X‘HE ESQMTION AND WENTIFICATEON' OF
CYANIDE-EC‘LERANT MICROQRGAMSMS
FROM A ‘E’RECKLSNG FEEER

Mai: ’56? {in Degree eé 3A. 3.
MECHEGAN STAKE CGLLEG§
Char!” Kmaar Shams

W55

”‘9’

EH53!-

 

 

LIBRARY
Michigan State
University

 

 

"‘ 7Q" i: ‘ ~/' 4“.

THE IJCIJITICT CT) IDETEIFICLLICN CF CYAUIDE-

'I‘CLEQTLTT ICICT-L‘C {GAITIESTLS TEE-CC»: A TRIS ZLIHG

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A TI“! :3]: _

Submitted to the School of Geo Mu te Studies of
Iich1:an State College of Agriculture and

ale plied bcienoe in partial fulfillrent

of the requirenente for the degree of

11.313143 CF 33 EITJE ILT 3113.10 ICLC: ”Y .2233 rUB L13

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l;a—J-£I.J1‘n

Becteriolory Section

‘1

The author wishes to express his the he to Dr. n.L.
Eallmsnn and Dr. C.F. Gurnham for their thoughtful
suggestions and a i in the experimental work that
forms the basis for this thesis.

Grateful acknowledgement is due the Desertment of
Microbiology and Public Health for supplying the
necessary media and reagents and estecially to my
fellow students in bacteriology.

Deep anoreciation is given to the National

7"

Institutes of health for the grant which previaed th

(1}
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as ship for the writer.

(73}

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Historical

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Experirental

Fig.

Table

Table

Conclusio-

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Proceiure...........................

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5011813910 £10.23? Lz’ Ca3i€7:o o o o o o o o o o o o o

Over-all View f the lrickline Filter

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or]. zinP.l:.'f_‘-i$ CJ. :keSUl-‘C’Cooooooooooooo

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1

‘\ .': 1. ° .0 ’1 n J‘ - ‘
geluccion o; cyani e anu tne

Proiucticn Cf Nitrite...............
309 of Jettlei iewoge enfl Trickling
Filter Effluent.....................
Utilization of Soiium Cyanide a: a
Sole Nitrogen bource................
Growth of Kicroorgflnicrs in a Keiiur
Containing Joiiur Cyshije as the
Nitrogen fiource.....................
Varible Growth of Licroorgflniszs in

£31113...o.................

Different

Cysniie Tolerance of Iicroorgsni ms

8 r‘v_- ‘
8111-1. ~- A!‘ {d .

Cysniie Tolerance of Iicroor

.-

Feviiefi Iyntheti neiium FC?nU13----

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Appen ix I Iientificstion of Iicroorgani:mc........

ABS“ “CD

A scale model trickling filter was put into oner-

ation with a cyanide concentration of 10 ppm. The con-

centration of cyanide was raised to 25 and finally to
56 ppm.

As the microor 3&3 nisxs increased the utilization of
cyanide, the production of nitrites was increased while
the nitro.te concentration rerained near a constant level.

Cyanide interfere with the measure of the BOD of the
settled sewage 1L high concentrations. The 3CD of the
effluent from the filter, tho ughc affected by cyanide,

did not show as rarked an effect as did the BOD of the
settled sewage. This nav be a tributed to the presence
of cyanid e- tolerant Wiccooroan are and the clarifying
action of the filter. It is believed that with the
addition of cyanide—tolr ant r:icroor3anis szs to the BOD
dilution water, the interference of cyanide on the BOD
will be diminished.

Through chemical analysr is it was determined that

1

cyanide was converted to nitrate, nitrite, and a.ronia.

p.

A medium containing dextrose was more favorable
for growth of micro rgonis 13s in the presence of cyanide
than a mediur without dextrose.

Soriium cyanide can be utilized by sore wicro-

oroanisms as a sole source of nitrogen but fail to grow

‘4

r
I

(l)

when used as a carbon source.

(3

The cyanide-tolerant microorganisrs were i olate;
from the trickling filter and i entified according to
the procedures outlined in Bergey's "Hanual of Deter-

V"
’ O

'._J

ninetive Bacterio o

02

A
R)
\I

J)

I

P},

T
‘1

INTRODUCTIC

Cyanides are by-products of many industrial processes,
particularly those associated with the metal industry.
Cyanide wastes of varying concentrations result from the
purification of gold and silver rem their ores, in the
manufacture of water gas, and in electroplating pnocesses.
These wastes are generally discharged into city sewage
systers or directly into streams and lakes.

The cyanides in industrial wastes, when introduce
into streams and lakes, nay destroy microbial and fish
populations. When introduced into sewage plants, the
cyanides may affect adversely the biological activity in
digesteas and activated sludge systews.

Because the cyanides are poisonous, they must be re-
moved or detoxified before being dumped into a water or
sewage system. The rethods of resoving cyanides from
industrial wastes include the following: oxidation with
chlorine or permanganate, removal of cyanide as the free
acid, conversion of cyanide to thiocyanate or cyanhydrin
compounds, hydrolysis to ammonia and forsate, precipitation
of gyanide as the insoluble iron conplex, and electrolysis
of the waste liquor. The chemical methods involved are
both exoensive and tire consuming, and in addition, require

space for the collection of the cyanide-containing wastes.

A less expensive method might be the use of a trickling

\J

(3)

filter syste: w th utilization of the cyanifie by micro-

organisms.
The éiscovery of a sicroorganism which will utilize

raniie as an energy source was the ultimate goal of this

(4)

'i

The tolerance of

W

azsis 1 c3ns ni. e

.1.

+ ~ ,5.
U C‘1tltraa 1“ u... 3

way depen7 upon various factors, inclufing conce Mt us
of cyanifle, the nediun usei, pH, growth factors avail bl
anfl the corp lex nature of the cell. A siZjle n-flium

may be use to test the cy'i‘..ni<7e
0:3 ll although other: requi

The majority of ricnoorganisms

‘.
119311.111

\rv

grown on itfle

altlou

’ "r’l

uents ions, , a

literature

(3).

n-ea“

conditions The

concerning ium require ents

tol sconce.

sta .tezents inv

The general

of: icroo gsnisss are scattered

arasvanh. In orfie
l A

A

+‘A a
UL;

a.

k.

"l“‘CC‘: nisr will be con

he order of thei-

Rae and “a0 (13) founi ths?

Nitrosoronas were inhibited by

O
snirle

16) found that calciur cy

C
bl.

5—}

oil by Yitroscronas n:

Cozic
ras

relatively cyaniie

without

-tolerance of some Licro—

re s>ecial nutrient

founi nay be

special nutrient constit-

kl':

few require anaerobic

is lacking in infornation

associate“ with cyanide

olving cyanifie—toleéance
a-i could not be covered

r to “pcfluce continuity,

:ifiered infivifiually and

aSIlflcaticn.

 

t both ID} ”was cte~ ans.
C3HL 1.3 e. Tam and lark

inr ib it ed nitrification
Uitrobacter.

 

to cysni~e concentrations

J.
U

(7) found tha Acetobaotep

-toles:nt, however no

stotenent of the ie3ree of tolerance was given.

The gram-nocitive cocci a tear to be the poet cyanide-
toleront “ionoorganicne although the" are not founi in
ebun once in :eJ13e (8). Cenrenter (6) foun: that at :-ep-
tocccci tole*et ed 5,000 ppm of cycnifie and Heroine lutee
wee cyanide tolerant but to a lee: r degree. The act-
ivity of virulent cocci was not inhibited by cyanide
while the ac tivity of evi"ulent cocci was decreeoefl (7).
‘lcelireies fe;elie was founi to be inhibited by
low concentration: of cyenide (lfi).

Burnet founfi that Evchericherie coli (5) resisted
1,000 ppm of cyaniie which agreed with the results obtained
by the e thor. Bessy (9) renorted that g. 33;; was
killed in six hours by 100,000 n-n of s Fiur cyanide.
Shirelle tolerated low co.centr2tions of cyanide while

l_'ore‘ i +v‘hooc was cor“let ely inhibit 1(5).

The spores of Bacillus subtil 3 are nore resistant
to cyanivie thin ere the veeetetive cells. The spores
survived five days in 1,000 pin of cyanide whereas the
vegetative cells eurvivefi only in concentrations of 100
ppm or 1 es (9).

The rereinin~ ricroorgeniszs will be considered as
one group eni incluced in this group ere roL::: , fungi,

algae. Axon: the eirae, Nitelle is cyanide-tolerant

:‘J
:3
Q J
I

( I

(l ) and Chl orella on SUSS” free nenia is not inhibitei

C)

1 - f '0 A ‘0 fl .
oy cyanide (9). holds appear to oe more res is tent to

cyanide than any other organism. Holds resist ten

('5')

percent so iun cyaniie for rcre than L4 hours (16).

" . a" . M n a
coil lunii Wm-

by the cyaniie-

J

H

Literature citing the po.eibility of sodium cyaniie
as an energy souece for vicsoorganisne could not be founl
although cyoni e reluction wws notei in many cas s but

was not attributei to a direct utilization of the cyanifie

(7)

31:21? 32:12:. P2100 EDUI-‘E

In onfier to develope a ricrobisl flora that will
utilize yen {We as a sounce of energy, it was necess ry
to fievise a thicrling-filter oppe*etus where the filtrate

q

COUll be reci‘culet 3 to buil

IIJ

up the nic;“ooie l nopule tion
on the filter substne te. A laboratory trickling filter
was constructei with proportions sixilan to those of a

pilot plant filter loce Ute: on the site of the East Lansing

U]
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{D
{3"
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Sewage Disposal Plant. The hoFel we
with a three inch diureter. The settlinr tank and feed

u

1 had five an: one liter capacities respectively.
A roller corpnessei a length of nubbe? tubing and force;
the liquii fror the feed ten; to the filter at a rate

of 30 rillili ten

(,0

per ninute sni a recycle ratio of five
to one. The Pate of flow was controlled by the rubber
tubing on? b? a screw clsrp Jhich ce3ul ates the flow

of the liquid to the purp. Figure l is a schenatic flow
new of the system. Figures 2 ani 3 are View: of the

overall Cyttef an? the putp fiesnectiveiy. The l-

The sewage fed to the filter was obtainei fTCJ t
effluent of the pnirery settling basin in the East Lansing
Plant. The ioxentic senage, ffiee frov in'u thiil wastes,

4.

7 _ ,7 _ ~ V a a :: 1
often containen géoun- garters which Lfl; not settl i.

*SCDdF-‘P'TJ

 

 

   
  
  

 

Settling

Tong

 

A

 

——J¢—_.—_.

Feed
Tank

 

 

A

Over-Flow

 

 

FIGURE 1

 

 

 

 

  
 

 

SCHEYETIC FLOW DIAGRAM

(9)

 

JL

Settled

DGI'TEL 9'9
V

 

 

 

$15-7- .317 2
L JL‘.t——l

 

 

FIGURE 3

 

(10)

The filte' was put into Opehition mi h the dOLeatio
sewage effluent flue an fiTiition of ten part: p r million
of soiiux cvanide. A"e: a five-week adaption period,
the CVLHile egocentrhticn 1:3 raise; first to 25 ppm

' ‘\ " :‘fi: ‘-\ ~ v‘ ~O"'\»\'_ ‘- V r: at " LIX. " ‘
on! then no ,u ‘3... here lo .'..'..52 .;C-..J.1'1uo.lnGtL UG‘CU £101.21";

‘- - ~ -, ,'-\ ~ .- -. ,‘ - . v . .1 ‘-— .-—‘ .fi. J. ‘
ueote Jene pebftfi e . u_ hie: Lene t; en iuily: at tie

n I- .o 3 ,i 4.4. v. ~. .0 . .4. £..- 4.
OUtLCu oi tge ceuuilfl: tan“ (efil‘equ); at a Junction

} ’1
('1-
O
C

(-1
,5.
S+
(1)
*4)
F10
FJ
C;

- 9 o -\ y _0 ‘ 1 ‘
c! 1 1yn ’5‘ l‘ - .-. qt . n A n-\ r‘ _- r1 (1 V
vor-a‘.-L-L... Vv.r\ '4.-.“ ye Id 1.1....Vl.‘ ‘.“‘"“ nC U ULIL.’ .—
m‘r‘unfie 63"“‘w'inrt- an "no +n"‘-e-
.h--Vl. .51.“, dwv.. .Iu-v xiv-«U

oxygen ierdnd (BOO), nitrites, eni nitcotes (phenol-

n -A - ‘.,_ :3 \ ’3 . n - a ,.. ‘: .., , V
lionie iet on) (1;. Lioctte; uni nit ites neée

P-
C‘.

deteniinefi colo'inetrically e floying a Co_e;an, model

33'
Ho
C"
' 5
H.
D
'3
:3
*4.
CT
[‘3
cf-
0)
7
d
(D
J
' b
O
2’
D
' 3
O
O
O
3
H
D’
‘2
c.
0
cf.
6

phoeedure outline? in gtandard Le'hoTs (
concentrations wefie ietecrine; bx histillation of the
The two cc: cn gethohe of aete':inin: the oonoentrnticn:
Mo c iiotnv “itnntion with
silver nitc te with potassium ioii;e a: tne iniicato?

was U"efi for the Qet e*‘in.uion of cyani e. The enijoint

Q A "V -9 I L1 ' v- v. .‘I p \
in 103 cc ncent;1t tn; aCS rather ”33L but Wluh expe" ence,

V

the enlooint was tore eviient. The procefiune was taken
foo: 1 Hook pebfonref at Yale University (15). Qual-
itative tests weue nonrefrei for the onesence of ferro-

cyanife, fecricyaniie, en’ thicoyanate as possible cyaniie
conversion pfioducts. The qualitative tests ”enformed were
taken fron a qualitative analysis renual (ll). The
streex-nlate rethoi usin: Nutrient Agar ($1fco), Dextrose
Agar (Difco), and a :ofifiei Bckofney's srntnetic agar
as Trowth meiia were exployei as a neon: of isolating
the microongmnioxs fron the filter. The plates NGPG
incubatei at room temperature. The ricroonganiors isolated
were iTentifie; accoriing to iescription: outline? in
Be"gey's Lanuel g; jetefirinotive Bacterioloix (3).
CynniGe-tolerence stulies uece perfODne; using soiium
cyaniie in various concentrations as a soucce of carbon
anfi nitrogen. A :efiiuw containing both carbon and a

J- u - v r .’ .» ,- 1-
ni'oogen source 113 use; as a conchol.

Those hicnoonganiors which wouli utilize soiium

'1

cyani e as a fOle sounce of nitrogen weée then stufie“

(

In,

to letennine the :ayiflity of cyani e ne’uction. ine
uicuoorganicms were ~epeate§ly inoculatei into synthetic
broth with cyanife as the nitrogen source. Those LiCDO-
organicxs which ouevivei were then inoculated into a
nutben of flasks containing th synthetic LQllUK with
cyanife. A Fistillation every six houns was perfornei to

fetertine the a cunt of cyonife utilize: by the th?O-

(12)

Stujiee wene else ngie on the effect that different
Finds of reflia woull have on the rnowth of a nicnoorganism

.:
v

in the presence of cyanifie.

“jfiifl'fl?’71 ' "-1 'T"" T ‘5 T '\ f“ ‘7' '. T ‘7" ' f’. l:.'_" V T.
13.1“ “3...... ‘ .-..'_ ....\. II "11...) ’ " ' I ..I ‘wF :- .

H~J~ a..-“ IA—-'-~— ‘

H

Graph 1 renresents the reaction of the lice,o"""rib.s
to the shock of crani e a: LGQL uoei by the cyaniie re—
duction ani nitrite pcoiuction when the concentration
of sofiiu: cyanife was cairei to 56 ppm in the filter.

The initial reaction of the ricroorgani rs to the hi.?. h
concentnation of cyanife had a stiwulatory effect that
reduce; the cyaniie concentration and elevatei the nitrite

pfoiuction. In the three-hour perioi following the aid-
ition of cyanife, the cyanifie refuction p:u act ica lly

ceased with a COF‘e‘“on_i 3drop in the nitrite pro-
uction. The nicroor3anicns unfierwent a "low a aption

periol in the final stages, until the reiuction of cyaniie

J

vat nearly ccxplete a 6 the pic uction of nitrites

:5

extrerely large. In the final analysis for cyanine and
nitrite, the microorganisms reiuced the oianide con-
cent'ation to one ppr ant were p o ucin3 ahproxigately
5,000 ppm of nitrite measurei as nitrite nitrogen. The
nicroorgani rs KU‘t reiuce the cyanife to pro uce nitr te
while the nitrate concentration re ainc relatively con-
:tant. As reportei by Pettet enfl Thomas (12), nitrate
proiuction is inhibitefl by high cyaniie concentrations

which wouli account for the builfl-up of the nitrites in

the effluent

(l4)

0.\

3d

00!

a;

or“

can

a:

mhdb HM CH. mflfl H.

is

 

 

—'——0

 

  

 

 

*3 s: 02 \x .2 as. e.-
\ /
. soapodé 4 ceflcmqno .
I; /
/ \ /
. / z
x x 1‘ N .\ x.
z \ /
o / \
mmfibfil I l./ \

z \

141.4.1‘2.
H HHA: . .L

HUS--DHH ho HQHuobflow 1“ .FHD cum... mm ham.

0 rmo HuHmo.,_.H~.,

 

 

 

 

—~.

.0\

as

v‘.

.3

ks

a.

 

mi .4
tr an:

2mm

(15)

n

I.)

,..
A-.

The OD,

sentefl in Graoh 2,

K
_..

tainin: cyanifie anfl

filter, bota of ”tic
toxicity of tEe cyan
evage no fin3 throu

ezsure of rfic

C

I.)

i‘coia activity, i

" r‘ lfiL
lkkh

which c. the BCD of sewage

the 303 of the effluent frcn b
h ”are greatl lowefiel due to the
lie on": he cln rificatioa of ”the

:h the filter. The inhibitory action

4.- I F. U f‘ '1 . - ~ 1- v ‘ J- ‘1. , 1‘ I _- I
of the ovar* e ”at ev1.enc Jaen t.e ave: 3e 30) values
J_ . -- _ . o J- I r " ‘ v~ I (’1
o caiiew from the fil eh containing cyani e were coxpasea
~v'L‘ “\ ‘ 1 +1 .3 9— 1+ . JI' ,- L w - .‘
«lofl cuo.e of a concrol lilce* file out cyani e. lhe

average 303 of the re w :eta3e nlus cvini e was hr; the
average B0? of the raw ceva3e w thout ycni:e was 101.
The atter value was obtair e ffior the 103 of tre East

('.
,Je

. vr‘ ~
.IK.’->’~
‘J

3'3

v

re ‘e 111

1

-u*ition of cyaniie

lr1:-

“'31.
1 LA: 'Iir

rio“oor3an in the

to cyanide; therefore,

or3anisre ani lower-e~ the 303. “he ~03 of he effluent
f?Cl the filter was lowered, not only by the pfie ence of
cyanife, but also by th cla‘ifiC'tion action of the ilter
CUth“Tt , 'hic;, so te- ”ith a zoo:leal vat. of LlCZC-
.c3ani~1s, actel ’9 the cl“ ifyin? agent uitr in the

llte”. She clanifiCCticn on re ova of organi c Latt- r
fro* the senate "culF teni to lower the EU). The

average BOD of the effluent ffio; the filter conta nin3
cyanifie was 9.8; a control-filter effluent without cvoniie
avera3e? arproxirately 2C, the latter fi3ure ‘avin3 been
taken fro: a sinilar Liece of work ealoc' by 3.0.

madam Cw osflm

d. ‘3 3

 

mme.om voappmm

 

Nwfimfiw

iiflqjfifiwiaz. .3 {#1 4. 3.. 1.1. .3“! .11 1.1.... .4
. . UL foyer. n. ncitm

PL. pr!» r FrHrL}... LEPL .r rvr

 

 

(l7)

(l), :tu. I sewi3e is

h
(J

an inoculu: of :icro-

1 ‘ f ‘- v _-t o J. 4. 3’
e--e to one 30: tilucion wave: as

. r~ . .V 7'0 ‘A ‘ V ' ‘ " . v I. ‘
eb3eniere. ii the new vessge was nub titut . Ly on

1 w. .t'b W, 3. .001 A. .1. ,n 4.2.. -0 1 4. .. L. .2 3,
inoculu“ iro. the eilluen. Ci one liltej, the inter-

ference to the 303 by cyeni e wouli be Lactially

’ .3 1- ? A. L. -'\ J " 1- ' ,
eli-inaoel by one Léeeence of c‘.ni e—Uole'ant :ic'o-
~“ H. '1 .- I v 0 ~‘< ‘A . L t" .L g -‘ . A L '1 :‘ . ‘- ‘ O_ n J-
C geni-;3 Wiupln once inoculu . Th BC; ciluoion mater
. .. ,.‘ " - J-' 1 H 7 . J- m-S -
was tee.ed x on tno e cyuni e-tolercnc .icroo:3ani-rc;

cnfi, with li ited results, the 303 of t
pluc cyanife we eixilcn to the Rid of the 16: o .33
The r croon33nisrs re;ucci or criiizei the cvehiie
in the fil We to other focme which were fetennine: by
quclitctive tests. The effluent iii not contain thie—
crtrete, ferricyenife, or ferrocycnide but iii contain
cofiur cyanifie, niteet " nitnit es, anl engonia. The
a pnoiuction

I . ' IV v - ’7‘ f‘l .V L ‘l r‘ A: .
1107CO?78n1 Ls cectfoy i one cycni-e Jitn

v

hen con1ilex cyanohyicin

(:3
(1'
O‘
(l
C+
:3:
(D
I J
J
{‘3
ci-
e
c!-

of 313319
coriounis.
The microorganicxe isolatefi an? iientifiei as
cyaniSe-tolerent are oe-o-ibe on; near: in Apten ix I.

The nicnoocganisxe which wene isolatei w he teetei
for t‘: 3e coility of «Zing, :oiiun cyaniie as a source

-' -.

of canbon an? nitrogen. The xicncon3eniznc woulu not uee

coiiui cveLifie as a carbon source. Ez3che ' checie coli,

(18)

a)

l

 

Achoorobacter

0%.“

‘1 L4...

A orobecter fielrtcvae utilized

 

sole source of nitro: en. Table l oho

thetic neiiun containing sodium

concentration: as a nitrogen un
concentrations of cvana_e of ECG ppt

reducel in the lower conc

ay have been Gue to the higher conce

as a source. Jutnient

nitrogen

solium

pnoducei

booth which was

cyanide a: a

we that a syn—

cyaniie in varying

growth in

phencnenon

ntzetion of c; aniie

“\I

use;

a control nefliur enfi contains? the care concentration:
of cvorife as the :yithetic neiiuw, iii not proiuce
grout in tLe 200 ppm concent-3tion 3 of cyaniie.

:e nicnoonganizr

fers of synthetic Ketiun

eguoieo

24

source of nitrogen wehe then

was re-ucei foo: 16.6 pom to l pom by
Celicctulun. AchPOKCboctec ielrervee

In 30 hours

 

 

9 ppm to 2.25
presentefi in Graph

The grow h of

(19)

in the

the

Q

ieterninc
hours the cyanide
1 pp: by

the cyanile

 

Achrorobacter
reiucefi tho cyanide

Lie refiuction is

pre;enoe of

m '1": 0 Kit 1’1 11‘ e \: iu r: {Zing}.

,?
s-J

wece gluco e

I?

   

'l

l‘

‘8,

 

fi’)" .Tv
(.7 - ;Pf1

J

"“ TTZAgn‘Pr-‘r P77. «,rwr‘Ivrw pupa“ «an A 1 y Iw-s
U;I.—..-.L —Q-.L‘- -1 K.‘ 1\ I an, V....'AL,.. H24 :3. - A. _.C '4

'4

"IT‘FQT" :rr cw

‘.. MH¢I

 
  
  
  

9...Achrorobacten ielicotul

lO...Achrorobacter ielnarvae

 

6 ll.

(20)

5...Alcnli:enel metalcdligenee

U"
l I
o..

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Jo

I \X‘k‘k I

\L

l‘kl‘k
l\k

I
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I *
xxx \I‘\L.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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PkrkW‘hF/ pw I. ,1 A ”TV . rILku

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(21)

IO

41‘ DJ [\J l-‘ \vi

f0

L‘l

*\ *\ fix
‘\ ‘k. *\

Tryptoce Broth

,1 /

‘x
*x *x
‘\

Tryptose Broth

All Reactive

n 7-31- '-
1191.13 3

f ‘ - -_ A - ‘ . o
CLAJIJE ADAPEATTbI

.. *V¢I

PPX
NaCN 500 550 600 650

\N h) H
‘N \ ‘|\
‘k ‘K I
"\ \ I
‘K \ I

Nutrient Broth

PP' '

UaCfi ICC 200 36C 400 1C0
Org. 89 89 99 89 89

Time

‘\
.0 fl
-thyt)

I
\\\\

— ,1

\\\

\‘K\\\

(24)

‘K \‘k\

\\\

k)!

Egj

C)

I--'

{'9'
.91
F-I

P.)

F9504. o o o o
T~TTJ ('
;.;-4)2 01‘.

lucoze...

Watev.....

h

m‘fi‘rr‘
.I
--L-J

: *H'r-r :p’fi
A

I
. ‘ .
'1.L-- _

[A
I“
x

(25)

5

. . . 9 Git‘cr 3
0 V (
. . .CU GI‘CVK. ‘

.. l G

'I1’NV
-Lvln

. .30 GH‘SIS

@000 Killilite?e

2"”? "VT-H.331” ‘rv-I-.'T Tao
V

’14-‘05;an

-'-'a—l.'.l.aL .L.

broth, tryntoee booth, ani nutrient broth. The 110?0-
0?ginifi"f grew begt in gluco:e bnoth with try: toe e b“oth
the sec on} be€:t ye iu: anfi nutrient broth last. With
the knowleige that tryptose broth eni nutrient broth
contain no ougfic as e ihbcthnete SOU?08, the glucose

in glucose booth ray ail in the photection of the miceo-

organicre from the inhibit cnv effect t8 of ovaliii e.

(D
.r':
(‘3
:1
(D

Tables 2 an} 3 sunjort the glove steterents. To
tfible: else pee eLt Late showing the adaptation of a

ricroorganiex to high coneen itretione of soiiuy cm! niie.
Cver a per iori of three tenths, the Li"“"ortenicii were

transfene? to higher eni higher cone entnations ‘1 sofiium

cvaniae. FlevobecteriUi aquatile was eventuell y W..e otei

{'4

to 550 ppm of cyaniie. After a period of three tenth
refit, this afaptaticn to the h :h concentration of

cycniee wee lost (See table 1, -ioico“”8niou 3 ).

(26)

CITCLUeICNQ

(2) Nitrates rcintcinei a constant level eni
enoeorefi e have a definite correlation with cyaniie

in hijh concentcntions.

Ill

(3) The pboiuction of nitnites i: ccnrelatei with

(

.14.

etion of the Kicroorgnnicrs to the cyaniie con—

(f
7')"
(D
I

C

oentnition in the filter. The nicrcorjnnigne will not
onoouce nitfiitee in high quintitiee until this adant-
etion he: occune .

— (4) Cyanifle in concentrdtions of 10 to 56 ppm
interfeés with the BCD of the new :ewage anl of the
effluent.

V!
O.

0.)

'. O . , -.
.nt LlC?COPgfinlJLS

f-’

(5) The afliition of cyaniie-to
to the ECU witch Score: of the intenfenence of cyeriie
on the COD.

(6) Qualita‘ive tests seen to verify the belief

thit cyanide is conventel to nitrites, nitrates, in;

"'H a .. :0 '1 ,.. ,., . J. A - , 4..
(7) lne pre_ence oi gluco e in CwLDiufleiyn “ion
. .. . 0'. f . r yr ,-' x ‘9' 1 .i. . ,—‘ (—- ‘L‘fi A
so LU. on? i_e in a no it“ it “one fovohctne

.l
‘4‘

‘ ' ' .\ I I" - 1- ‘ - - ‘n y_- —‘ .. ‘ -I— ‘
gooxtn of a :iccoosgani 3 than a nutrient neiiui without

(27)

1 1

(8) So7iuy oyenize any oe ut li ei 88 a sole soucce

N

F...

t- -. .. 4- r. v" 9 .- 1 :-
-e i ointei Liccoorqzniins, out

V

of nitrogen by core of t
not a: a oounce of ccnbon.

h.—

(9) Th :iccoorganieno can be claptei to :oiiun

.~ .3 .. .1. V”: .4. .‘ ,.. n , 1.. i .— 8 - J. , ‘ 4.
cy.ni e in hi;h concent ititn , but the e “jtqoitn .0
YA . “V ’ ‘ .. r‘ ‘ ‘ r‘ ‘ q. ,- L . 5 ~ ~
the l o CLHUOntfiwtiCP cln 08 lo 8 in a po ici of

T-rv‘srfi "1"r ~7'v 7711-“, ‘ :17: a"
O 5 -

.wu_A.-J.._...‘-\..L-

54 \J

(l) T or ine the phyaiolocieel activities involvin-
the oyanifie—tolenent TlCPCCT33nlSLS eni : Ziur cycniFe.
(2) The effect of cyaniie uncn the enzyze oyster:

of the cyanije-tclefiont :icrooroenifime.

(

1' T\ x. .. .2. L. .8 .. ~. 4- ., ~‘ . -"
(9) Jetefiine ope -eeo.;o¢ition pce_uete pcoiuoei

.L

4 -1 L fi - -. 4- v -. . ~ ”1" s . ,-.. :V N‘ u y 3 ~ - . y.- rfi ' -:‘
k" Cl'rjn... 99- oC;-3 .n 7" ‘ C -.‘CO .1; 41;" ‘- ['C'Jn in LC..1U..‘:. 03"..nl‘_.e o

OJJ . --U __'_ ._, "J

"i - L' «4- ; .. 1.1. at, 4. ' "1 .. .5
(4) eenecic itu ied on one omspteoiiiey oi any
v - v I . ‘ ' W J " ‘ . ‘
-iC‘CC?:.L1 - to LC luv c" n1;e.
'— \ '. j M an L ’ T‘ v‘ '9 - 3 L v-n -: -_ 4- ‘ r1 1;-
(3/ leeiin;3 i the QC} mate: with cyenlie-colercne

' ' r " ‘- u" ~V 'r. v‘n ' ,A g ‘- 1- r. f‘ . ‘ - - g n
pierces; n ti: “leb r1318 mon the the intesfehenee cl

7-! .. ."_ 1. "PW
coilu: oycni.e on the 3L4.

TOllUX cyani e a:

A
Ch
\1
:p
'3
c+
£3
(‘2'
F"
n
:3
0
I4)
(7"
23‘
(D
H)
}.J.
I. J
C?"
(D
' J
(t
O
m

(7) Develonrent of a nitrate tect in the pee ente

of cenbohy‘?3t 8.

(8) Study the respiration of cycniie-tclennnt
aicnoonginior: in the presence of varying concentration:
of cycni?e u in: he Wonburg technique.

(9) Developnent of a wirple test for oyani e in

the pne"ence of intenferin: icnr.

r" ‘- ~ . . . - #1. T - . a y '1.' 1- r-P~ 7‘ M
(lo) The po-'ibility oi 8.3o3115 8 ioli b0 it-iuh

O

cyanife to be use: in th- filter a: the negtructive

r\ M . L
'-.;~’enu O

(29)

LID: 1T”“E CITE?
F H . I ‘ ‘~ ’3 :- 3-

(1) nncn., gtan 33, Lccrc. 14:? cue +v" L“ ticn of
- ' ‘ '-‘ l" M I' A v ' '. '1 q “I ' :4.

Jetec an: Ser1 e , n ecic n luclic Lecltb Ac n.,
New York, rivet? 31., 1946.

(2 Bcnfon, G., 3 ~ Frieircnn, T.E., Stu1iee on
Bicl c HlC‘l Cxidcticnr. J. Biol. Cher. 13], 593-610,
1941.

(5) Bersey'c. Kenucl of Weten~inctive “cccc1iclc~v
Beltifcre: The Uillicgr en; Jillinc Ccnpany, 1948

Qixth E1.

(4) Bone, G. Acticn cf Pot1eeiur Cvcni e on Bacterial
Ccnpt. tend Lcc. Biol. 122, 96-8, 1955.

5) Burnet, F.K. The Acticn of Cyani1e upon Bacteria.

‘D

.L

filth. EQCt. 23-, 21-:4’8’ 19

6) Carpenter,
n Btreptocecci.

(

J

( C.A. The

c Biol. Bull.

(7)
the
Botcn.

( )

Cozic, T. The Influence
fezpircticn of here 1ceti
48, 212-14, 1956.

. ‘

LU

'5

Q I

c Deuc, D.G. Effect 0 J

‘0

Affect

Cvani1e unon Trickling

56.

1'4" 4'

of RespiL1tcny Inhi
fig, 227-35, 1931.

bitore

lfihr-‘fi
ULJ~+

of Pot
C B‘thelqi‘ag

Cyanide on
.::‘:eV 0 Ger} 0

UT.”

Filte‘

hicncorgenismc. A Thecic, n. .C.; 1954.

(9) wecy, G. Action of Sodium Cycni1e on Certain

Tic; OO‘”“U11C3 in .teleticn to their Place in the

finincl 03 Veget cele Kingicx. Leerirentale 78, 87-96,

1924.

(10) Ererecn, 3. he Effect of Cectcin Iaepinctory
In ibit one on the iespi ‘ticn of Ch: .03ellc. J. Gen.
Dhyciol. 10, 469-77, 1927.

(11) IcAlpine, 3.x. an1 poule, B.A. Funiamental: cf

Qualitative Chemical Analysis. LeJ 10‘7: D. Van

Noctncni Ccmncny, Seccnfi E1.

(12) Pettet, A.E.S., ani Thcrcc, 5.1. 1nalvrie of

Tniciling Fil‘e- Effluents. Inct Sewage Purif. J. and

Free. nt.2, 61-68, 1948.

30)

(13) 330, G.G. a 1 R30, H.V.S. lechanism of the
Licvobiological Cxiflat’cn cf Argonia. J. Ingian
Cher. ice. 16, 681-9C, 1939.

(14) R :9, 7. Inhibitevs of Eric ling Filter Micro-
?gvnirrz. Proc. Soc. Exp. 3101. Led. ;g, 69, 1934.
(15) Sevfnzc, E.J., Freepan, 3.3., Doflge, B.F., ani
Zabban, f. Analytical Lettofiz for tke Seterrination of
Cyaniies in Plating Wartes ani in Effluent? from
Treatrent Prcce: er. Plating 2;, 267-273, 1952.

\ ' v‘r- .- 7' .1 ,7 r a". -— f ' x v.-
(16‘ T3; 3.3. an; 0111 , H. . Ehe Actlcn of CalCiUm

’
Cyanifie as 3 $011 Disenfectant. 5011 :01. 51, 359-65,

1944.

Kicrocrgcnizm
Icrphclcgy

cheliun--- Ccnicia in chni‘z, branching,
riall elcngntei cell:

Cclcnie:--— Unite on Intrient Agcc, potato,

J

r) mm 1‘)
Lar- .,..__
v

u ,1 . ..
inn“ L.C.T.__'C

,.
I

FlcEey white on Kutrient Brcth
Phycicicgy
AG} :"Cbi. C
Ieluce: nitrates tc nitrite:
Peptonize: litruc nilk
Actively pccteclytic, liguifies gelatin
Cataluze positive
Isclote” cn ""nthetic age“ in the presence of 200

L—‘\
U

Characteristics 5 t beat those of Streptc;ypec albus,

U3
(D
(3
(D
4
)2
r‘
\,J
V
'd
(‘3
1
(D
\O
l
4:-
o

O
5
1:3
73‘
O
H
O
3
’4

Mctile

’1 -, V-“ N .+ "‘
Ufa; -ne» 3‘. give

PhyClClOSV
Aenobic

,~ .3. .- ° .1. .-
Tit;:te, re_uce. to nitricec

p:

9—1

Mt u: nil: $011
M1

(9

tin not liquifiei

Lictoce ferientei to aci: an; 3;;

H '1 v . ‘ 4. n 0 r“ _‘ .

UJUCC e fergeit : to coin enn 532

EC profucef characteeietic colonies
Isolate: fro? the effluent of the filter containing
upto 5 ppm of cyaniie.

Chccecte i1t ice fit be: t tho: :e ofE1e'iche ia coll,

Bergey'e (3) page 445.

.0

n H 7
unionism ,

Kornhology

-I
,-

Rofs, long an; occucing sing-

.._J
(.4

Kotile

Gran-negative

Yellow pigrent not water soluble
Physiology

Aerobic

filtrate: not fieiuoei to nitcitee

Iitrjurs mill: alkaline

Gelatin liquifiel

No @011 on gas from lacto:e or glucose

.. . .. ,. A . 4:- 1, .0" . y... .2 .3 ‘
Ilolctel frog the effluent oi tne lilte: containiig

J

J
(J
(a
fl
0

«J.
P)
O
(i'
(D
'4

H.
(.O
1

CLauacteWisticc be:t lit tn: e 0; ii-

ggpitile, Bergey'e (3) p130 428.
Iicrocrginifx 4

Icnobology

-'«' .. 11 -. '1 - ,_ -. '1
ions, stall, liencer, occurring singly

cohol

{J
[.4

Re? pi"fent not soluble in watec or
Physiologt

Aerobic

Nitrite: proiuce; fno: nitrct :

Litru: milb alkaline

1? gym: from carbobyfcate medium
Izclfitei for: new a wage containing SC on: of

O

cycni7e. Chcctcterist c: fit be;t tho e of Be i111?

A
\M
b

2

m
(A
.p
$-

‘Y ' ‘. I". q 1 ‘ " ‘ ' '-
. Cicliinuc, Berqey e

B

1
(3
O
’4.
(D
O

Hicnoorganicfi 5

Iorpbology
Rods, occurring cingl
Eon-motile
Gran-negative

Phgciology
Aerobic
Kitrites not pro ucei fcon nitrate;
Litmus r111 unchangei op alfoilne

No action on corbCthrate neiium

(:34)

 

./ ’ V
1 .- ,. .,— z - ..- f .
-_iC~1Ck/.Lflg~ n; ‘all. 0’ ’ \_4¢.& & 9
1-- -.1
--C 73110 4.07/1“;
.- \JU

-_ .2 . 1
occurring singly

noii nuciucel in glucoce anl l

’40

.1
.L

quifiei

Felicctulur,

..
4n... - N is . O
-..A.ULCI _' AoLur: U

r.

v‘. :3- .4-
no -, occucin:

J - Vi'd

‘ J- .2
n . ._
.--. :4 iv 3

11‘.

id ictile

.oetive

‘1

1““ ‘ \.\ :g "Cv 'raw"- 1". I .1, ‘ 1 A ‘
‘LD._; " ".-.V.~oU fik-fi-u L3"; VJ

V

- .-
A

Physiologj
Aerobic

1T 1-- .1. -.

fill .n. 'J -

pt‘casu

R Q

J. .., ‘ ‘. , y- .. .. o
Lie u; l i. @011 inn

.1.

”I: "‘
;,’. ‘..- L

in glucoee

(:5)

. m I -
- g, Decrev U
-4 U
l
’
I—r
actose

.- 1 w
11 alconcl

 

Q‘J‘ - ,0 O f
JGl.uifl liguiiie;
‘- . ..-,J..1 .°-,..:---, .. fl ‘M I.
-.G_‘.‘ 11.1.? U1 .6 ti ‘1). , 2e1~ey
.’. ‘ .a\ . .11 ‘o‘
“lC?CC?:Mnch lO
9- V
novpnololy
V
v :r‘ ‘1 ‘.,' - . .2 a 1 r M
-L,.c, :r;.1t, occnxéhlnr {finialy
-‘~r ,4 .4. 9"
“Cn-nOulle
~->- rat 1‘
enig-negxoive
1311 "r.“ 7 r917
*‘QJ «'J-chud
7;; «fl 5 r1
.I—V- L'ID-h|\l
1W 1.._ ,_. - .'.‘ 1 .0 0.. ., J-
-itfiite :umaixna. iTCL nitiet

 

Iitru: rilk reiucei, ccii
Lci? fecr dlucc1e fin licto e
-elatin not liquifitl
Ache Cb“cte? Fel FWVFe, Becgey'e
Licnoo1gcniir ll

_ O o ‘..
_pher :, occucfinc in pcc etc
’1‘- - v -v 1&1
econ :tcin varioie

. ‘-
flu

Yellow pigne not
Phyeiology

Aerobic

Nitnitee not prolucei from 1
Litm s nil? achline
Gelatin slowly liquifiel
Yon .. e1n *1 ~~.v'c ( ‘) c~
in cin love, ne aeJ , , pn,e

 

of

"““t
-U-L.

288

O

422

four anl move

e"
L.

Lionoocrdnicm l4

7- "155,- 1 M
AAC~ -dL-LC..-CI\~13r

r)
{‘I
V
(W
(3
C.)
C
J
Q.
.
’
11
I").
’
(I
F...)
t:

Lotile

Gran—ne3etive

Yellow pigrent nct water soluble
Phy:iolo:y

Aeccbic

‘T 4.

a u r r n. p . - n
i crite_ not pro ucem incl nitrc‘e-

*4
0
a

L
(J

u w o- w _ , - . w
nci: 13o- glucose uni

 

!

Bergey 9

Characteristic: fit best tho:e of P eu’o1cnce neviucifc,

 

 

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31293 02504 9937