites of attachment in the ribosome

Mechanism of uptake

Consequences of the presence of aminogl cosides in bacterial cells

H e e , he de a f ac a e e be de e ed.

Phenot pic suppression b aminogl cosides

Importance and danger

Persisters in biofilm

The The

ATP level

A

Aminogl coside persisters

How bacteria adapt to changes and water movements?

Discover of mechanosensitive channels

The

Structure and gating of mechanosensitive channels

The A h e a c ea f he c e g a c e f he cha e f g a e f he de effec f he f e ce e ag.

Visuali ation of MscL

Number of channels in the cells

Aminogl coside interaction with MscL

Voltage-gated channel vs. mechanosensitive channels

The I

Microfluidic in persister stud

MIC measurement

E citation and emission

A (Tab e 4-1). I c f ed ha cha g g he a f a b c ec e e ce c ca f he e ha .

C 5 interaction with cells

FACS

Conclusion

Super resolution microscop

A

Conclusion

Mechanosensation and Neo-C 5 accumulation

Centrifugation and cell resuspension methods

Ce

Mechanosensitive channels role in aminogl coside accumulation

I ha e e ed he e he effec f e f echa e e cha e E. MG1655, a he e cha e ha e bee e ed ha e aff e c . e che e e a .

Strain construction

MIC of cL and cS

F

C toplasmic accumulation of

Conclusion

Sorting out cells with lower ribosomal RNA e pression

I

Dilution effects on persister recover time

Conclusion

Design

Coating

Conclusion

Strains used in this stud

E a MG1655: W d e a
F-, a bda-, G-, fb-50, h-1

MG1655 : BP- ( )
S g e a :

MG1655 A MG1655 C MG1655 L MG1655 S

MIC measurement

The 1935 1942 (2006).

Chapter XII Resum en Fran ais

  Abbreviation BF: B gh F e d CCCP: Ca b C a de -Ch Phe h d a e C : Ch a he c C 3: C a e 3 C 5: C a e 5 D BAC 4-(3): b -(1,3-d b ba b c ac d)-e h e DMSO: D Me h S fO de DNA: De b N c e c Ac d EDPI, II: E e g -De e de Pha e I, II FACS: F e ce ce-ac a ed ce g GFP: G ee F e ce P e LB : L a-Be a b h LPS: L P Saccha de MIC: M I h b C ce a MS cha e : Mecha e e cha e M cL: Mecha e e cha e f a ge c d c a ce M cS: Mecha e e cha e f a c d c a ce Ne -C 5: Ne c -C a e5 OD: O ca De PCR: P e a e Cha Reac PDMS: P d e h a e PI: P d I d de PMF: P M ce F ce RNA: b a RNA STORM: S cha c O ca Rec c M c c TA: T A TIRF: T a I e a Ref ec F e ce ce c c TR: Te a Red RNA: a fe RNA VBNC: V ab e b c ab e

Figure 1 . 1

 11 Figure 1.1 Antibiotics target the functionall important regions of ribosome 1 .T b a b a e d a ed: 30S b ( ef ) a d 50S ( gh ). RNA ( e a d g e ) a d b a e ( b e a d b e) a e h . I each b , he e f a b c b d g, h ch ha bee de e ed b b che ca a d ge e c ech e a e d ca ed. The de a e e b a ed b X-a c a g a h .

Figure

  Figure 1.2 E amples of aminogl coside chemical structures. O he ef a e e a g c de f he e c g (4 5 g II g I, g II g III age ), h ch ca be d ec c a ed he ge a c c e he gh (4 6 g II g I, g II g III age ). R g a e be ed a d he che ca c e f a ab e d ca ed. R g II ade f de e a e, h ch e c e ed 9 .

  he ab e ce f he d g (F g e 1.3).

Figure 1 . 3 A

 13 Figure 1.3 Rearrangements of rRNA helices h44 and H69 in the near-cognate state upon binding of the aminogl coside paromom cin (PAR) 29 . Pa c (g ee ) b d he dec d g e 16S RNA. The RNA d a ed ed a d RNA e . The d ffe e ce be ee c e e f ed h ea -c g a e RNA e e ce a d ab e ce f a c a e h gh gh ed da a d gh b e.

Figure 1 . 4

 14 Figure 1.4 Overview of binding sites of neom cin in ribosome 32 . (a) Ne c (Ne ) da g ee 30S b , he 44 (b e) a d e c H69 ( gh b e) f he 50S b (g a ). RNA (g ee ), RNA ( ). (b) Ne c e ac h he H69. Da hed e d a e c a d RNA

Figure

  Figure 1.5 Gram-negative bacteria cell wall envelope 37 . (a) P a h a ( e ed a g c de a e) (b) Se f-ed a g c de a e h ch ca a d che a ca d ca e d a e ca f LPS.

Figure 1 . 6 .

 16 Figure 1.6. Model for the role of silver in aminogl coside uptake 51 . S e a g e a g c de a e. S e ca d ce e b a e b cha g g e b a e e . I ca a a ge b e , h ch e he f f ded e .

Figure

  Figure 1.7 Model for effect of chaperone on earl aminogl coside e posure in bacterial killing or survival 59 . A a a e f a g c de ac e e e a d b d g b e e c ded e de ( a ge a ). M ead e de a e b a e f e e a cha e e . L ed cha e e ca ac e e f d g, ead g ce dea h d e a c ea e f e b a e e eab . O he c a , h ghe ca ac f cha e e , a c a G EL/G ES, c ec he f d g h e e a d c ea e bac e a a .

Figure1. 8

 8 Figure1.8 c-di-GMP role in biofilm formation in presence of aminogl cosides 60 . I P. a g c de a d ec e ac h A d ec h gh cha g g e e f c-d -GMP.

Figure 1 . 9

 19 Figure 1.9 Suggested mechanism of h dro l radical formation and cell death 54 . (A) M f ded e b a e e a e ca ed e b a e a ca c e e (e.g., SecYEG) b cha e e e (SecB). M f ded e a e e e he e b a e a ca ed he e a (B) M f ded e ead h h a f C A. (C) Ac a ed C A h h a e C R, h ch eg a e e e f e e e e -e e e (DegP). (D) C A a ac a e A cA, h ch eg a e e a a d e ab c ge e . The e cha ge ce a e d ca e f ee ad ca f a , h ch ead ce dea h.

Figure 1 .

 1 Figure 1.10 Model for Ap4A alarmone action in aminogl coside killing 68 . A g c de ca e he d c f f ded e a d e b a e d a d ROS ge e a . ROS d ce he e e f L U, h ch e d c f A 4A. A 4A he eh h b ace a e a ce ( e a ed be e ed f he ), h ch ca e e d ce e b a e.

Figure

  Figure 1-2-1 Profiles of antibiotic killing kinetics of resistant, tolerant and persister cells 84 .The e f add f a bac e c da a b c d ca ed h a ed a . A h ge e a f e a bac e a h h b e e. Re a bac e a a c e g h. A h ge e e a bac e a a h ed gh g e a e ge e be ed b a a b c ha a ce b e a h da g e e. I he e ge a , he e e ce f e e h b a b ha c c e ( e ). Af e a b c ea e , he e a e d f a d g f ce b e ce f ed b a e dec ea e c f g (CFU).

Figure

  Figure 1-2-3 TA modules activated b stress responses lead to bacterial persistence 101 . S a a a d e ce f H A ead he e e ce f cha ged RNA he b a A e, h ch ead acc a f ( ) G h gh Re A a d f a f e e . ( ) G a d SOS e e ac a e ec f c e I a d e II TA d e ha ead e e f a .

Figure 1

 1 Figure 1-2-4 The model suggesting that lacking ATP leads to persister formation 100 . A dec ea e ATP ed ce he a b c a ge ac , h ch ead a b c e a ce.

Figure 1

 1 Figure 1-2-6 Mechanism of persister eradication ia the action of metabolites 107 . E ad ca f e e h gh ca ab f ca b ce , h ch ead ge e a f NADH. The d c f NADH de e de f he PPP, EDP TCA c c e. NADH d ed he e ec a cha f e b a e, h ch c b e c ea e f PMF. H ghe PMF ch ce fac a e a g c de a e.

Figure

  Figure 1-3-1 turgor pressure and osmolarit of E. c i 109 . E. g a ed , h ch ha 240 O (M a ed LB c a g 5 g/L NaC ). C a a e 10 a g e e a d he c a c e b a e. The ed e e f ab 6 a a d he e e b a e a d 4 a a d he e e b a e. MS cha e ga e f he e a cha ge f h e b e e b a a c ea e f 0,1 a f a d e e.

  e g h , h ch a e ed ge he b h e de . The e g h d ffe e ce a d . I ha bee f d ha he M cL a d M cS a e ac a ed 3 5 ec d af e ce each he ac a h e h d e .

  he c d g ge e f M cL, h ch he de c be a a e d ffe e

  ed TM1 a d TM2. The TM1 he ce a e a g he a -e b a e cha e a d he TM2 he ce f he e he a a f he cha e . TM1 a d TM2 a e c ec ed b

Figure

  Figure 1-3-2 Structure and gating of MscL 110 . (a) M cL c e f M . Th cha e c f 5 e . Each e ( ef ) c ed f -he ca a -e b a e d a ca ed TM1 a d TM2, c a c N-e a (S) a d C-e a eg a d a ce a e a c d a h ch he TM1 TM2. The f e TM1 d a a e a gh ac ed h d h b c b d e h ch a e he ga e f he e ( gh ). (b) 3D c c f M cL f E. he e b a e ( ) a d f ab e ( dd e). The cha e h c ed ( ef ) a d e ( gh ) a e . G a ba d ca e d h ha e f e a d e e b a e. The e c f g a h a he b

Figure 1 -

 1 Figure 1-3-3 Iris-like gating of MscL 121 .A ef he d ffe e age f ga g f M cL d a ed. The cha e c ed he c f a (C). Whe he e e he e b a e, he C ed-E e (CE) f . The e a e cha e eache he e c f a (O) h gh he e e f e c f g a cha ge (S1, S3 ) A he gh a ca g e c e ace f M cL ec ded a -50 V. A he d c f ed he ga g de a d gge ed ha M cL cha e a e e g a -e e a a d f a e g b e b a e e . I h d he f d ha he b dg g be ee C 20 a d C 36 f TM1 he ce ead ab ed e cha e 122 .

Figure 1 - 3 - 4

 134 Figure 1-3-4 MscL form clusters when all monomers were labeled b mEos3.2 in E. c i 125 . (a) Whe a d-de ed M cL-E 3.2 e e ed he e e ce f a e ch a M cL b , a he b f e cha e a e agged. I h ca e M cL cha e f ed e c e . (b) If a he b a e abe ed he ce , ca ed ce he a a f eed f E 3.2 a d ead f a f a f c a M cL c e .

Figure 1

 1 Figure 1-3-5 Cell level counts of MscL in different conditions with different techniques 126 .T d ffe e a f E. e e e ed d ffe e ed a ( ch LB a d ed a h d ffe e ca b ce ) de d ffe e g h ha e. The e cha e e e c ed b d ffe e ech e , (A) We e b h M cL a b d a d (B) We e b h GFP a b d a d f e ce ce c c ag g.

  b d g), e he cha e a g he a b c g h gh he e cha e he c a . The cha g g he M cL a e he K+ a d g a a e ea . (F g e 1-3-6) 132 .

Figure 1

 1 Figure 1-3-6 Interaction model of streptom cin to MscL 130 . Whe e c he ed a e ac h M cL cha e h ch ead ac a f M cL. Th cha e ga g a K+ ea age. S e c ee he M cL e a d he cha e bec e a a h a f he e f he a b c he c a .

Figure

  Figure 1-3-7 Calcium gated channels are induced b mechanosensation 134 . (a) I a c f d c cha e ce a e c aged he d ed a a d d g he echa ca a b h ca c ac h a ece f 1% -e aga e ad he f he ce . Ce g ee a e e e g a e e ge e (c GFP-ba ed ca c e ) h ch h he ac a f ca c ga ed cha e . (b) S g e-ce ca c ec d g bef e a d af e echa ca a . Each c f ace ec ded f g e ce bef e a d af e a .

Figure 1

 1 Figure 1-4-1: E ample of a microfluidic device used for stud ing persister cells 95 . Ce e e a ed a d g e a cha e . M c f d c a ed f e cha ge a d ca ed a he b f he f e ce ce c c age . A he e f add f a ed a c a g a c ( a ge ec a g e), f he ce ed e ce e e ( ed a ). Af e e g he a b c h a e cha ge f he he c f d c e , e e e ed g h. Pe e f ca a b a h a ce a h f e e c e f cc e ce a g g f 10 -6 10 -1 . O he he ha d g he e ge c a ce a e f he a fac f e e f a , e e a de e a de ce ab e f ch d 136 . I ece ea , c f d c c b ed h f e ce e e (f ed h T -A e e ab c e e ) e e ed cha ac e e e e f a 97, 137 .

  bac e c da ac f Ne -C 5 Ne -C 3, af e each he , I ha e ea ed he MIC f each c ga ed a b c c a he a e a d g e c . A h gh ha MIC a e ca be d ffe e acc d g he ed a h ch he ce a e g g, I e ed he MIC he ed a MOPS-G. The e e c e e ed e da a f ab he e he MIC a e ed ed a M9-G a d LB. We b e e ha MIC h ghe M9 ed a. The a d ffe e ce be ee he e ed a ha 58 M9 c a h ghe c ce a f h ha e. I ha e e ed e

Figure 3

 3 Figure 3-1 E citation and emission spectra of C 5 and Neo-C 5. Da a e e ec ded

Figure

  Figure 3-2 Activit of Neo-C 5 is PMF dependent. S a f E. MG1655 ce af e a 4.5-h ea e h 1.5 M e c 48 M Ne -C 5 h a d h 30 M CCCP (2-h c ba add f he a b c). P ed a e a e e a e ea ed c c e. E ba a e a da d e f he ea a d e e ca c a ed f h ee de e de e e e . S g f ca d ffe e ce , ca c a ed b he Ma Wh e e , a e d ca ed (****P < 0.0001).

  he MIC f C 5, a d he e h ed ha C 5

  a e a d a a ed b FACS (F g e 3-3).

Figure

  Figure 3-3 C 5 vs Neo-c 5 interaction in FACS anal sis. E. MG1655 ce e ed 32 M C 5 Ne -C 5 f 5 e a 37 C. Ce e e a hed a 0.45 -e-e HAWP e b a e f e ( ed ce e e e , Me c M e). B d g f Ne -C 5 a c ea d c f e e f ce a a f e ce ce e d a b d g f he d e C 5.

Figure 3 -

 3 Figure 3-4 C 5 does not enter live E. c i cells. Lef : a ed gh age f a f e d f E. MG1655 ce e ed 32 M C 5 f 1 h a 37 C. Ce e e a hed a 0.45 -e-e HAWP e b a e f e ( ed ce e e e , Me c M e). R gh : f e ce ce age f he a e f e d f e b a ed h a e ec -g ga f 50 a d a e e f 50 .

  Figure 4-1-A

FigureFigure 4

 4 Figure 4-2 Neo-C 5 killing curve. E. ce e e ea ed h Ne -C 5 (32 M) a 37 C a d a each e ce e e a hed f e a d d ded a , e a d ed e e a d ed LB MOPS-G a e f CFU c g. The ec d a a ed f c c . S f C 5 f e ce ce e each bac e ea ed b I ageJ.

Figure 4 - 4

 44 Figure 4-4 Effect of cell densit on antibiotic lethalit . A) Ce a OD: 0.2 e e ea ed h 32 M Ne -C 5 a 37 C. A each e , ce e e a hed f e a d aga ad d ec . B) C a g he Ne -C 5 acc a d ffe e OD (0.2 0.02) a d ffe e e . Ce h h ghe OD ha e e acc a .

Figure

  Figure 4-5 EDPI and EDPII inhibition b chloramphenicol treatment. FACS a a f ce ea ed h Ne -C 5 a d ffe e e b e a d he ce ea ed h ch a he c Ne -C 5 ed. A 60 a d 150 , EDPII cc ed he ce ea ed e b Ne -C 5; h e e , he acc a a c ea g ce e-ea ed h ch a he c .

Figure

  Figure 4-6 Septum accumulation of Neo-C 5. Whe ce e e ea ed f h a f e h a -e ha c ce a f Ne -C 5, bef e ce d ce e beca e b gh e C 5 g a . Af e ce d , e e e e ch b gh e C 5 g a ha d e .

Figure 4 - 7

 47 Figure 4-7 Super resolution images of E. c i cells treated with tagged aminogl coside. Ce a OD 0.02 e e ea ed b 4 M Ne -C 4 f 2 a 37 C, he a hed he f e . Ce e e aced d ec aga ad f ag g. N e ha ce he c c e he e a a de a f ab 15-20 e bef e ac f he e age . Pe he a acc a f a b c h ge a d he e a e Ne -C 5 c e de ec ed he e b a e ( d ca ed b e a ).3.3. Number of cell washes

Figure

  Figure 4-8 Removing Neo-C 5 e cess b filtration. E. MG1655 ce (OD600 :0.02) e e c ba ed 5 e h 32 M Ne -C 5 a 37 C he f e ed a 0.45 -e-e HAWP e b a e f e ( ed ce e e e , Me c M e). Ce e e a hed h 100 L MOPS-G ed ( e a 37 C), ec e ed f he f e a d ed a e a a ed b FACS. The a e age f Ne -C 5 g a f each a e a ea ed. Th ee a h g e e e e ed e e he e ce f Ne -C 5.

Figure

  Figure 4-9 Membrane saturation b Neo-C 5. A) FACS a a f ce a OD 600 : 0.02 ea ed h d ffe e c ce a f Ne -C 5 a d he a hed he f e ed a d a a ed b FACS. The c a c acc a he gh a f he a e a e ce age. B) Pe a c C 5 g a ed f each c ce a . F 32 M a d ab e Ne -C 5 b d g eached a a . I e ed a e e f d ffe e c ce a f Ne -C 5 he ce a d he e de a ed ha he e b a e a each g a a a 32 M f Ne -C 5 (F g e 4-9). I e e g c ea g he c ce a f Ne -C 5 64 M d d cha ge he e b a e

Figure

  Figure 4-10 Kinetics anal sis of accumulation of Neo-C 5 and PI b FACS. E. ce (OD 600 : 0.02) ea ed b Ne -C 5 a 32 M a 37 C ef a e ea ed b Ne -C 5 a d a PI a gh . Ce e e a a ed a d ffe e e . P e g a e ce age f C 5 a d PI a e d ca ed f each e e e .

  ce af e ea e . I ab, a c had bee e ab hed h

Figure

  Figure 5-1 Accumulation of Neo-C 5 in e ponential phase cells. Ce e e c ba ed MOPS-G ed a f 2 h b-MIC c ce a f Ne -C 5 a 37 C a d he ce e e a hed 3 e b ce f ga a d cha g g he ed a. The e a e d c Ne -C 5 acc a a e : e he a a d c a c. C a c b-a e ha 10%. Se f ce d ha e a h ghe Ne -C 5 g a . Sca e ba 5 . I e ea ed he a e c a d a a ed he ce b FACS (f g e 5-2). The C 5 e ea c e d he e he a ( e ) a d c a c (h ghe ) acc a e e . The e ce age f each ea de e d he e c d .

Figure

  Figure 5-2 Anal sis of Neo-C 5 accumulation patterns b FACS. E e a ha e ce a OD: 0.3 e e d ed 10 e a d 100 e . Each ce c e a he ea ed h b-MIC c ce a f Ne -C 5 f 2 a 37 C. Ce e e he a hed 3 e b ce f ga f cha g g he ed a. Ce e e he a a ed b FACS f C 5 g a . The ea h e f e ce ce e h g he e he a b d g f Ne -C 5 a d he ec d h ghe ea h g he c a c a e. The e ce age f each ea h he ab e.

Figure

  Figure 5-3 Measuring membrane potential in the diluted cells in different media and comparing to non-diluted cells. E e a ce e e d ed MOPS ed a h h d ffe e ca b ce he ea ed h D BAC4-(3). A a c f e g a , ce e e ea ed h e ha 60% de de a e he ce he a hed a d ea ed b D BAC4-(3). G: G c e, CAA: ca a ac d.2.1.4. Different Neo-C 5 concentrations and time of incubation

Figure

  Figure 5-4. Patterns of Neo-C 5 accumulation in e ponential phase cells. A) E e a ce OD: 0.3 e e ea ed h d ffe e c ce a f Ne -C 5 f 2 he a hed b he ce f ga c . Ce e e he a a ed b FACS f C 5 g a . B) E e a ce OD: 0.3 e e ea ed b 0.4 M f Ne -C 5 f d ffe e e f c ba he b ec ed he a e c f a a . P: Pe he a , C: C a c acc a .

Figure

  Figure 5-5. Cells with c toplasmic accumulation are PI positive. E e a ce e e d ed 10 e he ea ed h 0.4 M f Ne -C 5 a 37 C f d ca ed d a . Ce e e he a hed b ce f ga he c ba ed h PI (15 ) a d a a ed b FACS f C 5 a d PI g a .

Figure

  Figure 5-6. Resuspension methods trigger mechanosensation in cells and change c toplasmic accumulation of Neo-C 5. Ce e e g a OD: 0.3 he d ed 10 e . Ce e e c ba ed a 37 C h b-MIC c ce a f Ne -C 5 f 2 . The ce e e ce f ged a 8000g f 5 a d e e d he e ed a e he b a g ge be ed b e . Each e e e a d e e . The c a c e ce age ( d ca ed he f g e) cha ged gh a d he ea f e he a Ne -C 5 b d g a ed a e . C: c a c, P: e he a .

Figure

  Figure 5-7. Centrifugation should be followed b pipet mi ing the cells to trigger accumulation of Neo-C 5 in the c toplasm. Ce e e g a OD: 0.3 he d ed 10 e . Ce c ba ed a 37 C h b MIC c ce a f Ne -C 5 f 2 . The ce e e ce f ged ( a d ca ed) a 8000g f 5 a d e e ded he e ed a e he b a g ge be ed b e . The c a c e ce age a e d ca ed he f g e. C: c a c, P: e he a .

Figure

  Figure 5-8 Rifampicin treatment inhibits accumulation of Neo-C 5 in the c toplasm induced b centrifugation washing steps. E e a ha e ce (OD: 0.3) e e d ed 10 e . Ce c ba ed h 30 M fa c f 10 e e he c ba ed h b-MIC c ce a f Ne -C 5 f 2 a d a hed h ce f ga c bef e FACS a a . P e he a , C c a c acc a .

  c d g f F ec b a e. The a e e he e f ed b PCR (f g e 5-9).

Figure

  Figure 5-9. Verification of the construction of the mutants cL and cS b PCR. S g e-c PCR af e a c c c a h WT a h he a e a f ec f c DNA e . The ba d a d he e c e d g he a f ed DNA f ag e a e d a ed.

  Neo-C 5 in cL strain I c ba ed he WT a d L a h Ne -C 5 ( b-MIC c ce a ) a d he a hed he ce b ce f ga a d ed he be ha e de ce e

Figure

  Figure 5-10. cL has smaller percentage of cells with Neo-C 5 c toplasmic accumulation comparing to WT. Ce e e a ha e f g h e e d ed 10 e a d c ba ed h b-MIC c ce a f Ne -C 5 f 2 a 37 C. Ce e e ce f ged, a hed h MOPS-G MOPS-G h Ne -C 5 a hed. The ce , a e a e a a f he ce a hed h MOPS-

Figure

  Figure 5-11. Cells with MscL are l sed due to Neo-C 5 sub-MIC treatment after centrifugation. E e a WT a d L ce e e d ed 10 e a d ce f ged f 5 a 8000 g. 30 ec d af e ce e e e e ded he MOPS-G ed a h Ne -C 5 (0.4 M) b g ha g. Sh af e ce e e aga ad a d b e ed b f e ce ce c c ( ef ) a ed LB aga ( gh ).

  f e e g e e a d a e f each bac e a. The ce h

Figure 6 - 1

 61 Figure 6-1 Persisters for oflo acin have peripheral accumulation of Neo-C 5. S a a ha e ce e e ea ed h f ac , a hed h PBS a d ea ed h a b-MIC c ce a f Ne -C 5. A e e f e-a e he f e d he e a e f e e h dead ce . A e a e e e a g a d ha h e de f ed he . Pe e ha e e he a acc a ( e a ) acc a (g ee a ). N -e e ce d g ( ed a ) a d c e d dead ce VBNC . I h e e e I ed a b-MIC c ce a f Ne -C 5, a a e f e e ce a ce g h. H ghe c ce a f Ne -C 5 d ha e e e e e he d a a e.

Figure 6 - 2

 62 Figure 6-2 Sorting cells with FACS according to their si e and Neo-C 5 signal. E.

  he GFP g a dec ea ed a e ec ed (f g e 6-3-B).

Figure

  Figure 6-3. Ribosomal RNA e pression as a marker for persister rich sub-population. A) ch e e f ( ) h BP1 acZ ge e E. MG1655. B) K e c ea e e f GFP(ASV) e e . C) GFP f e ce ce e e ea ed b FACS WT a d ce h c d g GFP(a ) e e a ha e f g h.

Figure 6 -

 6 Figure 6-4. FACS anal sis of Neo-C 5 accumulation b less tolerant mutants and WT E. c i strains. S a a ha e ce e e ea ed h 128 M f Ne -C 5 f 3h a 37 C. The ce e e a hed a d a a ed b FACS. Ne , de cha ac e e be e he e ac f Ne -C 5 h he e a e e ga ed b d g ce e b a e. I ha bee e ed ha PMF e e he e a h ghe 108 . H e a ed e b a e a a e e he a b d g a e a c a c acc a . I d d a e e f FACS a d c c e e e ce f d ffe e Ne -C 5 c ce a a d d ffe e c ba e f each a . T a a e he e he a b d g b f e ce ce c c , I ea ed a a ha e ce h (2 ) h Ne -C 5 a 32 M. I b e ed ha Ne -C 5 acc a a e he a a d h ghe A

Figure

  Figure 6-5 Anal sis of membrane accumulation of Neo-C 5 in aminogl coside less tolerant mutants. S a a ha e ce ea ed h 32 M Ne -C 5, c ba ed a 37 C f 2 150 . Ce e e a hed a d a a ed b c c ( a d b ef ) FACS (b gh ). F e c a c c age a e e e ed f d ffe e e g . G: Ga , E T: E e T e.

Figure 6 - 6 .

 66 Figure 6-6. Kinetics of accumulation of Neo-C 5 (128 M) in less tolerant cells. S a a ce f a a d WT e e ea ed b 128 M Ne -C 5 a 37 C. Af e 2 a d 180 , ce e e a hed a d a a ed b FACS.

Figure 6 - 7 .

 67 Figure 6-7. Neom cin persisters e it the dormant state when cells were diluted. S a a ce ea ed h a c ce a f e c f 500 e he MIC ( ef ) f ac ( gh ) a 37 C f 6 h . Ce e e he a hed a d d ed f e h MOPS-G ed a. Ce e e d ed e e : 2, 4, 8, ,128, 256 e (C: Ce c ce a ). The ce OD a ea ed g a a e eade f 16 h .

Figure 6 - 8 .

 68 Figure 6-8. Persisters were not in carbon source (glucose) limitation during their recover . Ne c e e d ed 16 e MOPS ed a c a g d ffe e g c e c ce a . The e e g h ca de (OD) a 600 a ea ed h a a e eade f 20h. I ee ed ha he e a a fac che ca he e a a f dead ce a d e e ha a e e af e ea e h e c . Whe h fac a e d ed, a ed e e a e ea e a d he a e c ce a ed a a ed e e he d a a e. Whe e e e e d ed h e a a he d d a e a a . I ca ed h ed a: e-c ba ed ed a, h ch a f e h MOPS-G ed a ha a c ac h dead ce a d e e f 15 (F g e 6-9-A). Ye aga , h e a ec f c e e f e c a d f ac (f g e 6-9-B). O he he ha d, he e a a f e c e e had h e a e b g h f -e e ce a e a e e f f ac (f g e 6-9-C).

Figure 6 - 9

 69 Figure 6-9 Pre-incubated media with neom cin persisters perturbed cell growth. I ef c , ce

Figure 6 -

 6 Figure 6-10 Pre-incubated media with neom cin persisters did not perturb cell growth of neom cin resistant cells. I ef c ce d ed f e h MOPS-G ed a ( d ca ed h a a e bea e ).

Figure

  Figure 7-1. Design of microfluidic device. A) a f he c c . I ha 2 a d 2 . B) Bac e a ce e e aded he c c f e 1 a d he ed a c a g Ne -C 5 a added f 2. C) Ce e e a ed he cha e a d he f a b c a ed c ac h he bac e a a 1-c de cha e c ec .

Figure7- 2 .

 2 Figure7-2. Neo-C 5 binds to h drophilic surface causing a ver high background. Lef a e : B gh f e d age f c f d c cha e . Bac e a a e b e. R gh a e : f e ce ce c c age f he a e f e d f e . D e h gh bac g d e e , Ne -c 5 e ac h bac e a b e.

Figure7

  Figure7-3. Plasma treatment makes the surface h drophilic for a long time. A) C e a d PDMS bec e h d h c af e a a ea e , h ch ead f ee -OH g . Th a f g c a e b d be ee he PDMS a d he g a . B) P c e f c ac a g e ea e e . C) E f he h d h c e f a c e af e a a ea e a a f c f e.

  , S., Wa e , S. E. & F ed c , K. Re e e T a ca f RNA he R b e. M . C 24, 931 942 (2006). 16. B a a, M. A., Sh , S., H , J. M., F ed c , K. & Ca e, J. H. D. A S e c B c dc c , J., M a ed, D., Ca , M., Da e , J. & N e , H. F. I e ac f a b c h A-a d P-e-ec f c ba e 16S b a RNA. EMBO J. 10, 3099 3103 (1991). 20. M a ed, D. & N e , H. F. d g A S e. S 9, 647 658 (2001). 23. L ch, S. R., G a e , R. L. & P g , J. e, J. M. Rec g f C g a e T a fe RNA b he 30S R b a

  e . FEBS L . 587, 1848 1857 (2013). 34. M e e , J. P. & M e , P. F. Bac e a U a e f A g c de A b c . MICROBIOL RE 51, 19 (1987). 35. R e a, M., Ha c c , R. E., Sa e , J. G., Ha g, A. & McG a , E. J. E ha ced b d c c , R. E., Fa e , S. W., L , Z. S. & P e, K. I e ac f c e a ce. P . N . A . S . 111, 355 360 (2014). 39. M , M. E., Va Hee c , R. S. & Wa ace, B. J. Effec f de b E che ch a c .

f

  e, M., D e ge , Y., Ma -Ve ae e, I., Ba a , F. & E a , B. Me b a e P e a d T -C e S e Ac a T gge A b c-Med a ed Ce Dea h. C 135, 679 690 (2008). 55. L g, J. . P e Agg ega Ca ed b A g c de Ac I P e e ed b a H d ge Pe de Sca e ge . M . C 48, 713 722 (2012). 56. Va B ge e , R. A. & Ne dha d , F. C. R b e a e f hea a d c d h c E che ch a c . P . N . A . S . U. S. A. 87, 5589 5593 (1990). 57. B a ch , A. A. & Ba e , F.

  A., D e , D. J., Ha e e, B., La e ce, C. A. & C , J. J. A C Mecha f Ce a Dea h I d ced b Bac e c da A b c . C 130, 797 810 (2007). 62. F , J. J., De ad , B., W e , J. A., C , J. J. & Wa e , G. C. O da f he G a e N c e de P U de e Ce Dea h b Bac e c da A b c . ch a c . J. M . B . 57, 403 422 (1971). 64. Ke e , I., Ya a W , I ce c , J., M cah , L. R. & Le , K. K g b Bac e c da A b c D e N De e d Reac e O ge S ec e . S 339, 1213 1216 (2013). 65. D e , D. J. . A b c d ce ed -e a ed h g ca a e a a a f he e ha . P . N . A . S . 111, E2100 E2109 (2014). 66. Baha g , Z., K , E. & Ma e , D. R S P a a Ce a R e he SOS a b c a d e ha ce he bac e c da ac . P . N . A . S . 201822026 (2019) d :10.1073/ a .1822026116. 69. Pa e , E., W he , J. M. & She a ,

  d e da e d' e g e, a e e ha e d e da e d' e g e I e II (

Figure

  Figure 1. M canisme propos d'absorption et d'action des aminogl cosides che les bact ries Gramn gatives. Da e e e , 'a g c de ( e e a de a g e e ) e da 'e ace a e a a e de e e a e e a e e e f a de che a e e a d ab a d LPS. La de e a e de 'ab e 'EDPI, a c de a e e 'a b e e da e c a e a e e c e. Ce e a e d e d d ea d' e g e bac e e (PMF) e d a a cha e d' ec e da a e b a e e e. U e f e 'a b e e da e c a e, e e a b e, ce e a e a h e de e a c d e . I e gg e ce e a c d e e a e e e be ' g de a e b a e, ce c d a de e a e de 'acc a d'a b e (EDPII), a a d ff d'a b e e g a de c ce a da e c a e, ce e a e a ce a e. E ce c ce e ' e ac de a g c de a ec e bac e , bea c de ch e e e f e . Pa e e e, e a a c e e a b e a g c de a e e a e b a e c a e a e d e e c b e. Le a e e e de a g c de da a e b a e ce a e e e de f e .

Figure 2 .

 2 Figure 2. Comparaison de la r sistance, de la tol rance et de la persistance au antibiotiques et de la r ponse des populations de cellules bact riennes au antibiotiques bact ricides. Da a a bac e e a a ce (e ha ), e bac e e b e (e g ) a de e e a e a e e a b e. Da a -a e a ce (e e ), e e e a a e e a b e e e e g e e a e b e a a b e . Le bac e a e (e a ge) e e e e a e a e e a b e e e bac e a e (b e ) e 'a b e. L' b ec f de ce e de d c a e de c e de ea d e ' e ac de a g c de a ec e bac e . Da e ab a e, g de a c e a e a f h e, a c a e 5 (Ne -C 5). L' b ec f c a de e de h e a d'ab d d' d e e de ca ac e c e e ce c g a b e e d' d e e f de ' e ac de ce a b e a ec e ce e bac e e a ea de a ce e e. Je a a e ce e h d g e d e 'acc a d'a g c de da e ce e e b e c a ce a de da de bac e

Figure 3 LFigure 4 .

 34 Figure 3 L'activit de Neo-C 5 est d pendante de la PMF. S e de ce e E. MG1655 a a e e de 4,5 he e a ec 1 M de c e 48 M de Ne -C 5 a ec e a 30 M CCCP ( c ba de 2 he e a a 'a de 'a b e). Le a e d e de a e e a e a c e a e. Le ba e d'e e (e e a da d de a e e) e ca c e a de e e ce d e da e . De d ff e ce g f ca e , ca c e a e e de Ma -Wh e , d e (****P < 0,0001). E e, a b e 'acc a d'a b e a da de bac e c a ce a de a c c e f e ce ce e a a e FACS. Se e e ce e c c e f e ce ce a ec d ff e e d' c ba a ec 'a b e, a de ha e d'acc a d d ca e da e ce e . Da a e e a e, a e

Figure 5 Figure 6 .

 56 Figure 5 Courbe de mortalit de la Neo-C 5. Le ce e de E. a e a ec a Ne -C 5 (32 M) 37 C e cha e e ce e a e f e e d e e de a e , e a e d e e e e a e de b e LB MOPS-G e c age de CFU. La ec de a e a e a c c e. S e de ' e de a f e ce ce C 5 da cha e bac e e e a e g c e I ageJ. A a d e d ff e e a e de 'acc a de a b e da e ce e a age e b e c a ce a de, ' b ec f a de e e a Ne -C 5 de

Figure 7 .

 7 Figure 7. cL pr sente un pourcentage plus faible de cellules avec accumulation c toplasmique de Neo-C 5 par rapport au cellules de t pe sauvage (WT). Le ce e e ha e de c a ce e e e e d e 10 f e c b e a ec e c ce a -CMI de Ne -C 5 e da 2 e 37 C. Le ce e ce f g e , a e a ec d MOPS-G d MOPS-G a ec de a Ne -C 5 a e . E e, e ce e , a e e agea de ce e a e a ec MOPS-G, a a e a FACS. La a de ce ca a c d e e acc a c a e de Ne -C 5. I c e de e e e ce a a c a e e f e e e da a a a de ce e e c b g e. J'a d c ab e h de a e a a a de ce e a d c e che e ca a ca e b e e d e 'effe de a ce ca a 'acc a de Ne -C 5. (F g e 6)

1.5.4. O idative stress

	h d be e	ded ha a ( b	e a b g		a	) E.	a	ha ha e	g e
	a	-ac d		a	he b	a	e		S4 a d S5, h ch ca e c	a ab e e e	f
	ead g a a	g c de , a e ab e 63 . I	he ef e c ea he he	c d g ead
	ce dea h		da e e		.
		I	e	h h	b e a	,	a h		ha Fe-S	e	e a	g c de
	d ced ce dea h	h	he	e e	f he eac e	ge	ec e (ROS) e	e 48 .
	Ke e		., a		ed ha ROS			ed a b c ed a ed ce dea h 64 . M e
	ece	, a d ha		he ROS e	e	h	e b a a	a e	ed 65 .
		I a		ed ha a	g c de e e a	gge h d	ad ca f	a	h ch
	ead	ce dea h 54,61,62 . I had bee	gge ed ha	da e e	gge ed b	e b a e
	e	aff c,	a a	a d	f d g f e b a e	e	he a ea	f ce
	dea h af e a	g c de ea e . A h		gge ed ha a	-c	e	e f	e -
	e	e	he ce e e e	a e	a e	h	ce a d a ed -e	e	-
	c	e		e , a	e , a h	ha e a c	ec ed	e 54 (F g e1.9). H e e ,

. Terminolog : Antibiotic Resistance vs. Tolerance vs. Persisters

	1.7. Resistance mechanisms 2. Persisters 2.2
	I bac e a, e a ce 2.1. Discover		a	g c de	f e d e	e	a c ac a	b
	h h a fe a e , ace M e ha 70 ea ag , H bb a d B gge b e ed ha a fe a e a d c e d a fe a e (ade	a fe a e ) 72, 73 . b e e e
	O he bac e a c echa	c de b e b bac e c da a b c . H bb a a e a a d c ea e f e b a e e eab b e ed ha a a a	. f a
	A S	g c de-e a a	a a	f e a ea d e ed e c	e	g a d-b e ge e e c d g ea e 78 . Fe ea a e B gge
	a f c ed g c de-d f g e fe bac e a ce		e 74 . F he ha e e h gh a b c ea e . He a ed h e, a f he e ge e a e a c a ed	h
	a b	a	, h ch he fa d ff f bac e a e	f d g e a ce ac e 79 . I h ee g	ec e b , he gge ed ha he e ce da e 75 .	e e
	a 1.7.1. Adaptive resistance -d d g. I add h ch e e e e e c a he g a c , he d c e ed ha he e e e.	e c ea ed ff	g,
	S e E. Ada ead Pe h d a e e cha ge d e a d c ca e e e he bac e hage T4. Th a e e a ce a b c d eg a f ge e ha he he e d c e ed he e a cha ge e f a e e a d e ha bee ca ed he c fe he ce a ad a age he e e ca be def ed a he a b c e a b a f bac e a e e c be e ed ha he ca ac f a bac e a a ada e a . Pe e ce e a d a c h g ca a e a d a b c e e he e e ce f . I ca a be he e he he e e a e. 76 I he , a ce e a a fa c a g e a e f he e ge e a g e ce e e , h ch d e he ge e c c he e ge e e e a 38 . I ha e ha e ce ha e, a he e ac e f bac g d. Th he e ge c a be he ca e f fa e f a b c he a 80 .
	ce he he e , a I ha bee f a he ec a c d ha e e		af e e ha e a h ge c a a f he d g 69 . e e f e a ce d g . A f E. (UPEC) a e he a	ge he , ca e f
	A he e a eg e ca ha e g c de-d ced bac e a a ac fec		a	ead g a e e c d c e d g c ca ea e e de g 70 he he ha d, acc d g e fec ece . The e ce ca e af e a b c ea e d e
	d , b a c d ff c c e. he c he e ge e . Th a e		f e a e e d h ed he	a	c d . A e a ce f h	, he a h ge e	ec a e
	ba cc e ce f he e ge e f a ge e c d ea e I ha bee h ha	ch a c c f b b-h b a d h b ce a 81 .	e a e a a c ce a	f a	e	a g c de . I ha
	bee de P Se e a ece a ed ha	b a c e h b a d he ae b c G a -ega e bac ead-h gh ab d e h ha he e be ee he a e f e d ced ada e a e e e e a ce. a e ce a d e a ce
	c d Th ee	71 . Th e d f e a ce ha bee h e a c ea def ed a e	h he g ba he a e c a eg , h ch c be he e f d -eg a f a b c h gh . Pe e ff g had a ge	f	g a e a e
	a EDP II ha e 77 . g c de e h b ge e c	a	e g		he e g a b c e a ce 82,83 . a f e	he	a d e e f -e g h
	f c	a		e .

.2. Persisters in macrophage

	2.4	2.3Mac hage a e e		b e f	hag c	a a a f	e . The e
	b g ea e e g f a d d ge a h g ha d e	ha e he	e f	e	ec f c hea h
	b d ce	e f e g		b a ce	ch a	c be . A he a e	e a h ge ca
	a		a e h		e e		e a d dec ea e he cha ce f a h ge c ea a ce 89 . F
	e a		e, a	a h ge , c d g M.			, L	a d S
	., a d deg ada	f	g hag c		b h	e ce . I ha bee f d ha he e
	a h ge c bac e a h b he a a	f hag	e 90 .
			I ha bee h	ha b f	e ce		g e-ce a a	,	b e	de f
	S		e	e	fec ed ac hage. Pe	e a e	e ca g ce f	ed	a
	af e bac e a	a e	he ac hage . I ha bee	gge ed ha e	a ce a d ced
	he ce beca e f f	he ac d c e		e	f he ac e	ac hage a d ac f
			. The e c d	a		d ce	e ce ge e e e	S	86 . H e e ,
	e e f ac d a d ac f		a a	d ffe e	ac hage ce	h d ffe e
	hag		e , h	d c g he f	a	f e	e f	e bac e a a d	f	he . The
	e ac echa	f h	he e e		e	d ce he f	a	f e	e a e	a a e
	f deba e 89 .
			I ha bee h	a e ha he 14		-a	d e a a e	he e	e
	f	a	ef e, beca e b f f S ac hage 86 . d ce f	a	f e	e	b-	a	h	a e he
	e ad ca	f e	e	e d ff c .

. Wh do the become persister? 2.4.1. Dormanc and t pe I vs. t pe II persister

		2.4
		D		a c	a a e f a bac e		h ch bac e a d e	g	a d ha e
	ac		c	a		h g	g e	e a ha e	a	a	ha e ce . D	a
	bac e a a e		e	e a g a a b c beca e he	e ab	e e	, a d
	he		g	g 87 . H e e , e a ce a d e	e ce a be ca ed	h	d	a c . I
	ha bee f	d ha be g	-g	g a e	ac e e ab ca d	a	e e	e
	f	a	, beca e	d	a		b	a		e e	e	e . I h	d a h
	c a ed ha d	a c		e gh f	e	e f	a	91 . Acc d g	he	d	f
	f	a	f e	e f			ac hage , he e	-g	g ce	he ac hage
	a a a ac e a e 92 .
		P e e ce f e	e	a g e		a	ca be e ed a a	ec	a eg 93 .
	Whe a		a	g e h gh a e f c d	, e	e	a	e a d a	he
		a f he c		. M e e , ha bee	gge ed ha e	e	a be a a eg b
	h ch bac e a ca f ee	g a	ed ce dea h h ch c d be d ced b a b c ea e .
	Whe a a b c e ac	h	a ge	ce , ead	h b	f ce g	h a d e
	ha he ce a d	g a	ed ce dea h 94 .
		I ha bee		ed ha he e a e		e f e	e ha e e	d	a c	a e
	e he	he a	a	ha e ( e I)	d	g g	h ( e II) 95 .

.2. To in and anti-to in modules

	e	e ce 95 . A TA	d e d eg a	a ead g	h a e	ha bee	gge ed ha
	Ge e c ba he ea Be a d 96 . The h h gh e e ce he h	f a b c e a TA a ed a f E e 102 (F g e1-2-2). e ce a e d e ha e bee ea ed f , h ch	he f a ed b ge e c c ee f e b M ed a d d ce a h ghe e ce age f
	e	e . Yea a e , Ba aba a d c eag e 95 ed a a ca	g	A7 a e e, h ch a
	e f he e ga -f-f c	a e e	ac e	e	a c f d c de ce. The h ed ha
	A7 e	e a e he	-g	g ce . La e , H A a f d	be a	a d	h
	c g a e a		H B a g a		a		(TA)	d e.
		A	g a he 10	e II TA	d e	he	de bac e a E.	, H A	e f he
		d ed	a d ha bee		e	be	ed	e	e f	a	.
		I e		e a g	g E.	ce , he de e	f 10 RNA e d	c ea e ( TA10
	a :	a F, ch B,	e BE, d J/ afQ,	efM/ eB, h BA,	F/ haV,	afNO,
		RA, h cAB) dec ea ed bac e a e	e ce 97 . H e e , 5 ea a e ,	a h	ha
	he	e e	e f	a		TA10 a	a d e	he e e ce f he 80 bac e	hage
	98 . Rec	c	f he	g a TA10 a	h ch a	c a	a ed b bac e	hage
	80 h ed ha he e 10 TA	d e d	affec e	e f	a	e -ch
	c d	99 . I		a ed	, he 10TA	a	ed ce	e a ce f c	f ac b
	ha f a	c	. I ha a bee h	ha e	e f	a	10TA a	affec ed
	b c	e e h b	f	e	he (ch a he c	e ac c e ea e ) 100 . H
	RNA e fe a e a e c	ec ed	e	e f	a	e a	c ea .
		We		he e f TA	d e		c ea e he he e ge e	he	a
	e	e d ffe e	ce f e e 101 . I ha bee	gge ed ha he ac a	f he e TA
	d e	a f ac	f he	a		affec he ce g	h, h ch	ead g bac e a

Figure1-2-2 persister cells frequenc can increase due to stress responses 101 . Pe

	ead g	c ea e f e	e ce	he	a	. ( ) G ac a e	ec f c	e I a d II
	TA	d e h ch ead	e	e f	a	(f	a ce,	e I	H B). S	a , he
											e e	a
	he e ge		a	a d af e ac a	f e e	e he f e e c	c ea e .
		A	g d ffe e		ce f e	ch a		c e , H e , NaC	e a d
		ge e	e	he a	e ed TA-de e de	e	e f	a	100 . Th he	e
		ec f c	f			e . N effec a de ec ed	e a ce	a	c	100 a d
	e	a e	d ha d e	a	g c de.
		S ge e	e he	a h ge		a	he h	d	g h	e c d	. U de
	a	ac d a a		ca	b H A		, cha ged RNA	he b	a A e
		e he acc	a	f a e e ge	c e de ca ed g a	e e a h ha e
	e a h ha e (( ) G ). Th	ec e b d		e e a	e	a g g f	DNA
	e ca	,	e	a c			a		e ab c a h a	e 103 . ( ) G
	a g e a		h c d		h gh Re A, h ch de ec	cha ged RNA	he b	e

Table 3 -

 3

	1 MIC c	a	f e	c a d d ffe e	e	c c	ga e	d ffe e	ed a.

Table 4 -1 MIC measurements at two different optical densit of cells

 4

. MIC c ea e he ce ha e h ghe OD.

Chapter X Manuscript of submitted paper

P ea e c ac D e.FOURMY@ 2bc. a -ac a .f f e e g he a e a c .


Sylvain Bourdel

Yannick Bachelet

Member, IEEE Jean Gaubert

Remy Vauche

Olivier Fourquin

Nicolas Dehaese

Member, IEEE S Herve Barthelemy

R Vauché

A 9-pJ/Pulse 1.42-Vpp OOK CMOS UWB Pulse Generator

Keywords: Pulse generation, FCC, Transmitters, Impulse Radio, Ultra-wideband (UWB), Pulse shaping circuit. A 9pJ/Pulse 1.42Vpp OOK CMOS UWB Pulse Generator for the 3.1-10.6 GHz FCC Band

come   L'archive ouverte pluridisciplinaire

I. INTRODUCTION

ince the FCC has authorized communication in the 3.1 GHz to 10.6 GHz frequency band, several technologies have been developed to satisfy the communication market. These technologies are more or less suited to the numerous applications generally encountered. Typically, Orthogonal Frequency Division Multiplexing (OFDM) technique appears to be good candidates for high speed data communication whereas Impulse Radio (IR) is a good solution for low cost systems or positioning systems. In the case of IR systems, due to the applications, the most limiting characteristics of the system are the circuit cost production, the power consumption and the operating range. For the emission stage, those constraints mean that the transmitter must be designed using low cost technologies such as CMOS and must consume a small amount of power while generating pulses having large magnitude.

UWB impulse radio designs often use baseband pulse generator. Those baseband pulses are then used to produce Manuscript received May 21, 2009. This work was supported in part by Aix Marseille University and CNRS.

S. Bourdel, J. Gaubert, R. Vauché, O. Fourquin, N. Dehaese and H. Barthelemy are with Aix-Marseille University, IM2NP, and CNRS, IM2NP (UMR 6242), Campus de Saint-Jérôme, Avenue Escadrille Normandie Niemen-Case 142, F-13397 Marseille Cedex, France (email:sylvain.bourdel@im2np.fr).

Y. Bachelet was with Aix-Marseille University, IM2NP, and CNRS and is now with EDITAG SAS, 100 route des Houillères BP2 13590 Meyreuil Cedex, France. more complex pulses to match regulation frequency masks. A first technique to generate baseband pulse consists in using reflections in a transmission line (or a stub) to produce a pulse from a step function. It is difficult to integrate those generators in a CMOS technology due to the use of transmission lines or due to the need of discrete components such as SRD diode [START_REF] Han | Ultra-wideband electronically tunable pulse generators[END_REF] [START_REF] Eng-Choon Tan; Chia | Subnanosecond pulse-forming network on SiGe BiCMOS for UWB communications[END_REF]. Currently, delayed edge combination is a good candidate to achieve baseband pulse and is well suited for UWB systems since it can be fully integrated in CMOS [START_REF] Wentzloff | A 47pJ/pulse 3.1-to-5GHz All-Digital UWB Transmitter in 90nm CMOS[END_REF] [START_REF] Kim | Fifth-derivative Gaussian pulse generator for UWB system[END_REF] and consumes a small amount of power.

Different methods have been presented to generate FCC pulses from one or several baseband pulses. A first technique is to use transversal filters achieved by artificial coupled transmission lines [START_REF] Zhu | A 0.18 µm CMOS distributed transversal filter for sub-nanosecond pulse synthesis[END_REF]. The baseband pulse propagates along the artificial line and is recombined at the output to achieve a more complex pulse. The artificial line is synthesized with lumped elements which results in the use of a large number of inductors and limits the integration. A second method is to transpose a baseband signal in the frequency band of operation using a mixer and a Local Oscillator (LO). This method is attractive since it uses few inductors and it performs well when it is implemented using LO switching. In spite of LO leakage problems [START_REF] Xu; Jin | Power-efficient switching-based CMOS UWB transmitters for UWB communications and Radar systems "Microwave Theory and Techniques[END_REF], this technique currently appears to achieve the best performances in terms of consumption and pulse peak magnitude [START_REF] Sim | A CMOS UWB Pulse Generator for 6-10 GHz Applications Microwave and Wireless Components Letters[END_REF] [START_REF] Phan | Energy-Efficient Low-Complexity CMOS Pulse Generator for Multiband UWB Impulse Radio[END_REF]. Other methods based on elementary pulse combination have been presented with good potential for the pulse tuning and for achieving biphase pulses [START_REF] Kim | Digitally controllable bi-phase CMOS UWB pulse generator[END_REF] [START_REF] Wentzloff | A 47pJ/pulse 3.1-to-5GHz All-Digital UWB Transmitter in 90nm CMOS[END_REF]. Today, those techniques achieve pulse magnitude around 1Vpp but their power consumption dramatically grows with the pulse duration and it exceeds the consumption of LO switching topologies [START_REF] Bourdel | Low-Cost CMOS Pulse Generator for UWB Systems[END_REF] [START_REF] Norimatsu | A UWB-IR Transmitter With Digitally Controlled Pulse Generator[END_REF]. Another way to produce FCC compliant pulses is to use filtered combined edge [13] [14]. This method is very attractive for its low power consumption potentiality but is penalized by the filter implementation. However, due to the large width of the FCC frequency mask, the order needed for the shape filter is low which leads to acceptable loss (around 3dB), in standard CMOS technologies [START_REF] Bachelet | Fully integrated CMOS UWB pulse genrator[END_REF].

The generation method presented is derived from filtered combined edge technique and consists on simply exciting an integrated bandpass filter using a baseband pulse to generate its impulse response. The filter excitation is achieved using a digital edge combiner coupled with a current driver and appears to be well suited for low cost and low power systems. To demonstrate the efficiency of this generation method, an UWB FCC pulse generator is designed in a 0.13µm CMOS technology for medium rate applications around 36Mbs -1 . The power consumption is 3.84mW@100MHz Pulse Repetition Frequency (PRF) and the energy consumption is 9pJ per pulse. The pulse generator produces FCC compliant pulses having a measured voltage magnitude of 1.42Vpp. The digital edge combiner used in the design is able to generate pulses with less than 50ps width. It is designed with differential delay cell and high speed logical gate. The bandpass filter is a third order Bessel filter with less than 3dB loss and the whole die size is 0.54 mm 2 . Using a sizing method based on Power Spectrum Density (PSD) analysis, the generator is designed regarding the application needs. This generator achieves unipolar pulses and can be used in On Off Keying (OOK) modulation, Pulse Position Modulation (PPM) or Pulse Interval Modulation (PIM). In OOK mode, the generator can operate up to 38Mbs -1 to cover a large variety of applications including video streaming.

The paper is organized as follows. Section II deals with the theoretical concept of impulse response generation needed to compute the main parameters of the generator. This analysis is made for the FCC standard taking into account the impact of the modulation type and the impact of the bit rate on the design. The design of the pulse generator is presented in Section III. The on-chip filter and the buffer which consist in a current driver are first described. Then, the design of digital edge combiner is detailed. Simulation and measurement are presented and commented in Section IV.

II. PULSE GENERATOR SIZING

A. Pulse generation Principle

The method of pulse generation is depicted in Fig. 1. It consists in applying a short pulse at the input of a filter to generate its impulse response (h(t)). If we consider that, instead of a dirac, the filter is excited by a square function π τ (t) having a magnitude A and a width τ, the output (g(t)) and its Fourier transform are given by :

( ) . ( ) * ( ) e A g t t h t l τ π = (1)

sinc( ). ( ) ( )

e A f H f G f l τ πτ = ( 2
)
where l represents the power loss of the filter and H e (f) the normalized function transfer of the filter. Computing G(f) allows the determination of τ and H e (f) which leads to FCC compliant pulses. But, knowing G(f) is not sufficient to size the baseband pulse magnitude (A) and the emitted pulse peak to peak magnitude (g 0 ). Because these parameters impact the range of the transmission, they must be taken into account in the sizing of the generator. It is then necessary to evaluate the average and peak PSD to size A and g 0 and other characteristics such as modulation type and bit rate must also be taken into consideration.

B. Average PSD

The Fourier transform G(f) is necessary to calculate the average PSD of the emitted signal. In the case of linear modulation such as OOK or BPSK, the emitted signal is given by (3) :

0 ( ) . ( . ) N k s k s t a g t k T = = - ∑ (3)
where T s is the symbol time and {a k } is the sequence of the transmitted symbols. In the case of OOK the value of a k is simply 0 or 1. If the symbols of the sequence {a k } are independent the PSD is given by :

2 2 ( ) ² ( ) a s a i s s s G f i PSD f f T T T µ σ δ ∞ =-∞     = + -         ∑ (4)
where µ a and σ a 2 are respectively the mean and the variance of the sequence data {a k }. For BPSK modulation, µ a =0 and σ a 2 =1. The PSD is then composed by a continuous spectrum given in V 2 /Hz. For OOK modulation, µ a =0.5 and σ a 2 =0.25. The PSD is then composed of a continuous spectrum and a discrete spectrum where the magnitude of the lines is directly given in V 2 . Because Ts << 1, the spectral lines are much higher than the continuous spectrum and the ratio (η given in [START_REF] Zhu | A 0.18 µm CMOS distributed transversal filter for sub-nanosecond pulse synthesis[END_REF]) between the spectral line level and the continuous spectrum increases with the symbol rate (D s =1/Ts). These spectral lines reduce the capability of the generator to achieve large pulse at high rate because the only way to decrease the PSD (for a given pulse and a given rate) is to reduce the magnitude of the pulse. In the case of FCC regulation [16], the PSD is measured in a Resolution Band Width (RBW av ) equal to 1MHz which reduces η and so reduces the impact of the spectral lines on the maximum pulse magnitude allowed by the standard.

µ η σ = = (5)
With ( 2) and ( 4), we can express the PSD as a function of the main parameters of the pulse generator (A, τ, l, Z 0 the output impedance, D s , H e (f), µ a , σ a ) and taking into account the measurement method imposed by FCC.

( )

2 2 2 0 2 2 2 0 ( ) ( ) 2 . sinc( ). ( ) 2 . . s s av a s a i s e s av a s a i s G f i PSD f D RBW D f Z T A f H f i D RBW D f Z l T σ µ δ τ πτ σ µ δ ∞ + =-∞ ∞ =-∞       = + -                 = + -           ∑ ∑ (6)
Equation ( 6) gives the unilateral PSD as it would be displayed by a spectrum analyser with its RWB set to 1MHz. The Moreover, it appears that for a given filter and a given baseband width, the maximum value of the PSD (PSD max ), which is fixed to -41.3dBm/MHz by the FCC, linearly depends on A and Ds. If the frequency for which the PSD is maximum (f 0 ) is close to the frequency for which H e (f) is maximum, the maximum value of A allowed by FCC (A max ), can be approximated by using [START_REF] Xu; Jin | Power-efficient switching-based CMOS UWB transmitters for UWB communications and Radar systems "Microwave Theory and Techniques[END_REF] in the case of BPSK (µ a =0 and σ a 2 =1) and OOK (µ a =0.5 and σ a 2 =0.25) as follows :

1/2 1/2 0 max 0 max max 2 2 2 2 2 2 2 0 0 . . . . min , 2 sinc ( ) 2 sinc ( ) s s av a s s a l Z PSD l Z PSD A D f T RBW D f T τ π σ τ π µ         =               (7)
In these conditions, A max mainly depends on D s and is independent of the filter transfer function.

From A max it is then possible to compute the maximum pulse magnitude (g 0max ) imposed by the average PSD limitation for a given symbol rate. Because g 0 depends on the filter used (h e (t)) and the baseband pulse (π τ (t)) it linearly varies with A and can easily be evaluated as follows :

0 ( ) * ( ) ( ) norm t h t A A g g t l l τ π α = = (8)
where g norm (t) is g(t) normalized to g 0 . The α coefficient represents the magnitude efficiency of the filter response to a square function of width τ. However, to avoid excessive peak magnitude, the FCC also limits the peak power. It is then recommended to verify the FCC compliance of the peak power of the generated pulse.

C. Peak Power Limit

The peak power limitation is specified by theFCC as follows: "there is a limit on the peak level of the emissions contained within 50MHz bandwidth centered on the frequency at which the highest radiated emission occurs, f 0 . It is 0 dBm EIRP." The FCC recommends the use of a spectrum analyzer with a 50MHz RBW and a peak power detector. For rates lower than 50MHz, this measurement corresponds to the peak power of the emitted pulse filtered by the RBW filter. Under this condition, if the pulse spectrum is flat in the RBW and if the RBW filter is supposed to be ideal, the magnitude of the pulse at the RBW filter output for which the spectrum (G pk (f)) and its reverse Fourier transform (g pk (t)) are :

( ) ( ) 0 0 0 ( ) pk pk pk RBW RBW G f G f f f f π π   = - + +   (9)
( )

0 0 ( ) 2 . .sinc .cos(2 ) pk pk pk g t G RBW RBW t f t π π = ( 10
)
where G 0 is the magnitude of G(f), π RBWpk (f) is a square function having a width equal to the spectrum analyser RBW (RBW pk ). The peak power is then given by the level of g pk (t) as follows : If the frequency for which the PSD is maximum (f 0 ) is close to the frequency for which H e (f) is maximum G 0 is given by: ( )

0 0 sinc A f G l τ π τ = (12)
By using ( 6), the PSD max can be expressed as in [START_REF] Smaini | Single-chip CMOS pulse generator for UWB systems[END_REF] as a function of G 0 . This enables the evaluation of the peak power, as in [START_REF] Jung | Pulse generator design for UWB IR communication systems[END_REF], as a function of the symbol rate in the case where the average PSD complies with the FCC mask for BPSK (µ a =0 and σ a 2 =1) and OOK (µ a =0.5 and σ a 2 =0.25) modulations.

2 2 2 2 2 0 0 max 0 0 . max , s a s av a D G D G RBW PSD Z Z µ σ   =     (13
    =       (14)
Equation ( 14) is only valid for rates lower than 50MHz for which the RBW filter only responds to one pulse. Moreover, in the case where the rate is decreased to maximise the pulse magnitude, [START_REF] Jung | Pulse generator design for UWB IR communication systems[END_REF] gives the minimum rate below which further increasing the magnitude is prohibited.

D. Generator Sizing

First, the filter and the width of the baseband pulse must be dimensioned by using [START_REF] Eng-Choon Tan; Chia | Subnanosecond pulse-forming network on SiGe BiCMOS for UWB communications[END_REF] or directly by computing the average PSD given by [START_REF] Xu; Jin | Power-efficient switching-based CMOS UWB transmitters for UWB communications and Radar systems "Microwave Theory and Techniques[END_REF]. A large variety of filters can be used to be compliant with the FCC standard if their bandwidth and center frequency are tuned to match the FCC mask as shown on Fig. 2. Moreover, whatever the H(f) considered, a 3 rd order is sufficient to achieve FCC compliant pulse due to the large width of the frequency mask. This limits the loss in the filter and enables integration in CMOS technologies. The choice of the filter relies on a complex compromise. First, the designer must bear in mind that this choice can be reoriented by the filter design presented later on, especially if the value of the filter components can not be achieved in the targeted technology. To choose the filter, it is relevant to consider the value of τ needed and the magnitude efficiency represented by α. As shown in Fig. 2, where only 3 rd order filters are considered, the pulse2 achieves almost the same efficiency (α) as pulse3 with a wider baseband pulse which relives the constraint on the edge combiner design. Also, for similar filters (pulse2 and pulse1) lowering τ decreases α and increases the design constraint which is not helpful. Moreover, the pulse4 shows that using large baseband pulses increases the sinus cardinal function influence which degrades the pulse spectrum and also α.

Once the pulse (g(t)) chosen, it is then possible to compute with [START_REF] Sim | A CMOS UWB Pulse Generator for 6-10 GHz Applications Microwave and Wireless Components Letters[END_REF] the maximum pulse magnitude allowed by the average PSD, regarding the modulation type and the rate. Fig. 3 reports A max as a function of D s for OOK and BPSK modulations. Fig. 3 is plotted for pulse2 with 3dB filter power loss.

In the case of OOK modulation, for high rates, the maximum value of PSD is given by the discrete spectrum. When Ds is lower than Ds lim =RBW av .(σ a /µ a ) 2 , discrete spectrum is lower than the continuous spectrum and the maximum value of PSD is fixed by the continuous spectrum. In our case study, a rate of 36Mbs -1 is targeted. Such a rate can be achieved with OOK modulation if the baseband pulse magnitude does not exceed 2.56V, which leads to a g 0 =1.73V due to the 3dB filter power loss and due to α=0.95. Moreover, depending on the application needs, the computation of A max can be used in a different way. For a given pulse magnitude fixed by the budget link, the maximum rate can be evaluated for OOK or BPSK modulation.

Lastly, for a RBW equal to 50MHz in the case where A is set to A max the peak power given by ( 14) is plotted in Fig. 3.

Under this condition, the peak power limit is reached for OOK at Ds=1.23Mbs -1 and it is not possible to reduce further the symbol rate to increase g 0 . For BPSK, the limitation is reached for Ds=380kbs -1 but it leads to the same value of g 0 . For our case study where g 0 =1.73V, the peak power is -29dBm.

E. Generator specifications

This theoretical section presents the specification of the main parameters of a pulse generator that uses filter impulse response technique in the case of linear modulation. It shows that a generator can be achieved with a low order filter. A 3 rd order Bessel filter is chosen here, especially because it has a good response in terms of magnitude (α=0.95) to a baseband pulse that has a width equal to 75ps. This lowers the constraint on the design in a 0.13µm CMOS technology. Moreover, to reach 36Mbs -1 rate, OOK modulation can be used with a pulse magnitude of 1.73V leading to a good detection [START_REF] Lee | System On Package Ultra-Wideband transmitter using CMOS impulse generator[END_REF]. To achieve g 0 =1.73V with 3dB loss and τ=75ps, A max must be equal to 2.56V. Such a value can not be directly obtained in the technology targeted because A max >1.2=Vdd. As a consequence the filter must be driven in current mode as shown in the next section. Last of all, there is no limitation due to the peak power in RBW pk for OOK modulation having g 0 =1.73V.

III. PULSE GENERATOR DESIGN

Fig. 4 shows the entire topology of the proposed pulse generator. The generator consists in a edge combiner that uses varying differential delay cells and a fast logic gate. A transistor is then used to drive the filter in current mode to produce the magnitude needed (Amax=2.56V) at the filter input.

A. On Chip Filter Design

The design of the pulse generator is mainly determined by the performance of the passive devices of the technology used and especially the inductors. Due to the metal thickness and the substrate loss, CMOS technology provides inductors with low Q factor. The low order needed to achieve a FCC Fig. 4. Pulse generator architecture Fig. 3. Maximum baseband pulse magnitude (Amax) and peak power limit as a function of Ds for OOK and BPSK compliant pulse is a great advantage for the design of a filter with low Q devices because it limits the power loss (<3dB). Moreover, the values of the Q factor and of the Self Frequency Resonance (SFR) vary with L. It is then necessary to evaluate the technology to determine which values of inductors achieve admissible Q factor (around 10) in the overall bandwidth. This means that the range of inductor values to be used is limited. For the 0.13µm CMOS technology used for this design, the range is around 400pH<L<1.5nH. Lower values of inductors leads to inductor with Q<10 and higher values of inductors leads to SFR lower than 15GHz. Due to the large relative bandwidth of the FCC mask, the dispersion on the inductor values needed to implement a FCC compliant filter is low and filters can be achieved with such a narrow range. For the 3 rd order Bessel considered in this design, the values are listed in TABLE 1 and are in the range. Moreover, different filter structures can be evaluated to find the best inductor values. In our design, as shown in Fig. 4, the filter topology used is a modified ladder structure.

B. Filter Driver Design and Matching

In our case study, the magnitude of the baseband pulse needed exceeds the voltage supply. Obviously, the filter is driven in current mode. The sizing of MN3 depends on the filter input impedance and A. In the case where g(t) is a square function with magnitude equal to V DD , the current given by MN3 to the load during the time τ can be approximated using the MOS current law I D (V GS ) with V GS =V DD . Under this assumption the width of MN3 can be expressed as follows:

( )

2 0 . . . IN ox DD t L A W R C V V µ = - (15)
where R IN is the filter input impedance and is equal to the load impedance in most of the passive filter if the loss is ignored. The current needed to drive the 50Ω impedance is then I 0 =(A max )/50=51mA which implies a large transistor. The output of such a wide transistor influences the filter transfer function and can involve matching loss. During the conduction time (τ) of MN3, the output (Rds and Cds) can be approximated using small signal hypothesis leading to the electric scheme given in Fig. 5. To match the driver to the filter, MN3 must be sized to have Rds equal to R IN . Because both Rds and I 0 depend on MN3 width (W MN3 ), it is not possible, with such a simple topology, to size MN3 to have I 0 =51mA and, at the same time, Rds=R IN . It is then necessary to make a trade-off between the driving current (I 0 ), the matching (Rds), and the power consumption (W MN3 ). For the considered design the width of the transistor is set to 100µm which leads to the intrinsic characteristics given in TABLE 2. Due to the mismatch (Rds=122Ω instead of 50Ω), the value of I 0 is increased to 58mA (instead of 51mA) in order to ensure that g 0 =1.73V as shown in Fig. 10. A wider transistor would have a lower Rds and would better match the filter but would increase the power consumption.

To avoid transfer function distortion, the effects of Cds can be easily cancelled by taking Cp1 equal to its ideal value minus Cds. As shown in Fig. 6 the main effect of Cds on the filter transfer function is to reduce the 10dB bandwidth.

The drain of MN3 is biased using the first filter resonator to avoid the use of additional inductor. The gate is not biased to operate in C class. As a result, the filter does not consume DC power while no pulse is transmitted resulting in good power consumption performances.

C. Digital Edge Combiner Design

The technique used in the baseband pulse generator consists in propagating an edge through a delay cell. The pulse is then obtained by combining the input and the output of the delay cell using a logic function. Generation of accurate delay requires a phase locked loop and is not suitable for low cost systems. The delay variation mainly impacts α which is equivalent to additional loss. However, a small variation does not impact the output magnitude very much and only tunable delay cells are used in this design principally to compensate process variation. The delay cell used is the modified differential MOS Current Mode Logic inverter [START_REF] Marsden | Low power CMOS re-programmable pulse generator for UWB systems[END_REF] depicted in Fig. 4. The control voltage (Vctrl) and VDD are respectively applied to RFP and RFN pins. In this configuration Mp and Mn act like resistors and no additional capacitor is needed. The width of Mp and Mn is set to 0.5µm. The output voltages are : VOL=0.3V and VOH=1.2V and the delay achieved by the cell varies from 17ps to 300ps which gives to the cell a large tuning capability. The combination of the two edges can be achieved by a large variety of logic gate. The speed of the gate is very important to generate short pulses having sufficient magnitude. The logic function used here needs only 2 transistors to operate, allowing high speed transition. The cell achieves A and not B, which generates pulses only on negative edges.

Due to the difference between the transistor size of the delay cell and of the filter driver, buffers are needed to ensure the signal integrity. The buffers consist in serial inverters sized with the fan out law to make a smooth transition between the delay cell and the driver transistor sizes.

IV. MEASUREMENT AND SIMULATION RESULTS

The pulse generator has been designed with a 0.13µm standard CMOS technology from ST Microelectronics. The die is shown in Fig. 7 and its area is 0.54mm2.

A. Digital Edge Combiner Results

The performances of the triangular pulse generator depend on the load of the filter driver MN3 and accurate measurement is difficult to achieve. Simulations using the design kit and post layout extraction are presented here. The impulse output signal of the gate and its Fourier transform are depicted in Fig. 8 and Fig. 9 for different values of the delay cell control voltage. The gate is able to generate impulse with a minimal width of 46ps having 1.2V voltage magnitude. For a 0.4V control voltage, the pulse width is 75ps and the Fourier transform is almost flat in the FCC band which denotes good spectral efficiency and good pulse quality.

B. UWB Pulse Measurement

Measurement achieved with a 12GHz real time oscilloscope (Agilent DSO81204B) and simulations concord well as shown in Fig. 10. Taking into account the cable loss, the peak to peak magnitude is 1.42V instead of 1.73V. This results in a lower detection but increases the rate capability up to 43.8Mbs -1 . The width of the pulse is 460ps, and its 10dB bandwidth is 6.8GHz. As shown in Fig. 11, the compliance with FCC mask is good and the bandwidth of the pulse is very large. Due to this very large bandwidth, the PSD of the modulated signal (s(t)) must be lower than the PDS max imposed by FCC to avoid emission in the forbidden frequency band around 1GHz. This limits the rate and finally, the maximum value is 38Mbs -1 . As shown in Fig. 12 the spectrum of a 36Mbs -1 OOK modulated signal complies with the FCC mask.

C. Power and Energy Consumption

The major interest of the presented structure is its very small amount of energy needed to generate large pulses. The energy consumed by pulse is estimated to be 9pJ, as represented in Fig. 13, which is a very good performance regarding literature. This very good performance relies on two aspects. First, the pulse generated has a very wide spectrum which leads to a very short duration and limits the time conduction of the transistors. Second, the filter driver which consumes the major amount of power, operates in C class mode and has no DC power consumption. The DC consumption is 3.2mW and is mainly due to the differential delay cell. The total power consumption is 3.84mW@100Mhz and is linearly dependent on the rate. Moreover, the DC power can easily be decreased to zero by using classic CMOS logic gate in the edge combiner especially for very low power applications like RFID or sensor network.

For low cost systems that use energy detectors, an important criteria is the energy by pulses relative to the pulse magnitude :

0 p E g η = (16)
When the consumed energy is compared to the pulse magnitude, the presented generator also performs well since it achieves 6.4pJ/Pulses/V.

V. CONCLUSION

The design of a low power and high voltage pulse generator has been presented. It uses a bandpass filter excitation technique for which a sizing method, based on the average PSD, has been presented. The generator is fully integrated in a 0.13µm CMOS technology on 0.54mm 2 . It consumes 9pJ by pulses and generates a 1.42Vpp pulse. It can be used in the FCC band with OOK modulation and rates up to 38Mbs -1 . Regarding its performances, this generator appears to be very well suited for low cost and low power applications.

Fig. 1 .

 1 Fig. 1. Principle of pulse generation

Fig. 2 .

 2 Fig. 2. Relative pulse spectrum (|G(f)| 2 ) comparison for different filters and baseband pulses.

Fig. 5 .Fig. 6 .

 56 Fig. 5. Electrical scheme of the driver and the filter.

Fig. 7 .Fig. 8 .Fig. 9 .

 789 Fig. 7. Die Photographe.

Fig. 10 .Fig. 11 .

 1011 Fig. 10. Time simulation and measurement of the pulse generator output

Fig. 12 .Fig. 13 .

 1213 Fig.12. PSD of a 36Mbs -1 OOK modulation using the proposed generator

Hervé

  Barthélemy has received the MSc degree in Electrical Engineering in 1992 and the PhD degree in Electronics from the University of Paris XI Orsay, France in 1996. In 2002 he received the HDR degree from the University of Provence, Aix-Marseille I, France. From 1996 to 2000 he was an Assistant Professor at the Institut Supérieur d'Electronique de la Méditerranée (ISEN) in Toulon, France. Since 2000 he joined the University of Provence where is has been a full Professor in 2005. In 2007 He joined the University of Sud-Toulon-Var. Since 2005 he headed the Integrated Circuits Design Team at the IM2NP. The team counts 12 Researchers and PhD students involved in several research projects with industry. His research interests are mainly in the design of radiofrequency analog integrated circuits. He authored and co-authored multiple publications in international journals and conference proceeding.

TABLE 1 :

 1

				FILTER'S COMPONENT VALUES
	Lp1	Cp1	Cp1	Cs1	Ls1	Lp2	Cp2	C3
	(nH)	(pF)	(pF)	(fF)	(nH)	(nH)	(fF)	(fF)
		(ideal)	(Real)
	460	1.43	0.5	677	1.15	1.08	420	260

TABLE 2 :

 2 VALUES OF THE EXTRACTED NMOS TRANSISTOR HAVING 100 µM WIDTH, 0.13 µM LENGTH AND BIASED TO VGS=1.2V AND VDS=1.2V

			{ Cideal1	Cs1		Ls1
	I0	Rds	Cds	Lp2	Cp2	Cp3	Zant=50Ω
			Cp1	Lp1
			Idson	58 mA
			Gds	7.59 mS
			Cds	240 fF

TABLE 3 :

 3 PERFORMANCE SUMMARY OF PROPOSED GENERATOR AND COMPARISON WITH PREVIOUS WORKSSylvain Bourdel received the Ph. D in microelectronics of the National Institute of Applied Science (INSA) of Toulouse in 2000. He was with the LASS laboratory of Toulouse where he was involved on radiofrequency systems modelling and he was particularly focused on spread spectrum techniques applied to 2.45GHz transceivers. In 2002 he joined the Integrated Circuit Design Team of the IMNP in Marseille where he works on CMOS RF IC design and integration. He particularly works for low cost applications including RFID, UWB and narrow band systems. His area of interest also includes system level specifications, system packaging and interconnections.YannickBachelet got into preparation for the competitive entrance examinations to French Engineering Schools, Dijon, France, in 1999. He received the M.Eng degree in microelectronics and telecommunication from Polytech' Marseille and the M.S. in microelectronics and nanoelectronics from Aix-Marseille University, Marseille France, in 2004. He received the Ph. D of microelectronics and nanoelectronics from Aix-Marseille University in 2008. He works now with EDITAG. His research interests include UWB radio transceiver design and RFID systems development. Jean Gaubert received the M.S. and the Ph.D. degrees in applied physics from Paul Sabatier University, Toulouse, France, in 1985, and 1988, respectively. From 1989 to 2001 he was an Assistant Professor at the Ecole Nationale Supérieure de l'Electronique et de ses Applications, Cergy-Pontoise, France, where he was involved with high-speed GaAs and InP bipolar devices modelling. In 2001, he joined the University of Provence, Marseille, France, and the Institut Matériaux Microélectronique Nanosciences de Provence (IM2NP). His research focuses on the design and integration of RF/Microwave CMOS integrated-circuits and systems for wireless communications. He has published more than 60 refereed journal and conference papers. Rémy Vauché was born in Epinal, France, in 1985. He got into preparation for the competitive entrance examinations to French Engineering Schools, Metz, France, in 2003. He received the M.Eng degree in microelectronics and telecommunication from Polytech' Marseille and the M.S. in microelectronics and nanoelectronics from Aix-Marseille University, Marseille France, in 2008.He is currently a Ph.D. student of Aix-Marseille University with the IM2NP laboratory. His current research interests include UWB radio transceiver design in 0.13µm CMOS technology for low-power and low-cost applications. Olivier Fourquin got into preparation for the competitive entrance examinations to French Engineering Schools, Marseille, France, in 2003. He received the M.Eng degree in microelectronics and telecommunication from Polytech' Marseille and the M.S. in microelectronics and nanoelectronics from Aix-Marseille University, Marseille France, in 2008.He is currently a Ph.D. student of Aix-Marseille University with the IM2NP laboratory. His current research interests include CMOS UWB radio transceiver, RFIC packaging and interconnections. Nicolas Dehaese received the Ph. D in microelectronics and nanoelectronics from Aix-Marseille University, Marseille France in 2005. In 2006 he joined the Integrated Circuit Design Team of the IMNP in Marseille where he works on CMOS RF IC design and integration. His research fields include UWB, 2.45GHz RF systems, spread spectrum techniques and system modelling.

	Ref.	Vpp (V)	Vdd (V)	Width (ns)	BW (-10dB)	CMOS (µ µ µ µm)	Power Cons.	Ep (/pulse)	η η η η (pJ/p/V)	Area (mm2)	Technique	Modulation
	[19]	1,24	2,2	1,75	1,4GHz 0,18 29,7mW@36MHz	nc	nc	0,4(core)	Digital Pulse Combining	DBPSK
	[20]	0,18	1,5	3,5	500MHz 0,18 1,82mW@100MHz 18pJ	100	0,39(die)	OL switchning	OOK
	[21]	0,9	1,8	2,5	1,2GHz 0,18	1,8mW@10Mp/s 180pJ	200	1,21(die) / 0,57(core)	OL switchning	BPSK
	[10]	0,65	1	3	550MHz 0,09	nc	47pJ	72.3	0,08(core)	Digital Pulse Combining	DBPSK
	[8]	0,16	1,5	3,5	520MHz 0,18 1,68mW@100MHz 16,8pJ	105	0,3(core)	OL switchning	OOK
	[7]	0,673	1,8	500ps	4,5GHz 0,18	nc	27pJ	41,5	0,11(die)	OL switchning	OOK
	[13] 0,35 to 0,45 1,2 0,6 to 0,8	2-4GHz 0,13	10mW@160MHz	nc	nc	1,56 (die)	Filtered Edge Combination PPM/BPSK
	This work	1,42	1,2	0,46	6,8GHz 0,13 3,84mW@100MHz 9pJ	6.4	0,54(die)	Filtered Edge Combination	OOK