The bacteriology of cheddar cheese

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Material Information

Title:
The bacteriology of cheddar cheese
Series Title:
Bulletin / U.S. Department of Agriculture, Bureau of Animal Industry ;
Physical Description:
52 p., 1 leaf of plates : ill. ; 24 cm.
Language:
English
Creator:
Hastings, Edwin George, 1872-
Evans, Alice C ( Alice Catherine ), 1881-
Hart, Edwin Bret, 1874-
Publisher:
U.S. Dept. of Agriculture, Bureau of Animal Industry
Place of Publication:
Washington, D.C
Publication Date:

Subjects

Subjects / Keywords:
Cheddar cheese -- Microbiology   ( lcsh )
Genre:
federal government publication   ( marcgt )
non-fiction   ( marcgt )

Notes

Statement of Responsibility:
by E.G. Hastings, Alice C. Evans, and E.B. Hart.
General Note:
"October 2, 1912."

Record Information

Source Institution:
University of Florida
Rights Management:
All applicable rights reserved by the source institution and holding location.
Resource Identifier:
aleph - 029612187
oclc - 22231567
Classification:
lcc - SF623 .B14 no.150
System ID:
AA00018895:00001

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nt 6 prable -tondf ib
1 9-04, tw-ici'a Pr6duoedhav,,e pryi=UJ 'I ta)
f *dther 4*11
aytv'sepa, ted,'h:Rqoh a dist&w, that Uiv ]p
o d4 not the Ahm each colony theit is;
"'t were'the -on 'boloiiy ati afe-` Hence forms ajppw bul I
1 47
4 -pfates, Wat, AQ not A6n thow cr 0-wded with
orgunsm that fi-ndg most f;tv'orable w"tiaos kmfly''
UUMeo SOMP other --.-form, t4e latter'VV naver appeAr, 64
ixlro or W- $0 Meons6ni #8'ri6t to attract the, atteno, '.Q
th
tel,(A A".rado bfL11000 t4d J'oreven -500 to I m4
6 b
the 'ordinary'' plate-oultare- jiieth'
qrgamsms piese4 ini 01er numberk. This statewiat-AW' -,:j
er"uhs: obt-aija6d,,bY'Pr'e*ious' inveuPtois"O.
C
B b 1 t
.896 notomemfivo.lbac emorlojecal 6xaminitiozia hQ,'.
a, rip..emng: 10hed4ai. 6hew. The first- detaikd *o& aW
t tjV' 40f *hich the writers-are, acquambW w
RUse11*1 Thi x* as largely' with a-*qqantftA 'a,
2: Wo W. iv
fh6 chees' du "g, t iipe
mation. of t e_ rm, _hk ningprocess, by means'16f,'
ablished 1h' fa lorming'o
plates. e Ct tk4tacid- rgRM$M SL
09 t d- r: Of fhdb ctefia thus determined; It ma
iderrid thai th a6d-f0wing baAeria Ion to,- he B
gr(ip. The*ork.of Lloyd: on En 'Cllwddvx
led to: Uar reWt. The mbk- ext emi -work, on 'the b
ra '6ddar cheek Js.. th t:of Hardi dPtu Chal
Ao of Ch
1AW th differ6nt typb of 6_rp.nish4s ippe 1 ri4g
h: on cultures
from 0, ar' eeses during... e en ire pe
wrmd Ched.d ch
'The, e'th 300. cultdres thiisisolated "were studied in a4ft
reduded to 38 groipp, Of thepe' 38 4 beloigtd" to tlm,*
ftvn, Iddl acidi the only one which 'Was always'IOQUI
it PTROioilly a1Wa u.-ded: over 99 er nt of the
Rassow Ja The rim atid faU of bacter14 In Ched4ar'chesso
turai Expertment Station, Wrteenth Annual Report,,PP. 95 1.
Hardlng,, H'. 'A, and, Prueba, U., J.,- The, bactertil, Aorfl,- :fl"
Yoft Agridultural Experiment St ation, Technifti Buitetht- Ge, i















































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1-'order w.-Aawoftstrat& 4t;f WtIk *IL
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Cut& C44 aw, -a'&- IL Pirfioli-7 of ft
_44t
I fay,44,
fep&rate:"ve9sd' Aciditi deie6ninatims, weft ma& v,
i ts iA r4bleA..' Itwill be tiW- -that
are, gveii
the,acxti -,6f flie`_ whey mi, conifxct', with the, cutd, filk
wore rapidJy -than -fliai'-of t1* whey im -in coiitar*
Axi ra.-POv46Pvwnt at- acid in'wh Wonc and in wh

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NV hey and ourd.-o'..o. ..12 .14 .32,
1'016 4:
VV Rey -------------
Wjiey afid zur& ........... :09.. .10 .18 .200 49


It v, ris thought af of- sampk of milk was ctirdled by re*e
the cu eut and iflo. iied. to settle to the bottom oft deep
tI* Wney diffreni 16yels -Amild. show vary-Mg dweg of jw`
it the were so. proteeted that con-Vedibn currints, -did'"''
1. tend id miix the. fiquK In. 'other words, th6. acid whey expeW '

foa&ulaulate Mi the 10ver portion 04 the Oontainer.''
In orderAo ieA this,.&ep beakers were fdled with milk, the
Ukdw. with vennet,'And-thd curd eut:and allowed to -settib.,,",
beakers were keptin uth at' at 95 0 thus no wo
.rents, were present.. ince csr6 was taken to heat the milk to.
temperature c#rdI*g.. The samples of whey WOW,,
in such.a, manner OuA mix the different layers.' A:
resiAts'of a number trials are given. it Will U
a d' dit of the bot tom lay- ers iticreases, much bst6r ft4
Piper jqers, the, digat"ce, being in,
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Bulletin 122. Washington, 1910.
Babcock, S. M.; Russell, H. L.; and Vivian, A. Influence of rennet on cheese ritgen-
ing. Wisconsin Agricultural Experiment Station, Seventeenth Annual Report, pp. ;d-S
122. Madison, 1900. .
8Jensen, Orla. Studien fiber die Enzyme im KWse. Landwtrtschaftileh :flrb&!;iJ
der Schweiz, vol. 14, pp. 197-233. Berne, 1900. .: .. "?
Van Slyke, L. L.; Harding, H. A.; and Hart, E. B. Rennet-enzyme as a faeto# ,i ,
cheese ripening. New York *Agricultural Experiment Station, Bloetin 28&
June, 190. .8.. *

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the vurix
ox b
of cheew. bf reopet; extrad iA
ibaence of lactic'' organisms is, demoustr ated., im
i
t-: A4 per copf soli#ibii of 'agar w'aa-W&d to Oex le,
'Prop",ortion of'l'to 1, A Portion, bf t he milk a r
ind th p
*ted 444, w lad,101,11 orgWUSM
thwother, portion was not oeu]AWO, At tbk
Paw -W 4
,
Of in e
Mbition, $4%p Of 11ter
+
1*, L,
aeQ,, tbe',plates, whi
4,4 *_ forv "jDur h ogra h *as ea bt
41 pw,,k , i., 'T-h,& p ot p 1ak,
not*, d" in, the prw*nm, of ihe',
U I 'U t
Y_ the
Mia s
P4 1mv, 4b*,,, Uw, (wgauWIM no SU6 M*'rked'
""The h0 caseln'has r6sulted ii
r
*f, 4o''Opoity of the-n4lk egar o that -a nurAei,
Ahov'4* oa, #bo riead,,, wWi e, in, t1w emie, of the,=flk
no trwe
00046 we 14040, Orgamm*6 Wtre a-beent
ow bq',diAiupisheA.' In, the, mi avar A A 81
va A o

W, 80uwlthat if a cheese was made fro'm td 'febsk,
rming organisma the rate of ripewog
Aw`test tl-ds hypothe" a cheese, was' ft
Slex-reely aoy aoid-produein' Ov
Aedi xb, arid- what the
ver was formed, dump.
ur v 0 04 e
T% -m, ,, dter sfivndg 24, hmm at '209 0, ha(j 14
M M
per In order to iu8ture proper cut BaL'AT:*i,
0 nulk was,, raised, to ,O-.M per ceM by, the addition
or&r to follow the rate of acid.formation
ere Made sit'
th446,gar present.," the ckeew, w
vlm Aft giVen in Tae'

OOW of rheese 404o ftom m4lk cmtaining few. RMUO
'bacteria.

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w6&,,, The, as tommwed fba.
UkWre and, develop e6r,=,q)Wdingly,
70&be inmths: the ie a, Vona toldx M, Aiw
4MY eh-eese. Jlsvor.:.,
P rj
TME.P.ROTECTIVE ACTION OF ACID.

Vh -putr6factive, bacteri&:.of--ihe thit, are "Mn
M:Mijk7 findlence in
ideid, rexdion_vhich is ma r.]Ln -'he,
intained du ig't entir 'Pen
pro ve taefift 16f avd, w", first, dem-onstr
Sta6616 br;Bboo&,And',Rsus611J4
by va gihiinig iii
'mo"d %thw sugar: fram, cui rd a- t r,," T hie';
A
,velbped, oI 'd69iraMe,,6&rs and, -ixs;b6s;4ndba.teri
howed. li lying baUria -t* be: nlu 8. if I
ad oifts s qne merou
u or cane sugarveref aid&&,to, 4,he washe&, turd the ri'
P"
promw wag rftuch more nearly fiorinal. beca-use rthe aciA, reftetkil Wio".
themby restore, duelo, the, &rmontation of -tI ,:idde,&1,SU9jW, '19
t4o xLt!r0fftttiv hibited-,. as in f i mormal: hme,
p: e, organisms were in
erimental:w-orkdn flavlo'r devekpmentin Chqddar
eeses; were- prepared. fiomcbrd1iom which'the SU
1`by washing. In Plate I, figure 2, a nvi-baal, she
1' ese, arre:,illustrated, The vashed.curdl bhewe *asi
&Aure, ibeing a soft, plastic mum lk
odor. and taste.., On," tli,
mp t de.lv' the: -same, waghed, cuid,46e' old, hAd
e rmness was, kestor
'ith 1he result, thatith- fi'- ist
TMi r816 of organisms of, Ahe Bacte2ium laotigam.di_ group J41*1-
ChMdAr.aheese: may be'summarized as follo. 'gv i.:Y,
'AL They favor the rdloing.
u proms,
he'expuliion of thej !71 Ad
2.; They- favor t 81
They permitof, the'fusingof the 'paxti.
4. They activa-te the' siai of 1he renneti.extracL,
5. They have a prte ve.aption against4he putrd-
teria.
It is certain that this group 4:i'tn essential factor, in the rip'
of Cheddar cheese. Their perioa of growth bA
.12
short, since it has not been
after the disappearance of the r.
Babcock, F3. M.,:,4ussell, H. L., VlviaA, A., and Has
the nature of, the f4mentati6no occurring in. lnilk,:
.Experiment Station, Eighteenth Aninual R"061, DO.




































Fie. 1.-THE ACTION OF RENNET EXTRACT ON CASEIN SUSPENDED IN AGAR IN THE
PRESENCE AND IN THE ABSENCE OF ACID-FORMING BACTERIA.
.The same number (902) was placed beneath both plates at the time the photograph was taken.
The digestion of the casein in the presence of the acid-forming bacteria has rendered the
medium transparent and the number apparent, while in the absence of the acid-forming bac-
teria the digestive action has been almost nil.


FIG. 2.-THE EFFECT OF ACID ON CHEESE.
o"" n the left is a cheese made from normal milk by the usual process; in the center a cheese
,* .made from the same llk, the curd being washed free of sugar; on the right a cheese made
S from the same w drd to which acid~d been added. The washed-curd cheese had no
boad.. was a t ic mass. The adMtion of acid restored the firmness of the body.





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4mt ."YAfg*ed Adice in O&W Bnft 62&
A
ft'ergrowtil, W;64,1jdMg
founded. Indfed, some facta would iiadiQat6

Wth wrdAoprotwI* and- develpp-
otjques loogafter *14 jsipposed, tobe the per"Jd,
large, amo" %vailabl asto. the
clata. is,
0mg, Q11: M 'T'VeMWcheesc but, untw,7"UOY
am V".'A
S4CIW 14 jh. pqmpm6un4,.C,0,iIta*
xan Aug. !The content of tho
14ging, d espwigy
n
*,d4gire# ao4sv Tooxplain them chapWes
jv, `,tq_ enzpw- 7 "0, MostgroWAX to thooe Oalpo-
-V m-s, or to, qt]rtyWr gri;a tha
p, of b4d
h" 4,ca*d the, pexiod of bst

PXVWted &ow, that'Up pmid of activity is
kY dpterin 'the Or4inoxy types of bacwria.,
=0,,Usalerowth 4(rs no of course,
fQrmedlby tho Bactegium Iwtic acildi
tjAt 4yxing the early life of t1w, cheese, make up

*ATE* OR' '9A'CTkPdAt'GR0WTTf iv CAR I 199V.
w of the cooperative work cariiedloi 'y the, Dairy,
PepartimentqfAgricultwe and the WL5consln Bx-
loTgenumber qf,ebeeses have been examine
Som ,Wm in at frepent'intervals, dgring
*PJA" alul, (Iurmg, the, fa# potiola of, the ripening,
,vve akwwned, at lew fivjuejt rvals, duTin th6

IK()M 01P JMA3"A7TOf.'

*0*0& of" 1,00 used was the gTinding with stuik
,mod of- umplo of the cheese taken frOIR VariiDUS parth- of Sk
IS "_id :VodAr Condition& LV)Culationg in vary-'
-,wem vAde from, ther cheew gc4emion into lactose-agarr
3?4IA*k Ahk theveriod.of iumbationof the Istes, was,48-hours
UtOULO *eabotion 4t 70 was. supplemented by a fw6er
Tkis, mothod of incubation allows the develop-,,























The maximum number of bacteria as measured by the laotose-agar s :'3
plate cultures occurs early in the ripening process. In 8 of the 11*i
cheeses examined, as shown in Tables 9 and 10, the greatest number': ..
of bacteria was found within 48 hours. In the case of the examina- I"
tions detailed in.Table 11, it will be noted that the maximum numbers' ::
of bacteria as determined by lactose-agar plates have been found '.:-:
much later, namely, at the end of the fourteenth, forty-fifth, and '
seventy-seventh days. Thus, in no case was the maximum number 'i
found at the first determination. It is not believed, however, that **... ::
these cheeses, selected especially to illustrate certain points to be dis-_
cussed later, give a .true picture of the initial development of bacteria. '
In the case of 9 of the 13 to be mentioned later, the maximum number
of bacteria was found on the first, second, or third examinations.
Harding and Prucha 1 obtained similar results. Seven out of 9 Jl
cheeses examined showed the maximum 'number of bacteria at the
first examination, 1 at the second, and 1 at the fourth. k
It is impossible to determine the time at which the growth of any ;:\
particular type of organism in cheese ceases. As the fermentation
in the cheese progresses and the accumulation of by-products in-' ;
creases, cell death begins to occur. As long as the process of cell ;i4
division is more rapid than death of the cells, an increase in living i..
bacteria will be shown by the plate cultures. Soon death of the cells > |
is more rapid than cell division. At this point the decrease in ap-
parent numbers begins, although growth may continue for a much it
longer period. It seems probable that the growth of the bacteria of
the Bacterium lactk acidi group continues until the sugar is cr
pletely fermented. In milk the cessation of growth is due to t
1 Harding, H. A., and Prucha, M. J. The bacterial flora of Cheddar cheese..
York Agricultural Experiment Station, Technical Bulletin 8. Geneva, Dec., *0...19l
.. . : /....







t I
ofblin fIii bpeq n ahganf
,,rmwa, snw o|h~ame fcls m
gr tii s'cIihta h~p h
batrI o ia
'J e
th raim f h N~c~tf j j
ef f mitce'sniss6
Co8 f
-m iiiiiii iii~o~ c e as
A!-tkiw r lt trs
fIbn x~fd nJ~los
ilA
lss
2 ~~ 4 6 7
Hi7
1172




!2




Tx 71-1
ARA

n-,-Yvjnws of bacteru Pei pro* U oftom*, 0400W4-1

lNumbm.explew4d in VUbM.1'

CheM No. V. Ohe"&WO,54. Mom No. SL,,

Gel CkUtta Lactose- (1datin 'Lactose- qejo ft
Oultum
culfum. mituro. culture. Odt- '2,

.......................... .. 2W '144,
............ ........ ........... ....... --------- --
.- .. ... . 3
.......... .......... ......... .. I .... ........... ......
14 ....... .... ... 32 il 40.0:,
22.4 ......... w ........ ..........
............. .10
30.. 2 70 .......... .......... ..
37.4 .................. .......... .......... .......... 270 480 ......
451 -------- .......... 40 25 ...... 1,400
....... .......... I ........... .32 1,300 250 225
.......... 51 40 ........... .... iiy ... ......
r ...... 86.5 24 ......... ...... 128 250 84,
10D ....... ......... ...... . .......... 8.5 12 .74 65 loo 120
113 142 W 67
VA_ .... .......... 12 50 2& 5 146
143 13 13 12.5 Is 23.
............ 22.5 W 5
.................... .......... .......... ........... .......... 10.1
..... 6 ......... ...... .......... 17.5 6.5
176 ................... ._5- .......... ...... i3. 9
187 ................... .......... ------ 6 ........... ------ . ....


According to the determinations of MacNeal, Latzer, and Karrl
bout 5,300 billion colon bacilli
there are a in a, In f d
gra 0 rY 01W
Figuring. on a naoisture content of 90 per, cent, ther' :would beA530
on' in a' determ' nation madi
illi gram of the moist growth. From
by weighing out 1 gram of the moist growth of a coccus, formfrom
,,cheese, making a uniform suspension of this in, a known volume of,
water, and taking an equal volume of this suspension and of nor mial
blood and counting the number of bacteria and red corpuscles we
h ave obtained an average figure of 1,150 billion cells per grhn of the
moist growth. The basis for this method of counting is the fact that
from a known number of red blood cells one z can figure the V,01UM8
used, since the red cells are practically a constant quantity. The
average volume of the colon organism is 1.13 cubic: microns; of the,
coccus 0.5236. The agreement in: the determinations is thus very, dbse.
It is not to be supposed that, so far as the number of bacteria is
concerned, all cheese will be similar. 11he results shown in Table 11
emphasize this point; two of the cheeses, Nos. I and 54, showed atno
time over a few million bacteria per gram, while the remaining. two
had a consistently high germ content. Such differences may-be due,
to many causes. The period of m. imum numbers is followed by'la
decline, rapid in some instances, rlov "in others. At the time, the
cheese is fully ripe millions of living lactic bacteria are usually pres-

MacNeil, Ward J. ; Latzer, Lenore L. ; and Kerr, Josephine E. The fecal buterlA'ot, Av
healthy. men. Journal of Infectious Diseases, vol. 6, No. 21 pp. 123-169,. Apr. 1;
571-6091 Nov. 26. Chicago, 1909.










7

A-
Ndt, to,
AX7 V$w- oeiit, waslp 41a 1%,
-he koskra W49 M"W e
ho u a! f" ed
S !yFn J &Ples 0 MR Pesrv'
thp rnatiou,,,of M* fti4 4Y
ly enzyMS, of tile mfik.
Orooo
-In -'the'. bottles of --raw i ilk in- th t A-W, 'AC444"L
se oT, s xriigbt, be pre;efit:- 'First, one forming an acid -frpm
in Psbon f the, Wtio 'a,,ae Ahe-
dAho bacten ti" on
Up f lea -only the -iatter, could be acti*p,
'4 g 6f h inhrent
ree ,eat was sufficiewt to de9troy all -the
milk..:, "Dw thiw differently treated should shO r i
I elico m- increase
ve d" ff6r acid. it the bactrial 'e'pryps wem
R; Th -bo
-siwh 'Ppcb 4' td, of.. the ra'w-wilk series
miniTnum: numb, a _8 M-ce the milk wa" but a few ouii
Am &r'clean. con ions. The ity t1i* b6ttls-
an -wn M dit acid m I is
Cfi ,
:-the': fresh-milk. '.Three f a p#p, 1, orpn,
t' orl th
emao & f e it 'lie
tely. 0 Ole, 401i tUM
--Ww Nvas filled= afbr th additiou H
JUjt,W n MC d -and A-11VIa bottlle4, we M
the V e e,
rying
101 Added. Th fiiA:'aeid A nation. vere mi&At
afteraddingAhe toluol to ea& Wfle.:,:-It will benoted in T4
..'e raw milk varied from OJ84 tp',,*
.14 the., case lof 11
cent.' The *increasing neadityindidA61:.-A great inerease,1411
bers6f bacteria., which wis also shown: by microseop"I
ion de from each bottle At, the time thepreserv tiv,6,v
Set.of bottles filledmith, h at d Pdailk were treated 'b
'kZ,*A
mannpr s the, a milk. The firstWtk of Ns set contalned,,P'v
1i "' g bacteria. The geidity inthecase of the
ticAy 110 VIU
d hrom 0.203 to. 0.456 Per cent.
TAPP
In -v "e,
In order to dekpnine 6W long the cells Persisted an acti
dition tub6s oflmilk:were heavily inoculated from the varou4 b44" i't",
7% -vesults are given m Table 12.




. .. ... ...

































4m<







t t_4 _,4


At
t_1
V
W
given hlTible
-in dot4i!

I&--4W 6dSe bf 'Od4ity'hz, mflk pi-egeeV04 PO
A AV

When' DSM
PrWerv-
save
W199
SddfKL 59 90, 1,21
j4tla No. 1: PerMIZ, Per Cent. Per aW. per Cent. PjCrWW.
per mt. Ott Qmt,
184 0. 19 0.234, 0.273 0.29 317 6.868,
._01* .05 Oft AK
le NO. 21
--Acidity..
WS 0.052 .122 .14$ .21
-No.
261 IM .4
W U4 -136 .177 .20t, -
Bottle No. 4:.
.266 i.261,
-A-ciftY... W : .391 .414 .492
..... ...... ~tO05 A% .148 426-
Bottle fio.,5:
4 'i'M M.
Addity. .4W .45 492 .423 *,644
Jnbre9m., ....... .......... ------- .05 W .168 .21 .141 aft
BotUe No. 6:'
f W .356 -.408 .40' 1515 W8 .644
M IN
33ottle Iqq. 7;
'Acidity. .408 .40 561: 08 -.740 1;
ID81 ..10 171 337

13[RATED:NnX

Mot tie No. & 7F
cidity .................. 0. M 17 0.
im 0. 18 CL 212 O..W 0.262
... .............. ......... OD4 -.009 OM .098 .09 0
IkKtlo No. 9;
............. .203 .2M 225 ..258 -27 .278
003 _.OK .022 .066.1
I)OWe No. M .. ...
..........
Acidity .................. .256 .285 .317 an .338: SW .$m
......... .... ......... .029 .001 -075 .082 .098,:" ..107 '19
Bottle No: 11:
Aoidity ....... ...... .308, 3M .368 M A04 A23 4491 ,.4k"'
.......... ... A)a .136 -.087 MO L15 .124
botdo No. 12:
Acidity .................. .349 .47 .423 .46 .08 .492 M Ito
....... ......... ..078 .074 .111 ..079 .191 to
Mottle No. 134.
Acidity..,. .......... .41 .45 .46 407 *915
............. AW .04 -.087 IM .151
Ha No. 1k
All ..... ........ .4k .519 .5m -M -M .08
.... ........ M -ON ..M M21 .143 .=4 *;W-


From the'conditions of the experninent Wdis:: to be ex*,ted
the. *increase of acidity in the raw milk. would be greater thinjo,
,.1eated. milk, and that if the bacterial. eazyms were operk
increase .in ''acid would be directly proportional to the,
enzym, prewnt or to-. the mass, of. Ce4 It will be no
a ",. ffii, muditions:.showx to, b Pimut by the figmm
table. te dAta, fkm tbree bottles of es& -0- f f6,
Are graphical form,
given in














EL














_-L ItS


Alt




AL














Iw *ft- kom *ftll per ftnt toluol. Itotao It
Of bactedOv- 6064 7 the MAdiuura, and bowe




Viet




ii !eke 14.*





14
a






44_T_











fAd oft PVved Iflow











wtth a P" omt
b6ftle 14,the S"'botUe 11 AM
nj




1 14

























Raw milk. Heated milk. (MS ".
efzms ..::.:ie
of d.k) . .

Bottle!I-............................ 0.0012 BottleS.............................. 0.0005 0.0807
.Per cet. ,Per cent. PWCL ":..: :j
Bottle 2 ............................ 0014 Bottle9............................. O .0
Bottle 3............................ 0018 Bottle 10 ............................ .0007 .0t : A:
Bottle 4- - ....... .. ....... ..... 0015 Bottle 11 ............................ .0010 .000 .. Iq
Bottle ......... .... ........ 0020 Bottle 12 ............................. .0013 .0001 .
Bottle 6 ............................. 0023 Bottle 13 ............................ 0017 ... .
Bottle 7 ........................... .0027 Bottle 14 ............................ .0017 W16 .1 A
Bote--------------------------.002-Botle1-------------------------------.01 .4301
..: .+! fi
It will be noted that the daily increase is proportional to the *
amount of bacteria present, and that the daily increase due to the
inherent enzyms of milk is as constant as could be expected. The i
comparison has been made bottle for bottle in the two series. This i
introduces an error, since the amount of bacterial cells present in ;
the compared bottles of raw and heated milk was not always the .::
same. In the case of bottles 2 and 9, which are compared, the initial :
acidity was identical, while in the case of bottles 6 and 13, which are .
also compared, the initial difference in acidity was 0.083 per cent. ..
The results seem to leave no doubt concerning the presence of an acid-
forming enzym in the organisms of the Bacterium lactis acidi group
that acts on the milk sugar. It might be thought that the increase ,
in acidity was due to the production of amino acids by a proteolytic -IN
enzym. In this case the soluble nitrogen must be augmented. That :'
this does not occur has been shown by numerous investigators.
The experiment was repeated in full detail with identical results. :'
The increase in acid can not be asserted to be dimue to the formation of 4:
lactic acid, since, of course, no qualitative tests could be made in tel 7
presence of the lactic acid present in the milk at the beginnnp'.isl
the experiment.
Ii ". .:.Kj ':'ii [
j y ',V
.. :: .:.. :W : 0:







u I
pI fyd6nt tnm:I -----
I0;u"~4i-OYbdra jvi fot-
nOYRbl U eODM i hv *"
on|ihhdaewth-h fraino
thtviudat fnysaeenA A
iAk" mson osieal wt f1e*s
4! OidVfte"so hb ognss hc lwy



4 1
an tatte ~lsofth lde aceia a,



Aim!eiiyjunind .k
nFATRAI HKD* H S.
to!l httevredceia hne ht






























produce in average milk an acidity ranging from 0.7 to 1.25 per t^,1
The lactic bacilli produce an acidity usually exceeding 1.25 per cent.'
It has been the practice to infer that every flask of milk showing an : .
acidity above 1.25 per cent after one month's incubation contains thdt- -
lactic bacilli, either in pure culture or mixed with organisms of the .
Bacterium actiNs acidi group. In the flasks showing an acidity less
than 1.25 per cent it can not be inferred that the lactic bacilli are .
absent, since some cultures are found that produce no more acid than '.
Bacterium lati acidi. Many microscopical examinations were made' ,
of the flasks to establish the accuracy of the conclusions drawn from '
the data obtained by titrations. Microscopic examinations were also -
made of all the flasks from which the titrations did not give con-. :
elusive evidence concerning the organisms present. The smears were
stained with Gram's stain and decolorized with a mixture of one part ".J.:
of anilin oil 'and two parts of xylol. This method of decolorizing s:
removes the stain from the casein, but not from any of the organisms :*i
that have been found in cheese. I |
The dilution method thus gives information concerning the ab .'
solute and relative number of acid-forming bacteria of these tw.0i nR
groups. It may also give information concerning the P rSet?
still other types of bacteria when they are present in gr
S than those of the groups already referred to, since they .
. : appear in the flasks in pure culture. 4 17 ;









&&I-lt-A--
04
W4le i" '
d, 4mit
lt
V!
mme4
)e04 iNte1
c op et
ffrii fr
| ~ i~.iito'id" ''~t
z u 4ePF,-
I1
&IltZ W19 )O NW feoiiqo P
assoIikInouaewt hg iuV'a





1 4 i t Il l l 1 ,i f I I,1
L[mbf exrm!inm oJ
MeI~o
L tmMTia3-Gl-













































tounat at tne time wnen tne group nrst to appear-me /nacenumr
lactis acidi group-has attained its maximum development, but be-
fore any considerable number of the cells have died, and at the t.ime
when the second group of organisms finds favorable conditions for

growth in the cheese.

TAmaLz 16.-Numbers of Bacterium lactis acidi anl of acid-produoetg bacilli
cheese as determined in milk cultures.
[Numbers expressed in millions.]


Cheese No. 1. Cheese No. 54. Cheese No. 85. Cheese No. 91. Cheese No. 0&
I

Age. Batt. Acid- Bact. Acid- Bact. Acid- Acid- Aei&
.ea pro- ICU8 Ppro- laCt pro- Badct. o- pro. Bat. bla- pro.
,cii during cti during ,t d ucn toacid ucing t0acidi. dfulg
acid bacilli. jidi. bacilli acid bacilliillilli, bailli.


Days.
21.. '-
4....
8...
11....
14....
22....
30...
37....
45....
*56...
77..
86..
100...
113..
124.
143...
155.
165.
176.
187..
208...


10

100


. . *
10


1


1I000
100






10


1,000

10
..,.6...


10
10


10
10


100


... .. .. .. ... -A


10



1
1.... ..
1
10
10


i..... C i"
........ 100


ltoo
........
1"000


10
1,000
100
1i 0
10
10
10
-.. -... --.
10
10


10
10

10
10
100
10
ID



10
10
10
10
10
1,000
10
10
10
I0
10
1,o00
""ioo"


1,000
10,000
1,000
100
10,000
1,000
.m.......e.


**"ilooo
100
100
100
100
10
100
10
100


100
100
...e.i. i ..J



100
100
100
100
.100
100
loo









10
10

10
10
10
10.
loo


1,000
100
1,000
100
10,000
1; 000
100
1,000
1090
1001
100
100
100
100
100
10I
100
"100
10!

r .." "" "


1 4. .. .... .'
.i .!E p~E .:",i


:..:: *:..,:


: "i .. ::.








S %
t ;"......
" ......

7,:1
'"* ? '."' ^E .."


: .....s,::
i: ..".
.' ..,:"





,'1
.* .|i


10,
1
i1
10
10
1

. w.


:... ...: .':-

*r*: i, !



h. :*{ : :*" -*"! : JH
.. ', : .::r
4: ; i
.. "" :::. !'A
s e :i
; t;:: t


",' *...i'"





I~IN
-A&


W
adi i f
bo T
Ct .C.P f.'.C C .


90.
.. . ...... ......
9091 0'4 v,9
RIA 909 5 5 9.9 9.
0 ', iiiiiiiiiiiiiiiiiiiiiiii9ii
|0, 9 0. ,1 9
so
11 0 Q s o 909 919-141
(w .1 '0 5:,9'9 "
91. 0 6 .9 01 9.T 9 ,0
Akj : o i;1 9 0;9 '.I s 0' W *
4 0 '0 9. *91 0 5






















































Age.


Milk before ren-
net was adaed
When put in
press .........
When taken
from press....
2 days ........
3 days........
4 days.........
Sdays.........
days ......
7 days.........
9 days .........
11 days........
14 days........
18idays.........
21 days........
22 days........
28 days........
33 days........
35 days........
42days........
40 days........
5d(ays .... ...
56 days........
64 days .........


Cheese No.
307C.


Cheese No.
308G.


_______________ I .5


Biet.
lactid
acid i.


Lactic
bacilli.


Bact.
kcrti4
acidi.


Lactic
bacilli.


'Cheese No.
309C.


SBact.
lacdti
acidi.


Lactic
bacilli.


Cheese No.
Bl30oc.


Beet.
acidi
,acidi.


Lactic
baciali.


Cheese No.
3110;.


Biet. Lactic
acidi bad,


'5 I I I I I I I I I
F.cL Pat


P. ct.
100
100
100
100
100
100
100
100
100
100
100
100
100
100

100
100
90
90
go:


P.ca
0

0
0
0
0
0
0
0
0
0
0
0
0
'"01
0
0
10
10
10


P. t.
100
100
100
100
100
86
100
100
86
90
100
80
100
86


29
40


P.et.
0
0
0
0
0
14
0
0
14
10
0
20
0
14


71
60


P.ct.
100

100
100
100
go0
100

100
100
100
100
100
70


89
"71"


P. t.
0
0

0
0
10
0
0
0
0
10
0
0
s0

10
0
30

10


29
!


P. et.
100

S100
.100
4. .


P0t.c
0
0
0
.o.


..i66i""
100 0


10i06
100
100
33
1006


0
0

66
0.
O.


PF. cit.
T1
100
100
100
100
100


100

lio
-100
90

"90"


P.r ct
0
0 .
0
0
0
0
0


-0
0

10

. .10


.... J1 ,

A.
Bfc. T. :,- ^%


J "f 'i E."' .ft : .":.:.E:.
100ch '.,:p- ...;^^,
ac. ds. :,


P. Ci.P. I ,.
100 o ..."9,

100 ":, iQ



........... ,' fr"v'{-i-fa


* *** ^ *fi 1' "4^^
*I AF.

... .. .. .....i. ;I
:.. .... .... s . : ; .i 'a
100 .


V.' 151~)
...h %.....

.. ..... : . ,.i.,y .:. i",l
.... .
.00 6 .M..
"
4 ; ":


F
,. .. .. . ..*;. :.,.
i
** . ., .. ,* ; :* / ...*.* . -, W
, .^ :,, ;::", l:"**,;",i : :^....


I


". C : ... N.. Ai
:: ~ ~ ~ ^*" ***-.**"-,,,^^


. ....
I:. ,


= J


: r

















































































10, 1



-1 'v




44U --
*45.|






lji

. ..j!





1log


icosk !0



![-ii











I-- gI
7
0 j-, X,--
A W -
4o,
OCT,
4e- -

,It, 4
4i t
AJI










4
(Ahtr
Wa The 'dirmi
1acteriiti tsinibilAifivn'k' tG1 bfifioth9k
c+, in *cheei*
ealff be still greater,
It -is oertainly true thilt m"'normal MeMar choise 0A
',et 'of the ordinary lactic bact4ii--
re have been belieVed to be.-the only
dheese of' their constafit presence w great
jevel of anotber of organ, MIS.,
4o:*d by the ovment OrRaIng"nat
while th ferni6nt ilk ku pr6duc* lactic
of ck'ilo -in -r
i0m6,:,6ther: source w hmse on amount -of, &6
pear4e: of the sugar- before their maximum period of devell
The influence. of -the first has. been pointed lout
paper. The ro 'lie: of th cond group. ean not atthis time 1**
b.utbecause -of their constant p" resence, in h u ibers, XDP
ti if n6t those of the first group, -their influ
ofthb ch6em'can notbe a: i1ftor, om:
DETAILED STUDY -OF LACTIC BACILLI.
The work of Von Fre er IV
udenreich Jind.. numerous lat ir is
bdicates that the lactic bucilli represent a group with'certalm,
=on characteristics, such as morphology, optimum temperatluro.0'
*growtb, and to some extent, *in the production of compounds
Buggr fermented. In all fields, of bacteriology:,it is di&ult'A,-
present time to draw the boundary. lines of any group 4.
The placing of the lactic bacilli foundin cheesein a single
.may be incorrect, but bectu,96 of the.faci that. all thecultures'L
have verta win characters in wimmonit seems best to discuss &W
oup, although more detailed stud ight divide theni
a number of groups. A coimparstive study ha's been made of
cultures isolated in the 'course of the work. The main licts''d
study am here presented. All the cultures were isolated ffir4o) n i
with the. exception of No. 1601 which wag 'obtained from milk.
tures 58A, and 58.B represejt different colonies from p4tw9l,
insure the p4tityof the.. cultures to be, used 'the, di4A
The.-differences noted in the.. study of these two au
from the' same Attire 'illstfate the differences 4A(s,
obtain in cultural *ork.
ho act* of ion, he d1#,erentcu1tures;'i4
aoh tkey have, bew,
IV7" Red 'r
r4 "COO
'J





















































































TIN$






-44









.4f 1








gk,, ald
Zro, W=WXWW9"RWW
to,
N It
OaL, T



'Per Ct. Piff d.
0 -324 0.'91 ..........
-1.12 RMCOVe.
132
..........
n
.......... ........
Alf oictive ......... 4 11f), I
do. ...... 96, -1180
104 1.1 ....1.46 1?vxtro+jnaetive
'HiBmanaun wnd, M[4eisO. t
...Mpor the, f6rimtWIn
ac4_i,: whil a, e: WhA6 Ad An 2 f6u-nd that- the or,91aniBMS
a. larze amount o 3 per
AulcOd con-t IOM14A
-A I I an, ift-
vhe a I in
reas the ures at formed'
Q ary po' he'eind' 24
show.el a lev'o-rot wtr at f of but, in
it ni ely. Ual 1evo
S em were: appro i at i eq
ve cids.
The rotary pay r f 9, of the ciAt I was determin4l.
e. 0.. ures s dW
k. M.' Curriliel. Four'of these pr'od ced inactive:aci -1 Pro
ifia -with, 9;.: pkedonllnanco: of the inactive
ctive and levo a6d,
A
-and ina' tiv scids
poduced a mixture of 'dextro c e With a
:-,,nance.. of the dextror-rotatory f9rm'.. and the remaining 2
dextro acid. One cultute isolated from milk of which im
purl e
tU4 was made, produced pure levo aei*dL : With 11 cultures' `6+
from other source,% than mi,1k or cheese, only the pun dextro or
M & ive, .. ...
ct* acids were found,
This.type' of organiam, then, caA produce inaotive 4dd, or.,
vv
acid of either modification or a mixture of the' miactive,
either of the active acid[s. All ofthe forms 'tre 'found +in r,
Asolat6d from Cheddar cheesewith the exception of the PUM

TyrE or CUM PRODMIP.

In most of the more active cultures the red on 6f mu
return of color proceeded in exa0ly the same+ mamjwr as ii a,
milk:culture of BactorsUMU*8 4iA.. Onlhe_'other
took p W1 only.asli or partial reduction. of 'Vim
e. euiture&
2 ladinewomn, P., G.A and Ileftratt, Mary. + 'staO +
TafoUoUS DISeas", Vol, 6, N6. 3, PP, 304-8191 + cbleiLvo
t 4S)Whjt*# BeUjELjntz44x4 ATM
4f tho 00_WW ftjgarjeo
U, fVW""rA"hOiU%.
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V AA
v*t
A;
totie., In- Tabld L& ther
v---thatn6i -u; Woo'
ji -3 bOtor mediujn tl4fi 6r 64
This was espeeiallr
Ui4ally cuialed-milkAn-aboAt 11
*i1k2la,*whiefi.,poptone -had boom -adde&` -,Ili
th4itlhe mo4active cultures, curd p, nu m,
.-.lwhsshopten& by the Styr iath*' led*,
an(! I -da in exiltureg Nos, c9S- and W' 01
_d _ys. was.,oonsideraly_ kmatvrAwt, i1i Of
Ity- at 424z
.4 tl* MI01 nta" V 'I
to Mung lrk!m
milk:eontamem"gir OX'gram-6f peptone.,.
nif&htallyj thi6 expeiiment shows, -that the*'ti6on 6t
in eertain perM4 ge`OT a whi
ta ddity- Nudhel Ch,
peftentagr I parbjeWar cull ute, jana b
1841ylli Int e for eac
t. by th ndseptic action of. th.0 ae, i -but, by a' lack of Sa
w6genous

T4=4 22-77-Peromtage of acidity.. produced bv lactic bacilli in mft,$6
_A
Popt6ne has been'ad V

Adfty In 10 duji ACM7 ih 412 &Y8.

'No, Uflk with MIlk *ft VJ& Vith M-Uk with
PIWn .-PIMU..
G.5 gr= lv= "gram I grkmn
milk.
Peptonew PeRPN246 Peptone,

Pff CCOL per COW. Per .79*. PrT 04.1 -Pet Cew. Pff 4011C
5 ?:324, 139 L 3V.: "0.866 1.35 2,34-
92 24 1.4, 1.58 1.4 1. W 1. 77 ol ,41
91 1. 68 2.1 .....242 2.29


..The action of the enzymspresent in thethee'sle curd'brinw,
0 $0, It it
on, ncreasing amouAt of 'soluble, n us tpoun& ta'
_..:;6,Oe.jn4jnth o lpen per cent of: the totatiitro
f rv ing about 18:
Jheform of water-soluble Compounds." This'action mWC,
curd a medium fav6kable to the-growth of theIketio,
44
rticularly to those strAins similar to,'cuJture W
A.slawlyIn milk, or. I&A to grow, therein, as
The: of this. group of Orguni M'S'l-, POAW
urces- w shown by growing
other 6' t
tlmvr, OP
I A
YAW ond UST4 x Ui'%
jvfP*OjA New T,&)*
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16, eAn

W, to cam
-w 16 Pw -M
Rum
s dotormi Tho 's
increase ate in: the rvew g per 40
--,OUt2,itas: ossible to, 40er4lmle, w the t*,4t-
-ism for*bich nomews of 'differe4i UQA
be6n found whether gwwt4 i tak'g
A 4
1tjS,:. characteristic, cd the: co=:O*t t,1Wy-persiA1)1
090=1
poiibjy grow. WW4 coiidition':S' wjli4, destroy less
are present butltll:im Whir
and in which, thewpistuiv rese urated
r1elp Ptkv 9,,$Ot
ibhloril :.'Efrorts weremik to -deterifi their number,
t S, PA tiu COn'.:.,JMDg 3, or 4 pxppnt:of sodium ci-
remflta were. quite, satisfact(?r as; far aA, the ebromogenic,
Occu'' Pot,
concerned, biA....since many other C', a forms, dic
Jt U.0-e was.
. ..... .. In, some cheees .very few olromoganie fornas bAvv 1*PA1
.-ihilip in otherivf ilar q&Iity they,,ayio bqqAprese
Wiltlym.clonsidemble Aumber, In -do
Ithanthe us'4al sa*iwt:*f salt they made up, a, r
on of the Aora'th= 'in other cheese

MQUMFYXNG ORGANISMS.
-Atienti(m has been: tbse organisn*thitt-
-direoted:Ao
Bounded liquefying'action M Olatij and'.eaSein. Tbe
oonfirm to' of -a statement w&-ewdkk-fl
c6owered Of IMPOrtAfte the, ripewng of. lad 0*
Ue nik- wialwidy. Presebtia'awkiot,'


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