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■■OlimON TKT OMIT
(ANSI ond ISO Tf ST CHAUT No. 2)
1.0
I.I
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■ \-
8 GEORGE V
SESSIONAL PAPER No. 38a
A. 1818
REPORT
EXAMINATION OF AFFECTED SALMON FROM
MIRAMICHI HATCHERY POND, N.B.
By 7. C. HABBISON, D.8c., 7.S.8.C., etc., Principal of Xaodoukld Collefe,
Ste. Anne de Bellevne, P.O.
(With one Half-tone Plate.)
OTTAWA
J. DE L.ABRCXJUER1E TACHft
PRINTER TO THE KINO'S MOST EXCELLENT MAJESTY
mi
2836S— lo
• QCORQE V
6EMI0NAL PAPER No. 38a
A. 191t
VIII.
IXAMniAnOM OP AFFICTBD 8ALM0V. MIBAMICHI HATCHMT
nw BBmrswicx.
KeUininn Pond at the Miramiehi Hatchery, .South K.k N B
ported that n disease had broken out amongst the «alm.,n ''"'"^'"•/f'^n'ly ^
180
DUfAwrMtyr or thk .v4V4t kk^vics
• OEONQE V. A. Iflt
until th« «p«wiiinc time i-oidm amuiid. It happnnMi that the Su|ii-riiit«ndnit
of Fiaheritt w«» in the Maritime Provinmii when thi» iiifnniuitioii wa* reMired,
and I had him iiiitruoted tu vinit the poml and look into the matter.
There were on Tucaday of thi« week lomcwhat over 2,400 Mimon in *ha
pond, between 300 and 400 of which were affected. The dineaae taken the form
of a fungn*. The fint indication ia the rerooTal of the iiralea from the back
of the neck. They are evidently eaten off. Then a white funRUit d«>veloi», which
rapidly tpreada down the head to the eye* and raakea the fish Mind. It tub-
aequently appear* on dkff eat part* of the body and on the estremitie* of the
fina and tail. The fiah di*ea*ed were betrinninir to die, which indicate* that they
will not hwt more than a week or ten day* after they become affected.
An examination of the pond revealed no reason fur any unhealthful con-
dition*. Neither did there teem to be anythi.iR through which the water wa*
ricwing before it reachol the pond to cauHo it to be unhealthful. Some fish that
were in the towing pontoon* which had recently been taken from the fishermen's
nets tu be placed in the pond, were examined, and on a few of them the first
stage of the disease nbove r««ferred to was in evidence.
As it seemed possible that the scale* might have been removed from the fish
striking the top of the pontoons, one of the fishermen's nets was visited and
w''.en lifted there were three salmon and a grilse in it. Two of the salmon were
large fe '>ale* weighing about fifteen pounds, and they were perfectly Wealthy,
but the X lird. a small male weighing S or 6 pounds, was apparently affected, as
the scales were eaten away from th^ back of the head and he had an unhealthy
app« irance.
It would appear from the above that an epidemic ha* broken out amongst
the fish in the river, and in view of the importance of the matter it is deairable
that a capable bacteriologist should be immediately sent to the pond to thoroughly
investigate the whole matter. I may add that this pond has beer in operation
for many year* and in no instance in the pest has any such trouble been experi-
enced. The tide enters the pond, and at each high tide the water is slightly
b^ckith.
I shall Ik> obliged if you will give the matter immediate consideration and
wire me whether the Biological Board can at once arrange to send a properly
qualified man to investigate the matter. If it cannot, it may be possible for
the Department to arrange with that of Agriculture to send an officer from the
laboratory at the Experimental Farm here.
N.B. — Since writing the above a report ha* just been received from the
oflicer in chai^fe of the Port Arthur hatchery, in which he state* that a disease,
apparently of a similar nature, has broken out amongst salmon trout in the
Nipigon river. This is the first time that the department has heard of any such
disease there.
A few days later I received a statement from Dr. Huntsman, the main points of
which are contained in his report on this outbreak of salmon disease, now being
published.
On the arrival of tlio si)ecimens of fish sent by Dr. Iluntsman, tliey were inimiHli-
ately examined. They arrived in good condition, packed in ice, and w; le ■ -med in
the usual way. After examination of the organs and the flesh near tlie 'ed spots
or where the fungjs was growing, pieces of the various organs we.e excised with a
sterile knife, and cut open with a second sterile knife, and a portion of the pulp, etc.,
of the organ removed by mnins of a sterile platinum loop. In a few cases pieces of
the organs were taken out, seized with the forceps and scorched in the flame, and then
cut open with a sterile knife and u portion iemoved to sterile i)etri dishes. In all
cases the m.-iteriai was mixiMl with beef peptone salt-water agar, and from the various
.ttrsvTku sAmux, uiK.tunm ii4rvMBKy
w
SESatONAL PAPER No. 3to
IS£J: * ' """^ """"""^ ""•• '"""•* "'"'""' •" »»•-* •»•""""«« »'■" '•"•^' ""j
.kin «*,!I.„^";J" /T'o"'* ""™''' ,*'*'• ""' •"'•'P«i"" "* • ^-w |-iteh« of di^„^
AfUr thp plate* hnd not thry werp kept at SOT. Reniilt* :—
Milt.— About 60 polonim.
Liver.— About 1(H) coloniett.
Swimming Madd^r.-ConUined a r,uiintit.T of li,|uid. V..ry lartro i.i. ibcr
of o,.lnnie«. loo numerou* to count.
v.T.v''n'!ilar "'"^•~^'^"* **•> «''°"'«* <° <»'«' "«•• All th«^ c^lonie. were
A'W/(»'#/t.— About 00 coluiiim.
Four gpeciM wer.- inoUted from thi>. finh. nmrked Al. A2, A.J, A4
i-lesh near di«eatc(i okiii normal in appearancs-.
fun^i^.i;;:;'r \:nn:art :;:::!:i;t '•>:r::;'!:«;;::r;.:f ^
eolou,. .o^ewhat friable, intestine, empty, caeca empiy Jgbt o" rv ti, '-'l "
InnL^T'^:^ n.uch darker in colour. almoriivc;X"r^ l'" F^h 'normll
nnd „oo<l clour. Same te<.hni.,uc. One oe.c from each of ,hc part, nuntion.l
A iH^rn."^''* ****:!. *'""' '""' ''""'*"^ ^«« '^ "' -»"" <•<•'"■'"'« developed.
A I8rg^.r number from the one cru^hinl atg from the dark red left ovar-
Ltver.~2i} colonies.
//far<-. Woorf.-eO colonic, per oe-e. all practically identical
Isolations Bl, B2. B3. B4.
disJr^'vilT/i?.*'£' l!-'^"''^ "t """""'*"»' ♦ «'^«'»i"" "f " ^ew small areas
aiscoioured jisible in the »kin. Hesh normal in appearunc... Interior orwi.> annar-
ently normal. Smears from the rarious organs showV^ laeteria. ^^
Heart's blood.-About 250 colonics to the oese, all similar.
^ffi7«.— Innumerable colonies. Two s|)ecies.
Ltver. — '20 — .30 colonies per oese.
Kidnet/n.—HO—uiO colonies.
Two isolations — CI, C2.
.hi ^«"^^' ■*:7'^,^*'?* ^^'- '""'■•' gelatinous slime around the tail. .'■ areas of
coLst^"^ "^"' *?" '"Tl- ^'"^ ^"""''^^ "W^-'-d -«lth.v. testing. sHtlv
rr> '^'^'r'"* ^••'"^•'r'- -'"P'y- •'^"'""s from the h...rf. bloo.l liver „lTd"ev
showed a number of organisms:— Kiune>
//car** t/oorf.— 30— to colonies, all similar.
Lirer.—io—li, all similar.
Kidneys.— i^\ colonics, all similar.
«:ff9*.-About 1.50 p,.r CKK. This is an estinwte, us ,. larpo ^muth had
occurred in the vicinity of the crushed part of the egg.
One isolation. Dl.
188
DEPARTMByT OF THE VAVAl SBItrWE
8 GEORGE V, A. 1918
FiBH No. 5. — Skin between the eyes and the back of the head was bruised and in
pla:cea dirty white in colour. Microscopical preparations showed the presence of
fungus. Fleah nomud. All organs normal. Intestines empty. Smears from the
Lillt, liver, heart's blood showed a number of organisms. Plates : —
Hearfa hlood. — Numerous colonies.
Liver.— 40 — 50 colonies.
MUt. — ^A few colonies.
Three isolations— £1, £2, £3.
Fish No. 6. — Skin bruised between eyes, fungus present in this area. Flesh
normal. Orgai^s normal in appearance. Intestines empty. £gg8, salmon pink in
colour. Intestines slightly congested Smears from heart's blood, liver and egg
showed bacteria present Plptes: —
Hearfa blood. — ^About 80 colonies, all similar.
Liver. — 30—40 colonies, all similar.
Esg»- — One egg about 200 colonies., all similar.
One isolation, Fl.
Fbh No. 7. — ^A large amount of diseased skin from which preparations of the
fungus were prepared. Flesh normal. Intestines empty. Organs apparently healthy.
Kidneyt. — About 30 colonies, all similar.
Liver. — About 50 colonies, all similar.
Heart's blood. — 30 — M colonies, all similar.
One isolation, Gl.
Fish No. 8. — Large amount of diseased skin from which fungus growth was
easily demonstrated. Liver pale in colour. Ovary deep reddish. Intestines empty.
If any whitish eggs in ovary. Spleen normal. Plates: —
Egg. — ^About 150 colonies to the egg, large masses of bacterial growth near
the crushed portion.
Liver. — ^About 250 colonies.
Heart's blood. — ^About 150 colonies, all similar.
A number of diseased portions of skin were cut off and examined in a variety of
ways. Very good prepartions were obtained by teasing portions of the diseased skin,
triturating the material with 40 per cent potassium hydrate. After removal from this
reagent they were washed in water and transferred to Lugol solution, or else (staiiied
with safranin, eosin, or fluorescin, dehydrated and mounted in balsam. Such teased
partii'loa of the skin gave, as a rule, better results than sections.
These preparations show that the fungus was a Saprolegnia, and I presume that
full particulars of this fungus have been already given by Dr. Huntsman. A very
full account of the salmon disease probably caused by Saprolegnia is given in the report
of the TTnited States Commissioner of Fisheries for 1878, the artii-lo having been
reproduced from the proceedings of the Royal Society of Edinburgh, written by A. B.
Stirling, of the Anatomical Museum of the University of Dublin. A very conipre-
hcnsive paper by S. Walpole and Prof. T. H. Huxley entitled "Disease among the
Salmon of many Rivers in England and Wales" appears in the bulletin of the United
States Fish Commission, vol. 1, 1881, and was a reprint of a pamphlet contained in the
"21at Annual Report of the Inspector of Fisheries for Eiigluiid and Wales for the
year 1881 presented to both Hou:*es of Parliament by oonimand of Her Majnsty."
It seemed peculiar that injuries, which apiH-nrod «t first to 1k' mere abrasions, and
vhich subsequently bei^ame infoctid by the fungus Saprolegnia. should have such
AFFECTED SALUOX. MIBAMWBI HATCBERT ^fg
SESSIONAL PAPER No. 38a
tw^T?!!? ^^* uiK.,. the fish as to produce slnggi.hneHs and death in the short
penod of time mentioned by the officer of the hatchery and by Dr. Huntsman, and it
tnerefore seemed important to make a thorough examination of the diseased fish to
«!J!;. •* ^^ ^^' '"^" producing disease, and to ascertain if the funttus
wrrh,.™*"*liT * P"?^ />' » secondary invader. Unfortunately such investigation
was hampered by the fact that no live salmon wer« avaiUble for inoculation, and the
L T„^?». ''*«"^"'»'« the pathogenicity of the organisms isolated was to attempt
to infect the common gold fish.
n.»5^"l **!? course of this examination I obtained a publication of the Fishery
l^«8^ f""'?'""' entitled "The Life-history of Salmon in Fresh water. Glasgow^
Jw,-^'' T^i ' '"'^' ^J- J^^"*"" Patterson. Assistrnt Bacteriologist of the
Corporation of Glasgow, on "The Cause of Salmon Diseases", and I am indebted to
live* '*irfish ""'*'"'"^* '•'•'''' "'^'■^ subsequently used for the inoculation of the
A f»?^K™ ^^^ '^^^ fish could be inoculated it was necessary to work out in some
detail the various organisms which were isolated from the salmon. The principal
biological and cultural characteristics of those were as follows —
A. 1.
«„H tnTf-""^ "."^^ '»«*'>»»» 7'**» '"""•l^d «nd«. occasionally bent, which occurs singly
Gelatine Plates:—
H hours, colonies just visible to the naked eye.
shaoif *rrt'r'rir ^ f"°- •> •*'""?*"' """''^' ^^^^ » "quefying centre saucer-
shaped. Centre of the colony dense with a mass of deposited bacteria.
within 'fi' "'T**''^ t'jff "f the colony seemed slightly fimbriate, and the mass
within the centre might be seen moving.
«;„,;f.,*'''*' '"'^°'''*' ^"t ^^"^ *° ^^"""^ ^ »"«> » ™«»- in diameter, but with
similar appearance to that at 48 hours.
-J A»ff», geletine completely liquefied.
Gelatine Stick:—
Growth is beat at the top. Line of puncture filiform.
mra.1i depth ^''*"^^'"'*'°° '^*'"'' ^^tending to the .sides of tube and about 2
witl/fir"' f''^«'*J» ""if"'™- line of Puncture a cloudy area 10 mm. in diameter
with small outgrowths into gelatine forming a cloudy cylinder. At the surface
liquefaction is stratiform to a depth of 4 mm. sunace
of Tlfr^"' .'h ^T**? '"" 'TT"^^- «f«tified liquefaction extended to a depth
of 7 mm. and tlie cloudy area looks like a saccate cylinder.
8 days, liquefaction to a depth of 8 mm.
10 days, there is a distinct dark stratum underneath the liquefied area
IS daijs. very slight increase.
Beef Peptone Agar, iS hours: —
Colonies 1 - 2 nnn. diameter, round, raised, entire ed^o. listening white
SXrwith^^'nati £:'Zr^ ^^ ^ -^'^ ^'^^^ <>-. and
whit^ft Srtln ir^n^'"^' -°""'' '""'^ ™-^'- -"^ •^--' — '
1M
DSPAttTHmfT or TBE NAVAL BBKVIOM
8 GEORGE V, A. 19t«
Bhped Beef Peptone Agar, Blood Heat (S7.5):—
Litde change after three cbtys' growth.
The organism grew fairly well at blood heat
g4 hours, spread over about half the sloped surface.
48 hours, growth denser, spreading, flat, glistening, smooth, semi-opaque,
whiteish. No further change.
Olueoae Agar Slope: —
i-k hours, at room temperature, smooth, vigorous, whitish, moist and spread-
ing. Cloudiness near the growth.
6 days, colony more cloudy, considerable gas production and the column of
agar is burst aiMrt in the middle.
Olucose Agar Stick: — 24 hours. Growth vigorous over surface and pronounced
cloudiness from the surface to a depth of 10 mm.
48 hours. Increase in growth and a few gas bubbles appear on the line of
puncture.
No further change occurs.
n»ef Broth:—
B4 hours, strong, cloudy.
3 days, much heavier. Sediment flocculent.
7 days, yellowish-green appearance in the upper layer otherwise no change.
JJunhams's Solution: —
The organism grew well in Dimham's solution, and at the end of 5 days at
room temperature was tested with Ehrlich test, allowed to stand 20 minutes and
the results then recorded. This organism was negative to this test. No Indol.
MUk:—
24 hours, no change.
3 days, coagulated with extrusion of slight amount of whey.
5 days, curd has become firmer, and a cheesy smell developed.
7 days, slightly more whey extruded;
No other change, although observed for some twenty days.
Litmus MUk: —
24 hours, no change.
4fi hours, no change in constituency, but colour is changed to avellaneous.*
5 days. Colour uniform, slight digestion with separated whey, soft curd,
yellowish ring around glass, smell disagreeable.
3 weeks. — Curd still undigested, whey yellowish, yellow ring, curd avellaueus,
few gas bubbles un shaking.
Potato : —
24 hours. Moderate, dry, slightly raised, cream-coloured growth.
4S hours Increase of growth, dry, raised, slightly rugose, cream-yellow colour.
6 days. Abimdantly raised, massive, rugose growth, cream colour at margins and
pinkish on top. Odour unpleasant and slightly pungent, resembling that on
milk.
3 weeks. No change.
A. 2,
Small bacillus with rounded ends, short, often in pairs, actively motile, stains well
with methylene blue, and is gram negative.
I Chroniotaxia <eu NomencUilor Colorti-m. P. A. Saccardo.
AFfBCTED SALMOtf, MtBAMICBI BATOBBBT
1H
SESSIONAL PAPER No. 38a
Gelatine Plate»: —
«4 hours. Colonies just visible to the naked eye.
Z^^^'rll! '"'"' ""''"T^ " "'^ °* 2-3 mm. in diameter; round, saucer-
area around. W.th 3 objective interior of the colony is glumose Edges
clearer, but less distinct than A. 1, ' « "'"o"- ^oRes
« d«*.. In moderately seeded plates there is complete liquefaction.
Oelatine Stick: —
** ^aZ;, ifjT'^f ""'JT- u'"" "* P"""*"'^ " ^•'"••''y "«» « ""»• 'n <li'«nete'
3 Z. in depth." " ^'"' '" ^* *"""'• "'*"'"^"« *° «'*^^^ "^ *"»^ ""<*
^ ^toTdelthTfl !!!Jf™T-^ 'I''* fimilar appearance. a„d stratified liquefaction
to a depth of 5 mm. Liquefaction gradually increases.
6 S' ii„ul i^ f "'^ ^l"^ remainder of the tube saccate liquefies,
o aays. L,iquefaction to a depth of 4 cm
10 days. Liquefaction of the gelatine in the tube complete.
Beef Peptone Agar: —
^ *tr.".! ^^ temperature. Colonies 1-2 mm. in diameter, raised, glistening
WUhloW^tivV^"'*^ ""' ^^"'^'^ "P"'-**"* by transmifteJTight'.
all around "^ ' "'"*'* *"""^" "'^^ " "'«" ^^''^^^ "^^^
^ ^Tnd ^^1°"'*'*' '"'''^'"^ I" "'*^ """^ '""'^ •" *^«°^"y- Colony becoming whiter
and more convex, somewhat resembling a yeast colony.
Beef Peptone Agar, at S7°C.:—
Very slight growth at 24 hours, after which there was no further growth.
Olucose Agar Slope: —
"/r^lthtTthTe'^or' """"* ^"' ^"''' ^'•^^ -^-^ "«" "—
Olucose Agar Stick: —
S ?""'■*• ^'"T**!..^"*"™ "" «"'*'*«*'' *•>'" ""d spreading. Not character!, c
48 hours. Gas bubble on surface and below- Afterward^ no further chnge.
Beef Broth:—
H hours strong clouding, which increases, with abundant sediment,
•^o further change.
Dunham's Solution: —
^ *r;^'j;?r* ^^^f'f^^' '^^.^^ "Jt^ EhrHch-s reagents; allowed to st„nd
for 20 minutes and then recorded. No Indol.
MUk.—
*4 hours. No change.
5 ^ys. Coagulated with extrusion of slight amount of whey.
6 days. Curd becomes firmer, and cheesy smell develops.
^"Ta*n^. ^^^^ '""^''^^ "P t«> wventh day. after which there is no further
las
DBPAKTMBST OF TBE HAVAh BgSYtOK
8 QEORQE V, A. 1918
Litmut Milk: —
a houra. Ko change.
iS houn. No change iu colour or consistency; on shaking numerous small gaa
bubUea appear and form a foam on surface.
6 day: Coagulated, moderately firm curd, liliacinus in colour. About a quarter
of the tube is whey, and much darker in colour (atro-Tiolaoeus).
5 weekt. There is a reddish ring at the surface, considerable digestion, whey
occupying three-quarters of the tube, isabellinus in colour. Curd flocculent,
aTcllaneus; oduur slightly cheesy.
Potato:—
e^ hou.a. Growth moderate, filiform, slightly raised, cream-yellow colour. This
increases, and in
6 day* growth is moderate, raiaed, rugose, moist, shiny; dirty cream-yellow,
darker in centre where growth is most massive.
S weekt. No further change.
A. 8.
Medium-size bacillus with rounded ends, resembles A. 1 in appearance. Active
motile^ stains well with methylene blue, and is gram n^ative.
Gelatine Plate*:—
H houn. Just visible to the naked eye. Growth rapid.
is houn. Colonies are 2-6 mm. in diameter, round. Liquefaction saucer-shaped,
inner ring dense, caused by deposited bacilli. With i objective thd edges of
the coI<mies are fimbriate centre grumose and flocculent. Masses of the
bacteria can be seen in movement.
S days. Colonies increase to 12 mm. in diameter, SRU'jer-shaped liquefaction,
whitish in centre, more transparent at the margin. To the naked eye the
edges are entire, but with a microscope slightly fimbriate. There is a cheesy
smell on opening the plates.
4 days. Plates are liquefied.
Gelatine Stick: —
SJ^ hours. Resembles A. 1, but slightly less growth.
48 hours. Line of bacteria is filiform, smooth on surface. Liquefaction strati-
form, 4 mm. deep. Liquefaction continues.
10 days. Licuefaction is 1 cm. deep with medium beneath darker in colour, but
clear.
Agar Plates: —
48 hours. Colonies are 1-3 mm. in diameter, round, raised, yellowish-white. With
} objective edges are entire, dark in centre, granular, gradually becoming
lighter to margin, which is clear.
3 days. Colonies are round, white, edges entire, brownish in centre. Convex.
J, dayii. No change.
Agar f>lope, 37°C.:—
2Jt hours. Very slight growth, filiform.
7 Jayg. Nu further change.
Olucose Agar Slope, 20" . —
A spreading, flat, white, shiny growth; agar beneath very cloudy. Cream yellow.
No gas.
ttsr
Cloudy
ArracTKD SALitoy. visamicbi hatch kkt
SESSIONAL PAPER No. 38a
Olueote Agar Stick:—
Betf Broth:—
H hours. Clouding moderate. Sediment.
S day». Qrowtb heavier, slight pellicle.
S days. Ring and pellicle.
7 days. Yellowi(/h-green colour in upper layers.
Subsequently no change.
Dunham's Solution: —
^"ra/d'sS m,wL'* 7T t«»P«Jt"'«. tested with Ehrlich test, allowed to
stand 20 minutes and then recorded. No Indol.
ililk:—
^^'IS*' 1,^" '*!T' "J?*" *^« fifth d«y. when there is coagulation with soft
curd, cheesy odor Curd gradually becomes harder and the whey greenish
m colour. Dig«,tion takes place to about half the volume.
Litmus Milk: —
The colour is gradually bleached and in iS hours is avellaneus.
!•• ?°'*S,'*'°° *^* P^*^ ^° '^ *" « '^'^'' » *"**' fi''* «"«i which gradually
Agests^ Blue rmg at the top; separated whey is isabellinus in colour.
S weeks. Greenish-blue colour; whey thick, curd avellaneus, odour unpleasant.
Potato:—
«4 hours. Growft moderate, raised, filiform, cream-yellow in colour.
T' , ^"''^ 'f»"'«' <l'rty «nd ochraceus slightly rugose. Growt* gradu-
ally changes to ferrugmeus in colour.
3 weeks. No change
A.4.
A. 2. Actively motile, stains well with methylene blue, and is gram negative.
Oelatine Plates : —
3Jt hours. Just visible to the naked eye
iS fco«r». Colonies punetiform (less than 1 mm.) white and gli.tenhig. with S
objective they are seen to be round, with entire edges, and granular.
3 days. Colonies slightly punetiform, white, plietening, oonvex. oupitat.. With
* objective edges entire and granular.
No further change.
Gelatine Stick: —
H hours. Growth unifrom, line of bacteria filiform.
fAaT\^'""'-^^ *"^°™ ^ .''""""• '•■""^ ^''^ '>""'»'■- "■' 'i»o <'f bacte.ia
M 2 \r'L'!,"°'" ?■""''■'*'• ^'"" "^ ^'"''"'« ^•'"°"'' to Papillate.
faciiof '^^P'***""' «t the point of puncture may be noticed, but no lique-
13 days. . Liquified area around the line of piuictury.
1M
DEPARTMByT OF TBE ItATAL BKKVtOB
S QEOROE V, A. 191S
Agar Plate*: —
i8 \our». CoIoniM are filiform, glistening, r»i«ed. With I objectiv.) the coloniee
are round, dense in centre, and granular, clearer at margin, edge* entire.
8 dayi. Coloniee slightly larger, opalescent, white.
No further change.
Oelatine Agar Slope at ST C—
Little, if any, growth observed. Continuous observation for 7 days.
Olucose Agar Slope: —
Growth moderate, moist, shiny, slightly raised, whitish.
3 weeks. Agar ie brown beneath the slope.
Olucose Agar Stick: —
Growth filiform, thin surface, growth spreading. Gas bubbles along line of punc-
ture.
No further change except the agar becomes brown beneath the surface to a depth
of 1-2 cm.
Beef Broth:—
H hours. Slight clouding and sediment.
S days. Clouding and sediment increase slightly.
No further change.
Dunham's Solution:
Grown for five days at room temperature, tested with Ehrlich test, allowed to stand
20 minutes and then recorded. Indol positive.
Milk:—
5 days. No change visible.
6 7 days. On shaking tube a gassy foam rises to the surface.
10 days. Milk had coagulated, hard curd, whitish whey.
Litmus Milk: —
No change in appearance in H hours.
iS hours. Abundant gas which rises to the surface in small bubbles. This was
noticed each day up to the sixth day, and the foam was very heavy. The milk
gradually coagulates and forms a blue ring down one side of the tube, remainder
is a firm curd adhering to the tube. Bleached cream colour.
Potafo:—
g^ hours. Moderate growth, filiform, slightly moist, cream coloured.
J)8 hours. Becomes slightly rugose.
days. Growth slight, slightly raised, and a dirty yellc (mellcus).
S weeks. No further change.
B. 1.
This organism on examination was found to resemble in all respects A. 1,
B. 2.
A small size bacillus about IJ times as long as wide, rounded end, frequently in
pairs. Actively motile, stains well with methylene blue, negative with gram.
ArraoTMo salmon, mimamicbi batobsht
IN
SESSIONAL PAPER No. 38a
Gelatine Platei:—
84 hours. Visible to the naked eye.
iSAoun. Punctiform. colony raised, glistening, whitish, i objective shows ronnd
dense, granular colony, entire edges. oojecuve snows round.
No further change.
Oelatine Stick:—
U hour». Growth uniform, round, filiform; no liquefaction.
f ^our». Growth uniform, no liquefaction to surface
3 day,. Slight depression at the boint of bacteria. No liquefaction.
Agar plates: —
48 hours. Uniform, 1 m.m. in diameter, round, glistening colonv Win. a „kj~.»
.ve round, dense, shading to lighter; granular, S'e^^- '"' ' '•''''^'•
3 days. Colonies are glistening and bluish white.
No further change.
Agar slope 37° <. ■ —
' Tt ol\TjS^;*oS!^.""* °' ^^ ^"•"" -'-- -^^--« - the surface
Olucose Agar Slope: —
Moderate growth, spreading, flat, moist and whitish.
flZ7' A "^ T ''"''"^ "PP*" ""«^ «"«»»' ""^"^"^e in growth.
3 weeks. Agar is brown underneath the slope. B^wui.
Glucose Agar Stick: —
No'Sh.?'*?* "'""^^ on surface, ga.s bubbles along line of puncture.
No further change except for browning of the agar underneath the Surface.
Beef broth: —
Si hours. Moderate growth, moderate sediment.
a days. Growth slightly heavier.
6 days. Clearing.
No further change.
Dunham's Solution: —
Grown for five days at room temperature, tested with Ehrlich test allowed ia
stand 20 minutes and then recorded. Indol positive. ^"
Litmus Milk: —
24. hours. No change.
ooa£atio7' ^ *"' ^°"" "" '^' ™'^"'' ''^"" '"^^ •* '^''^'''- C°l°« ««aceous, no
6 day,. Much gas in foam form. , No coagulation. Colour liliaceous Colour
dSestS ■ '""'' ""' '"'"^ °" *"^"^^- ^'-''^ wZy but littTe
Potato:—
24 hours. Filliform, dry, raised, colour niveus.
e days. Growth becomes slightly raised and more massive
S week.'). Xo change.
B. 3
Resembles in all respects A . 4.
190
DBrAMTUKST Of TBM IfAVAL gMMTtOt
• OEOROe V, A. 1»1t
B. 4.
BeMmUet in all respeota A. 4-
0. 1.
Small to medium bacillua about twice aa long as broad, riightly rounded endi.
ActiTely motile, aUina somewhat unevenly with methylene blue, gram negative.
Gelatine Plaiea:—
£1 houri. Colonies visible to the naked eye. , ■ • •
4« hours. Colonics punctiform, round, white, raised and glistening. 3 objective
round, evenly dense and granular with entire edges.
No further change.
Gelatine Stick: —
Growth uniform, line of puncture filiform, 4 gas bubbles along line of puncture.
10 day*. Depression at the point of puncture.
IS dayt. Line of bacteria has liquefied.
Agar Plate*: —
iS hour». Coloniea are punctiform, 1-1 J mm. in diameter, round, raised, white,
glistening.
With 3 objective colonies are round, dense in centre, clear margins, granular,
entiip ledfres.
No further change.
Agar. 37' C.:—
U hours. Moderate growth, flat, slightly spreading, smooth and translucent.
No further change.
Glucose Agar Slope: —
Flat, moist, spreading, whitish growth, few gas bubbles.
No further change except browning of the agar beneath surface.
Glucose Agar Stick: —
2i hours. Filiform, growth spreading on surface.
^ hows. Few gas bubbles along line of puncture.
No further change.
Beef Broth .—
21t hours. Moderate clouding, floeculent, abundant sediment.
5 days. Clearing.
No further change.
Dunham's Solution: —
Grown for five days at room temperature, tested with Ehrlich test, allowed to
stand 20 minutes and then recorded. Indol positive.
Litmus Milk: —
21f hours. No change.
i8 hours. Slight amount of gas, colour somewhat lighter, no coagulation.
6 days. Much gas in foam form. No. ooagvlatio... Colour lili.iceous.
' Subsequently milk coagulates, blue ring, surface clear, whey on one side, curd
adhering to two-thirds of the tube; bleached to a cream colour and of firm
consistency.
ArracrtD tALnox, niRAMicm batcbmt ^
MMIONAL PAPER No. iU
Potato:—
V^^f ^^*^; moderate growth, cream-yrflow.
a wetks. No further change.
D. 1.
.hh ^fthZl'wut!;!::'. ^Z^r' '""•"'•^ ^"•'- -'^^^ -»"- -ai„, wen
Gelatine Plate$:—
Colonie. Tijible to the naked eye in U hour,.
'' *r.e tlfl-^iril^'^"'*^"^- -'-- ' "^-"- round, granular.
x/T" . ^"'o"'** '^t^rae mow dense. Convex
Ao further change.
Gelatine Stick:—
Ap'ir Plates: —
'' X.]^r.:-itri^ - - -- ts;
3 days. Slight increase in sizes; otherwise no change.
Agar slope, 37° C: —
No growth at this temperature. ~
Olucoae Agar Slope:—
"""'tJ::; fr^ThaS::"' "'-"'• ^^^ '— •^-^ ^-th the growth.
Otucose Agar Stick: —
Line of puncture Jiliforni. spreading on snrf.mn tv,,^ r
nppcar in 48 hours and slight Zr.Z i Sowth otL™ " " T^^ '"'^'"''^
browning under growth. ' "*''<"™'«'' "" <-hange except
Beef Broth :—
2i hours, iirowth moderate, sediment moderate and ilocoulent
o days. ( leuriiig.
No further change.
Dunham's Solution: —
Crown for five days at room temi)erature, tested with Fhrlieh t^t „ii a x
stand 20 minutes and then recordcl. Indol positive. ' """''''* *°
Litmus Milk: —
'' 'Tie surfaT""' ^■'""'^' '"' "" '""^'"^ *^^ '^"^ -«" «^- bubbles rise
IS hourx. Gag more pronounced. Colour liliaceous
6 days Foamy gas. No coagulum. Colour liliac^us.
S weelcs. Blue ring on surface cleared away alonjr one side rpmo.-„J„. «
adhering to the tube. Bleached cream colour ■"•'■nainder firm curd
m DBPAMTMmiT OW TMM VAJAL aUWWM
• OIONOE V. A. lilt
PeMo:—
ModerBte growth, raii«d, ruftoM, waxy, crwiin yellow in mbur.
E. 1., E. 3. «nd E. S.
Rnemble A. 1.
F. I.
Medium iixe bacillus with rounded end«. Actirdy motile.
On ataining with methylene blue there are two or three dark granulea in moat
of the organiinu. Gram negative.
OelatvM plain: —
24 /io«r». .Tust visible to the naked eye. Round, white, gliatening. I ohieet-
ive brown, edge* entire, granular.
Subsequent liquefaction.
atUttint Stick.—
Growth uniform, line of puncture filiform, growth becomes slightly lioaviet
and on the <tth day there i* a slight liquefied depression.
10 day». Liquefaction is infnndibuliform.
iS dayt. Complete liquefaction.
Agar Platet: —
Agar dope, S7' : —
Very slight, if any, growth (7 days).
Gluco»e agar dope: —
Filiform, non-spreading growth.
Glucose Slick: —
Filiform growth, nothing on the surface.
e day». Slightly heavier, subsequently no change.
Beef Brolh r—
Slight clouding, flocculent sediment
3 days, clearing.
No further change.
Dunham's Solution: —
Grown for five days at room ten.; crature, tested with Ehrlich test, allowed
to stand ^ > minutes, and then recorded. Very weak Indol.
Litmus Milk : —
6 days. No change visible until 6th day, when colour becomes darker. This
increases.
6 weeks. Colour is atrocyaneus. There is progressive digestion without
coagulation.
Potato : —
Whitish growth restricted and filiform.
3 weeks. No fuHher change.
-. L
Medium size moderately thick bacillus with rounded ends, very considerable
variation as to size, actively motile, stains well with methylene blue, gram negative.
AmCTMD MALMOir, MlMAMIOMt nATOMMMt tfc
8CMIONAL PAPER N». St*
Hkoun. JitftviaiUe to the naked eye.
4* hour: Punctifonn.
« dayt. 1 . 6 mm. in diameter, round, aaut -w-ihapeu, liquefaction. Whitiab
J^r ?i!'' "*r* *"^. "f ' **"*'*• R**'**** Jin- like the q>ok.. of a whed
from the centre oonaiatinf of depotited bacteria. With I objective e<kea ara
entire and interior granular to grumoM. ^^
4 dav$. Pktea have liquefied.
Qtlatint 8Uek:—
fo™^*?"*k J'''"'?'»«*i°n •»«»»i«[. « mm. in depth. This increaie. and U strati-
form to Mcchato. In ten day* tube is completely liquefied.
Agar Platet-.—
Agar »lope. S7' : —
Very slight growth, in 24 hou-ra.
No further change.
Olueoie ag-.r $lope: —
Moist, white, spreading, smooth. Gas in condensation water
growth"""**' ^'^"'•''*"'"' ~''"" «t *••« »»«' »' tlM» 'lope, and centre of surface
Glueote Stick :—
un<Jure°™' '^'"^^^^ "Preading on surface, 3 or 4 gas bubble along line of
No further change.
Betf Broth:
H hours. Strong, cloudy, moderate sediment
S days. Pellicle over enti.3 surface.
7 das/s. Yellow-cream colour in the outer layers.
No other change.
Dunham's Solution: —
Grown for five days at room temperature, tested with Ehrlish test, allowed to stand
20 minutes, and then recorded. Indol very strong production.
Litmus Milk: —
iS hours. Colour is lighter.
i days. Alkaline digestion commences.
e days. Almost complete digestion, remaining curd, in fine particles, dirty viola-
ceous m colour. Whey i of tube. Semi-transparent and avellaneous in colour,
no odour. Blue nng at surface. '
Potato:—
H hours. No apparent change.
4S houri. Slight growth, filiform, yellowish.
6 days. Moderate growth, slightly raised, moist on the moist part of potato and
dry at the top, femiginious in colour.
S weeks. Colour is redder, otherwise no further change.
23366— 2a
DtPAarMnr or ran hatal anficB
t OtONOE V, A. 1011
H. I.
A ihort to modium •tout bMiUiu, actirely motile, •taiiw well with methylene blue,
{■ gram necative.
OeMJiMPiiiiM.—
t\ koun. Jut viiible to the naked ejre.
4 day*. Punctifonn, later liquified.
QtUiintaHeh:
Growth uniform, filiform.
4« Atfura. Slightly liquefying, 3 nun. in depth, »tratifonn. Liquefaction increaiw
and ii slightly Mcchate with flocculence.
to da)/$. Liquefaction become* infundibuliform.
16 dan*. Whole tube is liquefied.
Agmr Plate;—
iS houn. Round, uniform, glistening, '•Mony. With I objectiTe round, edge*
■lightly crote. Slightly granular coiony.
5 day*. Colony becomet more maHive and bluiih white; otherwiae no further
change.
Agar Slope, ST :—
Very slight growth, one or two ^jlonie*. Increaaea along line of puncture.
7 day*. No further change.
Olueote Agar Slope: —
Thin, translucent, moist, film in 24 hours. No further change.
Olueose Sticle: —
Filifor.n No surface growth.
Be' Vroth:—
£4 hourt. Slight clouding, fiocculent and abundant sediment.
5 dayi. Clearing.
7 days. No further change.
Dunham'* Solution: —
Grown for five days at room temperature, tested with Ebrlish test, allowed to stand
20 minutes, aud then recorded. InJol very strong.
LUmu* Milk: —
ti hour*. No ch'inge.
iS hour*. Tubes become darker in colour, Htro-violaceous. No coagulation.
Subsequently there is g-adual digestion. Whey first with a violet shade,
throughout, vhich gradually concentrates as a deep blue ring on top, and curd
becomes semi-lransparent, isabellinus in colour, and thick but not viscous. A
little undigested curd at bottom of tube. (3 weeks.)
Potato:—
U hours. Very slight growth.
i days. Growth moist alijrhtly raised, smooth. Colour brown, light testaceous.
S weeks. Colour changes somewhot between rosaceous and testaceous. No further
change.
ArnCTBD 0ALMOK, MIKAMWHI MATOMBHT
SEtSIONAL MWR N*. M»
■UMMARV or CIIARACI
CliAlM(TERwms OK TMK Ml< «..-.«.,* Ms M (Hacillu, Mlmonin pe»tis Pattewon).
Morphohgital Characters.
A *hort. .. \.k bacillu* witi, nuindod end-, vnr^ing i„ lo,.K,h. m-ourring .iugly and
oV™''" Tf,''"'^ *" *""?■« ^"*''^''^ •""*"••• n""-*l'"'i'-l>t.',ri,.K, doo« not\tain with
or nT^^-lt ; ^°7 '""l"^ ?'\f "J'^^T'-^ "* *'■*• '°""' »«>'nP<''«t"'e, hut 8how8 little
or no growth nt 37 C. and u killed at this temperature in ttk-ut mx days.
♦l..t "^ °'K"""'™ «P°s«l to » niixture of ioe and «alt for a week not only survived
that low temperature, but grew profuMy while in the mixture. Involution forms were
only observed in itlucose media. It appeurs to 1* :< Mriot aerob.-. Pathogenic to fish
non-pathogenic to frogg, mice, and gi'iaea-pig.. * '
CuHurt>*.-Room Temperatttre.
ring of liquefttction around them of a tra«..parent greyish colour, which ripidly
rXr ♦^%P;•»•^^«^•°">""' -n'P»«>tel.v li.,uefied ,„ about :$« hours after ?£
appearance. The de„.e pin-point centre and transparent a... of li.,uefactio,!
around is markedly characteristi,- of the bacillus, together with the verv r«3
liquefaction of the gelatine. • ^ "
'""t^iis:;;;::S£x^'>' """"■ '" " ^■°"" •■"■« ■"-"• "•* ""'^
CrM Othlne. .OS ,,r „nl J(«<,._Tlio »rowth i. more prot,,.,.
CarUl CMim. .OS „, ,„(, a„J._v,.j p,„f,„e j„,,i,.
DBPAHTMWST OF TBK NATAL aBRVWB
8 QEORQE V, A. 1»1t
Atmr Streak :—
Daow, nrufcae, crMunooloarad moiiit •hininv growth klong needle tnck in
18 noun, with irregular margin, whicn gradually
iipreada over th«
Growth barely viiible.
•urface of the agar.
Af/ar Smear .—
Small pin-jioint oream-oloured oolonie* at the end of 18 houm with iirrguUr
•prMtding transparent margini.
Agmr Olueom Slab :—
Profuae oream-ouloured growth along needle track for about half an inch at
the end of 24 houra, •preading on the rarfaoe. The agar gradually
beeowtf elo'idji /torn the eurfiue and paralltl to it, and cxlemde for about
kalfnpinoh down tke media. Ko gat prodaetion.
Agar OlM-ote PlUe :-
Crvamcolnured colonie* with moiat ehtning anrfaoe and white eloiidineu
around taeh Colon]),
Blood Serum :—
Bouillon: —
At the end of 18 hours the bouillon become* cloudy throughout, with a
marked skim on the surface and clinging to side* uf tube, with a slijht
depoeit at the bottom.
Bouillon iulueoee) :—
Similar to ordinary bouillon, but growth much more profuae,
BouilUm Tauraehdate Ulucote : ~
8l«ht growth, turnini, the media slightly red. No gas formation
Litmue Milk :—
In about 48 hours them is a distinct acid reaction, which gradually increases,
and in about seven days the milk become* coagulated and gradually
diitcsted.
Peptone Water. -
Markml cloudinea* throughout at the end of 18 hours. Give* no mdol
reaction.
Potato:- , J
Very profuse yellowiah brown growth at the end of 18 hours, raised on the
surface of media like blisters, with moist shining surface.
Agar {Anaerohinallil) . —
No growth
Ko iierceptibli! gniwtli.
Very slight growth.
No growth.
No [lerceptible changt*.
Very slight clondinnw at
the end of 48 liouf.
Givexnoindnl n-actlcm.
Vnry sliglit growth in 48
houTa.
No growth.
The organism also withstands the effect of ordinary water, sterile water and sea-
water for a considerable time, as flasks of those inoculated with it and kept at the room
temperature for over a month gave profuse growths when reinoculated on agar. It
does not, however, survive more than a week in distilled water. It also ki-eps well on
sub-cultures, as tubes of agt inoculated from sub-cultures about a year nld gave pro-
fuse growths in about 18 hours.
I ! '' The chief rharacterintirn of the bacillus are those: —
Actively motile, non-spore-bearing bacillus.
On .-ub-cultiire it grows profusely in IS hours at the room tompornture.
On sub-culture it grows profusely when exposed to deg. C. for a week.
Shows little w no firoyth at 37° C.
■ • Is killed at 37° C (98-6° F.) in about sl.t days.
Liquefies gelatine with extreme rapidity,
('oagulates and digests milk.
Forms a cloiidinexs in flhirosf ai/ar in tlie uoighhourhood of the urowth.
(Irows well in sea water.
Strict aerobe.
Involution forms only obeprve<l fin glucose media.
Does not stain with Gram's method.
Pathogenic to fish.
Non-puthiigenic to frogs, mice, and guinea-pigs.
'
ArracTBD baluok, uiramwbi batobmt ^„
SESSIONAL PAPER No. 38a
CONCLUSIONS.
fJ! li^ ^?'"^» .^J^^^^ania ferax h not the cauHe of the salmon disease.
^.•«..?^- ^*" ^'"!?,'"' *"''" '""""'^ ^'""K*' abrasion or ulceration of the skin, and the
r4rZSr"? not contracted when the skin of the fish is in a healthy t^te
dea/Sh^ntrsirter'" """ "^ *'""'""**^ ^'^^ ''-'» ^-"-^ «-H to other
(6) The fact that the bacillus grows profusely when placed in a freenne mixture
t^^ r™::^fatu':aLn':^rt°h^ ''-'■ - ^-- ^^' showTthaTtrs
'%:^i ''r ""1 ^'^^.-^'^ ^'"^ '" "*">"» ^8 ■"><» 35 days, respectively.
not irow *'7^r/i? r** ^'1-^ *^7* *^" "' ^^ ^"t*'- "^^^^^ Saprolegnia does
not grow r a 1; therefore a d-seased salmon entering the sea, and returning to tb^
..ver apparently free from fungus, cannot be said to be free fr^m S,e Zase
OOLO-FISH EXPERIMENT.
tankYn'oie o^7r!T \"""''^'n.°/ ""^^'^'^ "''^ P"'*'^^^** ""^ placed in a large
ank m one of our laboratories. The change of water resulted in a few dying, so to
Thev rf T"" '^"^ !^ management we kept them for a month before inocu atU
They were then removed from the aquarium and two fish were placed in each of eigh
third day. and the fish fed every alternate day.
„..» Illl irJ!^**i°" was carried out in the following manner: The fish was taken
? KW .u ""J^ "'"^ *'"' *°P °^ *•>" ^"^'^ ""'1 P"'* °f °n« «i<lo near the gilk gentlv
rubbed with sandpaper until there was a slight effusion of blood, and this abraded
ZyV^"' 't*^ ^'*^ " P'^''""'" "^'^ "^ ' •"""• '^''"'^•^ ^itl" -nate""! Sen
from a 24-hour-old agar 8 ope culture. A separate piece of sandpaper was used for
.ia<-h ftsh Several loopsful of the culture were added to the water of each jar
In this way organisms Al, A2, A:{, A4, Bl. B2, Cl. Dl. El, Fl, 01 wereinocu-
K.ted in duplicate, and four fish were rubbtxl with sandpaper but not inoculated The
hsh w-cre observed daily, and the inoculated water was ohauged on tlio third day
Ihe control fish rubbed with sandpaper and not inoculated are still alive.' and
of the inoculated fish ne m each of the jars inoculated with A, A2, B2 Cl and Dl
died 22, 30, 34. 27, 4 days oftcr inoculation. , ^ i. ana ui,
Bacteriological examination was made of these fish, but in no case w„s I able
to obtain from the dead fish the organism which was inoculated. Evidently these
organisms were non-pathogenic to gold-fish. One fish in each of the jars fro.n which
the dead fish were taken remains alive, and, at the time of writing (May 10) appear
•luite normal Of course there is the possibility that some of the organisms isolate.1
niight lie pathogenic for salmon and not for gold-fish.
Patterson states with reference to his B. salmon is pestis that:—
"Dace inoculated with this bacillus died as the result of inoculation in
from two to seven days. Dace, river trout, sea trout and gold fisli inoculated
tee
OKPAKTMSST OF TBS WAVAL 8BMTI0S
8 OEORQE V, A. 191S
with Saprolegnia remained healthy. Dace, aea trout and one gold fish inocu-
lated with Saprolegnia and B. salmonis pettia died in various periods of time
(S to 18 days) except the gold fish which died after inoculation and showed signs
of the fungus on the giU covers. No attempt was made to make cultures from
the dead gold fish."
Patterson concludes that: —
" Saprolegnia grows on live
breaks down the superficial tissw
grow on."
h in the presence of the organism, which
id forms a suitable nidus for the fungus to
I had no Saprolegnia to try similar experimMits.
The difficulty of obtaining and keeping fish for experiments in a laboratory
unequipped for such work, and the difficulty because of lack of laboratory equipment
to carry out experimental work at the hatchery, will have to be overcome before any
decisive experiments can be undertaken.
It is, however, significant that all organs apparently healthy in the salmon
examined contained bacterid in large numbers, and of comparatively few species, and
I am unable to state or find in any literature or obtain information as to the bacterial
content of the normal organs of fish, or how soon after death, and to what extent, these
organs arc invaded by bacteria. Very large numbers of bacteria were found in the
eggs from a number of the fungous-infected salmon, and under normal conduions one
would scarcely expect to find so many bacteria present.
All that can be stated at present is that Patterson's organism, B. salmonis pestis,
was not found, and that the large number of bacteria present accompanying the
Saprolegnia may have some pathogenic role, but the rules of proof (Koch's postulates)
would have to be worked out where fish, the means of keeping them, and laboratory
facilities are provided.