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Distribution of Pseudomonas fragi

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DISTHIOTTIOH m W M f . . *****
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UMI Number: DP12868
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SABLS OF OONTSHFS
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IKTRQDUCfiOM.......................... ..........................................................................
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SXTIXf Of UfKBiSQaS.. ...................................................................................
3a
METHODS........................ . . ...........................................................
5
C o llectio n o f sa sp le s...............................................................
M ilk s ta p le s ..........................................................................
B a tte r im o I mwo o*a ...............................................
a a e wa e e a a a a a o a o a e a a e a e o a a a . a a a a a a a a a a a
B aliy p la n t m $ 1 h ..............................................
t e n sam ples.......................................................
5
5
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Jf
5
5
Iso la tio n o f e a lta ra s ..........................................................
Id e n tific a tio n o f c u ltu re s
........................................
5
6
UPEHIM Sm i................................................................................
7
General e sa sid o m tie ss o f is o la tio n p ro ced u re.
..............
Oaftoral ooouroos o f s a a p le s .
..........................................
D istrib u tio n of Fa. frairit in Iow a.
...................................
D istrib u tio n
D istrib u tio n
D istrib u tio n
2M .strika.tioa
S titiita lia i
In normal d a iry p ro d n o ta. .........................
in d e fe ctiv e d a iry p ro d ucts..........................
In d a iry p la n t e<jaipnant.............
in d a iry p la n t s a to r s u p p lie s ............
an farm s
....................................
7
8
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8
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18
lk
Ik
D istrib u tio n of F s. f r a a l in E entu d qr.....................................
21
In normal d a iry prodnota.................................
in d e fe c tiv e d a iry p ro d n o ts. ......................... 21
In d a iry p la n t equipment
...........
21
in w ater su p p lie s...................
23
on farm s............................................
D istrib u tio n
D istrib u tio n
D istrib u tio n
D istrib u tio n
D istrib u tio n
D istribution o f £&. fron t in the united S ta tes
...............
Id en tifica tio n o f cu ltu res o f £ g . fra u ti.
Sons fa eto rs o ffse tin g aroma prodnntion.......................................
sisobssios of
21
msmm........................
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$2
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kS
mmm............................
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ACKHOWLEDOmEST...............
531
LHESASQFS OISSD.
52
...............
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
23
IHfROHJQflQI
»n»«y
i f b a c te ria produce
t« «4iic u «th i»
d a iry p rod u cts ra p id ly , a co o p erativ ely m a ll masher o f sp ecies oanse
s o s t o f th e p ra c tic a l d if f ic u ltie s ,
o f th e se .
jpftffiffim ** f r a a l appears to be one
I t s is o la tio n 2ms tee n rep o rte d fro a suae reu s sasple* o f a ilk ,
o res* , t o t t e r and e th e r d a iry p ro d ucts.
Many o f th ese s a b l e s had a
d is tin c tiv e e s te r - lik e odor, s t e l l a r to th a t o f th e May ap p le,
(Podanhrllw a n slt& taa) o r were ran cid ,
Paftadonfflaas fr» * i i® o f esp ec ial in te r e s t to th e d a iry in d u stry
because, in a d d itio n to I t s a b ility to hydrolyze f a t , i t i s p sy ch ro p h ilie
and ab le to grew a t tesp eratu re# o f 5°C* o r low er,
f h is a b ility ren d ers
i t s presence p a rtic u la rly im portant in h o tte r o r o th er d a iry p ro d ucts
sto re d a t tem peratures considered low enough to p re re n t growth o f many
sp ecies o f a ia ro o rg a a isa s,
fhe d e te c tio n o f
f r s * i in naey s a b le s o f d airy product*,
some o f ffhioh o rig in a te d a t w idely sep arated p o in ts g eo g rap h ically ,
suggests th a t i t i s widely d is trib u te d .
I t s freq u en t occurrence in a llk
in d ic a te s th a t i t probably Is found c lo se to th e source o f the m ilk supply.
In o rd er f o r the d a iry in d u stry to combat o r c o n tro l t h is organism more
e ffe c tiv e ly , d e ta ile d inform ation is needed reg arding i t s h a b ita t and
d is trib u tio n ,
m is in v e stig a tio n was undertakes p rim a rily to supply srach
d a ta .
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
B sfz st o f h & w m rn
E iohhols, in 1902 (H ), is o la te d an org an ise fre a MUc h eld 8 days
* t 3*5^ be 7®C, which he describ ed a s & noa-spore form ing, e h o rt-red
th a t Has s e t t l e by asaas o f a sin g le p o la r fla g e lltw ,
B a c te ria l f r a * i.
He nased i t
L ater In v e stig a to rs ( ? ) , (10) t ( U ) , (1 2 ), (15) h are
c a lle d a tte n tio n to th e f a c t th a t t t should have te a s p laced La th e g an ts
Pseudomona s because o f i t s sin g le p o la r fla g e lln s .
She s e a t s trik in g b io ­
chem ical c h a ra c te ris tic * t th le organ! i» was i t s a b ility to prodace a
s t r awb e rry odor and fla v o r on a l l media u sed exespt p o ta to .
She ease y e a r, S rab sr ( 6) ,pu blished a d e sc rip tio n o f an organ ise
producing a straw berry aren a,
St was is o la te d f r e e b e e ts and was a ls o a
non-spore form ing, s h o rt-ro d ,b u t was m o tile by eeeae o f one t e n in e p o la r
f la g e lla .
Be named i t Pseudomonas fy»M *rlaa.
I t d iffe re d fre n Bacterium
f r a r i in th e ty p e e f colony and in th e production o f a flu o re sc e n t p ig n aa t.
In 1905, G raber ( 7) is o la te d f m
was ee sim ila r te Fsendnaonas
fra a a rla s I I ,
p a ste u rise d a ilk an organism sKdoh
th a t he nested i t Pseudomonas ,
I t prodnoed a straw berry a re s» b a t was not flnhsw see& t.
She co lon ies on g e la tin were la rg e r and wore ra is e d than those o f h ie
e a r lie r organism.
Sassong (11) ie e la te d fre a a sample e f ran cid b u tte r an organism th a t
produced an odor sh ich he describ ed a s resem bling th a t e f a flo w er o f the
Bgy a p p le .
When he attem pted te p u rify M e organism by p la tin g methods,
he fennd sev eral colony v a ria n t* .
a sso rtin g to sin g le e e ll is o la tio n
technique, he found fiv e d is tin c t colony ty p e s developing frees c u ltu re s
green fro a a sin g le e e l l.
Be designated th ese a s 8 ( sa e o ih ), $ ( in te r ­
m ediate) and H i, 82 sad *3 (ty p es o f rough) c o lo n ies.
Xasseng concluded
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
th a t th e th re e sp ecies d essrih ed h r Siehhol* and Srufcer were in r e a lity
v a ria n ts of a sin g le sp eo ies.
l i e in v e stig a tio n in d ic a te d th a t
B aaterixat fre e d M ekhels corresponded to h i t 1 . ty p e, FsoafeBMaias
frcwBftTiae Sruher te M e 0 type and Paendoaoaas f ra sa»rl se | | 6ruh er te
M s S typ e.
i t B e rg er's Manual o f D eterm inative B acterio lo gy , 5*h
e d itio n , f t ) th e organise i s lis te d ae
(B ichhols)
Etasseeg, lo n g and Hammer.
An organ ise Aeeerihed hy Conn, I s te n end Stoeklag f 3) eg?*®* wwy
c lo se ly w ith t^imdmaQpaH fraw i and prohahly should he included in th e
eaee group.
I t met is o la te d fro a a ilk preserved f o r se v era l weeks a t 1*6.
and was a sh o rt m onotrichic re d , which, la a il k , gave r is e to a pronounced
o d sr th a t was wet described f e lly .
I t was eased
L nati■
SSpjlMSjf.fh e e a rly in v e stig a to rs who is o la te d and described Pseadeaowgiff fr^ td
d id s o t g iv e d ata on th e d is trib u tio n o r h a b ita t o f th is org aaiae.
Ih e
sources from which I t wee is o la te d were raw s i l k h e ld a t lew tempera­
tu re s , (3) (H) p a ste u rise d a il k (7) sand sugar b e e ts ( $ , A fter encenm ter*”f yfffflffWllffiiy fraw i l a ran cid t a t t e r , Hoseon* (11) and Basseng, lo n g
and la m e r f ig ) ew salsed a v a rie ty » f a o ra sl and abnormal d aisy prodnota
to determ ine i t s presence l a thaw .
St was is o la te d fre n occasional
samples o f nonea! m ilk, cream , h a tte r aad ie e eresaa.
ah&orm&l d a iry products a ls o y ield ed th e organism .
Many samples o f
l a se v era l oases
Pseudomonas f r a r i wfeg Is o la te d fro a samples o f raw o r p a ste u rise d a il k
aad cream which developed e ith e r a Ik y ap p le odor o r raa o id fla v o r aad
odor m b eing h eld a t low teey eratw ro s.
Many sasqples o f re so ld h a tte r o r
h a tte r w ith a May sp ple o d or, e s p e c ia lly tho se w ith a low s a l t c o n te n t,
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
ware found t a rnwtaUr th e o rg an ise.
f r a e l p e r g ra s were weiy M # t.
O ften th e numbers o f 21EB8SBHB6B
Cottage oheese and ta l k condensed m ilk
w ith d a fe eis suggesting 19m a c tio n e f Eaeadaaonas ftnjud. were fonad to
co ntain th is organism l a sev eral e a se s.
The is o la tio n o f PjMradogaaaa
f r e e ! was a lso rep o rted frow inch sources a s hoaogeniser packing, w ater
fre n a lak e on th e Iowa S ta te College campus and sheep f a t th a t had been
h eld l a th e lab o ra to ry f o r sev eral weeks.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
C o llectio n o f Saaples
»* M ilk tammXmm ««re tskew fey Boos* o f s t e r il e a ilk th ie f p ip e tte s
d ire c tly fro a cans o f a ilk feeing d e lire re d to a ilk p la n ts fey producers.
They were p la c e t la s t e r i l e , co tto n stoppered to s t tu b es.
b.
B a tte r samples aero ofetaiaed fey reaowing a a a ll p ieces o f b a tte r
fre n a p r in t o r l o t o f fe a tte r w ith a s t e r i l e w ire loop o r s a a ll k n ife .
ft* B alry s la n t samples included w ater and ewafes fro a v ario u s
piooos o f p la n t equipment.
sore* cap g la s s fe o ttle s.
Water saap les wore oo llooted la s t e r il e ,
Saaples from equlpae&t were ofet&iaed fey swab­
b ing p o rtio n s o f th e su rface o f equipaent w ith a o iste a e d s t e r il e o o tto a
swabs.
4.
Farm smnoles laoluded w ater, s o il, bedding, feed and bajr aa4
swabs from sueh th in g s a s a i l k u te n s ils , a ilk s to o ls , feara f lo o r s , du sty
lo d g es, stanchions and fla s k s e f cotra.
s t e r il e , sersw cap g la ss ja r s .
The form er wore c o lle c te d in
U olstened s t e r il e e o tte n m ake were used
In e o lle o tin g a a te r ia l f r e a th e equipaent and cows.
Is o la tio n e f C ultures
a.
m enrio b aeat process was re g u la rly used because e f th e h ig h
b a c te ria l co ntent o f nsay o f th e s a b l e s .
As suggested fey & ssen g , long
and Hawser (la ) , a ilk and e re aa sas& les were kept a t 5° te 1O®0. f o r a
p e rio d o f 7 te 10 days b e fo re p la tin g .
Sw ell p o rtio n s fro a samples e f
w ater, s e l l , hay, bedding and lik e n a te r ia ls were added te tubes o f
l it s o s a ilk which, in tu rn , were su b jected to th e enrio h aeat p ro cess.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
Xa th e case o f swabs, th e e n tire swab was p laced l a a tube e f litm u s
a il k fo r enrichm ent.
b.
Ed® enriched saap les were p la te d ea a ile blue s u lfa te a sd ia a
w ith f a t em ulsion added a f te r th e method o f Hammer aad C o llla s (8) and
e f Long and le a s e r <13) .
u sed.
E ith e r beef in fu sio n o r beef e x tra c t ag ar was
B utt e r f a t m e employed re g u la rly fo r th e em ulsions,
H a te s were
incubated a t room tem perature fo r 2 to 3 days.
c.
C olonies o f lip o ly tic organisms which were suggestive e f
g e. fra x l were picked in to tubes o f litm us m ilk.
C h a ra c te ristic co lo n ies
do not take up much dye fro a th e medium and appear alm ost s h ite when
examined fro a th e b et ton o f the p la te w ith a hand lo n e.
c u ltu re s were incubated a t room tem perature.
The litm us a ilk
U sually a f te r 2 o r 3 days,
c u ltu re s o f g£. f ra x i developed a Say app le odor.
CJoite commonly they
developed a p in k o r a e id rin g a t the su rface o f th e m ilk next to th e w all
o f th e tu b e.
On extended incubation th is acid a re a deepened somewhat.
C ultures e x h ib itin g c h a ra c te ris tic s o f go. f ra x i were preserved f o r more
d e ta ile d id e n tific a tio n stu d ie s.
Id e n tific a tio n o f O ultores
C ultures were f in a lly Id e n tifie d on th e b a s is o f shape, s is e , gram
s ta in , m o tility , la c k o f sp ores, lip o ly s is , g e la tin liq u e fa c tio n , produc­
tio n of a May apple odor in m ilk and production o f a c id fro a arab in o se,
d ex tro se, g a la cto se , g ly c e ro l, la c to se and lev o lese.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-7 TETPTgBTWTiaWAli
S eaaral co n sid eratio n s o f is o la tio n procedure
While th e is o la tio n procedure used in d e te c tin g £*,. & S & ^
*•!**-
tiw ely sim ple and, judging frost th e r e s u lts , reasonably e ffe c t le e ,
undoubtedly t e « ■ » in stance* i t f a i l s t e d e te c t th e o rgan ise,
n e c e ssity , t b* assa& ts e f m aterial examined e re w a l l.
@f
la «<itr f mtpi ■«
which co n tain very f e e £§,, f r a a t. th e organists say he s i seed e n tire ly .
Sampling ef h a tte r and cheese p re se n ts d if f ic u ltie e because o f th e
probable uneven d is trib u tio n e f organisms throughout th ese p ro d u cts.
S e ll and sim ila r m a te ria ls p re se n t d if f ic u ltie s because e f th e la rg e
numbers e f e th e r organisms p re s e a t.
Many e th e r sp ecies e f b a c te ria ,
e sp e c ia lly those p re c a st i s t o l l , a re capable e f growing a t th e tem­
p e ra tu re s used i s the enrichm ent pro cess f a r Eg. f r a g i.
A fter h o lding
a ilk in o au lated w ith e th e r m aterial a t 5° te 10®0. f e r | to 1# d ays,
p la tin g * u su a lly must be aade a t d ilu tio n s e f 1- 1 , 000,000 e r h ig h er i s
o rd er te o b ta ia p la te s c a lla b le f e r p ick in g o o lo a ie t.
Hass th e propor­
tio n o f th e t e t a l o rig in a l f lo r a r e p r e s e n ts by go. f r a a l i s ia p e rta B t.
On th e o th e r hand, th e d is tin c t, p le a sa n t aren a produced by th in
organism i s a marked a id in determ ining i t s presence, b o th l a m ilk and
on ag ar p la te s ,
fh e presence o f n ile b lu e s u lfa te la th e m sdlm i s e ls e
advantageous because i t ten ds to in h ib it th e growth o f se es o th e r
organisms w hile ap p aren tly I t does n o t g re a tly in h ib it gg. f r a d .
In
o oasid ering th e r e s u lts rsp e rte d in th is stu d y . I t should be recognised
th a t f a ilu r e t e is o la te th e organism from a sample dess n e t preelude i t s
p o s s ib ility o f i t s presence in th a t sample.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-
B -
Se&sral m m i o f s a b le s
Samples examined to studying th e d is trib u tio n o f
fg a g j came
Hem ra th e r w idely sep arated geographical aren a. Most o f too samples
o rig in a te d i s to e s t o t
and a i m
to
a
con sid erab le mother se re obtained
o th er s ta te s .
to
X esta^gr
Because o f th is general rolatioM feip to e
d a ta to d is trib u tio n e f g&, f r a s l a re p resen ted to t o n e se c tio n s.
D istrib u tio n o f |& . f r a a l to Ie to
to a s M
to U s:
Samples o f m ilk fro a th e
re g u la r supply d eliv ered hy p atro n s to th e Iowa S ta te College m ilk
p le a t to re examined fo r th e presence o f Pg.. fr a /tl.
d a ily fro a lk farm s w ith to a rad iu s o f ? s tile s .
Milk was receiv ed
A tru ck ro u te served
th e m ajo rity o f th e se farm s w hile a few produeers d e liv ere d th e ir
products in d iT id u ally .
fro a 5 to SI samples were obtained a t in te rv a ls from th e m ilk
su p p lies o f th e v ario u s farm s, as shown in fa b le 1 ,
Samples o f m ilk
fro a 10 {JX.k p e r cent) o f th e lk farms y ield ed £&. f r a r i to one o r so re
to etan o es.
fh e p ero to tag es o f saap les fro a th ese 10 farm s th a t y ield ed
? s . f r a e i ranged fro® 7 .7 p e r cen t to b2 .9 p e r cent and averaged 20.6
p e r c e n t,
fh e organism was found to 7 (7*7 P«r cent) o f $1 saaples
obtained to O ctober, to 16 (22.9 p e r cen t) e f JQ saap les c o lle c te d to
Boveaber and to 6 (kO.G p e r cent) o f 15 saap les obtained fro® December
to May.
y ield ed
Of th e to ta l 1?6 s i l k samples examined, 29 o r 16.5 p e r cent
fraidL.
too sample of raw cream, separated in the Iowa S ta te College milk
p la n t from a ilk delivered by some o f the above producers y ield ed th e
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-9 -
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1
I
organ ise,
m strlfe atfo n fjft, d e fe c tiv e d airy p ro d u cts.
Samples o f d e fe ctiv e
b u tte r are o ften received a t th e D airy In d u stry S ection o f th e Iowa
A g ricu ltu ral Ssperim eat S ta tio n fo r exam ination.
were examined f o r the presence o f £&. f r a e i.
S everal sash samples
Some o f the» had been
subjected to keeping q u a lity te s ts and c r itic is e d a s feeing ran c id {
o th ers had feoen c r itic is e d a s feeing p u trid , ta llo s y o r a s having some
o th e r d e fe c t.
included.
Several saap les o f d e fe c tiv e experim ental feu tter sas
D efective feu tter serum, p a ste u rise d cream and raw sk in a ilk
were a lso examined,
B esults o f th e exam inations a re p resen ted in fa b le 2 ,
Of th e 28
feu tter samples exaained, l6 o r 57.1 p e r cent y ielded £$,. fraugt.
She
organism m s is o la te d from th e sample o f ran cid feu tter serum, fro® feoth
samples o f ran cid p asteu rized cream and from th e sample e f raw skim m ilk
which had developed a Mey apple odor on holding In a re frig e ra to r fo r
some tim e.
S m s, 20 o r 62.5 p e r cent of 32 m ap les of d e fe ctiv e feu tter
and o th er d a iry products y ield ed gg. fneurit.
It was is o la te d from life
( 70.0 p e r cen t) o f 20 saap les e f d a iry products described a s ran c id o r
having a May apple odor.
Pp. f r a g i.
Presumably, th ese d e fe cts were caused fey
P u trid and tallo w y d e fe cts have not been a ttrib u te d te th is
organism and i t s presence in some samples o f feutter w ith those d e fe c ts
was probably o f secondary im portance.
While th e nan&er o f samples i s
not la rg e , th e high percentage of saap les y ie ld in g th e organism i s o f
sig n ific a n c e .
D lstrlfeatio n Ip d a iry -plant equipm ent.
p la n t equipment harboring
The p o s s ib ility o f d a iry
fra g i was stu d ied fey means o f swabs from
v ario u s equipm ent, e sp e c ia lly churns, used in cream eries and m ilk p le a ts .
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
Table 2 .
£&. f r a x l In d e fe ctiv e d a iry p rod u cts.
So. e f
D a iry
product
D e fe e t
Sampler
exam ined
Saaplas yielding
£&.*
Be.
l&t
A .
P e r eez
Bancld
i5
f
6©.©
P u trid
6
3
5 0 .Q
H
Tallowy
1
1
100.0
U
May apple
1
1
100.0
•
Be s p e c ific c ritic is m
5
2
* 0 .0
B a tte r serum
Bancld
1
1
1 0 0 .0
Cream (p a s t.)
Harold
2
2
10G.O
Skim a ilk (m « )
May apple
1
1
100.0
32
20
$2.5
B a th e r
»
Summary
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 12 •
Saaples e f t U i n atu re w ire obtained i s 39 Iowa, cream eries and in 1 leva
a il k p la n t.
As in d ic a te d in fa b le 3 , churns were th e main type e f equipment
examined but a few swabs were obtained f r e e p rin tin g and wrapping equip­
ment, r a ts and m iscellaneous p ie c e s.
Samples from 60 churns in 37
cream eries y ie ld ed Jhs. f in a l in only two
c &b s s
o r 3*3 P®r c e n t,
fh ree
saap les from p rin tin g o r wrapping ta b le s and one sample fro a a p r in te r
b e lt each y ie ld ed the organism .
o th e r sam ples.
I t was not is o la te d fro a any o f th e
In a l l , J s . fra g l was found in only 6 o r 5,8 p e r s e a t
o f 10b saap les examined.
More d e ta ile d exam inations o f a v a rie ty o f p ieces of equipment were
made in one crbaaery and one m ilk p la n t.
p ie e e s o f equipment in th e cre&nery.
Swabs were obtained from 2b
fhaee included n early a l l th e
p ie c e s o f equipment w ith which c re ss o r b u tte r comes in c o n ta ct.
However,
th e only swab y ie ld in g £ s. fra g l was one obtained from th e tcp o f th e
ta b le m which th e b u tte r was wrapped.
In the s i l k p lan t, 20 swabs were obtained from v ario u s p ieces o f
equipment and front v ario u s lo c a tio n s, such a s th e weigh room flo o r and
th e loading p la tf o r s .
fh e only swab in th is group y ie ld in g
fra g l
was th a t obtained fro n th e flo o r in fro n t o f th e weigh tan k .
Following the is o la tio n o f gg. f ra g l from two sanples o f ran cid
p a ste u rise d cream, a batch o f ersam was follow ed through the processing
o peratio n s in the p la n t where the d e fe ct occurred.
£*. f ra g l was
is o la te d from a sample o f th e raw cream in th e v at b efo re p a ste u risa tio n
was begun.
Hone of th e samples taken a f te r th e dream had reached th e
p a s te u ris a tio n tem perature (155°I'*) y ield ed the organism .
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-13-
fafela 3 * £&• tra^k i s d a iry p la n t equipaeat
t e . o f d afry p la n t#
fro a afcioh *a*pl*a
te . *f
piaaaa « f
war* attained
egoipaeat
te .
37
60
2
3*3
P rin tin g ta b le s
2
3
3
100.0
P rin te r b e lt
1
1
i
100.0
Friday p rin tin g equipment
1
3
0
0 .6
Wrappers
X
2
©
0 .0
?**•
2
12
0
0 .0
Puaps
1
£
0
0 .0
H isaellansaus equipment
2
21
0
0 .0
40
104
6
M
Equipment aacasisad
€&8B9til
Suaaaxy
Sasple# y ie ld in g
te . f te s i
Fer east
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
** lk «*
D istrib u tio n in d a iry p la n t w ater su p p lie s.
37 cream eries were examined.
Samples o f w ater fro *
She m ajo rity o f th ese saap les rep resen ted
tb s reg u la r w ater sapply e f th e creamery & ltheu#i f i r e were samples o f
wash w ater, one was w ater l a which parchment was soaking and two were
ferine used fo r th e same purpose.
Of th e 33 saaples fro a re g u la r w ater su p p lie s, 5 o r 15.2 p e r e ea t
y ield ed £§,.
(ta b le h ) .
However, th re e e f th ese saaplee were f r e m
one cream ery;
fra
l m s is o la te d fro a th e re g a la r w ater
w th e re fo re , P s. .W
WHgMM
supply o f 3 o r 10,6 p e r c en t e f the cream eries whose su p p lies were
examined.
One o f fiv e saagri.ee o f wash w ater y ield ed th e organism , a s
d id one sample taken from ferine used f o r soaking parchm ent.
In e l l ,
P s. fra g l was Iso la te d from J o r 17.1 p e r eeat o f the h i w ater samples
examined.
D istrib u tio n go f a r e s .
The presenoe o f g s . fra g l in m ilk su p p lies
from a considerable percentage o f farm s d e liv e rin g a ilk to the Iowa
S ta te College m ilk p la n t suggested the source of th is organism to fee on
th e farm s.
A ccordingly, th e farm s o f 6 o f th e Ik producers were v is ite d
and saap les obtained fo r exam ination.
Xa the absence o f d e fin ite inform ation regarding th e prevalence o r
d is trib u tio n o f £g. fra g l on farm s, th e f i r s t two v i s i t s were made t e
fa m e whose a ilk su p p lies had y ield ed the organ!as.
The r e s u lts o f th ese
v i s i t s as w ell a s of an a d d itio n a l v i s i t to each farm a re id e m in Sable
5-
At the f i r s t v ie i t to the fa m o f Producer 6 , 21 samples were ob­
ta in e d , 8 o f which y ield ed £&. fra g l.
I t was found in 13 o f 20 saap les
obtained on th e f i r s t v i s i t to th e farm o f Producer 8 .
The samples
which y ield ed the organism on both farm s were d i r t from the bam flo o r,
bedding, m ilk (not a s e p tic a lly drawn) and swabs from m ilk p a ils , m ilk
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-
15-
£*» A m * la w ater su p p lies o f d a iry p la n t* .
Saaplea
B agolar w ater eopply
lo . e f d a iry p la n ts
fro® which saaplea
«H * obtain**
X*.
Simple* y ie ld in g
Sas^le*
g e. fraud.
exaniaed Ho.
P ar e e a t
30
33
Wash, w ater
5
5
l a t e r f e r soaking p&rehsent
1
1
B rine fo r soaking parobaent
1
f
37
*1
Snwaaxy
*
15.2
20.0
©
0 .0
50.0
7
o f th ese saap les ease tr m mm creaao ry .
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
17.1
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sto o ls and led ges,
In a d d itio n , on th e form er farm , i t was found in
th e g ra in m ixture and s o il su rface e a te r fro a th e barnyard, and on th e
l a t t e r f a r a , I t mss is o la te d from w ater fro a a stock w atering tan k , d i r t
from th e barnyard, hey end swabs from c o ats o f
cow b,
a
stanchion and
a
f lo o r sc rap e r handle.
th e l a t e r v i s i t to each o f th e farm s re s u lte d la th e is o la tio n o f
g». f r a a i from f o f 13 samples obtained a t th e farm o f Producer 6 and
fro a € o f IS samples obtained a t th e farm o f Producer 8 .
fhe saap les
which y ield ed th e organism on both farm s a t th e second v i s i t mere
bedding and swabs from th e bam flo o r, a stanchion and d u e ty le d g e s.
On th e fa n s o f Producer 6, sample# o f w ater from a stock m etering tan k ,
barnyard d i r t , hay, th e co at o f a corn and a m ilk sto o l a lso y ield ed
iS,* fra**- w hile on th e farm- o f Producer 8, e th e r saapdee y ie ld in g i t
were g ra in and a sem p er handle.
P s. f r a g l was Iso la te d fro a 36 (5 ^-5 p e r cen t) o f 66 samples ob­
ta in e d on th e te e farm s.
A ll th e s in swabs from f le e r s and ledges
y ie ld e d i t , a s d id a l l th e fo u r saap les o f bedding, and i t was e ls e
is o la te d from &t.5 p e r can t o f th e saap les fre n m iscellaneous barm equip­
m ent, 57.1 p e r o ast o f th e samples o f d i r t , 5© p e r e e a t o f th e samples o f
fe e d , h2.9 p e r o a st o f th e saap les o f w ater, h i .7 p e r se n t o f th e samples
fro a th e anim als them selves and 22.2 p e r e e a t o f th e sample# from m ilking
u te n s ils .
fiv e v i s i t s were made to farm s from whose m ilk su p p lies
had n o t been is o la te d .
B asalts o f th e exam ination o f sa ap le s obtained
m th ese v i s i t s is given l a f&bld 6,
o f Producer 1 .
fra d
fe e v i s i ts were made to th e farm s
On th e f i r s t v i s i t 2k sam ples were o b tain ed , 12 of Whieh
y ie ld ed th e organism.
On a second v i s i t 5 o f 12 samples were found to
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-18-
o«
h h o j»
n w o jtd
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R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
i
h arb o r i t .
S a b le s fro * th i* fa r* th a t y ield ed £&. f m g i included w ater
from stock w atering ta sk s and from the b am flo o r* barnyard d i r t , bay,
bedding and swabs from stan ch io n s, flo o rs and manure.
On sach o f th e o th e r th re e farm s £&. f r a e l n » is o la te d from samples
e f barnyard d i r t and bedding.
On two o f th e th re e farme w ater from sto ck
w atering tan k s and swabe from stan chio n s, m ilk s to o ls and ledges a lso
y ie ld ed i t .
8sag>lee from which i t was is o la te d ea w ily one of th e th re e
farm* included w ater fro a s o il su rfa ce , hay, and swabs from a scrap er
handle and a dusty ledge.
She somber o f sample* examined from th e farm s
e f Producer 3 , 5 and lk was 11, 12 and 12 re sp e c tiv e ly and th e p e r e e a t
o f samples fro a th ese farm s y ie ld in g £ s . fraud, was 5^*5, 58*3 end k l . |
p e r e e a t.
In a l l , 71 samples obtained on fo u r farm s, from th e m ilk su p p lies
o f which gs,. fraud had n o t been is o la te d , y ield ed i t in 35 {**3*3 P9? cent)
o f th e In sta n ce s.
A summary of th e r e s u lts of eammination o f samples obtained on a l l
nine v i s i ts to the six farm s i s shown in fa b le J.
Of th e m ateria ls and
etjuipasni sampled, bedding, flo o rs and led g es, w ater, d i r t , m iscellaneous
barn equipm ent, fe e d , th e cows them selves end a ilk handling u te n s ils
harbored J&. fra g l in th e o rd er named.
Bedding, w ater fro a sto ck w atering
tan k s, hay, g en eral barn equipment and d ust and d i r t on ledges and flo o rs
in th e bam a s w ell a s o u tsid e ware e sp e c ia lly im portant a s sources o f
th is organism.
M ilking u te a s il* were r e la tiv e ly fre e o f i t .
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 20 -
TaMls 7 .
£&. freuti ia samples fw « s ix Zourn farm*. SaHsazy
and S T
Mo.
Sauapl#s y is ld SO.
!&P* o f sasp le
S asp iss la g £&. f r a a i
Saaples
exaained Me. F or e sn t tx a s is a l
o f T ables 5
S a b le s y ie ld la g £ g . fra g l
Vo. For cen t
l a t e r fro a
**&
i l a d : tank
Mara flo o r
cooling ta s k
s e ll su rface
D irt fro a
Marayard
bam flo o r
Feed
hay
g ra ta
sila g e
u
i
i
3
6
8
1
6
2
0 .0
J2 .7
100.0
0 .0
66.7
19
11
57.9
12
2
€
2
50.0
100.0
14
8
§7.1
8
5
17
8
47.1
3
8
I
0
3
62.5
37*5
0 .0
9
8
88.9
9
8
88.9
9
6
3
2
33.3
33-3
15
5
33.3
k
%
0 .0
50.0
0 .0
0 .0
0 .0
14
2
1 4 .|
29
16
55.2
Bedding
d a la s is
coat
■ till
M ilking u te n s ils
■11
...W ng
- *3 machine
'■ - - p a ils
s tra in e rs
o oo ler
caas
Mi so. Mara equipment
stanchions
a ilk s to o ls
so rap ers, shovels, e tc .
2
1
3
6
2
0
0
6
15
9
5
8
5
3
5 3 -|
P 'f
60.0
flo o rs
lodges
16
9
6
€
60.0
66.7
15
12
63.2
Manure
1
1
100.0
1
1
106.0
Somany
13?
n
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
51.9
- a D istribution of
fv**i to Earntucky
D istrib u tio n to normal d airy p ro d u cts.
th re e s e ts o f saap les from
a lto d e liv ere d by patron* to a Lexington, Kentucky, s i l k p la n t wax**
eaaaiaed f o r the presence o f £ j , f r a a i,
and th e o th er two mot* to Beeemhcr.
One s o t was obtotoed to Jon*
to o sample* rep resen ted too m ilk
sap p lies o f 26 farm* to th re e Kentucky co u n ties.
Six farm s su pplied one
sample e&ch, nine farm s to o samples each and. eleven farm* th ree sample*
a a to .
to a l l , 57 s a b le s wore examined.
Heme o f the seventeen samples obtained to Jto e y ield ed th o organism ,
a* shown in $abl* S.
ta in e d to Seeember.
I t woe found, however, to l€ o f th e *10 samples ob­
Inclu d in g summer and w in ter sam ple*,
frsurf. was
is o la te d from 16 o r 86.1 p e r cent o f to* 57 sasp lea examined.
Of tho 17 farm s rep resen ted by to e too* sam ples, only 12 were
represen ted by to* saap les obtained to December.
Samples from nine o f
th ese 12 d a ir ie s y ield ed Pg. f r a a l. w hile saaplee fro® only th re e d id
n o t.
fhe organism was is o la te d from samples rep re se n tin g m ilk su p p lies
from 13 of th e 26 farm s.
D istrib u tio n to d e fe ctiv e d a iry p ro d u cts,
tody a few saaplee o f
d efectiv e d a iry products were a v a ila b le f o r exam ination a t Lexington.
S ato o f two saaplee o f skim m ilk, which developed & May apple odor to
to e d i v e r s i t y of Kentucky D airy Department re frig e ra to r* y ield ed to e
organism.
I t was mot is o la te d from seven samples o f b a tte r follow ing
keeping q u a lity te s t* a t a Lexington creamery* non* o f th ese camples
were c r itic is e d a s being ra n c id o r having a May apple odor.
D is trib u tion to d a iry u la n t eqoinment.
A swab from a ( t e n to a
Lexington d a iry p la n t d id n o t y ie ld £&. f m d .
fern swabs obtained to
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 22 -
Sabla 2.
£&•
to a llk deliirored to a Lexington, Kentucky,
» ilk p la n t.
Bato o f d e liv ery
13-5-39 lfc-2i~3$
Producer
Sg-i. .0
23«
fto to
2%
«A
3"?
a#
2*
2!
Seaplo*
Ho, Sample*
y ie ld in g
•m to u A .
flU X B M l
3
1
3
1
a
3
2
2
1
1
1
0
0
0
0
0
1
6
1
1
2
+
*
♦
+
♦
♦
*■
♦
+
am
am
33
>i Ho.
y ie ld -
i» £ z&-£mek
I t.
For « eat
1J
ao
20
6
8
ito.o
8
HQ.Q
0 .0
1
3
3
3
2
2
3
1
3
3
2
3
1
1
2
2
3
0
2
0
1
0
2
1
1
a
0
0
0
1
0
57
16
28.1
•S aaplee n o t y ie ld in g go. * » « * deeignated
osarples y ie ld in g go. f r a * t designated +.
** Saaplee were a lto a tta in e d a t fa ra o f th ese producer*.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 23Jone and Beaesber from th e receiv in g room flo o r o f th e ease p la n t
y ie ld ed th e o rgan ixs.
I t vats a lso is o la te d tv m a s e a t obtained la
December from th e flo o r o f a covered tra c k d e liv e rin g m ilk to th is
p la n t, w hile a s t e ll a r sample obtained in Jane fa ile d to y ie ld i t .
B l& lrlhiatloa in e a te r ^ ap p lies.
p la n ts in Kentucky were n o t examined.
Saaplee o f w ater fro® d e ify
A sample o f pond e a te r reeeiv ed
a t tins Kentucky A g ricu ltu ral Experiment S ta tio n from a fajrmer who
questioned I t s f itn e s s f o r c a ttle to d rin k was found to co ntain
g s, f r a g l.
Another e a te r sanple y ie ld in g th e organism m s taken from
a p o rtab le ic e box in whieh severed b o ttle s o f s i l k end cream were
shipped from L o u isv ille to Lexington.
Water co ntain ing seme m ilk
s o lid s , th a t remained in th is shipping box a f te r removal o f s i l k and
c ro ss samples developed a Hay apple odori g e . freer! wee re a d ily
Iso la te d from i t .
3& fM a*U aft, m Uzm *
fo rty n in e samples rep re se n tin g w ater,
d i r t , feed , m ieoelleaeotts barn equipm ent, flo o rs and ledges were ob­
ta in e d os te n Kentucky fa re s d arin g th e summer o f 1939 (fa b le 9 ) .
two o f th ese samples ( b . l p e r cent) y ield ed
rep resen ted feed ; one g rain and one hay.
f r a a i.
Only
lo th saap les
Most o f the saap les were ob­
tain ed i s Ju ly and Jag o st when th e w eather was hot and d ry.
M usty nine saaples were obtained from nine Kentucky farm s in
December.
Of th ese 37 (J l.b p er cen t) y ield ed £ e. free*, (fe& le 1 0 ).
I t wee is o la te d from § o f 12 w ater samples obtained in December, as
eomparsd w ith none o f 13 obtained in th e summer.
SI even o f Ik d i r t
saap les obtained in December y ield ed th e organism , a s c o n tra sted w ith
none o f 12 c o lle c te d in th e summer,
th e re was l i t t l e d iffe re n c e between
th e percentages o f feed saap les y ie ld in g gg. f ra x i in December and in
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 2fc -
fa il* f ,
£&. f a t t l l a M aple* f r o s te n Keateokgr farts* during te a
m i f wr se aso n .
fjrpa o f aaapl*
fala p from
*%0<ek e a te rin g ta s k
Artot&ag cap
cooling
'weak issk
i a r a f lo o r
r ia s e fra * u te n s ils
M r! fro a
i &iiiP6hP&
i a r n flo o r
PftS*
g ra ta
hap
K lee. equipment
stanchions
d rlak tag cop
U ser
ledge
SawaTy
So.
So. Saaples
So,
Saaplee y ie ld in g
Saeples
y ie ld in g
Samples
g£. fra g l
examined gg. f r a f l
exaainsA So.
P ar coat
$
2
1
1
1
1
1
0
0
0
0
0
0
0
*3
©
0 .0
10
2
0
0
12
0
0 ,0
12
T
1
1
If
2
10.2
2
1
©
©
3
0
0 .0
1
1
©
©
2
©
0.0
2
*1.1
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 25 -
faBle 10, gfc. fragj in saaples fron nine Kentucky fares daring tfce
winter season.
type e f
le .
la , Saaples
X»;
Saaplee yielding
Saacplee
essntaed Jg. fn^al
m e lw t
Eaaples
tragi
So.
Water trm
. stock watering tank
dria&iag eqp
eroSfe
cistern
ced in g ta d :
Bern JX esr
rin se tr m u ten eils
Mart tr m
Barnyard
ta rn fle e r
nasgere
g aiter
Peed
grain
Bay
allege
1
1
1
2
1
1
P ercent
f
X
t
1
0
0
X
12
9
T9.®
X*
XX
7*.6
9
2
2
X
9
1
X
0
«
9
1
1
2
0
IE
3
16. T
€
3
6
3
5®.©
©
0
©
©
©
©
9
©
0.0
B ottler
B ottle
Misc. Barn e^dpoent
stanchions
edllE stools
scrapers
mangers
wash tank {dry}
3
1
2
I
X
X
8
1
%
2
X
X
2
X
©
0
a
*
If.®
Gelling
H ears
hedges
CoBweBs
X
6
10
2
©
*
3
©
if
T
36.6
99
3T
37>
Bedding
Uteneil*
s ilk p a iit
stra in e r
cooler
Saaoarjr
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
suamer.
presence o f th is organism m s demonstrated la three o f tho
s ix saaples o f bedding obtained daring Deoeaberj samples o f bedding were
not obtained daring th® summer season.
lik ew ise, s o c le s were so t
j
obtained in sooner from u te n sils for handling s ilk , hot o f nine samples
obtained from u te n sils la December none y ield ed the organism.
It m s
Isolated from foor o f 21 samples obtained from m iscellaneous d a iiy barn
equipment, such m stanchions,, m ilk stools* s t a l e r sad broom handles,
e t a .. in December, bat" not fro® ssay o f three obtained from sash equipment
in the summer. fee saaplee obtained f w i flo o rs and ledges in summer did
net y ie ld £a. f^a«c w hile i t m s found in seven o f nineteen sash saarplos
obtained in December.
Saaples were obtained from only one o f the fam e in both summer and
w inter.
Among fourteen samples obtained from th is fafm in Jane, only one
(grain) yield ed fa,. fmsad w hile i t m
obtained ia December,
found in seven o f eleven sarplee
The w inter saaples from ebicfe i t was iso la te d were
water <2 saap lee)* d ir t (2 saap les}, bedding and swabs fro® the flo o r and
from a dusty led ge.
Approximately the same m aterials and equipment were
sampled on each v i s i t .
Saaples were obtained in December a t the farms o f two producers
lis t e d In f&ble S.
Six o f ten samples from the farm o f Producer 239
yield ed g£. fragl .
These ineluded samples o f barnyard d ir t, hay, water
from stock tank, and swabs from a dusty led ge, bam flo o r and m ilk sto o ls.
The saaples in which i t was not found included feed , sila g e and swabs
free a stanchion and a sem per handle.
The sin g le sample o f milk obtained
m delivery to the m ilk plant yielded th e organism.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-27Three o f fifte e n saaplee obtained a t the fa r s of Producer 2^7
y ield ed g£. fr a g i.
These throe saaples included d irt froa the barnyard
and from the bam flo o r and rater from a c iste r n ,
Samples in which i t
was not found included grain , d irt t r m a manger, hay (gn m & ), water
fm
a coolin g tank and sw&bs from stanchion, cobwebs, dusty window
led ge, B ilk sto o l, s ilk p a il, stra in er, s ilk can and dry wash tank.
A
s ilk sample froa th is fa r s obtained on d elivery a t the s i l k plant in
June did net y ie ld the organism, w hile i t was iso la ted from a second
sample obtained a t the s i l k plant in December.
▲ sussary o f the r e su lts o f examination o f a l l saaples obtained on
Kentucky farms i s given in fa b le 11.
per cent o f the ihS saaples examined.
g s . fr a g i was found in 39 or g$.h
Bedding, d ir t, water, flo o rs and
led g es, m iscellaneous barn equipment and feed were found to bather the
organiss in the order given.
One h a lf to one th ird o f the saaples in
the f ir s t four groups y ield ed £*. fr a g i. but i t was not found in any
sample fro® s i l k handling u te n s ils .
These r e su lts are very sim ilar to those already reported fo r Xowa
although the percentages o f Kentucky saaplee y ield in g g s . fra g i are some­
what lower.
A p a rtia l explanation fo r th is may be because approximately
was third o f the Kentucky saaples were obtained i s the su sser season whoa
££• iss &
was found only very infrequently w hile a l l the lews fan s saaplee
were obtained during the w in te r and spring.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
— 28
$aMt* 11, 2a, fraarl in saaples tr&m 10 Kentuelqr tv tm Spring both
etssater and eia-ter, Swwary e f feble* 9 and 10*
80,
*ype -af sample
Water from
stock caterin g tanks
drinking cups
ereek
cistern
tap
cooling tank
sash tank
tarn flo o r
rin se from u te n sils
U r t fro«
barnyard
tern flo o r
usanger
f it t e r
Peed
grain
bay
s i lag#
tedding
Milking u te n sils
s ilk p a ils
strainer
cooler
can
b o ttle r
b o ttle
M iss, tern equipment
stanchion
railk sto o l
scraper, broom and
shovel handles
nsnger
drinking cop
wash tank (dry)
C eiling
flo o r
Ledge
Coteebs
Snsraary
Samples
examined
11
3
1
1
1
3
1
2
2
1f
u
2
1
80, Steadies
yielding
ffiagi
86,
E s p ie s y ie ld in g
Saaplee
£&, frsg l
examined 8»,
Per cent
5
1
1
1
0
0
0
0
1
25
9
36.0
9
i
i
26
11
H2.3
0
20
16
1
0
3
37
5
13.5
B
3
6
3
9&.Q
3
l
2
1
1
1
0
0
0
0
0
0
9
0
0.0
10
$
k
1
2
1
?b
H
16.7
2
I
0
0
1
0
1
0
21
7
33*3
ih s
39
26, h
7
11
2
2
k
3
0
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-
23
Distribution of £«. fragi In the United Stains
In order that the d istrib u tio n o f £ e . fra g i throughout the United
S tates could be studied, a request fo r a sample o f barnyard n i l nas sent
to the Dairy Department a t each State A gricultural College in the United
S ta tes.
Barnyard s e l l sa s requested because o f the frequency with which
Pa. fra g i had been iso la ted froa I t and the convenience o f c o lle c tio n and
shipment.
fh e requests were made in the la te winter or early spring
souths as th at seemed the most probable time for the easy detection o f
the organism.
Samples were received from 33 s ta te s, representing a l l
sectio n s o f the country.
Sxamination o f the samples revealed the presence o f {&. fra g i in
those from 23 sta te s or 69.? per cent (fa b le 12).
Saaples from 10 sta te s
fa ile d to y ie ld the organism; these samples were re-examined, but with
the same r e su lts.
By grouping the sta te s in to an eastern and a western d iv isio n , an
in terestin g comparison can be made (fa b le 1 J ).
In the eastern sectio n
are Included the sta te s from Minnesota to Louisiana and those eastward;
in the western sectio n , those free North Dakota to fex&s and westward.
She eastern sectio n ineluded 31 s ta te s, saaples from 20 o f which were
examined.
She presence o f f&. fra g i was demonstrated in saaples from 18
( 90.0 per cent) e f these 20 sta te s,
f h is in d icates & wide d istrib u tion
o f the organism throughout the eastern h a lf o f the United S ta tes.
In
the western sectio n are ineluded 17 s ta te s , 13 o f shieh furnished saaples
fo r examination.
Samples from only 3 ( 38.3 per sen t) o f these 13 western
sta te s yield ed g s. fr a g i.
While a sin g le sample from one p articu lar spot
in a sta te i s by no means representative o f the en tire s ta te , n everth eless,
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 35 -
fa h le 12.
£&. fra g i l a barnyard s e ll froa m rie n s sta te s la the
United S ta tes.
O rigin o f samples
y ield in g
Um I
Alahana
Arkansas
Delaware
Georgia
TWHnWft
lo se
Xmtwsksr
Louisiana
Maryland
Minnesota
Missouri
Montana
Sew Jersey
Korth Carolina
Worth Dakota
Ohio
Pennsylvania
South Carolina
ferae
V irginia
Washington
West V irginia
i
!
t
t
t
t
t
i
t
I
t
e•
t
:
Origin o f easplss
net y ield in g
fr a g i
C alifornia
Colorado
Florida
Idaho
Kansas
M assachusetts
Yebraska
Yeir Mexico
South Dakota
Ironing
e
•
:
r
t
i
•e
:
:
•
*
:
t
:
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 3 1 -
: 3 R S? ®
gi
Z
$ta
t
el % 5? « &
35
to•
it
%
4*
%
1
-I*
\
“S
el & a s ao
35
•
Im
«
3
| a
«Zd <
«3 *52
■£! *
ft
o
t
3
3
•
!l-t
*
«
®
«
*«
4*
-$3 3
1«
«M
«
*«
ft
1
mm
4»
S
»
&
*
*
I
£
<£
•
»
*
s
•
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-32tha m ajority of saaples (8 o f 13) from t h is se atio n of tho country
which did not y ie ld £ s. f r a g i probably In d ica te s th a t th e organism i s
n e t ae prev alen t in th e w estern s ta te s a s in th e e aste rn .
A summary of r e m i t s of examinations o f a l l saaples obtained on
fa re s i s given in fa b le lb .
y ield ed J s . f r a g i.
Of 316 samples examined, 131 ( b l.5 p e r cent)
I t was Iso la te d fro * s e re than h a lf th e saaples from
s ta te s oth er than Kentucky.
However about one th ir d o f the Kentucky
saaples were obtained daring the season when the r e la tiv e ly high tempera­
tu re s a re unfavorable to th e wide d is trib u tio n o f the organism on farmsj
i f these % sasples are exeluded, hS.3 p e r cent o f th e saaples obtained
on farms y ield ed th e organism,
f h is summary
o f th e fames a s a scores o f £ s , fra g i in m ilk.
Id e n tific a tio n o f c u ltu re s o f £ s . fram i
Zn order to e s ta b lis h th e presence o f £ s . fTagl on a p la te , c u ltu re s
were picked and studied in some d e ta il.
The f i r s t c r ite r io n used in
p icking colonies in to litm u s milk from p la te s containing n i l s blue
s u lf a te and b a tte r f a t emulsion was th e a b i l i t y o f the organism to
hydrolyse fa t*
f h is was in d icated by the presence of f a t d*qplets
sta in e d blue in stea d o f pink in th e v ic in ity of the colony.
As many
o th e r organisms a re capable of hydrolysing f a t , the appearance o f th e
colony in question i s Im portant.
In o rd er to d if f e r e n tia te co lo n ies,
they were examined fro * th e bottom o f th e p la te w ith th e a id o f a hand
le n s ,
sy p ic al surface colonies of £jb. f r a g i u su a lly were round and
opaque and from 2 to 5 mm. in diam eter.
A fter 2 to 3 days incubation
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
- 33 -
£&• ***** i& ail
oa*ploo ofelatnoa, on farns.
Swaajy
of f&m*s 7. 11 and 13
te\
i
i!
•# M -«*
*ll
i.ip
it &
l<\
&
i5
w
Ol
M
£
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s
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M
s
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it
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m
*"*
OH *0-
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r*»
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&
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s
its
r*+
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ill
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si
p
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+»•»
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l*v O
*
Jt
m 4
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m
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3
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a !< i i
&
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a
II
p
£
■0*»
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s
<S*«
£
a
i
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i5
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-
-
they appeared p ra ctica lly chalk w hite. w hile colonies o f many other
organisms had taken up enough dye to make them appear b lu ish .
With
some experience* the eolon ies o f £ b. fra g i could he distinguished quite
accurately.
Warn Pe. fra g i mas picked into litm us milk and incubated 2 to 3
days a t room t separators, a Hay apple odor became n oticeab le.
I f a drop
o f eth yl alcoh ol was added to the tubed milk before inocu latin g, the
aroma mas produced a fte r only 1 to 2 days.
U sually, a fte r 3 to ^ days,
a slig h tly pink (acid ) ring appeared a t the surface o f the milk next te
the w all of the tube,
In most cu ltu res, th is was follow ed in a few days
by coagulation of the milk with same reduction o f the litm us.
However,
in some c a ses, the milk did not coagulate, even a fte r incubating 2 to 3
weeks.
After r e fla tin g fo r p u rifica tio n on beef infusion or beef extract
agar containing M ile blue su lfa te and fa t emulsion, typ ical colon ies
were transferred to agar slopes fo r use in inoculating various media.
M o tility was determined by removing to a microscope slid e a loopful
o f a 1 day old culture from a tube o f neutral sugar medium and examining
the organisms under the high dry len s o f the microscope.
cultures id e n tifie d as £ s . fra g i were m otile.
A ll o f the
Smears fo r Sram sta in in g
were also made froa 1 day old cultures in a liq u id medium. AH o f the
cultures o f th is organism were Sram negative.
They were a ll rod shaped,
although u su ally short and medium length rods were found in each culture.
Most o f the organisms were sin g le or in p a irs, although la some cultures
chains o f several c e lls occurred.
Mo spores were observed in any o f the
cu ltu res.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
ffea a b ility « f d ifferen t cultures o f jgs. tr * x l t i liq u efy g e la tin
tw le d .
Host o f then did so with considerable rap id ity , mom slow ly oad a
fin seeaed unable to do so .
When liq u efa ctio n occurred, i t eonasnly was
o f str a tifo r a or craterifona type.
Action of cultures designated as £&. frauel o s the indicator media mood
was n aif ora except in tho case o f arabinose.
Whey a l l produced acid rapidly
in dextrose, hot somewhat slower in galactose, t o r mwm daye usually
elapsing before the galsetose beeaae acid .
®he m ajority of th e cultures
produced acid i s arsbinese although w ith w ay cultures nearly | dajr»
•lapsed before the anabineee became d e fin ite ly aeid .
Warn o f the cultures
produced acid in la c to se , levu lose or glyp erol.
Sane fa c to rs a ffe c tin g arean production
Several Investigators hare considered erase protection by b acteria and
hare attearpted to id en tify the oesyetmds reapeselhle fo r various a m i .
I s studying e fr u it ea ter producing organise, Bacillus pgagpalleas.
which he had iso la ted , Maassen (1*0 analysed the daeospositlon prcdaets
formed by i t .
Aaoag these he found e v a ler ic aoid e ster which sm elled
lik e the anyl eater o f v a le r ic a eid .
presence o f th ie compound.
Be attrib uted the areas te tin
In 1903, Orlaa (5) reported the iso la tio n o f
B solU ue aroaatlcas la a tle which produced a strong fr u ity aroma in m ilk.
He thought the areas was due to eth yl la e ta te foraed in the B ilk .
Seek
(1) studied the aromatic substance formed lay Mioroooooas e ste r ifle a n s and
found I t to he insoluhle in aloohol hut solu ble in eth er, chloroform and
carbon d isu lfid e .
OaellattsldL ( 15) reported that the addition o f leu cin e
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-3 6
-
to meat peptone agar stim ulated aroma production by Bacterica estero arom aticm . esp ecia lly in the presence o f a l i t t l e eth yl alcoh ol.
Be
soted so stim ulation from the addition o f propyl or amyl alcoh ols,
opinion leucine se e s p lit out o f protein and further
In Ids
to form
iso -v a le r ic acid and iso-am yl a lco h o l, vshich united to fora iso -v a le r io iso-a a yl e ste r .
He thought the presence o f eth yl alcohol merely In ten si­
fie d the aroma, as I t had he an found to do in a r tif ic ia l "apple esters" .
With most o f the aroma producing organisms described in the lite r a tu r e ,
the pleasant arose disappeared and mas replaced by an aanoalaoal or cheesy
odor,
Gmelianskl attrib uted the former to deaminization in the leu cin e
breakdown and the la tte r to the presence o f iso -v a le r ic aeld whioh had
been reported to be one o f the causes o f the odor o f old cheese.
In the studies m £ s . fra g i several media mere investigated from the
standpoint o f th eir e ffe c t on production o f the May apple odor. Butrient
broth <0.3 per cent
beef extract - 0 .5 per oent peptone), Uschlnsky's
medium, 0 .5 per cent beef extract solu tion and 1 per cent solu tion s o f
peptone, g e la tin , dextrose and la cto se were lnoaalatsd and examined fo r
-i •'
growth and aroma production a fte r 5 days* {toed growth occurred in the
nutrient proth, beef extract and peptone solu tion s and there was s lig h t
growth in g e la tin .
A strong Hay apple aroma was produced la peptone
so lu tio n , a f a ir arena in nutrient broth end g e la tin and a slig h t aroma
in b eef ex tra ct.
Bo growth or arena production occurred in Uschia sky’s
medium, dextrose or la cto se so lu tio n s.
When 1 ad. o f the peptone
solu tion was added to 5® ml. o f the la cto se solu tion considerable aroma
was produced.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
~ 37 For aroma production, Q aelianski (15) recommended a medium containing*
tap water 1000 a l. , peptone 5 to 10 g n ., potassium phosphate 1 ggu, agar
20 go. and a few drops o f eth yl alcoh ol,
Where plate* ware poured w ith th is
medium and 0 .1 a l. of a suspension of gs* fra g i d istrib u ted over the surface
with the aid o f a s te r ile bent glass rod* abundant growth and & wary strong
aroma production occurred} the aroma remained ty p ica l for several days*
A medium employed fo r cu ltiv a tio n o f propionic acid bacteria was ale*
need.
It con sisted of? d is tille d water 1000 ml,* sodium la c ta te 10 g * .,
yeast extract 7*5 £».* peptone 20 gm. and washed agar 12 go.
On m i* medium
growth was abundant in 2h hours but only & slig h t May apple aroma was pro­
duced, together with a slig h t aamoniaeal odor.
Growth was a lso abundant and
a very strong May apple aroma was produced in 2% hour* on a p la te to which
0 ,1 ad. of Id per cent eth yl alcohol had been added before pouring the agar.
Xn both these oases, however, a fte r
hour* the pleasant aroma hod been
e n tire ly replaced by a strong sismoniacal odor.
fh rss se r ie s o f s ix p late* each were used in studying the e ffe c t m
aroma production o f peptone alone and in combination with glycerol or bu tterfat*
Each o f s ix plate* in the f ir s t se r ie s was poured w ith 10 ml. o f 2 per
cent agar a fte r adding the follow ing amounts o f 5 per cent peptone solution*
none* 0 .2 , 0 ,b , 0 ,6 , 0 ,S and 1,0 ml,
Xn a second aeries glycerol (1 ml. of
5 per cent solu tion ) was added to each o f the p la tes containing peptone and
agar as above, and in a third se r ie s 0*5 ml. of a 5 per oast emulsion o f
bu tterfat In 0 .5 per ea st agar was added.
A fter pouring, a ll o f the plate*
were inoculated by spreading a suspension o f gg. fr a g i over the surface o f
the agar.
She p late* containing the larger amounts of peptone showed more
abundant growth than did those w ith the sm aller amounts in a ll three
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
eerie u .'' "Uw « » t abundant growth and aroma production occurred in the
se r ie s with added glycerol and the lea st in the se rie s w ith the peptone
alone*
Another se r ie s o f p la te s m e prepared to study the e ffe c t on aroaa
production o f glycerol e ster s o f several fa tty acid s.
The e ste r s used se re
tri-p rop ion ia , bri-b atyrin , tr i-n -v a le r in , t r i - i so -v a lerin , brt-®apnel»,
tr i-h e p ty lin , tri-o a p ry lin end tri-ca p rin ,
solut Ions or suspeae ions wore u c e i.
One tenth per cent ^queens
©a© per cent eth yl alcohol was added
to each solu tion or suspension in the hope that the e ster s containing the
longer chain a d d s would d isso lv e b etterj however, a ll except the t r iproplonin and tri-b u tyrin mixtures rem ised cloudy to m ilky.
One ml. o f
a solu tion or suspension was added to d ifferen t p la tes before adding beef
infusion agar.
A drop o f a suspension o f £$» fra g i was then spread on
the surface o f the s o lid ifie d agar.
A fter 2 days a fa ir amount of aroma
had been produced on each o f the p la tes containing the e ste r s, the
greatest amount being produced on tr i-h e p ty lin .
any ester had a fa in t aroma.
The check p la te without
The most typ ica l aroaa was produced on
p la te s containing tri-p rop ion ln , tri-n -V a leria , tri-h ep ty lin and i r i caprin.
The p la te eoataiairg tri-b a ty ri* gave a a lig h t butyric acid odor,
that with bri-eaprylin an odor resembling that o f pine and those with
tr i-iso -v a le r in and tri-oap roin were not ty p ica l.
Jkrosm production was also tested on Giaelianski*» agar without alcohol
end with m ethyl, eth y l, propyl, butyl end amyl alcoh ols.
After 2 days
growth on p la te s, the most conspicuous and most ty p ica l aroma had been
produced on agar containing ethyl a lco h ol.
A fe in t aroma had been pro­
duced on the control with no alcohol hut none m agar w ith methyl alcoh ol.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
fh # aroaa produced on &ga r w ith propyl alcohol was not ty p ic a l and ra th e r
f a i n t , while th e odors o f the buty l and amyl alcohols were so stro n g they
powered -up any aroaa th a t may haw® been produced by £s« f r a g i.
th ese
r e s u lts confirm those of Omelianeki who found th a t alcohols e th e r than
e th y l alcohol had l i t t l e e ffe c t in in creasin g aroaa production,
The ease
r e s u lts were obtained when 1 , 2 , 3 or h drops of the above alcohols were
added to c u ltu re s o f jgw. fra g i in tubes o f litm us s i l k .
More aroma was pro­
duced and i t was produced more quickly when 1 to h drops of ethyl alcohol
was added p e r tube than in th e check tube w ith no alcohol added o r when
o th e r alcohols were used,
When 0,05 p e r cent of methyl, e th y l, p ro p y l, buty l and amyl alcohols
were added to 0,1 p e r cent peptone so lu tio n s inoculated with P s. f r a g i . a
ty p ic a l aroma was produced only in the so lu tio n containing e th y l alc o h o l.
When a 0 .1 p e r cant so lu tio n o f la c to se was used with a sim ila r s e rie s of
so lu tio n s, th e r e s u lts were the same.
Various combinations o f peptone, eth yl alcoh ol, glycerol and la cto se
gave the greatest aroma production in solu tion s containing the a lcoh o l, and
there seamed to be no advantage in including glycerol or lacto se or both
over peptone and ethyl alcohol alone.
The a d d itio n of 0 .2 p e r cent of e ith e r leucine or e th y l alcohol to
0 .5 p e r cent peptone so lu tio n increased the aroma produced by £&. f r a g i .
and when both were added i t was Increased s t i l l more.
A s lig h t a ty p ic a l
aroma was produced by £&. fra g i when grown in 0.2 p e r cent leucine alone
o r in 0 .2 p e r cent leucine p lu s 0 .2 p e r cent eth y l a lco h o l.
The a d d itio n
o f 0 .2 per cent leucine to skim milk re s u lte d in the production of s lig h tly
more aroma than did th e a d d itio n of 0 .2 p e r cent ethyl alco h o l.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
When both
leu cin e and. alcohol sere added, the aroma produced sensed about th e same as
shea leucine alone was added.
fhe a d d itio n of 0 .1 p e r cent tryptophane to a 0 .1 p e r cent peptone
so lu tio n did not increase aroma production.
However, when 0,2 p e r cent
e th y l alcohol was added to th e above m ixture, considerably wore aroma was
produced.
Some aroaa was produced by th e organism i n a so lu tio n containing
3 ,2 p e r cent tryptophane and 0,2 p e r cent alaohol b a t not a s finch a s in aae
containing 0 .2 p e r eent peptone and 0 ,2 p e r can t alco h o l.
O lyceeel• alanine o r caleium a c e ta te , to g eth er w ith peptone so lu tio n ,
when added to p la te s before pouring w ith 2 p e r cent a g ar, did not stim ulate
arena production above th a t produced when peptone alone was used.
Use of
e th y l alcohol in t h i s Banner w ith peptone re s u lte d in a marked in crease in
aroaa.
When glycoool o r alan in e was used w ith alcohol but without peptone,
■ore arose m s produced than when combined w ith peptone but le s s than w ith
the peptone-alcohol combination.
Apparently | e . f r a g i th riv e s b est near the surface of s i l k as changes
produced by i t in litm us m ilk begin a t the su rfa ce .
At f i r s t the m ilk
u su a lly becomes s lig h tly a lk a lin e a f t e r which an acid rin g i s formed a t the
su rface next to the w all o f th e tube.
In order to t e s t th e e ffe c t of volufis-
surfaoe re la tio n sh ip s, the contents of fo u r tubes of litm us milk were emptied
in to fo u r s t e r i l e , 150 a l . cotton stoppered erleameyer fla s k s .
The depth o f
the milk in the fla s k s was about 0.25 cm. a s compared w ith about k cm. in
th e tubes,
fhe surface arm o f milk in each f la s k was approxim ately 28 sq. cm.
a s compared w ith about 1 .8 sq. am, in each tube.
One drop of 10 p e r cent
e th y l alcohol was added to one fla s k and one tube of s i l k and 2 drops t e
another fla s k and tube.
One tube and one fla d c were h eld a s checks to compare
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
-
H
-
the changes taking place la the o th er th re e flask* and tu b es, tdaloh were
each Inoculated w ith 1 drop of a suspension o f £ e. f r a g i .
A fter 1 day th e re a o tio a of the m ilk In th e Inoculated tubes remained
unchanged w hile s i l k in a l l of the inocu lated fla sk s had become a lk a lin e
(Table 15).
Mo arena could be detected in any of the tubes but th ere was a
d is tin c t aroma in th e fla s k s containing a lc o h o l, the aroma being most
conspicuous In the fla s k containing 2 drops.
A fter 2 days the inotm latbd
f la s k containing m alcohol sas s t i l l a lk a lin e , the fla s k w ith 1 drop was
s lig h tly a lk a lin e and th a t w ith 2 drops o f alcohol was about the same as
the uainoculated c o n tro l.
Xn a l l the inoculated tubes an aoid rin g had
begun to form a t the s u rfa c e , but the m ilk was unchanged below i t .
The
a re as had lessened appreciably in the fla s k s but had become apparent in the
tu b es, the most aroma being present in the tube containing 2 drops o f
a lc o h o l.
On the th ir d day the inoculated fla s k containing no alcohol was
about the same color as the c o n tro l, but the fla s k w ith 1 drop of alcohol
was s lig h tly acid and th a t w ith 2 drops was d is tin c tly a e id .
change was noted in the appearance of any of the tu b es.
SO fu rth e r
In the fla s k s the
aroma was f a in tly p e rc e p tib le only in th a t containing 2 drops of a lc o h o l,
while the aroma in the tubes appeared to remain the same a s the previous
day.
On the fo u rth day, milk in each of the inoculated fla s k s had become
more acid than th e uninoculated co n tro l and eventually a l l became pink (acid)
and coagulated w ith no reduction of the litm us.
turned as pink as did th a t in the fla s k s .
The milk in the tubes never
The aroma had disappeared com­
p le te ly fro a a l l fla s k s by the fo u rth day but p e rs is te d in the tubes f o r a
much longer tim e.
A pparently, the alcohol and abundant oxygen supply coja-
b laed to speed up the metabolism of £&. f r a g i so th at i t c a rrie d i t s re a c t ions
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
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through more quickly.
As a check on a c id production by £&. f a u l . th ree s e r ie s of so lu tio n s
s e re prepared, a s shown l a Bable 16.
Before in o cu latin g , 25 e l- p o rtio n s
o f the so lu tio n s in s e rie s 1 and. 2 were t i t r a t e d w ith 0.05 # sodium
hydroxide, u sin g phenolphthalein as an in d ic a to r.
A fter inocu latin g w ith
? » . f r a g i and Incubating 11 days a t roam tem perature, 25 ml. p o rtio n s w ere
removed and t i t r a t e d ,
th e v a ria tio n in th e amount o f sodium hydroxide
necessary in S eries 1 <0.1 p er cent peptone) a f t e r 11 days was r a th e r su r­
p r is in g , e sp e c ia lly because a l l th re e o f the solutions in S eries 2 (0 .5 per
cent peptone) required approxim ately equal amounts and these l a t t e r amounts
were n early th e seme as those necessary to n e u tra lis e the solutions contain­
ing alcohol in S eries 1,
However, the r e s u lts obtained w ith S eries 1 were
corroborated by a th ird s e rie s made up lik e S e ries 1 but w ith a few drops o f
broa oresol purple in d ic a to r added.
A fter 11 days the re a c tio n o f the 0 .1
p e r c a st peptone without alcohol appeared to be about n e u tra l, while both
so lu tio n s containing alcohol were a cid .
S leetrem etrie pH determ inations by the quinhydrone electrode on the
so lu tio n s in S eries 5 a f t e r 20 days were 7*32, 5*^0 and 5*&9» re sp e c tiv e ly ,
f o r th e peptone alone, peptone plus 0 ,1 p e r cent alcohol and peptone p lu s
1 p e r cent alcohol.
She so th ree so lu tio n s were l e f t a t room tem perature
u n t i l 12 weeks had elapsed.
Baring t h is time they became le s s acidf th e
so lu tio n containing 0 .1 p e r cent alcohol appeared to be about n e u tra l and
the others a lk a lin e , the so lu tio n without alcohol appearing th e most
a lk a lin e .
I t is w ell known th a t many substances which, have a very p lea sa n t and
agreeable odor when present in minute q u a n titie s have an e n tir e ly d iffe re n t
and o ften disagreeable odor when in concentrated form.
I h ls i s tru e o f mask
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
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R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
and indol which a rc used a g re a t d e al i n perfumes.
id ea o f th e odor of the aroma m aterial of
in o rder to o btain an
f r a g i in more concentrated
form, th e follow ing experiment was performed.
Two 6 l i t e r hatches o f 0 .2
p e r cent peptone so lu tio n were prepared and s t e r i l i s e d in 12 l i t e r fla s k s ,
follow ing s t e r il i s a t i o n , 12 ml. (0.2 p e r cent) of e th y l alcohol was added to
each fla s k as well a s 1 n l . o f a suspension o f £g. f r a g i.
The inoculated
fla s k s were incohated 3 days a t room tem perature darin g Which tin e a strong
May apple aroaa was produced.
About 1390 ge. o f aoditua chloride were added
to one o f th e fla s k s and i t s contents were than d i s t i l l e d u n t i l about 500
ml. o f d i s t i l l a t e had heen c o lle c te d .
The arom atic m ateria l cams over in
th e e a rly d i s t i l l a t e and none could he detected in the d i s t i l l e d c u ltu re .
The contents o f th e o th er fla s k were d i s t i l l e d without adding sodium chloride
and ahout the same amount o f d i s t i l l a t e c o lle c te d .
About 100 ml. o f e th e r
was added to each d i s t i l l a t e to take up th e aromatic m ate ria l.
The e th e r
was l e f t in the fla s k s w ith the d i s t i l l a t e s fo r 2 days, being thoroughly
d is trib u te d a t in te rv a ls during t h i s tim e,
i t was recovered w ith a
separatory fu n n el, d ried w ith anhydrous sodium s u lfa te and th en placed in
s n a il fla s k s which were not tig h tly stoppered and, accordingly, allowed the
e th e r to evaporate slowly.
Whan the e th e r had e n tire ly evaporated, th ere
remained in each f la s k only a very small amount o f m a te ria l.
In a r e la tiv e ly
high concentration the odor o f th is suggested p lu s blossoms, but i n re la ­
tiv e ly low concentration th e odor resembled th e May apple odor which i s so
c h a r a c te ris tic o f g£. f r a g i .
The r e s u lts In d icate th a t th e aroma m aterial o f
in e th e r.
f r a a i i s soluble
T his is a lso in d ic ate d ty th e f a ilu r e to d e te c t the May apple
odor in the d i s t i l l a t e s a f t e r e x tra c tin g w ith e th e r and allowing th e e th e r
odor to disappear.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
MSGSSSKW Of HESULfS
Th» existen ce o f J&. fra a i in the United States was overlooked m i l l
i t was iso la ted and id e n tifie d at the Iowa A gricultural Experiment Station
sons years ago.
the frequency with which i t was noted i s dairy products
th ereafter suggested that I t i s widely d istrib u ted in Iowa.
I t s iso la tio n
from a few samples o f dairy products from other sectio n s of the country
was evidence that i t i s not confined to that sta te .
fhe examination o f milk supplies delivered to p la n ts in Iowa and
Kentucky showed that £g. fr a e l was present in the milk in a considerable
number o f in stan ces.
In the milk su pp lies in which i t was found, the
method o f examination used did not detect i t in a ll the samples studied.
I t was not detected a t a ll in milk su p p lies from seme farms, even when as
many as 15 saap les, representing a period o f shout 2 months, were examined.
Perhaps, i f larger samples, p ossib ly a p in t or mere, could have been
%
studied, an appreciably larger pefeemthge o f the samples would have yield ed
it.
As i t i s e a sily k ille d by h eat, i t should net be present m w e ll
eared for m ilking u te n s ils ,
th erefore, i t s presence in milk may be seme
indication o f the care used in production.
However, because o f i t s
psyckrophillo nature, Jo. frasri i s more prevalent in the w inter when the
cows most be kept in the bam fo r longer period s, and i t may be mere d if­
f ic u lt for the dairymen to prevent contamination o f milk by i t during
th is season.
fhe m ajority o f the sp ecies of the gem s Pseudomonas appear to be s o il
and water forms.
Stridently £&. fr sg i i s no exception,
fhe iso la tio n o f
th is organism from a m ajority o f barnyard d irt samples obtained on Iowa
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*7 -
and. Kentucky farms to seasons favorable for Its growth indicates that It i s
commonly present to so il to those regions* failure to isolate i t from any
of 10 such samples obtained on Kentucky farms to summer emphasises it s
payehrophillc nature, Since the dirt samples sere taken from the surface
and sinter samples from some of the easts locations yielded the organism* i t
is probable that to summer it survives deeper to the so il where temperatures
remain lower then at the surface. Hone of the samples of tap water or water
direct from w ells to Iowa and Kentucky yielded J&. fra a l. hot water from
sources exposed to contamination by dirt* such as stock watering tasks*
surface of ground* e tc ., harbored it to a large majority of instances.
Sinee a ll of the general barn equipment, feed* bedding and even the animals
themselves have ample contact with d irt, the presence o f the organism on
them is easily explainable.
Of toe dairy equipment sampled, only milking
u tensils are given such care or treatment as to control this organism, This
is reflected to the data, since only 2 of 23 saaples from such equipment
yielded £ s. freed. fheee two samples case from farms whose milk supplies
had frequently yielded toe organism, and presumably these dairymen were net
earing for their ailktog equipment as w ell as seme of the others.
The m m frequent occurence of £s. frael to barnyard so il in the eastern
part o f the United States than to the western part is of in terest.
It is to
the eastern part that most of the nation* s milk is produced and most of the
dairy products manufactured. The fact that £&. fraud is more widely distrib­
uted to the heavy milk producing area increases the p o ssib ility of contaatoar*
tion of a large portion of milk and other dairy products. fh ile the reasons
for this geographical distribution are not d efinitely known, the weather and
especially the amount of annual rain fall may offer an explanation.
States
to the western prairie section and the Eooky Mauntains have only a coapara-
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- *m t t w l y small annual r a i n f a l l , mash la s s than those l a th e e a ste rn se ctio n ,
l a a d d itio n , t i n high tem peratures occurring l a summer throughout such of
t h i s t e r r i to r y probably a re detrim ental to th is psychrophilie organism,
fhese conditions no doubt influence the e x iste a se or trcurrival o f an
organism, such a s J&. f r a g i . whose h a b ita t appears to be s e l l and e a te r .
Conceivably, such a comparatively dry area a l ^ t form someehat of a b a rr ie r
to th e westward spread o f !* • issgkfhe e ff e c t o f alcohol on a re a s production is of i n te r e s t b a t probably
o f l i t t l e p ra c tic a l Importance,
I t may be o f use shea time i s an im portant
f a c to r in determ ining the presence of F s. f r a g i o r in th e prelim inary
id e n tific a tio n o f i t .
By I t s uso r e s u lts might be obtained a day sooner,
th e use of alcohol in th e enrichment procedure does not seam t e be p a r t i ­
c u la rly h e lp fu l because th e aim of t h i s procedure is to produce s u ffic ie n t
increase in th e organism to dominate the f lo r a so th a t i t s colonies can be
re a d ily d etected a f t e r p la tin g .
I t i s doubtful th a t alcohol would speed up
th e reproduction o f £ s. fraud appreciably a t tem peratures used in th e
enrichment procedure.
fhe arom atic m aterial produced by £ s. f r a e l appears te be formed from
p ro te in o r p ro te in fra c tio n s .
In peptone o r litm us m ilk made from skim
m ilk, where l i t t l e or no f a t i s p re se n t, no ra n c id ity d ete ctab le by small
i s produced.
When c u ltu re s a re kept in cotton stoppered tu b es, the May
apple aroma eventually disappears.
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grnftiawr
Bata a re presented on th e d is trib u tio n o f gg. fra g i in normal and
d efectiv e d a iry products, d a iry p la n t equipment and e a te r supplies and on
farms.
Samples examined fo r the presence o f th is organism were ohtaised
p rin c ip a lly in Zowa and Kentucky, although d a ta on samples of s o il from
barnyards in 31 e th e r s ta te s are Ineluded.
P a .frg g i was found freq u e n tly in sa ap les o f normal m ilk and cream
d eliv ered by producers to an Iowa m ilk p la n t.
I t m s not d etected in milk
d eliv ered in June te a Kentucky milk p la n t, but was found in bO.O p e r cen t
o f saaples d eliv ered to the same p la n t in December.
Several samples of d efectiv e d a iry products, e sp e c ia lly those c r i t i ­
c ised as rancid o r a s having a May apple odor, yielded the organism.
In g e n eral, th e d a iry p la n t equipment examined was r e la tiv e ly fr e e o f
it.
An appreciable percentage (10.0 p er cent) of the leva d a iry p la n t water
supplies examined yielded the organism.
A large proportion (51.8 p e r cent) o f d i r t saaples obtained os Iowa
farms and saaples of e th e r m aterials o r equipment lik e ly to cose in contact
w ith o r be contaminated by d i r t was found to harbor P s. f r a g i.
R elativ ely
few ( h . l p e r cent) sim ila r samples obtained on Kentucky farms during the
summer season yielded i t , but i t was found in a la rg e r proportion (37*^ p e r
cent) o f those obtained in December.
P s. f r a g i was found in 23 o f 33 samples ( 69,7 p er cent) o f barnyard s o il
obtained from s ta te a g ric u ltu ra l experiment s ta tio n s throughout th e United
S ta te s .
I t was found in samples from a la rg e r proportion of s ta te s (90.0 p e r
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cent) i s the e a ste rn h a lf e f th e country than from thoee in the w estern
h a lf (38*5 p e r cent)
fhe wide d is trib u tio n of £ s. f ra g i on fa m e emphasises the importance
o f fa re s as a source of th e organism.
Arena production by J&. f r a g i in m ilk, peptone b ro th and on a g ar media
containing peptone was stim ulated by th e presence o f e th y l alcohol but not
h r o th er alcohols tr ie d .
The arom atic m aterial produced by £ s . fra g i was e th e r solu b le.
concentrated, i t had an odor resembling th a t o f plum blossoms.
R eproduced with perm ission o f the copyright owner. F urther reproduction prohibited w itho ut perm ission.
When
i s g r a te fu lly sad* to J* . 1 . V. Bbsm* f o r h is h elp ­
f u l advice and suggestions in carrying e a t t h i s in v e stig a tio n aM i s
preparation. <*f the w a m se rip t, and to P ro fesso r k. W. Badnick e f learn
S ta te College and Mr. jfesss B. Smathere o f th e B n iv erslty o f Xeatadqr
t h e i r a ssista n c e in obtaining saaples In th e f i e l d .
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-58-
m u m s CI«B»
1.
Back, X, tfkiasr einen fra c k ta th a r bildaaen Mikrokofckas ___
e a te riflo ta n a ) A rt. a . ft. k a ia e r l. Sanrndlleitsaata
25&-2&3. 1906.
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Bergey'e Manual of Determ inative B acteriology,
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3.
Gann, H. W., k i n , ff. M. and Stocking, V. A. A o la s s if ic a tio a of
d a iry B acteria. 18th An. Sept. Gann. ( S t e m ) Agr. l*g%»
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M chhol*, Wilhel*. B rdbeerbaclllus (la o ta r ta a f r a e l l
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O rtas, X. Ubar a lu m aeoaa aroaablldenaa B a c illu s, nabst einige®
Beasrtamgea u b e r Beintoalturen f a r B xportbutter. Centbl.
f i f t h S i. p . iH l.
Gentbl. l a k t .
Bakt. AM. 2. f t
€.
Gruber, 9k. PsauioBKnaaa frm eariae. Xina Xrdfeeergerueli ersoagende
la k te r lo . GoatM. Bakt. AM. 2 . 2* 105-722. 1902.
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giaBlliffltmiMtf & l t t £ * a t II* GoatM. Bakt. AM. 2. | | t 122-123.
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S ak tarian . Centhl. Bakt. AM. 2 . 12,: 60I - 67H. 1§07.
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Iona I g r . B spt. S ta. Bes. Bui. 225. 1937.
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5313.
h&rn* s* *•
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Jour. Baet. gi
Reproduced with permission of the copyright owner. Further reproduction prohibited without permission.
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