Part Ill: Bacteriology

Brucella and Bordetella

Chapter navigation
Pathogenesis
• Bordete/lo

BRUCELLA
(
n,e gc-nu:, Bmcell<l cons1~ts of ,er) small. non-motile.
aerobic. grarn-negatin· coccobacilli that gro\\ poorh
00 ordina0 media and ha, e little or no fermentati,~

1:x.mer~.
Brucella :,pecie:. cause a zoonotic infection in
human:, called brucellosis. The organism primarih affecb
goab. sheep cattle. buffaloes. pigs and other ·animals
and 1:, rransrnitted to humans b) contact with infected
arr'Tlab or through their products. The human disease
Fig. 34.1 Bruce/la: gram-negative coccobacilli (Source :
\\as recognised along the \ lediterranean and is known Department of Microbiology, Pondicherry Institute of Medical
b) ,anous names such as Mediterranean fever. Malta Sciences, Puducherry)
influerz,e~ fe,er and undulant fe, er The bacteria ,vas named after
a Bntish army doctor. Da,id Bruce who discm·ered mistaken for cocci (as was done by Bruce. who called
ed natJ•~
Bnice/la 111eli1e11sis (Bmcella after Bruce: melite11~i~ b the them \licrococcus 111elite11s1s). In older cultures. they
Roman name for c'\lalta) Subsequently. B. abortus and ma) show irregular forms They are non-motile, non-
ful, U'cert. capsulated and non-sporing.
B. s11is \\ ere isolated b) Bang d.nd Traum from cows and pigs
respecti,el). lhese three species cause human brucellosis.

Other species c~JS :1g animal infections include 8. canis, isolated

I Classification
from cases o' e abortion, 8. ov1s from abortion in sheep and Based on nucleic acid homology and gene sequencing.
B neotorr,e rs;:1 woodrats. 8. canis may occasionally the genus Brucella is considered rnonospecific. I loweYcr.
cause a rr r sease, but the other two are not pathogenic due to their differences in pathogenicity and natural hosts.
for huma~ the traditional classification has been largely retained .
There are ten species that comprise the genus . The
organisms most commonly causing human infection and
~ Clinir their natural hosts arc: B. 111elite11sis (goats and sheep).
A50-year-c ·eJ with a history of intermittent fever tor
B. suis (swine), B. abortus (cattle) and B. ca11is (dogs) .
, 'Jf appeltte and nausea The farmer sold
the past th~
milk from s et nearby On exammatwn, no specific
laboratory mvesligation revealed only A second class1ficat1on of brucellae into two main species-
. inftlifn# signs .-.ere 8. melitensis and 8. abortus- is based on their CO 2 requirements,
anemia. G, •• " .,vnaf history. blood culture and standard
:r antibodies were carried out. The culture H2S production, sensitivity to dyes (basic fuchsin and thionin),
agglutmat,on ' agglutination by monospecific sera. phage lysis and oxidative
at this time w.,J Pout SAT titres were > 11,280, suggestive
metabolic tests with amino acids and carbohydrates . Many biotypes
of brucello., ~ 1 , "It responded to a course of doxycyclme and have been recognised in these species. 8. sws strains that produce
streptomycin. H2S are known as 'American' strains and those that do not as
Note: Please rrote the clinical presentatwn wllh non-specific signs 'Danish' strains.
and symptoms Diagnosis depends on a high degree of suspicion in
apatient of pyrexia of unknown origin with a suggestive occupatt0nal
history and a positive specific tests for brucellosis I Antigenic Structure
The somatic antigens of bruccllac contain tv..o main
I Morphology antigenic determinants , A and M, which arc present
in different amounts in the three major specie~.
Brucellae are gram- negative. coccobacilli or short rods,
B abortus contains predominantly A. while H. 111ditcn.,i~
O 5-0.7 x O 6-1.5 µmin size. (fig. 34 I) . They may be
322 Part Ill Bacteriology

contains M_. B. suis has an intermediate antigenic

pattern. Maior antibody component, absorbed A and 1\1 Bruce/la is primanh an intracellular h
affcc t"mg the ret1culoendothelial
·
monospecific scra are useful for species identification bv ,uteni pni . ogen
h
. f
P~c diI ec11on h . \\ ll J
the agglutination test. · or macrop . .iges. dt·ndntiL t'db I DC l ,ind
ti ophoblas1s. The b.ictcna can enter. sunne ..1nd re1,J1 · •
Antigenic cross-reactions exist between brucellae and Vibrio cholerae w1"th"111 Ihesc cc II s· and cause diseast'. This accouni f lw: C
. ,f . • s or n,
and persons receiving the cholera vaccine may develop Bruce/la 1c ractonness to chemotherap1 and the l'll·t'\iS!,
·. bl .. . . . .. . . tnce 0f
agglut1mns lasting for about three years. Antigenic cross-reactions also ,_1<1 e bac11l1 "ll~ high lei els of c1rculming an11b0Jit'S. 'flic

IfpidernioloQY
exist with Eschertch,a coll 0:116; 0·157, Salmonella serotypes group hpopolysacchande component of ihc Brucel/a cdl ,,al' .
N (0:30 antigen Kauffman and White), Pseudomonas maltophlfia, a Yirulence factor ' 11 ,. i ·dk1,1:--
Yersm,a enterocoflllca and Francisefla tularensis. A superficial L ll L'fl C
antigen resembling the Salmonella Vi antigen has been described. Transmission Organisms from tht' infoc1ed animal pu111,. l;,,at~. -.h
enter the human body through a wound, the rnnjunctJ\d .-
J1rcd1'·,tllff\:~"" • 1

I Virulence Factors by inhalation or b) ingestion of pruduLts from in'cctcd

animals.
1111,,n.
,.,n-~t,ll t. ' 11 '
''" ., , ,r,•;1d dc)C:--
.
h1l1t1l111
•

The virulence factors of bruccllae arc the following: Incubation period This is usualh ab0u1 I 0-30 da 1 , f·1'" · 1 · , 1r
The nh1:--t 11111 c
♦ Lipopolysaccharides on its cell wall, especially the but may wmetimes be wry prolong~d. · ·· \' lk pr,,duct:-, Ill
0 -polysaccharides (0-PS) of the LPS play a 1\iajor ,,J!,t,ibk, ,1r ,, atcr
Spread They spread from the initi.il siie of inh:1 0 n
role 111 , irulence .
through lymphatic channrls to the local I~ mph gland, in,... ,·1nkctcd ,101.
u ·,.,
♦ The bi ,d ing and penetration of the organism into the where the) multiply intracellular!) The) then spill 01c•
1 ·,i.,t,C1n j, c~pec1
h,, I e. 1~ h regulat ed by the genes B1·rR.1BFrS.
into the bloodstream and are disseminated throughout h arJ m I ctt'nnar
~ fh ..1!:- 1 IV tc 'nhibit fusion of phagosomes and the body The) have a predilection for the pla;enta. ird i, r:irtkul.irh c.
1, ">s , -; ,·nu rcphL ation in the phagosomes is a probabl) due to the pre sen ct of en thritol in it. which hJs .1 rariou, part, of In
,ec.und \ nulenc.: rreLlianism. stimulating effect on brucellae. Fever, sweats and C\treme to H. mditl'/1.\i!> nc
♦ Ot,.,c rt:r u ..1tory g.:.,e, fo r Yirulence are \firBI- VirB I 2, fatigue occur 2-4 weeks after the initial mfecuon.
""l:ic.h t<.1c1I tat.: the bacteria l intracellular growth in Types Human infection ma) be of three t1 pes.
ohagoc:ytic ard no, phagoc.vtic cells. i) Latent infection with onl) serological but no clinical
~, a bacteriophage Several bac.teriophages that evidence;
, Rr•,cel/a strai ns ha, e been isolated . These phages ii) Acute or subacute brucellosis; and
cd!ly similar. The Tbilisi (Tb) phage has been iii) Chronic brucellosis.
J a~ th e reference phage, and at routine test Acute brucellosis is generall) caused b) B. mclitrnsis
-i [)) ,~cs only B abortus. B. suis is lysed at I I is usuall) kno11 n as undulant re, er but this i, , .. 1 ,.J~t ).
'- J <.TI> \\ 11le B 111e!tte11sis is not lysed at all. misleading as only fe,1 cases sho,, the undulant pattern. •,'uJc culturt, 5Cru
It is associated 11ith prolonged bao.:tcrcmia and irregubr
fever. Symptoms vary with muscul.ir .ind articular pams, I Cultural char1
Resistance ,rvbc, and d
asthma tic attacks, nocturnal drcnchmg rn cats, e-.haus1ior. capno . . c, not
Pr..i.t.'lac are destroyed by heat al 60°C in IO minutes and by anorexia, constipation. nen·ous in~itubilit) and chill,. , ph1lic, rt·qu1ri
I 'i-c phe·1ol in l 5 minutes. They are killed by pasteurisati?n. •rtunun1
Common complications are articular, l1,sc,1us. 1iscer:tl vr , 0 ,. _ kn11't'r,1tu1
They remain viable for l O days in refrigerated milk, neurological. There could be m;irkcd 1hrombocytopc<1iJ u--, _4
♦ l'
one month in ice cream. four months in butter and for with low platelet count,, "hich ma, re,uh in hkcdmg 0 r •rl,\\ th i,
n1••·
•ul;:i ,
sk)\\ ,l
varying periods in cheese depending on its pH. They m~y thromboc) topenic purpura. ' Lllrrcnth <:
also survive for many weeks in meat. They tend to die Chronic brucellosis m;i\ be nun-b;ictcremic. "id1.110,i- er-uni J
tf\ POl,l!o infl
in buttermilk. B. melitens1s may stay alive for six days in gradc infection and periodic e,accrbations The ,,mp!Olll> Pto,c 1
Jnd . ' !!,tr. ·1h;
urine, six weeks in dust and ten weeks in water. arc generally related to a state of, I1) pcrscns111,
. · · ii) · in . th,· l \ t'I I
'de l t.-I\(' 0 lL'Xiinict,
patient. Common clinical 111anifc,1ati0n, nrc "'ratu'.g. p
'nth(' · · r, thrih
The species identification of Bruce/la strains is not ~traightforward. Strains lassitude and joint pain" ith minimal cir pcrk1Jic l')n',,a. ♦ l: gro\\th f b
that behave biochemically as abortus and serolog1ca!IY as melltens1s and The illness lasts for ,car,. I) uc 10 ,.mc' . J ,~mp tc,m,· ~nd r-v\~th o i

vice versa are often seen. Species and biotype 1dent1f1cati0n depends on prolonged fever, the· infect ion 1, c.:linicall) con,ickr,·,I 8 '
d
lld gl' on'' 0 I'd I Ill
a variety of other factors besides antigenic structure. dnd ''tc-ning c.01
one of pyre-.ia of unkno,~ n ori~in l PL Ol ., . ~ wT
1 '-h tough 1,
Jmmunity in brucellosis 1s maml~ cdl-mtd,at, d n ~ o1ngl', i • Pc, c
I Pathogenicity . I • II ,
tvpe of T helper cell response anu ~e -Ill<< 1.1 l r· ( -d
are required to eliminate bn11.:e II ,1c thr0u~Th J ' U\,lh
i1111nun 111
. r-d 810od n ,1ntigcn
111Rn · cu1i11rc
0 1
All three major species of brucellae are pat~oge~ic . . I ~I\ \Jf b
macrophages; tumour necrosis lactcir, a I' h•1 ' d 1;llllll'·l,
:m •' if ,1, ot R ru1..
to h umam. B• l 11elitensis being most . pathogemc. wh ile
and interleukins I and 2 arc i111rort.111t mcdw1,1r, '
1
t i, ' l f
. a or . ,and B• suis are intermediately pathogemc. th '
B b I 11~
Ut,
 rotcctive response. This is probably the most important
~echanism in recovery and immunit~ in brucelh,is.
Chapter 34 Brucella and Bordetella
-
Tissue reaction to Brucella con s ists of granuloma
formation with epithelial cells. giant cells, lymphocytes
and plasma cell s. Granulomas heal with fibrosis and
sometimes become calcified.

I Epidemiology
Human brucellosis is acquired from animals, Jircctl) or
indirectly. Goats, sheep, cattle, buffaloes and S\\ inc arc the
common sources. The modes of infection an.' ingestion,
contact, inhalation or accidental inoculation. Pcrson-to-
l----===---------
Fig. 34.2 Bruce/la suis colonies on chocolate agar (Source:
Public Health Image Library. ID 17132 / Todd Parker, Ph.D.,
Assoc Director for Laboratory Science, Div of Preparedness and
Emerging Infections at CDC I CDC)
person spread docs not ordinaril'.I occur.
The most important vehicle of infection is raw milk.
Bone marrow cultures }ield a higher rate of
Milk products, meat from infected animals and raw
isolation and rema in posi tive lo ng after the blood
vegetables or water supplies contaminated by the feces or
cul ture has become nega tive.
urine of infected a n1mal'i may also be respo nsible. Contac t
infection is especmlly nnportant as a n occu pa ti onal Automated blood culture systems like BacT/
hazard in veterina~ians, butchers and a nimal ha ndlers. ALERT a nd ID sys tems like \ ITEK and MALDf-ToF
and is particularly ,ommon during the calving season. In have g rea tl y redu ced the repor ting time to within 48 to
varioui. part!. of India, m o st human infections are due 72 ho urs, as compared to rn·o to three weeks required
to B. melitensis acquired from goats and sheep . for the conventional Brucella broth or Castaneda's
method s.

I Laboratory Diagnosis Castaneda's method Bl ood is inoculated into a bottle

of trypticase soy broth or Brucel/a broth in a biphasic
The clinical manife.,tat1ons of huma n brucellosis are blood culture bottle (Fig 34 3). Cultures are positive
variable, and only if a high index of suspicion is maintained only in about 30- 50 per cent of cases, even when
will the disease be identified . Clinical diagnos is is almost repeated samples are tested B. melitensis and B. suis
impossible and laboratory co nfirmation is essential are isolated more readilj than B. abortus.
lClinicu/ cuse 1). Laboratory method s for diagnosis
2 .. Biochemical reactions Brucellae are cata/ase-.
include culture, serology and hypersensitivity tests
111· t ~·· ox1~ase_- and urease-positive. No carbohydrates arc
I. Cultural characteristics Brucellae are strict
ordm~nly fermented, though the) possess o., idative
aerobes and do not grow anaerobically. B. abortus is capacity .
d'
cap~ophilic, requiring 5- 10% CO for growth. The
2 3. Serological methods These are . t .
?Ptirnum temperature is 3 7°C (range 20-40°C) and pH d' • impor ant in
fll \ IS 0,0-7.4. iagnos1s. Several serological tests have been d I d
,,, ♦ Growth is slow and scanty in ordinary media. The
• I d' eve ope
me u mg the standard agglutination te:,t (also known as,
Brucel/a agglutination test) and ELISA.
I 11 1 \I I llledia currently employed arc serum dextrose agar,
serum potato infusion agar, trypticase soy agar or The standard agglutination test (SAT) .
d · Ib is commonly
tryptose agar. The addition of bacitracin, polymyxin one m most a oratories. Equal volumes of serial dilutions
and cycloheximide to the above media makes them
&elective. Erythritol has an especially stimulating effect
• ~ the growth of bruccllae.
an~ on ~olid media shows small, moist, translucent
d Khstenmg colonies (fig. 34.2). Mucoid, smooth
: . rou~h types of colonies appear, associated with
ngea in antigenic structure and virulence. I
I
~OOcl culture is lhc most definitive method for the -----+--- Sohd I
- - . lia of brucello~is. In recent years. the isolation medrum I
of Brucel/u lrom lhcse specimens has increased
, ...___
, the automated culture and identification
lrqurd
mect,um
Fig. 34.3 Castanecta·s bottle f;-bk>~ ~lt:re
324
-- -
Part 111 Bacl1mology
-- - -- ---- --
ill a watll lmlh ;ii 70"C 1'01 ti() iO 111i111tll'~. ll a111ihodiL•~
ol the paliu1l ~ sc 111111 a11d the sla111l.ird1sL·d a11ligL'II (I, illcd ;irL' m :sl'III 111 till! 111i lk, the bi1c 1ll 1 illL' aggh1ti11.ill'd :ind
b11Splll5io11 ul a !.landard sl r;1111 ol /1 alwr/11 .\ ) :irl' 1111xnl 1
ri~L' wilh th<.. Ll'U1111 lo lonu :1 hlttL 1111g ut thL' lop, leaving
a11d 111c11h;1ll'd al i TC fol :.M hulll ., (II' ill"( fur 18 hours
till' 111ilk u11sta111cd .
♦ l'osihvc ~AT A l1l Ic ul 1{10 01 111ml is co11s1di:rul
,11'11d ll ,llil Ml>SI pal i,·11ts with m:ut, ln11ccllus1s dl'vclop
tilll'~ ol b•IO 01 lllllll i11 i 1 wcd.s ul ill11i: ,s. 'I 1ln.:s
1c11d lo dl'Ll111,· alt,•1 th\- au1IL plrnsl of till illness lluth
I Prophylaxis
A, tlK 1m11011ty of ln1111Htt i11fcct1011' arc au1uin.:c.l hy Ute
lgl\1 t1 1HI lgl ; a11lihod1,·s ,1ppcar in 7 IO days alter thl'
rn11su111ptio11 of L:01lta11111rnti:d 111i ll, prl.!vc11tio11 rnmi\ls \ Gra
011wl ol di111r;il i111l'cl iu11 . A, till' d1scasi: proµrl'SSL'S, fig, 34. 4 .
of chcck111~• dairy .ininwls loi bntL:i.:l losis. No suitable
lgl\1 :1111ihodil·~ ,kL111K whik Iv<, ;111tibodil'S fll'l'Sist coceobac1th w
vac<:111c j,, avai lable fo t hu111a11s. l low<..·VL I, vacL:ines have (Source: CDC
Ill illlll'ilSl' 1111i11c . 'I hi: 1,,c., rnn hl· d1f\ul'llli.ilcd lrom
bei:11 developed 1'01· use in ani111Hb. U alior/11,1 str:iin 1g gpecitics.htrnll
lgl\1 by lrca1i11g lhi: Sl'i\1 111 wi th 2-111cruq>lul·lha11ul
(21\11.). lgM 1s dc~110yl'd hy this agc:11l lill' liltl' vaccine is proledivc in catt le.
lulluwi11g the ltl'illllll'lll with 2M I· tl'llrds lg(;, llipobr 111
♦ halsc-negalive SAT Scr;1 oltl II co11lai11 'blocking' 01
'non-ngglulinaling' antibodies . I hi: hlod:i11g l'ilccl
I Treatment (In staining

111av M>11H:l11m·s he 1L111uvcd hy p11111 hL.iling ol !Ill'

,L 111111 a, > J l l,ir ,O 1111n11tcs 01 hv ming 4% snlilll.: as
lhuccllom in mlulb 1s lrl.!atcd wit h cJoxyl!ycline for at lea<t
45 day, .ilong wilh strl.!ptomy<..in I M dai ly for the ftrst two I Virulen
ti• ~l1lucnl I ,r thL llst I he 11 ,,·.i 1Lliahk 1m·lhml 101 wcL.ks A complete course is necl!ssary to eracJicalc 1hc 1/urdl'IClla p
b,1,1•• 1 111 biu ~ 1,1 c I L' ,rnd d.:ILLtmg imo111pil'tc' 111fcL.tion Childrcn an.: trea ted wit h w trimoxawlc w11h ♦ Aggluti11
.i 1I " 1, b1 n tl, Pn•hsl 1.:<I. rdm11pic1n 01 gLntamir.:in. ,igglutino
H,",\f
tn fci;.lion
• ;itiv~ 1 .iv occur due lo antigenic
~Ir.till~ t:
--------BORDETELLA -------
,ti r m ncgcitivc
v1rulc11l.C
111 t'll
epithelial
1hc genus IJ,m/etella is namcc.l after f ulcs Bore.let. ·1he
I 11 , i~~ JI rm~ and inc
spcLics responsible for human infec tion is fforc/etelfa
I t •• ,u I • f'ilame,
1
perlussis (per/ussis means intense L:ough). A related
• I• l l \Ill I r' l r the I, I 11 111 llq(,IIIVe uf thL· th
!JaLillus, B. paraperlussis, was isol.itl!c.l from mild cases cf
,a1 m l I g1\ a I ,I po· t v <,A I the othe
Y.hooping cough anc.1 IJ lmmd1iseptica may occasiunall)
f ~d,em l11a ('l)/1 l) I 16: 0: 15 7 to the c
111leLI human beings, prod ucing a condition resembling
~a/111<u11:lla sLroty,1c, group '\l (0: rn antigen trythroc
pertussis.
K.11111111,111 an<l Whitl!) FHA i,
Ac 11Clo1111mas 111altophilio With p·,
Yersi11io e11/eroculiticu
f ra11c1sella t11/are11sis BORDETELLA PERTUSSIS
Su/111011el/a Vi antigen
The complcmcnl fixation tc&I is more useful in ~ Clinical Case 2
1:hronil! <..a5l!s as 1l detects the lgG antibod • l A four-month-old child was brought from a per,pheral centre ,n d
11 h' . I y "so.
owever, I is is no ongcr prefe rred on acco unt of its remote rural area wI/h complaints of severe cough ending wdh a
cum bl!rsome technique typical whoop for the past 1O 12 days and apnea dunng the bouts
ELISA rs :ens!llv<:. and specific and can detect lgM followed by vomItmg The child was said to be sleeping comtorta/Jti
.
. . ,111t1bod1es
and lg(, &eparatcly· It i• lllei·el'ore, use 1.uI
0
,
between bouts. Blood counts were raised w1/h Jymphocy/Os1s There
ror dilkn:nllallon between '•icute aild c.111.omc .111 fect1on
. w:; no hiSlory of immumsalion smce birth The laboratory reporte~0
~h wth ofB · pertussis from the nasopharyngeal swab Adiagnosis ',
Diagnosis in animals oopmg cough was made. In the hosp,tal. /he chrld responded wt,
to erythromycm and supportive care.
Rapid method, such .i s rapid dip&tick 1 1 • d
B. I d I cs <111 Rose
cnga car_ tc,t lHVl! bel!n employed for the de . . .
bruccllos1' 111 herds or callle. lecl1011 ol
Milk ring tcM is uscd tu detect fJrucel/a a t'b d' .
I Morphology
. gt am -ncg.it ive L.oc~ 0 t,aciilU'le
milk. A sample of whole milk is mixed well _n ' o tcs m . . · · · i, "• sma, II , ovo1tl
H.. pertm1i1
. d s hape (Hg 14.4), It ,~ .,c:,r•"'
u. f uniform ·', ·1z.c, ,m ... vii
the
•
stained Bruce/la antigen (·• ,.
u ~oncentratcd su , ol .
wtth a, drop
. u1p~ulatcd h11t H.:nJ, c vSl' th'
killed B. a/Jurtus stained with h , . . · spenston
em<1toxy 1m) and incubated
of .. non -spori ng. 1t 1s
and
t:<1psule on repeated cultiv11tiu11 .
 Chapter 34 Brucella and Bordetella 325

,cmithing factor (I !Sf) (responsible for heightened

~cns1t1vity to h1'ti1111111C) an<l islet activating protein
( IAP) which induce excessive insulin secretion by
t11
the pancr1.;at1<.. 1,ld rells arc seen only in experimental
fl ' animab and not in patienb.
PT b a 117,OOO-molccular weight hexamer pro-
d"'"' tein composed of six subunits with an A-8 structure
Ji! 1hr f (/\ be111g the lnzymatically active moiety and B the
d fq ll
Ul<;lll\
I
l
1 lg J4,4 G1,11ll st,1111 nl ll p111111si;is s l10w111q q1,1111 11oqnl1ve
b 111 J 1ng component) It can be toxo1<lt:d . PT toxoid
b the mujor component of acellular pertui.sis
vuccinc~
, 011nti,111111 wh1rh nµpo,11 tis whoil:; 1051m1llli11q lhu111l1p1111ls • Horclet!'lla pert11.\s1., st rams <.-arry a novel allele for the
1Si111rco C:DC !11\lps //www nlc qnvl pnrlu ssIs1cli11Ical/dIsoaso-
pertussis toxin promoter, which confers increased
nt ,po1'1l11•s htmll)
pl'rtuss1s toxin (Ptx) production. Epidemiologic
llip,,l.11 111l'l,llhrtll11.llll' g1.111uks 111:1~ lw dr111l111,tratL'd data suggest that thesl' strains arc more virulent in
Pll s111ini11g ,,1th t,1l111dim· him·. humans
♦ Ade11ylate cyclase (AC) Known as the AC toxin
(ACI ). 1t acts by catalysmg the production of cAM P by
I Virulence Factors v,1rious types of t:clls
♦ lleat-labile toxin (HLT) It is a cytoplasmic
Jlo,d, tdl,1 f'l'l'/11 ., ., i., h.,~ ,r, l'1,1I , irnkm:l' lar llll''
♦ ~gluli11ogt•t1~ l\1Hlkl 1'lla1· Lill I\ surlal'l' prote111 present in all bordctellac. It is dermonecrotic
,1ggl11t 1m1gl'll" .1,~oL 1,1t1·d ,, ith '1 111hnaL ~ 11.1in, cm1'ing and lethal in mice. Its pathogenic role is not known.
i11il'rt1011 hdlHlg tll t\ l'L~ I . 2 nnd , hl'lll L' ,·at:Cllll' ♦ Tracheal cytotoxin (TCT) It is a low-molecular-
st1,1i 11s Cllllt am tlll's1• l.1l t111, \ gg l11t i11ogl'ns prnmotL' we1ght pep11doglycan whic.h mduces ciliary damage in
BOR0ETfill \II 11kl\l.l' '" hdrmtr b.1LI L'I Ill t,> att,ll h tll Il's pirator) hamster trachea Its rok m disease 1s not known.
rpitlll'li,1I u 'll· They urc u,dul 111 ,erotyping ~traim ♦ Lipopolysaccharide (LPS) LPS or the heat-stable
toxin is present in all bordetellae and exhibits features
und in epidemiologicn l studies .
♦ rilmt1<•11 tm t 1wggluti11i11 (fllA) 'I hi, is one of gram-negative barterial endotoxins. It is present in
lil' th1' thr~ L 1, , lutt11111s p11)d11 u •d b) H. pcr/11., .,i~. the whole-cell pertussis vaccine but is not considered
/\)/\ 'lll 0 till' 11thl't , , PI ,md a lipid lalllll'. It adhere~ to be a proteLtive antigen.
If'< rf1111' \: lo the cih I the res pirntor) epithelium and to

h ,111J 8 ~ I
crylhro9t, . 111bod1c, to FIIA arc protective ,md
HIA b m, u tn nccll ulur pcrtu,,is rnceinc, along
I Resistance
',r~\, rrJl , It is a <lclicatc organism. killed readily by heat (55~C
,,ith Pl m 1d11,.
for 30 mmutcs). drying and disinfectants. It survives at
FHA and PT hell' c 11 , r,s promote secondary mfec!lon by coating 0 - 4°C Outside the bodJ, B. pert11ssis survives for five
other bacteria 'L dS Haemoph1/us mfluenzae and S pneumoniae
V • days in dncd droplets, three clays on cloth and a few hours
and assisting t'l ·r binding to respiratory epithelium This phe- on paper
nomenon has bee'l termed piracy of adhesins.

♦ Pertac:ti11 It i, :111 outer mcmbranc protein (OMP)

anti11,l'n present in all viru lent strmns of B. pert11ssis
,\ntihlid~ to pcrtadin ran be seen in the blood or
d1ildrcn after infcrtion or immunisation Pertactin is
included in nccllulnr pcrtu~~b vaceinci..
I Toxins
• l\,rtussis toxin (PT) Thi, is present only m
B. pertussis . I' r is l'\.prl'SSl'd on the surfacl' or the
bac11lus and Sl'L'rl't1·d inlLl thc surrounding 1111:dium
The toxin e,hihits divnsl' bi,1k1gical lymphocyto,h
produeln1 factor (l.PF) and causl's profound
I mphocytosis in pcrtu"is patil'nts . '] hc hi,taminc-
I Pathogenesis
B. pert11ssis is an obligate human parasite and is responsible
for whooping w ugh or pertussis in humans. Infection
occurs through aerosols and close contact. In the initial
stages, the bacilli are confined to the nasopharynx, traL:hea
and bronchi. Clumps of bacilli ma) be seen enmeshed in the
cilia of the respiratory epithelium./\'::, the disease progres~es,
inflammation e,tends into the lungs , producing diffuse
bronchopneumonia with dc,quamation of the aheolar
epithelium (Clinical case 2 ). Afll'r an incubation period
of about 1- 2 \\.eek!>. the disl'a,c takes a protraded cour~c
rompri,ing three stage!>, each lasting approximatclv t,\o
weeks .
 326 Part Ill Bacteriology

1) Catarrhal stage onset is 111s1dious, wi th low-grade dct ~GCI

fever catanhal symptoms and a dry, irritating cough. 1 he di sease is more common 111 females than in males • sor I O"
• u11 - •
Clinical diagnosis 111 this stage is difficult. This is at all ages. Jt 1s worldwide 111 dislribution. It oc<:urs in • Jlc! ccphal
.ilso the stage of ma\i111u111 inlect ivit y. (If wspected, epidemic form periodically; the disease is never abscni unu
from any community . ·r11tof)
treatment can arrest the infection at this stage.) rc1 Pl
Whooping co ug h is o ne o f th e most infectio us of I rcu11I
1il Paro:1.yMnal stage leads to increase in coug h intensity • Ciu inrLll
,1nd occurs in distinctive bouts. During the paroxysm, bacteria l di seases and no n-immun e conta cts seldom rc11n'
escape the disease . 1hib1tory '
the patient e,perirnces violent spasms of continuous u I ·ii
coughing, followed by a long in-rush of air into the d111rco11 ,'
With universal immunisation, childhood pertussis is on the decline pefO\
iJc, •
,,
lmost v11pt) lungs, wHh a charactrriMic whoop
but adolescent pertussis is on the rise due to waning of immunity Plait'> url'
1hc1" t 11' 11ame v.hoop111g cough) (Fig. 34 j)
at that age Adults become prone to infection unless they receive
11 (om, lescent stage follm,s the paro\ysmal stage booster immunisation. Growthis
1c r , rd sevc ·1ty of cough grndually .Jlonil'' :i re
, c 1sJall} la, s b-8 wrrb, though Around 1980, pcrlussis cases started increasing ,151:id. grc
b r ot•aLtrc. grad ually and by 2017, it had risen lo 18,000 in the US resembling
(CDC). In India, around 25,206 cases of pertussis were (onllucnt g
reported in 20 I 5 ( EP I r-act Sheet 20 16-WHO) JppC,JJWlCl',
B. pertussis causes 95 per cent of whooping cough Jircd i1111nu1
c 1km ir, L0JT phL,l!Jom, could arist cases. About 5 per cent are caused by B. parapertussis b1ochc111ical t
~,sure effects (subconiunclival hemorrhage, and infrequently, whooping cough may be caused by
t, 1ro..1s .:n•physema)-thesc occur during the B. bronchiseptica. 4.Biochcmic11I
'L'Ut of coughing. ~ac11vr It prot!
H~\jllralory complications (bronchopneumonia. lung
LollapscJ these are self-limited, with the atelectasis
I Laboratory Diagnosis ; Pol) mcre)c
fl! morl' , rnsit h
resolving spontaneously
The bacilli are present 111 the upper respira tory tract mos t ,uhur~ ) idd i, p
♦ Neurological complications (convulsions and abundantly in the early stages of the disease. They ma} be
coma)-these may result in permanent sequelae o Serology Sc
demonstrated by microscopy or more reliably, by culture. ii not used roufi
such as epilepsy, paralysis, retardation, blindness or
deafness. I. Collection and transport of specimen Res pirator) 7 01her I
samples can be collected by per-nasa l swab. pos t-na,al
The infect ion is limi ted to the respira tory tract and
swab (dacron or ca lcium alginate swa bs) or using the cough
leucocyto~is j~ ~
the bacilli do not in va de the blood stream .
plate method. The swabs are to be plated without delay btcocy1ic cou ni s
per cent IYmpho1.:i
I Epidemiology
or tram,ported in a 0.25 0.5 ml Casamino acid solution,
at pH 7.2, in mod ified Stuart's med ium or l\lisc hulu\\ ·s
charcoal agar
Whooping cough is predomi nan tly a ped iatric disease,
th e incidence and mortality bei ng highes t in th e firs t year ♦ Per-na sal or nasopharyngeal swab A lle,ible
of life. Maternal antibodies do not protect against the nichrome wire is passed along the floor of the nasal
I I
di sease. Immunisat ion should, th erefore, be start ed ea rly. cavity and material is collected from the phar~ ngeal
wa ll . Nasopha ryngea l aspi rate collected through a ,ofl
lio)
ca theter att ached to a syri nge is a better source. It can 'ti!
be used fo r PCR also. I
♦ Post-nasa l (per-ora l) swab Secretions from the
pos terior pharyngeal wa ll are collected with a cotlon
swab on a bent wire passed through the mouth .
♦ Cough plate met hod J\ culture plate is held about
IO 15 cm away from th e patient ·s mouth during a bout
of spontaneous or induced coughing so thar droplets of
respiratory exudates impinge dirccth on the medium.
2. Microscopy Micros<:opic diagno,;s depend, ,lll the
demon stration of the bacilli in re,pira1 orv , ec retiom by
Fig. 34.5 Pertussis 1n a female infant who presented with
Gram ,tain or fluorcsccnt antibody techni~ue
paroxysms of cough (notice the indrawn chest muscles and
protruding tongue) (Source. Public Health Image Library, ID 3. Cultural characteristics It is an obligate anohc·. It
6379 / CDC}
grows best at 35 - 36°C. Comple\ media are neu•, s.ir, lor
primary isolation . The media <:ommonl\ used .1rc: ·
 ('\111ptiil J4 (1!1111111 ~ IIIHl flu1,h•lull11 327

l\,)~d C:,'Uf.•'" 1-~ ,,-rht.·•11 , 11111<• h\,,,,,1 .11-111

h,·,·11 11rn h,11,I) i1111111111lw1I. 1111•, ,IH11tld ,ti,,,
h1:
R~i11n l ,'"' ""·,1 1 " 1h h,,r,,• ,,,,,,, 11,\ , h.11 , ,,.,\ ~1,,·11 ,,,,,,,h,l,l\i S p1t'l,·1,1l>h 1·1,1l11,1111\,ill, N1111
111111111111-.·,I , , ,11 111, ts ,h, •11ld I r,·,·111.' ,·1, tltt ,,111~ l'in
', ,"C'1•h.1l,"-1'•", ,,, "' l11hibi1 th,' 1i.1n11.1l 11pp,·r
l'"'l''"'·"I ~ l,11 I\\ d.l\ ~ 1111,1 ,·,,11111,I \\ilh 1h1.• pitlil'III
"''plmt-,r~ 11,,, .1
Ch11~,}al bh•,-.1 llf.11 1'- ,h.11,,,,1\ ,,, 1,,11 , ,, h.111~,· h.1, l ,·.1,,·d .
l ,1111plk111h111, I ,1,·1,,1, , ,,11111l>111i11~ 1,, 1,,xiv11, "'
;, ' \0 , ,, ' ' '' ·'"" Ill \ 1111111,' 111,,h h Ill .,ti,,' th,·
, •l• h , ,nl,,t,111, , , 111 th, t\"'I'" •' ,, , , ,, , 1111111 , l'•"' '""·111111 ,·11< c·11h11l,,1 1nlll\ h,IH' 11,,1 h,·,·11 dl'l111vd ,
\11,,1h,·1 11,h,·1M' .-tk,I 11!,11 , 1111 \l\\'111 ·I~ h11111 .il11·1
,·h:tf\:,).'\\ l,ll\~ \\ ilh ,t.11,h. lh'\11 1.lh,,, ,I I\ h '"' .111,I
1•,·1111 ,1, ,,, lll'I 111,T 111,1th111 is 1h1• h)puc,111il'•
,'I\.'' ,k, ,, hi, h .1r,· 1, ' " ' h, /:, • ,I. 11 ,I
In p111 ,·,pl1t1,h ,, q,,,,,dc ( 1111 Hl I hi, is 11 1 -u 'I' 111· l'd
l' ,\t,> .w, 111- ub.11,,I III l11~h l11111H,h1 , ,11 , , ,,, ~'
,, 1,11 ll"lllli,111 Ill (11111111111,,l(lllll 1''ij)l'l i11\I) pl'l lll~Sh
\.;l\.\\\lh ' '"'" \ lt.-1 Ith 11l•.llh'II 1,,, ,~ ' h,•111,.
,,,11111111111!( ,,1,,11w, lh,·tl' " ,knc,tsl'd s1.'11so1y
,. ,,n ,, , "' , n1.1ll. ,l,1111,• ,h.11•,,I , ,111,,,,,h , •'l'-"I"'-.
1\\\,11,·111·" , ir l,,s:- ,ii l'11ns.·1u11s111.:,s U(1·,u11p,111h:d h)
\ ,, 1d f:1'1.'~ 1,h \\ hit, ,llh ,. ,,,, \\Ill~
11,111,11 1111d 1111",·k II\ p,l!l1111lil) . I his is ,1 ,c,crsihil·
,,, ml•hng ' b,,,·,·t,·,I p,· 111' , •r ' mn,·11n ,lr,,p, ·
11,h ,·1,,. ,., ,·111 l'.1111.•111 ~ 11.' l'11H'I' wit Ii s11pp1H li1 c
l',)nllu,·nt ~'''" I 11,,, •, 1 , ' ,1!11111111111111 p.1i111 ·
,pr,•,1i.m.:, n ,,,11,1 •1,•\\ 1h '·"' t-.· ,,,111'111111•,1 1,, 111,111 l!,!1'1111'111
t.\1111r11i111lk11tion, II s1'\l'tl' u1111plic.iti1u1s sm h
d11"1.-.:'I lll\ll\lllh'l1\l,'r,•,,, 11,, 11,111 1: ,,,, , 111, .11111,,·1,1 ,,1
,1, , 11,·,•11h,tl1•11,11h~ ( I i11 , IO 111illio11 , ,1n·i 11l·C~),
t-1,, h,-mh·,11 t,, b
, ,·1w 11·, , sh,,, k ,,1 II\ p1.·11') 1ni,1 d,•vclup lollo\\ i11 g
-l Biochemical n: .:11c,n, /i -. "·" '' 1, "1, ,, h,·1111, .1lh 11,·1111"'' 1,1"·111,111,,11 11h,cq11t•n1 dusc::; ul the 11K.:111c
\, '11,,. It pn..-.l ' ',,,,I.,,.\ 11,I ,.11.,1.,,,. 111,' .. ,1111,1111di,·,1tl·d Ruucinc pcrtu"h ,u,·dnulion h
'i llollmcras h: cllon (l'C R) 1'1.. ' 1' 1,.1,,·,I 1,•,1, m•I 111hh11hk nlln th,· 11~1.• ut w11.•11 )l'11r, u, 11,h·cr,c
.1 ,. m,, ,' ,,n,11 ,, "' .,,_,,..,. ,,,111111, 1 11\1 11,,·,l 11< 1\\ . , , th,· r,·11llh>11, ,1n• likch
1.ultur,' \Id,\ I" ' 11 , 1111"1' .: ln•/111/111· 1'11<'<'111<' \1·1·11111,11 1mci11,·s ~t>111111 11i11g 1hc
Laboraf l n:
11
l• ::,crolo~ S 1lug1,·nl rl111 i:t11"'' 1, '"'' h,·lpl11l ,1ml t'""''d11,· "'"'I'''"' 111 ,,l till' p1:1111ss1, ha, illus (P 1:
111 \ ni:i:l11tt11u)t1.'II' I , l, ; .11ul p1.·rC11l'lin) , lir~I
1, 11,,1 u,cd rou tmch for diu.:n•"''
~ Other rnt0r) p11 r1111\\'(1.•n \ l,11 kc,I ,k,,•l, 11•,·d in l.l(',111 . .11, '"'" used 111 111,,,1 u11111tr1cs
,1, th,·, ,.111,,· l.11· k" 1.'I' 1\',11.· 1i,>11s, p,1111,·111,11 h i11 ,,llk1
l,:,u,,K,t."t' 1~ ,•,md \\ 1th r--·1.H,,,. h1111 1 h,,, \l,,,1, u, 11.il
ku,·,,, , 11, ,'l'lllll' ,!l) th 'll ,ll, \ll\ll 1•,·1 111111 " ith l1ll :--ll , l11ld1,·11
Huth "huk ,·l'l l 111\11 ,11.·dlulur , lll'dlll'' huvc 11
p,·r ,, nth mph,,.. , h>' ,,l 11h1111t <)O pn Cl'lll. \\ ith h11k
pn111.•1.·li1H\ 11111.• II
,,·II , ,1 ..·111,·, th, ,,,._,,,., I11111 ,k l'11m · lu ,o p1.'1 Ll'III
l'hltmat i11 ,1h,1111 Ii,,· \l\lri. .111d is 11hsc111 ,1l1c1 12 )•'HI~ I 1c11
l11lh 111111111111,,·d ~11h1,•1.·1s Ill.II dcn•l11p pc1111ss1, h111
El /'< 1,,~1, 1, '"'"'pllbl,• t-1 ,,., ,•r,11 11 111i1111 111,, \'1111
not ~nidllin l \ 111i1111,·,.. ,t,,.1l tlw1 •'I') ,, h,·111•11,1,tl th,• d1'l',\Sl' b 11111d
,1 nh 11 1mt1.1t,·d \\lth111 th, I nl 1, II d,\\' , 11 tlw di.,, ,1,,· .
1

E111hrom~dn 1.1r d.1rilh\'\llll)t:lll ,11111 ,1tithn,111n•111 ,1n·

BORDETELLA PARAPERTUSSIS
the dru11,s of ..:h,ik,·
lhb i, :111 i11hn1111·111 u111s,· llf \I ltPupi11g u111gh I he
I Prophylaxls di"'•'''' is 111ild. 1h,' pnl11~s1, , 1111 i111.• du, s m•I p1 otn.l
llf,1111'1 II, I•,Im1,,•r111.,.,,., 111kd11>11,
M-'Jwl••N'II kill,•cl 1·,u·ci11t1 :-1'1', iii, 11111111111b.11i,111
"ith kilk·d /i raw.,~•'-' , ,1,·,·i1w l\;1, b,·,•11 f, 1111,d "'"
clll'\:11\l' Smu,11h ph,1,,· l ,truit1 is 11,,'d f,,, , ,1,·,·11w
pl\)\\U(li,,n l\·1 tnssi, , .1, ,·i11,• 1~ 11,11.11\1 .1d111i111s1,·1 ,·d BORDETELLA BRONCHISEPTICA
m 1:1.1mbmu11,,11 "ith di]'hth,•11.1 .111d i-·1,11111, t,,,,,id
I hi, i, 111, 1111,• h, ,,, 1111il'l1111c IIH ~t'll., 11 1s u11t1gc11i1.,1II)
Uriplc Vl\(\;U\l') /l /'< rtm.,i, .,b,, .,,·1, .1, .111 .1dj111 ,1111
tor the toxo,d,. 1,1,,d11,·i11~ \1,•tl\'1 ,111111 1, 1,h 1,·s11< 111,, "l.11,•d II• I/ f'• 1tn"1s 1111d /lru, cIla ,1/1mt11s II It., l>(,·n
l1llllld Ill ,,111,,• I I ll'I) ~111,ill 1•1u1 1 ,,111011 \ll I p~r 1.~111) or
Children under hH1t' n·nr, "1111 un· ,•untud, ul
patients should n·(,·in· :, h,1,1,1,·r ''"'" ii th,·, hnd , '"'' ,,, ""''•>l'i11g "'11rh
 Part Ill Bacteriology

m EY POlNTS ~
❖ The genus Bruce/la com prises ron r I L I, 1 ~l,•c, gr m n, .__dt ve coccobacill1 that grow poorly on ordinary medi a and have
little or no fer mentative propertks t , r .. t ,11 < , Js1.1" ,n the modaw, catalase ,rnd urease tests.
❖ Brucellosis is a zoonotic infect,o"l C.dr..c;ed by d sprrn_,c; of Bn,cella Bru,e1/a abortus prod uces a less severe form of the disease with fewer
sequelae than that caused by Bruce/Ja suis or Rrucella mel1tenm.
❖ The reticuloendoth el 1al system 1s affected in brucellosis. The Thl ce ll response and cell-mediated immunity are required to eliminate
brucella.
❖ Blood, bone marrow aspirate or biopsy material are obtained for culture . Serolog ica l t est s are th e mainstay of diagnosis .
❖ There are currently no licenced vaccines for human use against brucellosis. Animal vaccines are considered unsuitable for humans.
❖ Members of the genus Bordete/Ja are gram-negative, rod -shaped bacte ria which are aerobes, non-motile and catalase-posit1ve . Bordetella
pertussis is by far the most impo rta nt species.
❖ 8. pertussis causes whooping cough . It attaches to t he nasopha ryn x, and then grows and spreads to the ciliated cells of the bronchial
tree . The bacterium secretes several toxins along with adhesi ns t hat lead to cell damage and accumulation of fluid, which induces the
paroxysmal cough .
❖ For diagnosis, nasopharyngeal swabs or aspirates are collected . Bacteria can be cultured on charcoal agar or Bordet-Gengou medium to
yield characteristic bisected pearl colonies.
❖ Erythromycin is given to treat active cases. Vaccination of infants and children is done as part of rou tine childhood immunisation, and the
bacterium forms a component of the 'triple antigen ' (diphtheria-pertussis-tetanus) .
❖ In recent years, acellular vaccines have also become available. Aggressive vaccination to achieve herd im munity has resulted in a fall in
the incidence of the disease.

I QUESTIONS 0------- - - - - - -- - - - - - - - - - - - - -
Essays Short Answers
1 Describe the pathogenesis and laboratory diagnosis of 1. Serological tests for the diagnosis of bru ce ll osis
bru cellosis. 2. Epidemiology of brucellosis
2 Write in detail the pathogenesis, virulence fact ors and 3 Milk ring test
prevention of pertussis 1n children . 4. Virulence factors of B. pertussis
5. Pertussis vaccine